Association of bovine papillomavirus type 2 and bracken fern with bladder cancer in cattle. The bladder cancer syndrome that often accompanies chronic enzootic hematuria in cattle grazing on pastures infested by bracken fern has been experimentally reproduced in animals fed a diet of bracken. The experimentally induced tumors were histologically and pathologically indistinguishable from the naturally occurring ones and comprised two main types: (a) carcinoma of the urothelium identical to that seen in humans; and (b) hemangioendotheliomas of the subjacent capillaries. Often the two types of tumor occurred together in the same bladder. In animals experimentally immunosuppressed with azathioprine "bracken type" hemangiomas developed in the bladder lining. DNA of bovine papillomavirus (BPV) type 2 was found in 46% (7 of 15) of the natural cancer cases and in 69% (9 of 13) of the experimentally induced lesions, independently of histological type and including the hemangiomas of the azathioprine-treated animals, suggesting a close association between BPV and bovine bladder neoplasia. Moreover, BPV-2 DNA was found in experimental animals that had not been inoculated with BPV at all or had been inoculated with a different BPV type and had been kept in isolation, suggesting that BPV can persist in a latent state and be activated when the animal is exposed to the bracken cocarcinogens and to immunosuppressants.


Modulation of c-jun and jun-B messenger RNA and inhibition of DNA synthesis by prostaglandin E2 in Syrian hamster embryo cells. Fatty acid metabolites such as prostaglandins are important regulators of DNA synthesis and cell proliferation. However, the mechanisms involved in this regulation are unclear. We have examined the effects of several fatty acid metabolites on the expression of the growth-related genes c-jun and jun-B in Syrian hamster embryo cells. Treatment of cells with prostaglandin E2 (PGE2) resulted in the inhibition of epidermal growth factor (EGF)-induced DNA synthesis and c-jun mRNA accumulation, whereas PGE2 augmented EGF-stimulated jun-B mRNA and markedly stimulated jun-B accumulation in the absence of EGF. Treatment of cells with PGE2 resulted in rapid accumulation of cyclic AMP (cAMP), whereas prostaglandin F2 alpha did not stimulate cAMP formation and did not alter EGF-stimulated DNA synthesis or accumulation of c-jun or jun-B mRNA. Forskolin and 8-(4-chlorophenylthio)-cAMP mimicked the effects of PGE2 on DNA synthesis and on the expression of c-jun and jun-B, suggesting the involvement of cAMP. We have shown that EGF-induced DNA synthesis requires the formation of hydroxyoctadecadienoic acids, formed from linoleic acid by a 15-lipoxygenase, in Syrian hamster embryo cells (Glasgow et al., J. Biol. Chem., 267: 10771-10779). Inhibition of this 15-lipoxygenase blocked EGF-dependent hydroxyoctadecadienoic acid formation and mitogenesis but did not affect EGF-stimulated c-jun or jun-B mRNA accumulation. The data suggest that the modulation of EGF-dependent DNA synthesis by PGE2 is associated with altered expression of c-jun and jun-B in Syrian hamster embryo cells. In contrast, hydroxyoctadecadienoic acids appear to act downstream or divergent from c-jun and jun-B expression in the regulation of EGF-dependent DNA synthesis.


Premature fractures of platinum-cured Silastic shunts. Experience with one unitized ventriculoperitoneal (VP) shunt system has disclosed an unacceptably high rate of premature distal shunt tube fracturing. From March, 1986, to February, 1989, 114 new or replacement unitized VP shunts were placed; the patients were followed through November, 1991. Twenty-two (19.3%) returned with peritoneal limb fractures, with a mean time from shunt implantation to fracture of 31.5 +/- 11.3 months (range 15-62 months). All shunts broke in the neck 1.1-11.2 cm below the valve. Of the 22 cases, there were no instances of infection, previous peritoneal limb revisions, or recognizable iatrogenic shunt injury at placement. These fractures also occurred earlier than the usual time expected for biodegradation of shunt tubing. The Silastic tubing in all of these shunts had been manufactured using a more recent platinum-curing process. This tubing has decreased elastance and is more brittle than tubing cured with the previously used dichlorobenzoyl peroxide (DCBP) catalyst. Our studies suggest that the premature shunt tube fractures were related to changes in physical characteristics of the platinum-cured Silastic tubing. Therefore, the DCBP-cured Silastic is to be considered preferable for shunting products. Neurosurgeons are asked to report recurring patterns of shunt system failure.


TGF beta signals through a heteromeric protein kinase receptor complex. Transforming growth factor beta (TGF beta) binds with high affinity to the type II receptor, a transmembrane protein with a cytoplasmic serine/threonine kinase domain. We show that the type II receptor requires both its kinase activity and association with another TGF beta-binding protein, the type I receptor, to signal growth inhibition and early gene responses. Receptors I and II associate as interdependent components of a heteromeric complex: receptor I requires receptor II to bind TGF beta, and receptor II requires receptor I to signal. This mode of operation points to fundamental differences between this receptor and the protein-tyrosine kinase cytokine receptors.


Expression of a membrane protease enhances presentation of endogenous antigens to MHC class I-restricted T lymphocytes. We find that expression of the membrane dipeptidyl carboxypeptidase angiotensin-converting enzyme (ACE) enhances presentation of certain endogenously synthesized peptides to major histocompatibility complex (MHC) class I-restricted cytotoxic T lymphocytes. ACE appears to function only in an intracellular secretory compartment of antigen-presenting cells. ACE-enhanced antigen presentation requires the expression of the putative antigenic peptide transporters, TAP1 and TAP2. These findings demonstrate that a protease can influence the processing of endogenously synthesized antigens and strongly suggest that longer peptides can be transported from the cytosol to a secretory compartment where trimming of antigenic peptides to the lengths preferred by MHC class I molecules can occur if the appropriate protease is present.


Torso receptor activity is regulated by a diffusible ligand produced at the extracellular terminal regions of the Drosophila egg. torso encodes a receptor tyrosine kinase (torso) required for anterior and posterior terminal development of the Drosophila embryo. Injecting eggs with in vitro synthesized torso mRNAs revealed that torso activation is governed by an extracellular molecule, presumably the torso ligand, produced at terminal regions of the egg during early embryogenesis. In the absence of torso, the ligand shows no apparent localization, indicating that it is diffusible and normally bound by an excess of torso receptor at the egg poles. Mutant ligand-binding torso proteins can suppress telson formation in a dominant negative manner, suggesting that the ligand is limited in amount. Analysis of torso mutations indicates that torso functions as a tyrosine kinase and that gain-of-function mutations causing ligand-independent activation are located in the extracellular domain.


Evidence for free radical formation during horseradish peroxidase-catalyzed N-demethylation of crystal violet. Crystal violet (gentian violet) can undergo an oxidative metabolism, catalyzed by horseradish peroxidase, resulting in formaldehyde formation. The N-demethylation reaction was strongly inhibited by reduced glutathione. Evidence for the formation of a crystal violet radical during the horseradish peroxidase catalyzed reaction was the detection of thiyl and ascorbate radicals from glutathione and ascorbate, respectively. The concentration of radicals from both compounds was significantly increased in the presence of crystal violet. Oxygen uptake was stimulated when glutathione was present in the system and this oxygen uptake was dependent on the dye and enzyme concentration. Oxygen uptake did not occur when ascorbate, instead of glutathione, was present in the system. However, when glutathione was present, ascorbate totally inhibited the glutathione-stimulated oxygen uptake in the crystal violet/horseradish peroxidase/hydrogen peroxide system. Although a weak ESR spectrum from a crystal violet-derived free radical was detected when the dye reacted with H2O2 and horseradish peroxidase, using the fast flow technique, this spectrum could not be interpreted.


A comparison of the mouse versus human aryl hydrocarbon (Ah) receptor complex: effects of proteolysis. The differences in the molecular properties of the nuclear aryl hydrocarbon (Ah) receptor from human Hep G2 and mouse Hepa 1c1c7 cells were investigated by time-dependent partial proteolysis with chymotrypsin or trypsin followed by column chromatographic and velocity sedimentation analysis. The sedimentation coefficients, Stokes radii and apparent molecular weights of the untreated human and mouse Ah receptor complexes were similar. Treatment of the nuclear Ah receptor complexes from both cell lines with chymotrypsin for 10 or 60 min gave lower molecular weight proteolytic products which also exhibited comparable molecular properties and salt gradient elution profiles from Sepharose columns linked to DNA. Treatment of the human and mouse nuclear Ah receptor complexes with trypsin (5 micrograms/mg protein) for 10 or 60 min gave a minor low molecular weight (29.7- or 25.7-kDa) proteolysis product which was detected only with the mouse Hepa 1c1c7 Ah receptor complex. The time- and concentration-dependent proteolytic digest maps of the human and mouse Ah receptor were determined using receptor preparations which were photoaffinity labeled with [125I]7-iodo-2, 3-dibromodibenzo-p-dioxin. The human Ah receptor was significantly more resistant to proteolysis by trypsin or chymotrypsin than the mouse Ah receptor. At a low concentration of chymotrypsin (1 microgram/mg protein) the Hepa 1c1c7 receptor was degraded to two lower molecular weight fragments with apparent M(r) values at 71- and 48-kDa whereas the Hep G2 Ah receptor was relatively stable under these conditions. Although the human Ah receptor was more slowly hydrolyzed than the mouse receptor by trypsin, the major photolabeled breakdown products for the Ah receptor from both cell lines were observed at M(r) 48- and 45-kDa. The results of this study demonstrate that there were subtle but significant differences in the human and mouse Ah receptor complex; however, the proteolysis studies suggest that there are common structural features in their ligand binding sites.


Observations on the structural features and characteristics of biological apatite crystals. 2. Observation on the ultrastructure of human enamel crystals. In a series of studies to investigate the structural features of biological crystals, using an electron microscope, we examined the ultrastructure of human enamel crystals at near atomic resolution through the cross and longitudinal sections of the crystals. The materials used for this study were the middle layer of the noncarious enamel from freshly extracted human erupted permanent molars. The small cubes of the enamel were fixed in glutaraldehyde and osmium tetroxide and embedded in epoxy resin using the routine methods. The ultrathin sections were cut with a diamond knife without decalcification. The sections were examined with HITACHI H-500 and H-700 types of transmission electron microscopes operated at 125-200 kV. Each crystal was observed at the initial magnification of 300,000 times and at the final magnification of 10,000,000 times and over. Using this approach, the authors have been able to show the configuration of the hydroxyapatite in the cross and longitudinal sections of the enamel crystals and observe the basic hexagonal pattern of the unit cell viewed down the c-axis. The authors sincerely believe that the electron micrograph shown in this report is the first atomic image to be obtained from a hydroxyapatite crystal from the human enamel, using the sections.


Radiotherapy in the treatment of hepatocellular carcinoma and its metastases. A study was conducted to evaluate the effect of external radiation therapy on hepatocellular carcinoma (HCC) and its metastatic lesions. A total of 33 patients with cytopathologically proven HCC were subjected to radiation therapy over a 4-year period, and treatment was discontinued in 8 cases due to jaundice, severe discomfort, or early mortality. Thus, 25 patients with 28 lesions underwent irradiation with a total dose ranging between 3000 and 5600. Of these, seven were irradiated for liver tumors, and the results showed that two lesions decreased in size, the symptoms improved in 1 case, and another patient maintained stable disease for 4 months. Among the 21 metastatic lesions treated, only 2 patients failed respond to the treatment. Nine subjects were irradiated for bone metastases, and the bone pain subsided in all but one case. The survival for bone metastasis was as long as 23 months when the primary tumor was treated effectively. Three of the four cases of irradiated skin nodules disappeared and had not recurred after 5 months, 1 year, and 4 years, respectively. Tumor shrinkage or symptoms of relief were noted for three abdominal lymph nodes, one neck lymph node, one pleural tumor, and one lung tumor. Clinical improvement associated with a stable lesion was observed in two patients with brain metastasis. Follow-up revealed regrowth of the tumor or recurrence of symptoms in most of the patients. However, none of the patients died as a direct result of a metastatic lesion. Although external radiation therapy is palliative in intent, it appears to be useful in the treatment of HCC and its metastatic lesions.


Approaches to optimal dosing of hexamethylene bisacetamide. HMBA is a potent differentiating agent capable of causing > 95% morphological differentiation in cell lines in vitro. The induction of differentiation is dependent on both the concentration of and the duration of exposure to HMBA. However, acute toxicities (neurotoxicity and acidosis) have limited the maximal HMBA css value to < 2 mM, which is at the lower limit of effective in vitro concentrations. When HMBA css values have been maintained at 1-2 mM, thrombocytopenia has limited the duration of HMBA infusion to < or = 10 days. The present studies were performed to determine whether exposure to HMBA could be individualized and maximized without resulting in intolerable toxicity to patients and to determine which factors would predispose a patient to the development of acute toxicity during treatment with HMBA. For these investigations, patients were given HMBA at a target css using an adaptive-feedback-control method rather than at a set dose. Because HMBA administration produces large anion gaps, a simple maneuver such as alkalinization might enable the escalation of plasma HMBA css values to > 2 mM. HMBA was given as a 5-day CI to 14 patients (26 courses) at 2 target HMBA css levels near the maximal tolerated value in the presence or absence of concurrent alkalinization with sodium bicarbonate. Symptomatic acidosis occurred in one patient who did not receive bicarbonate. Neurotoxicity proved to be dose-limiting at the target HMBA css value of 1.5-2.0 mM in the absence of concurrent alkalinization and at a css level of > 2 mM, regardless of alkalinization. No neurotoxicity was seen at target HMBA css values of 1.5-2.0 mM in patients who did receive concurrent alkalinization. Alkalinization was not associated with any detectable changes in plasma HMBA metabolites. With the maximal tolerable 5-day HMBA css having thus been defined at 1.5-2.0 mM, we attempted to maximize exposure to HMBA by defining a tolerable duration of infusion. Individualization of the duration of HMBA infusion to a target nadir PLT was performed in patients who had received an initial 5-day CI of HMBA at a css 1.5-2.0 mM along with concurrent alkalinization. The AUC achieved and the thrombocytopenia produced during this first course were used to predict the duration of infusion that each patient would subsequently tolerate (at an HMBA css of 1-2 mM) to achieve a nadir PLT of 75,000-100,000/microliters.(ABSTRACT TRUNCATED AT 400 WORDS)


Radiation therapy in patients with unresectable hepatocellular carcinoma. From April 1978 through December 1989, a total of 17 patients with unresectable hepatocellular carcinoma (HCC) were treated with radiation therapy alone or radiation therapy in conjunction with percutaneous ethanol injection (PEI), transarterial infusion chemotherapy (TAI), or transarterial embolization (TAE) at the National Medical Center Hospital. The median survival of all patients was 13.8 months. The survival values determined at 1, 2, and 3 years were 58.8%, 26.1%, and 9.8%, respectively. Only the pretreatment liver function affected the survival value. Between patients who did not have liver cirrhosis (LC) as well as those who had LC of Child's class A and patients who had LC of Child's class B or C, the differences observed in the 1-year survival value and the median duration of survival were statistically significant (P < 0.05). The serum cholinesterase (ChE) level seemed to be a good indicator of liver function during the radiation therapy. A field size of 150 cm2 and a total dose of 5000 cGy (TDF 82) seemed to be well tolerated by patients who did not have LC and those who had LC of Child's class A. The field size determined whether patients with poor liver function such as LC of Child's class B or C would develop severe hepatic deterioration after undergoing radiation therapy.


Effects of multimodal treatment and hyperthermia on hepatic tumors. The therapeutic results of Lp-TAE (transcatheter arterial embolization in the presence or absence of Gelfoam particles preceded by the infusion of a mixture of lipiodol and an anticancer drug via the proper hepatic artery) or DSM-TAE (transcatheter arterial embolization with degradable starch microspheres and the arterial injection of anticancer drugs via the hepatic artery) combined with hyperthermia were evaluated in 30 patients with hepatocellular carcinoma (HCC), 5 subjects with hepatic cholangiocarcinoma, and 22 patients with metastatic liver carcinoma. Hyperthermia was performed using an 8-MHz Thermotron RF-8. Tumor temperatures could be measured in 31 patients (54.4%) with malignant lesions of the liver who had undergone hyperthermia. The mean maximal temperature (Tmax) was 41.5 degrees C in the metastatic liver cancers. The efficiency of heating in HCC was unfavorable, i.e., the mean Tmax was only 40.7 degrees C. The rise in tumor temperature was greater in either HCC or metastatic liver carcinoma associated with portal invasion of the lesion. The tumor-temperature elevation was also excellent in HCC that had been subjected to embolization with DSM in combination with hyperthermia. The response rate (complete response plus partial response) was as high as 40% (4/10) in the group in which the tumor temperature could be raised to 42 degrees C or more. Among the 52 patients who had shown a high pretreatment level of tumor marker, that value decreased in 34 cases (65.4%), and the decrease was greater than 50% in 22 cases (42.3%).


Radiotherapy of cholangiocarcinoma: the roles for primary and adjuvant therapies. A total of 22 patients with cholangiocarcinoma who had been treated with external radiotherapy between 1978 and 1989 were analyzed. Of the 22 patients, 18 had cancer of the hepatic hilus (Klatskin) and 4 had intrahepatic biliary cancer; all but 2 of the subjects had advanced disease. In all, 16 patients underwent primary irradiation for unresectable tumors, 4 were subjected to adjuvant irradiation after gross tumor resection, and 2 received preoperative irradiation followed by gross tumor resection. The mean initial irradiation dose was 52.0 Gy (range, 26-78 Gy). The TDF (time-dose-fractionation) for the entire course of radiotherapy ranged from 49 to 154 (mean, 100). The median survival of all patients was 10 months, and the cumulative 1-year survival value was 37.7%. The external radiotherapy proved to be effective in the treatment of cholangiocarcinoma in terms of palliation and survival.


Significance of multidisciplinary therapy for hepatocellular carcinoma. The effect of multidisciplinary therapy for hepatocellular carcinoma (HCC) was evaluated in 121 resected cases. The 5-year survival was 100% for absolute curative resection (12 cases), 59.1% for relative curative resection (n = 37) and 10.9% for relative non-curative resection (n = 59). However, none of the patients survived for more than 3 years after absolute non-curative resection (n = 13). The non-recurrence in the preoperative TAE groups was different from that in non-TAE groups undergoing absolute and relative curative resection. The 1- and 3-year non-recurrence rates for relative non-curative resection were 92.3% and 53.8%, respectively, for the preoperative TAE group and 56.1% and 28.1%, respectively for the non-TAE group. These data show that preoperative TAE is effective in relative non-curative resection. Functional disturbances of the coagulation-fibrinolysis system in cirrhotic patients were improved after PSE. All patients undergoing hepatectomy after PSE had an uneventful postoperative course, including well-maintained function of the coagulation-fibrinolysis system and a decrease in splenic volume. At 1 year after hepatectomy, cirrhotic patients with critical liver function and poor coagulation-fibrinolysis showed appreciable hepatic regeneration. One patient died of hepatic failure 1 year after the operation. In recurrent HCC, the 1-, 2- and 3-year survival values after reresection were 100%, 75.0% and 25.0%, respectively. The respective values following TAE were 79.0%, 42.0% and 9.0%. Three cases of recurrent HCC were effectively treated, i.e., two patients achieved a partial response and one showed no change, by continuous intra-arterial infusion of 5-FU and lentinan with intermittent one-shot injections of epirubicin using a subcutaneous infusion pump. These three patients are alive at 1 year and 7 months, 1 year and 4 months and 6 months after the treatment, respectively.


Long-term survival following treatment of hepatocellular carcinoma in Singapore: evaluation of Wellferon in the prophylaxis of high-risk pre-cancerous conditions. The present paper reviews several studies performed between 1977 and 1986 in Singapore on the 10-year survival outcome of treatment for stage I and II hepatocellular carcinoma (HCC). Of 801 HCC patients evaluated, only 2 survivors (0.3%) remained in complete remission for 13 and 14 years, respectively. One had received four weekly cycles of prednisolone, Adriamycin, vincristine and 5-fluorouracil for an inoperable HCC with a 10-cm diameter, and the other had received localised synchronised hepatic irradiation and Adriamycin. As follow-up, the use of localised hepatic irradiation consisting of 131I-labeled (30 mCi) iodised oil in lipiodol infused via the hepatic artery appeared to benefit patients with small residual tumours but did not affect larger tumours measuring 2 cm in diameter. Prophylactic, intermittent long-term administration of lymphoblastoid interferon-alpha (Wellferon) was carried out in pre-cancerous, high-risk hepatitis B surface antigen (HBsAg)-positive patients with cirrhosis, in immediate male relatives of liver cancer patients, and in persons who had undergone hepatic resection. In the untreated group, 10/162 (6%) cirrhotics, 3/18 (17%) male family members, and 6/10 (60%) post-resection cases developed single or multiple HCCs within 1 year of screening done at 3-month intervals on the basis of alpha-fetoprotein (AFP) levels and real-time hepatic ultrasonography. In contrast, none of the Wellferon-treated group consisting of 518 cirrhotic patients, 82 male relatives of HCC patients and 20 post-resection cases developed HCC. Two HBsAg-positive individuals who had not been treated with interferon (IFN) developed hepatic nodules which that showed dysplasia, AFP elevation and chromosomal changes. These studies demonstrate the poor results of late diagnosis and show that early intervention and prophylaxis with Wellferon can reduce the incidence of HCC in high-risk persons. In addition, transhepatic chemoembolisation and liver resection are suitable methods for treating small HCCs (single or multiple) that are detected by screening. However, some of these early-detected HCCs remain highly malignant. Prophylactic treatment of pre-cancerous conditions appears to be a better option as a long-term programme for HCC.


Clinical aspects and epidemiology of hepatitis B and C viruses in hepatocellular carcinoma in Japan. The incidence of hepatocellular carcinoma (HCC) in Japan has increased over the past two decades. Of the 379 patients with HCC treated at Shinshu University Hospital over the past 20 years, 112 underwent treatment between 1971 and 1980 and 267 were treated between 1981 and 1990. The prevalence of hepatitis B virus-associated HCC and hepatitis C virus-associated HCC was 54% and 34%, respectively, during the first decade and 31% and 60%, respectively, during the second decade. Major factors contributing to the increased incidence of HCC include an increase in the incidence of type C chronic hepatitis and an increase in the incidence of cirrhosis of the liver, which in turn are the result of blood transfusions received about 30 years ago. Donated blood testing positive for hepatitis C virus antibody is currently rejected from the blood supply. However, the occurrence of post-transfusion hepatitis with the potential to develop into HCC has not been entirely eliminated. In addition, there is an as yet unelucidated route of horizontal transmission of hepatitis C virus.


Prospective and randomized clinical trial for the treatment of hepatocellular carcinoma--a comparison of L-TAE with Farmorubicin and L-TAE with adriamycin (second cooperative study). The Cooperative Study Group for Liver Cancer Treatment of Japan. A randomized clinical trial comparing L-TAE with Farmorubicin (FARM) and L-TAE with Adriamycin (ADR) in the treatment of hepatocellular carcinoma was conducted from October 1989 through December 1990. In all, 192 hospitals participated in this study and 117 patients were entered. The patients were randomly allocated to group A (L-TAE+FARM) or group B (L-TAE+ADR). There was no significant intergroup difference in background factors. Additional treatment consisting of repeated TAE or surgery was given to 66 patients. Four factors were analyzed in this study: the percentage of reduction in tumor size, the change in the AFP level, lipiodol accumulation, and survival. None of these factors differed significantly between the two groups. The final evaluation of this study will be based on differences in survival after a long-term follow-up. Toxic effects manifested less frequently in group A than in group B, and the decrease in the platelet count in the peripheral blood was significantly lower in group A than in group B. These results suggest that FARM exerts a more favorable effect than does ADR in the treatment of hepatocellular carcinoma.


Liver transplantation for hepatocellular carcinoma: clinical results and future aspects. The treatment of unresectable hepatocellular carcinoma (HCC) by liver transplantation remains controversial. In our series, the 5-year survival value for 87 patients who underwent transplantations between 1972 and 1990 was 19.6%. There was no difference in the long-term survival of patients who had underlying cirrhosis and those who did not. In patients with early-stage tumors the long-term prognosis was improved, the 5-year survival in stage II disease being 55.6% according to UICC criteria. Even in some cases of more advanced tumour stage, good long-term results were obtained. In a review of the recent literature, we evaluated prognostic factors to work out criteria for a more differentiated indication for liver transplantation. Resection of increased radicality--which will keep its place as the therapy of choice--and transplantation should be performed complementarily. Further developments will reveal the value of multimodal therapeutic strategies, including chemo-embolisation, chemotherapy and immunotherapy.


Treatment of small hepatocellular carcinoma. Of the 692 patients with hepatocellular carcinoma (HCC) who were admitted to our hospital between 1976 and 1990, 60 (8.8%) had small HCC with a maximal diameter of below 2 cm. The outcome of these 60 cases was analyzed after they had been divided into 4 groups based on the therapeutic method used: operation group (17 cases), percutaneous ethanol injection therapy (PEIT) group (20 cases), transcatheter arterial embolization (TAE) group (13 cases), and oral anticancer drug therapy (per os) group (10 cases). The 1-, 2-, 3-, 4-, and 5-year survival values obtained for the operation group (100%, 87.5%, 87.5%, 87.5%, and 87.5%, respectively) were significantly higher than those found for the per os group (P < 0.01). The best therapeutic results were achieved in the operation group. Although the follow-up period for the PEIT group was short, the 2-year survival of this group was nearly equal to that of the operation group. Whereas the duration of survival tended to increase in inverse proportion to the severity of the underlying liver cirrhosis, the survival values did not differ between solitary and multiple tumors or among the different histological grades of HCC. In this series, 20 patients died; 9 deaths (45.0%) were due to progressive disease and 3 deaths (15.0%) were attributed to hepatic failure. Because the operation group included many patients who displayed relatively good liver function, we cannot rule out the possibility that their excellent outcome may have been associated with this background factor. Therefore, further prospective investigation is necessary to compare the efficacy of various therapies in patient groups with a homogeneous background.


Efficacy of combination treatment--(TAE with adriamycin and ethanol)--for hepatocellular carcinoma. Among 44 patients with hepatocellular carcinoma (HCC), combination treatment with both transhepatic arterial embolization (TAE) and ethanol injection therapy (EIT) was performed in 10 patients. Only two had tumors measuring less than 3 cm in diameter. In all, eight patients had solitary tumors and two had multiple tumors. The tumor was classified as stage I in one patient, stage II in six subjects, stage III in two patients, and stage IV in one subject prior to TAE, but one stage II case was changed to stage III after laparotomy. The clinical stage was I in two patients, II in six subjects and III in two patients. Five patients with tumors of stages I and II achieved either a complete response (CR) or partial response (PR). However, three patients with tumors of stages III and IV showed progressive disease (PD). Thus, the response rate (CR+PR) was 50%. For tumor stages I and II, the 1-, 2-, and 3-year survival values were 100%, 100%, and 83%, respectively. For tumor stages III and IV, the 1- and 2-year survival values were 75% and 25%, respectively. Combination treatment of HCC appears to be efficacious for tumor stages I and II.


Results of hepatic resection and postoperative arterial chemotherapy for hepatocellular carcinoma. To improve the outcome of patients who had undergone hepatic resection for hepatocellular carcinoma (HCC), we employed postoperative adjuvant hepatic arterial infusion chemotherapy (AHAI) in 23 patients. Patients showing various risk factors for the recurrence of HCC were given one shot of doxorubicin and mitomycin C suspended in an oily medium (lipiodol) and an infusion of 5-fluorouracil. The 3-year survival value calculated for patients who were treated with AHAI was 75%, which was significantly higher than that found for patients who did not receive AHAI (n = 156; P < 0.05). In addition, among the patients who underwent hepatic lobectomy, the survival of those who received AHAI was also significantly greater than that of those who did not (n = 46; P < 0.01). AHAI did not cause any severe complications. These results indicate that AHAI may be an effective therapy for patients with HCC.


Selection of therapeutic modalities for hepatocellular carcinoma in patients with multiple hepatic lesions. In the present study, we compared the survival of patients with multi tumor hepatocellular carcinoma (HCC) following their treatment with liver resection versus TAE. A total of 336 HCC patients were treated at Osaka University Hospital between 1980 and 1989. Of these, 140 patients underwent liver resection in the presence or absence of TAE and 173 subjects were treated with TAE alone. Our TAE protocol consisted of 50 mg Adriamycin, 3-5 ml lipiodol, and Gelfoam. The 1-, 3-, and 5-year survival values found for the liver resection group were 87.4%, 66.0%, and 47.4%, respectively, whereas the values calculated for the TAE group were 64.6%, 29.9%, and 15.8%, respectively. The survival of patients in the resection group was clearly better than that of subjects in the TAE group. Of the 140 patients who underwent resection, 36 cases were proven to have multiple lesions by histopathological examination. The 1- and 3-year survival values determined for this special group were 67.9% and 33.3%, respectively. Of the TAE cases, 113 were diagnosed as having multiple lesions by imaging examination, and their 1- and 3-year survival values were 59.7% and 24.9%, respectively. No significant differences in survival was found between these two different treatment modalities for these multiple-lesion cases. The results of this study indicate that it is unlikely that surgical resection is superior to TAE alone for the treatment of HCC patients with multiple lesions.


Transcatheter hepatic arterial chemoembolization using epirubicin-lipiodol: experimental and pharmacological evaluation. The experimental and pharmacological characteristics of various formulations of an anticancer agent (epirubicin, EPI) and lipiodol were evaluated in vitro and in vivo. Three forms of EPI-lipiodol, i.e., an oil-in-water type of emulsion (O/W type), a water-in-oil type of emulsion (W/O type), and a suspension (S type), were prepared and investigated for their stability. An O/W-type emulsion using a stock solution of Iopamidol as the solvent for EPI was the most stable form in the stationary state in vitro. In 16 patients with malignant liver tumors (14 hepatocellular carcinomas and 2 liver metastases), the three forms of EPI-lipiodol were injected into the proper hepatic artery. The plasma EPI level was monitored periodically and analyzed pharmacokinetically. No significant difference in the pharmacokinetics of EPI was detected among the O/W, W/O, and S types.


Assessment of chemoembolization therapy for primary liver cancer using a stabilized adriamycin-lipiodol suspension. We formulated a new lipiodol-Adriamycin suspension (ADM/lipiodol, 50 mg/10 ml) that remained stable for 48 h (half-life, 25 +/- 3 days). In five cases of hepatocellular carcinoma (HCC) resected after intra-arterial infusion of this agent, the ADM concentration in the tumor was quite high and the tumor necrosis rate was more than 80% on histological examination. Over a 5-year period, 180 patients with unresectable HCC underwent transcatheter arterial embolization therapy (TAE) in the presence or absence of this agent. The regimens consisted of suspension injection alone (A, n = 54), suspension injection + TAE using gelatin sponge (B, n = 29), TAE followed by suspension injection (C, n = 34), and TAE alone (D, n = 63). The estimated 1-year survival values determined for patients treated with these regimens were 70%, 73%, 43%, and 39% respectively, and the corresponding 3-year survival values were 27%, 31%, 15%, and 10%. The survival achieved using suspension injection was thus superior to that obtained using conventional TAE, and combined therapy with suspension injection followed by TAE seemed to enhance survival, although there were some biases in tumor size and in the stage of tumor progression. For patients with tumors measuring 5 cm or more in diameter, the survival obtained using regimen A was lower than that achieved using regimen D, but the combination of TAE and suspension injection improved the 1-year survival value obtained using regimen D from 34% to 52%. For patients with tumors measuring less than 5 cm in diameter, the survival achieved using regimen A was markedly better than that obtained using regimen D, although no difference was found between the survival value achieved using regimen A and that obtained using regimens B and C. On the basis of these results, our newly formulated ADM-lipiodol suspension was surmised to be effective by itself against relatively small HCC tumors, whereas it enhanced the efficacy of conventional TAE in large lesions.


Comparison of the anticancer effect of ADMOS alone and ADMOS with CDDP in the treatment of hepatocellular carcinoma by intra-arterial injection. A total of 135 patients with hepatocellular carcinoma (HCC) were treated by intra-arterial injection of an Adriamycin/mitomycin C oil (lipiodol) suspension (ADMOS) alone or of ADMOS plus cis-diammine-dichloroplatinum (CDDP). In all, 59 patients were treated with ADMOS alone and 76 were treated with ADMOS plus CDDP. A reduction of more than 25% in the tumor size was obtained in 13 of 38 (34%) evaluable patients in the former group and in 39 of 76 (51%) evaluable patients in the latter group. Serum alpha-fetoprotein (AFP) levels decreased by more than 50% in 10 of 17 (59%) and 23 of 33 (70%) patients in the respective groups whose pretreatment AFP level was estimated to be over 200 ng/ml. Overall, the 1-year survival value was 68% and the 2-year value was 41% as determined by the Kaplan-Meier method. No statistically significant difference in survival was observed between the two groups. The initial tumor response correlated with the survival value. No severe complication was encountered except for one case of liver abscess formation. No serious change in the laboratory data was observed following treatment with these regimens. Although the tumor response was significantly better in patients treated with ADMOS combined with CDDP injection than in those treated with ADMOS alone (P < 0.05), no significant difference in survival was found between the two groups.


Outcome of localized hepatocellular carcinoma treated with segmental arterioportal chemoembolization. When lipiodol is injected into the hepatic artery at a dose exceeding a certain level, it flows into the portal vein. On the basis of this feature, an emulsion of Adriamycin with lipiodol was injected into a segmental or subsegmental artery such that it was delivered to the portal vein of the same segment, and the artery was then embolized with Gelfoam. This segmental arterioportal chemoembolization (cement therapy) was performed in 50 patients with localized hepatocellular carcinoma. A posttreatment CT scan showed that almost 100% of the lesions were occupied by lipiodol. The cumulative survival values determined for the 50 patients were very high: 83.4% after 1 year and 62.7% after 2 years.


Clinicopathological study on combination therapy consisting of arterial infusion of lipiodol-dissolved SMANCS and transcatheter arterial embolization for hepatocellular carcinoma. Combination therapy (LpTAE) consisting of arterial infusion of a lipophilic anticancer drug, SMANCS, dissolved in an oily lymphographic agent, lipiodol (LPD), and transcatheter arterial embolization (TAE) for hepatocellular carcinoma (HCC) was studied with special reference to the pathological findings. A total of 32 patients were subjected to surgical resection after LpTAE. The pattern of LPD deposition in the tumor was examined by CT scan (Lipiodol CT, LpCT) at 7 days and/or 1 month after LpTAE. The resected materials were examined radiographically with soft X-rays and histologically. LPD was deposited in tiny daughter nodules with a diameter of less than 5 mm and in tumor thrombi as well as in the main tumors, which showed necrotic change. Part of the LPD flowed out from the main tumor via the drainage vein and was deposited in the capsular invasion, resulting in necrosis. LPD accumulated almost exclusively within the blood spaces of trabecular-type HCC, creating a pattern corresponding to a cast of the tumor vessels, which showed prominent necrosis. On the other hand, LPD was not deposited in scirrhous, compact, or well-differentiated HCC, which showed little or no necrosis. It was demonstrated that LpCT images, which accurately depicted the existence and the extent of LPD deposition and necrosis in the tumor, were useful for precise evaluation of the therapeutic effect. Our findings indicate that LpTAE and LpCT are valuable for the diagnosis and treatment of HCC and should play a central role in systemic therapeutic approaches to this disease.


Effectiveness of Lipiodol in transcatheter arterial embolization of hepatocellular carcinoma. The effectiveness of Lipiodol (iodized oil) in transcatheter arterial embolization (TAE) of hepatocellular carcinoma (HCC) was retrospectively evaluated using statistical analysis. A total of 343 HCC patients who underwent TAE at 5 institutions between 1984 and 1989 were divided into 2 groups: the GS-TAE group underwent TAE with Gelfoam sponge alone, whereas the LP-TAE group was given Lipiodol (LP) immediately before GS-TAE. The statistical T value calculated for the LP-TAE group showed that the administration of LP, the tumor size, intrahepatic metastasis, portal vein infiltration, and serum total bilirubin and alpha-fetoprotein levels significantly (P < 0.01) affected the patients' survival. Both the cumulative survival determined using the Kaplan-Meier model and the cumulative hazard calculated using Cox's proportional hazard model differed significantly (P < 0.01) between the GS-TAE group and the LP-TAE group (log-rank test). These results confirmed the effectiveness of LP used in combination with Gelfoam sponge for TAE of HCC.


Transcatheter arterial chemoembolization for hepatocellular carcinoma. The factors relating to the duration of survival were analyzed for 329 cases of hepatocellular carcinoma (HCC) treated by transcatheter arterial chemoembolization (TACE) between January 1, 1983, and December 31, 1990. The cumulative survival value obtained was slightly higher but not statistically significantly higher in these 329 cases as compared with the 129 cases reported in 1989. This improvement was probably attributable to careful selection of the patients on the basis of the clinical Child's status and to improvement of the superselective angiotechnique used in the present study. The overall cumulative 1-, 2-, and 3-year survival values in this series were 50%, 25%, and 15%, respectively. The median duration of survival was 12.7 months (Kaplan-Meier method). The 1-, 2-, and 3-year survival values determined for 190 patients in Child's group A, for 95 patients in Child's group B, and for 44 patients in Child's group C were 60%, 30%, and 20%; 35%, 20%, and 10%; and 35%, 20%, and 10%, respectively. Analyses were also carried out according to the tumor's type and size, the integrity of the tumor capsule, and the patency of the portal vein. Our results disclosed that a better outcome in terms of the median survival period and the survival value was favored by the following factors: a single lesion measuring less than 5 cm in diameter, an intact capsule, a patent portal vein, and a good clinical status. It is also essential that all patients who have undergone TACE be periodically evaluated by ultrasonography, CT, and angiography to determine whether repeated chemoembolization is necessary.


Arterial infusion chemotherapy for advanced hepatocellular carcinoma using EPF and EAP therapies. Arterial infusion chemotherapy of EPF (etoposide, cisplatin, and 5-fluorouracil) or EAP (etoposide, Adriamycin, and cisplatin) was carried out in 28 cases of advanced hepatocellular carcinoma (HCC) between January 1988 and December 1990, and assessment was made of the anticancer efficacy of each treatment method. In all, 13 patients were treated with EPF therapy and 15 received EAP therapy. The anticancer agents were infused through a catheter inserted into the proper or common hepatic artery. The catheter was inserted via the axillary artery or common femoral artery using Seldinger's method or the cut-down method. The results of each therapy were analyzed in relation to the tumor regression rate and the side effects encountered. The tumor regression rate was determined on the basis of two-dimensional evidence obtained by computed tomography performed before and after treatment. The treatment results were also compared with the results of chemoembolization therapy using a mixture of cisplatin (CDDP), Adriamycin (ADM) and lipiodol. Of the 28 patients treated with arterial infusion chemotherapy, 14 (50%) attained a regression rate of 50% (PR). In all, 46% of the EPF group and 53% of the EAP group achieved a PR. These results were superior to those obtained using chemoembolization therapy. In general, the side effects were relatively mild and transient.


Clinical evaluation of intermittent arterial infusion chemotherapy with an implanted reservoir for hepatocellular carcinoma. A total of 45 patients with advanced hepatocellular carcinoma were treated at Osaka City University Hospital by intermittent arterial infusion chemotherapy with an implanted reservoir. The treatment consisted of intermittent infusion of doxorubicin (5-20 mg/body), mitomycin C (4-10 mg/body) or degradable starch microspheres (600-1200 mg/body) plus doxorubicin (30 mg/body). In all, 26% of the patients received this treatment for disease recurrence following transcatheter arterial embolization (TAE). Among 43 evaluable patients, 4 showed a complete remission (CR) and 16 showed a partial response (PR) on computed tomograms and angiograms. For all 45 patients, the 1-year survival value was 41% and the 2-year value was 14%. Of the 20 patients who showed a CR or PR, 77% survived for 1 year and 29% survived for 2 years. Tumor regression showed a close relationship with the duration of survival. Intermittent arterial infusion with an implanted reservoir caused the least adverse reactions and seems to be appropriate for use in patients with advanced tumor extension or stenosis of the hepatic artery caused by repeated TAE.


Oxidation markedly reduces bilirubin interference in the Jaffé creatinine assay. Bilirubin causes underestimation of serum creatinine in the Jaffé alkaline picrate assay. We report an approach for preventing bilirubin interference by pretreating serum samples with peroxidase and H2O2. The dissociation of bilirubin from albumin and its subsequent oxidation markedly reduces the bilirubin interference and enables accurate determination of creatinine concentrations by the Jaffé reaction even in hyperbilirubinemic sera. Within-run CVs were 2.6%, 4.0%, and 3.8% at mean creatinine concentrations of 88, 165, and 349 mumol/L, respectively (n = 20). Day-to-day CVs were 4.0%, 6.3%, and 5.8% for mean creatinine concentrations of 87, 168, and 364 mumol/L, respectively (n = 12). Average recovery of creatinine added to serum in the presence of 600 mumol/L bilirubin was 97% (n = 15). This method requires only small serum volumes (70 microL) and is easily applicable to automated analyzers that can be programmed to add three reagents consecutively.


Phospholipase resistance of the glycosyl-phosphatidylinositol membrane anchor on human alkaline phosphatase. Alkaline phosphatase (ALP) is attached to the cell surface in mammalian tissues via a glycosyl-phosphatidylinositol (GPI) anchor and can be released from the membrane by GPI-specific phospholipases. In a range of cultured human cell lines, however, the sensitivity of ALP to phospholipases was observed to be variable in magnitude (approximately 20-90%). The mechanism of phospholipase resistance was explored with phospholipases of different bond specificities. The results suggest that phospholipase resistance is the result of acylation of the inositol ring in the GPI anchor. The occurrence of phospholipase-resistant forms of ALP may have important implications for the in vivo release and disposition of plasma ALP.


Decreased abundance of alkaline phosphatase and calbindin-D9K mRNAs in the intestine of the spontaneously hypertensive rat. Alkaline phosphatase activity and calbindin-D9K immunoreactivity are decreased in the intestines of spontaneously hypertensive rats (SHRs). To investigate the potential role of altered gene expression in these decreases, we measured, by Northern blot analyses, the abundances of alkaline phosphatase and calbindin-D9K mRNAs in the proximal regions of the small intestines of 14-week-old SHR and control Wistar-Kyoto (WKY) rats. Alternate 4-cm segments of intestine were used for measurements of the proteins (0-4 cm, 8-12 cm, and 16-20 cm from pylorus, segments A1, B1, and C1, respectively) and mRNAs (4-8 cm, 12-16 cm, and 20-24 cm, segments A2, B2, and C2). Calbindin-D9K (immunoassay) was decreased in SHR vs WKY rats by 27%, 64%, and 67% in segments A1, B1, and C1, respectively (P < 0.01); its mRNA was decreased to a similar extent (69%, 82%, and 80%, respectively; P < 0.002 by analysis of variance). Alkaline phosphatase activity was decreased in SHRs by 58%, 54%, and 51% in segments A1, B1, and C1, respectively (P < 0.01); the abundance of its 3.0-kb mRNA was decreased to a similar extent: 57%, 80%, and 69% in segments A2, B2, and C2, respectively (P < 0.02). The mean decreases of the 2.7-kb mRNA of alkaline phosphatase were statistically significant (P < 0.02) but smaller (38%, 40%, and 35%). The mean abundance of vitamin D receptor mRNA in the same animals was decreased slightly in SHR vs WKY rats (3%, 36%, and 20% in segments A2, B2, and C2, respectively), but the difference in the values was not statistically significant. Decreases in alkaline phosphatase activity and calbindin-D9K immunoreactivity may reflect decreased mRNA abundance and not decreased enzyme-specific activity or increased protein degradation.


The clinicopathological features of familial cylindromas and trichoepitheliomas (Brooke-Spiegler syndrome): a report of two families. We have studied four patients from two families and obtained the clinical data from four other affected family members who demonstrated the coexistence of multiple cylindromas and trichoepitheliomas. An autosomal dominant pattern of inheritance with variable penetrance was exhibited in both families, with a female to male ratio of 3:1. The clinical and pathological features of this rare association are discussed.


A hereditary immunodeficiency characterized by CD8+ T lymphocyte deficiency and impaired lymphocyte activation. An unusual form of severe combined immunodeficiency in children from two different families was associated with absence of CD8+ T lymphocytes and normal numbers of CD4+ T lymphocytes that did not respond to stimulation by non-specific mitogens, specific antibodies against T cell receptor or specific antigens. The defect in the CD4+ cells was bypassed by activating agents which are independent of the T cell receptor. The combination of an activation defect and selective depletion of CD8+ T lymphocytes suggests that the defective pathway is important in the differentiation of immature thymocytes as well as the proliferation of mature lymphocytes.


Cystic ovaries in women affected with hereditary angioedema. Polycystic ovary (PCO) syndrome is biochemically characterized by abnormal gonadotropin secretion and polycystic ovaries associated with increase in size and functional activity of stromal tissue; multifollicular ovaries (MFO) are defined by the presence of multiple cysts with no increase in stromal tissue. A central (hypothalamic-pituitary) abnormality, including high plasma beta-endorphin (BE) concentrations without simultaneous elevation of ACTH, was reported for subjects with PCO syndrome. Since we have found the presence of high plasma BE concentrations in hereditary angioedema (HANE) during attacks as well as during symptom-free periods, we studied, by means of pelvic ultrasound scanning employed to determine the prevalence of PCO and of MFO, 13 women of reproductive age affected with HANE who were not on oral contraceptives. We have found PCO in 5/13 (38.4%) and MFO in 7/13 (53.8%) HANE patients. Nine patients had oligomenorrhoea (five with PCO, three with MFO, one with normal ovaries), five (three with PCO, two with MFO) were hirsute and only one (with MFO) had weight loss. No patient was obese. Mean plasma LH, testosterone, prolactin, cortisol and ACTH concentrations were normal, while FSH was significantly reduced and LH/FSH ratio increased. BE concentrations were significantly high in all the patients studied. Our results clearly demonstrate that women with HANE frequently have cystic ovaries (polycystic or multifollicular) in the presence of high BE concentrations.


The B cell repertoire in patients with systemic autoimmune diseases: analysis of Epstein-Barr virus (EBV)-inducible circulating precursors that produce autoantibodies against nuclear ribonucleoprotein (nRNP). Peripheral blood B cells from patients with systemic autoimmune disease and healthy volunteers were immortalized using EBV and the frequencies of B cell precursors that produced immunoglobulin class-specific antibodies against anti-nRNP, a specific marker for mixed connective tissue disease, were assessed using limiting dilution analysis. The frequencies of EBV-induced B cell precursors that produced IgG anti-nRNP were correlated closely with the serum titres of the corresponding autoantibodies, which indicates that B cell precursors that produced potentially pathogenic autoantibodies could be immortalized from the peripheral blood of the patients by EBV. In contrast, the frequency of EBV-induced B cell precursors that produced IgM anti-nRNP in patients with systemic autoimmune disease was comparable to that in healthy volunteers and greater than those that produced IgG and IgA anti-nRNP. Moreover, many of the clones that produced IgM antibodies against nRNP reacted with other autoantigens, such as double-stranded DNA, single-stranded DNA and rabbit IgG. These polyreactive IgM antibodies are believed to belong to the 'natural antibodies', to be coded by the germline immunoglobulin V genes, and to react with evolutionarily conserved structural cellular components, including nRNP. Our finding that nRNP is one of the target antigens for this polyreactive autoantibody may lead to the elucidation of the origin of the pathogenic IgG and IgA anti-nRNP antibodies found in sera from patients with systemic autoimmune diseases.


Idiotypic mimicry of a cell surface DNA receptor: evidence for anti-DNA antibodies being a subset of anti-anti-DNA receptor antibodies. Anti-idiotypic anti-DNA antibodies (anti-anti-DNA) have previously been described in both patients with systemic lupus erythematosus and healthy individuals. Jerne's hypothesis predicts that such antibodies would bear a paratope reactive with non-sequence specific DNA binding proteins. Here we have explored the notion of a molecular mimicry between anti-anti-DNA antibodies and antibodies to a previously described 28-29 kD cell surface DNA binding molecule. It was shown that affinity purified anti-anti-DNA antibodies inhibit the binding of DNA to cells and that MoAb to the 28-29 kD receptor react with anti-DNA antibodies. These findings indicate that a subset of anti-anti-DNA antibodies are idiotypically related to antibodies reactive with a cell surface DNA binding molecule. It is hypothesized that anti-DNA antibodies may arise when a convergence of genetic and environmental influences favours an unrestrained anti-idiotypic response to cell surface DNA binding molecule(s).


[Cerebrotendinous xanthomatosis--a case of brain MRI abnormality and osteoporosis]. A 37-year-old male with cerebrotendinous xanthomatosis showed brain abnormal MRI findings and osteoporosis. His parents had no similar symptoms. He had mental retardation since childhood. Swelling of Achilles tendons was noticed at age 28, and gait disturbance appeared at age 34. Physical examination revealed bilateral cataracts and swelling of Achilles tendons. Neurologically, he showed mental retardation, cerebellar ataxia and spastic tetraparesis. Cerebrotendinous xanthomatosis was diagnosed by marked elevations of serum cholestanol level (24.3 micrograms/ml) and cholestanol/cholesterol ratio (1.81%) as well as characteristic clinical manifestations. On brain MRI study, T2-weighted sequence showed bilateral focal lesions with high intensity signal in the globus pallidus and cerebellar white matter adjacent to the dentate nucleus, and T1-weighted sequence showed low to iso-intensity signal in the same regions. These findings suggested demyelination rather than xanthoma or lipid infiltration. Radiological examination showed mild osteoporosis of lumbar bone. However, serum levels of vitamin D3 and calcitonin were within normal range, and renal function was normal. Osteoporosis in this patient possibly resulted from disuse bone atrophy for several years. The combination therapy of oral administration of chenodeoxycholic acid and HMG-CoA reductase inhibitor (pravastatin), and LDL apheresis slightly improved EEG abnormality and gait disturbance, but not brain MRI abnormality.


[A case of chronic toluene poisoning with myopathic changes]. We reported a 27-year-old man, who had inhaled toluene vapor for 11 years and developed both polyneuropathy and myopathy. He gradually developed progressive muscular weakness in the limbs 2 months after the last inhalation and became nonambulant. Neurological examination revealed distal dominant muscular weakness and hypoactive deep tendon reflexes, especially in the legs. The cranial nerves were intact and neither sensory disturbances nor cerebellar signs were observed. Laboratory studies demonstrated mildly elevated serum CK value, but serum electrolytes and urinalysis were normal. Electromyogram showed a neurogenic pattern in the limbs and also a myogenic in the gastrocnemius muscle. Motor nerve conduction velocities were delayed in the limbs. The sural nerve biopsy specimens showed demyelination, myelin avoids, and axonal swelling compatible with toxic neuropathy. The muscle biopsy of the gastrocnemius revealed necrotic muscle fibers, vacuolated muscle fibers, phagocytosis, a disordered intermyofibrillary network, and small group atrophy on light microscopy. On electron microscopy, two outstanding findings were observed; the first, dilatation and proliferation of the membrane system, and the second, appearance of irregular membrano lamellar structures and vacuoles associated with abnormal membrane system. These histological findings suggest that toluene may affect directly the membrane system of muscle fibers and consequently induce rhabdomyolysis.


[A case of herpes simplex encephalitis diagnosed by polymerase chain reaction]. A case of a 55-year-old male with herpes simplex encephalitis (HSE) was reported. He was admitted because of fever, headache and memory disturbance. T1 weighted MRI showed low signal intensity and T2 weighted imaging revealed high signal intensity in the medial portions of bilateral temporal lobes. Herpes simplex virus (HSV) antibody titer in cerebrospinal fluid (CSF) was not elevated. HSV DNA in CSF was amplified by polymerase chain reaction (PCR) and identified by the microplate hybridization method. The PCR technique would be useful for the diagnosis of HSE.


[A case of carcinomatous autonomic and sensory neuropathy]. A 61-year-old male with carcinomatous autonomic and sensory neuropathy was presented. The disease started with numbness in his lower extremities and loss of sensation gradually developed over the upper extremities within two months. Finally he became unable to walk because of orthostatic hypotension and dystaxia due to loss of deep sensation. Physical examination on admission revealed mydriasis with negative light reflex, orthostatic hypotension, sensory loss of below C-4, areflexia, positive Babinski's sign and sexual impotence. Laboratory examinations revealed serum antineural antibody, elevation of protein and oligoclonal IgG band in the CSF. Sural nerve biopsy specimen showed severe loss of myelinated fibers, especially in the large-sized fibers. The pharmacological functional tests of the autonomic nerves disclosed disturbance of parasympathetic nervous system as well as the postsynaptic fibers of the sympathetic nerves. An instillation test of 2.5% mecholyl solution demonstrated contraction of both pupils. Chest X-ray showed abnormal shadow at the right hilus and transbronchial biopsy revealed small cell carcinoma (oat cell type). Plasma exchange improved orthostatic hypotension and decreased the titer of the antineural antibody. In this case, the autonomic ganglion including the ciliary ganglia and the dorsal root ganglia cells were predominantly involved and such cases have been described as a variant of paraneoplastic syndrome in the literature.


Preliminary report on cytokine determination in human synovial fluids: a consensus study of the European Workshop for Rheumatology Research. The Cytokine Consensus Study Group of the European Workshop for Rheumatology Research. A consensus study involving several European research groups was conducted in order to assess the reliability and reproducibility of cytokine measurements in biological fluids. Six synovial fluids and one serum--some of them spiked with recombinant human cytokines--were aliquoted and distributed blindly to different laboratories. The samples were tested for tumor necrosis factor alpha (TNF-alpha), interleukin-1 alpha (IL-1 alpha) and 1 beta, IL-6, IL-2, interferon gamma (IFN-gamma), and the soluble receptors of IL-2 (IL-2R) and TNF (TNF-sR55 and -sR75), using various immunoassays and, occasionally, bioassays. The same ELISA used in different laboratories yielded comparable results, whereas different ELISAs usually detected the highest levels in the same samples, but yielded different absolute values. This finding highlights the necessity of establishing international standards for all immunoassays. Many other questions arose during this preliminary study, and further investigations are planned to clarify them.


Platelet alpha 2-adrenergic receptor responsiveness is increased in elderly men but not in elderly women. We ascertained platelet alpha 2-adrenergic receptor responsiveness in healthy young and elderly men and women by determining the minimum concentration of methylnorepinephrine, a selective alpha 2-adrenergic receptor agonist, required to initiate the primary and secondary aggregation response in platelet-rich plasma. We observed that platelets from elderly men required a smaller concentration of methylnorepinephrine to stimulate primary aggregation than did platelets obtained from young men. However, the concentration of alpha 2-adrenergic receptor agonist to trigger the secondary aggregation response did not vary with age. There was no difference in the responsiveness of platelets from young and elderly women for either the primary or secondary aggregation response. We conclude that platelet alpha 2-adrenergic receptor responsiveness is increased with age rather than decreased, as predicted from some studies of the density of platelet alpha 2-adrenergic receptors and their coupling to adenylyl cyclase. Furthermore, the increase in alpha 2-adrenergic receptor responsiveness is gender specific.


Psychoactivity and abuse potential of sumatriptan. Sumatriptan is a 5-HT1D agonist of therapeutic use in migraine and cluster headaches. To determine the profile of psychoactivity and abuse potential, a double-blind Latin-square crossover study was conducted in 12 male subjects with histories of substance abuse. The effects of subcutaneous placebo, sumatriptan (8 and 16 mg), and morphine (10 and 20 mg) were assessed on measures of subjective, behavioral, and physiologic responses including signs, symptoms, Addiction Research Center Inventory scales, onset of drug effects and miosis. Sumatriptan was psychoactive, was discriminated from placebo, produced a dose-related decrease on euphoria scores, elevated scores on measures of apathetic sedation and disliking, and lacked identification as a prototypic drug of abuse. There were no clinically significant effects on heart rate, pupil size, or blood pressure. In contrast, morphine (the positive control) produced expected dose-response relationships on measures of reinforcing and physiologic effects. The study suggests that sumatriptan has a low abuse potential.


Nail-patella syndrome associated with mixed crystal deposition arthropathy. This report describes a case of nail-patella syndrome in a woman with a strong family history who presented with effusions in her shoulder and knees. Microscopic examination of the shoulder fluid suggested the presence of calcium pyrophosphate dihydrate (CPPD) crystals, and examination of the knee fluid suggested both hydroxyapatite (HAP) and CPPD crystals. To our knowledge, the coexistence of these two conditions has not been reported in the past. Moreover, it brings up a new element regarding the inflammatory origin of the nail-patella syndrome.


Virus infection and hand papular dermatoses in young children. Four strains of enterovirus and 1 strain of adenovirus were isolated from stool and pharyngeal swabs by inoculation into primary human embryonic kidney cell culture in 25 cases of dorsum manus infantile papular dermatitis. Two strains of Coxsackie A-9 virus and 1 strain of Echovirus-25 were identified. With the isolated virus as antigen, a matched pair study by sera antibody test was carried out in 43 children with papular dermatitis and 40 normal individuals as controls. In 26 cases of Coxsackie A-9 virus positive papular dermatitis, a 4-fold rise (> 1:1.6) in the relative serum antibody was observed in 80.77%, while no high levels were observed in patients with Echovirus-25 and adenovirus dermatitis (< 1:16). The authors can reasonably conclude that epidemic Coxsackie A-9 virus infection is obviously related to infantile papular dermatitis and is probably the main cause of the disease.


[The Askin tumor].

[The Askin tumor]. The Askin tumor, a primitive malignant small-cell tumor of the chest wall, is mostly seen among children and adolescents. It is closely related to Ewing's sarcoma of the same location, both tumors showing a chromosomal translocation t(11;22). Its origin from neuroectodermal cells is deducted from several ultrastructural details and from the expression of specific markers like NSE. Pain and deformation of the chest wall are the cardinal clinical signs of the tumor. Chest X-rays will frequently show destruction of ribs and pleural effusions. Effective therapy consists of radical surgery, local radiation and adjuvant chemotherapy. This multimodal concept allows minority of patients to remain disease-free but the overall outcome is rather unfortunate.


Features of apoptotic cells measured by flow cytometry. The present review describes several methods to characterize and differentiate between two different mechanisms of cell death, apoptosis and necrosis. Most of these methods were applied to studies of apoptosis triggered in the human leukemic HL-60 cell line by DNA topoisomerase I or II inhibitors, and in rat thymocytes by either topoisomerase inhibitors or prednisolone. In most cases, apoptosis was selective to cells in a particular phase of the cell cycle: only S-phase HL-60 cells and G0 thymocytes were mainly affected. Necrosis was induced by excessively high concentrations of these drugs. The following cell features were found useful to characterize the mode of cell death: a) Activation of an endonuclease in apoptocic cells resulted in extraction of the low molecular weight DNA following cell permeabilization, which, in turn, led to their decreased stainability with DNA-specific fluorochromes. Measurements of DNA content made it possible to identify apoptotic cells and to recognize the cell cycle phase specificity of the apoptotic process. b) Plasma membrane integrity, which is lost in necrotic but not apoptotic cells, was probed by the exclusion of propidium iodide (PI). The combination of PI followed by Hoechst 33342 proved to be an excellent probe to distinguish live, necrotic, early- and late-apoptotic cells. c) Mitochondrial transmembrane potential, assayed by retention of rhodamine 123 was preserved in apoptotic but not necrotic cells. d) The ATP-dependent lysosomal proton pump, tested by the supravital uptake of acridine orange (AO) was also preserved in apoptotic but not necrotic cells. e) Bivariate analysis of cells stained for DNA and protein revealed markedly diminished protein content in apoptotic cells, most likely due to activation of endogenous proteases. Necrotic cells, having leaky membranes, had minimal protein content. f) Staining of RNA allowed for the discrimination of G0 from G1 cells and thus made it possible to reveal that apoptosis was selective to G0 thymocytes. g) The decrease in forward light scatter, paralleled either by no change (HL-60 cells) or an increase (thymocytes) of right angle scatter, were early changes during apoptosis. h) The sensitivity of DNA in situ to denaturation, was increased in apoptotic and necrotic cells. This feature, probed by staining with AO at low pH, provided a sensitive and early assay to discriminate between live, apoptotic and necrotic cells, and to evaluate the cell cycle phase specificity of these processes. i) The in situ nick translation assay employing labeled triphosphonucleotides can be used to reveal DNA strand breaks, to detect the very early stages of apoptosis.(ABSTRACT TRUNCATED AT 400 WORDS)


Immunoblotting technique to study release of melanophore-stimulating hormone from individual melanotrope cells of the intermediate lobe of Xenopus laevis. The melanotrope cells in the pars intermedia in the pituitary of Xenopus laevis synthesize and release the melanophore-stimulating hormone (alpha MSH), a small peptide that causes skin darkening during the process of background adaptation. Evidence has been found for a heterogeneity in biosynthetic activity of the melanotrope cells. In the present study two questions were addressed: (1) does the melanotrope cell population also show heterogeneous alpha MSH-release, and (2) can this heterogeneity be changed by extracellular messengers? Since dopamine is known to inhibit alpha MSH-release, this messenger is used to study the regulation of the heterogeneity. To quantify alpha MSH-release from individual cells, a cell blotting procedure has been developed for the binding and relative quantification of the small alpha MSH peptide. The immunoblotting procedure involves binding of the cells to a carrier slide and binding of released alpha MSH to a nitrocellulose filter. After immunostaining, the amount of alpha MSH per cell was quantitated by image analysis. Untreated melanotrope cells reveal a distinct variability in alpha MSH-release, some cells showing low secretory activity, whereas others are strongly secreting, indicating heterogeneity of alpha MSH-release. Dopamine treatment strongly inhibits alpha MSH-release from individual cells, resulting in a clearly less pronounced melanotrope cell heterogeneity. The effect of dopamine appears to be dose-dependent as a low dopamine concentration has only a moderate effect on the alpha MSH-release. It is proposed that dopamine is a physiological regulator of the degree of melanotrope cell heterogeneity in alpha MSH-release.


Phosphatidylinositol 4-kinases and the role of polyphosphoinositides in cellular regulation. Phosphoinositides play a central role in the transduction of signals for a variety of hormone and growth factor receptors. Multiple derivatives of phosphatidylinositol are present within the cell including phosphatidylinositol 4,5-bisphosphate, the phosphorylated derivative that is hydrolyzed by phospholipase C to produce the two intracellular second messengers, diacylglycerol and inositol 1,4,5-trisphosphate. The synthesis, degradation, and subsequent resynthesis of the phosphoinositides form a metabolic cycle known as the phosphoinositide cycle. The phosphoinositide cycle begins with the phosphorylation of phosphatidylinositol to form phosphatidylinositol 4-phosphate, a reaction catalyzed by phosphatidylinositol 4-kinase. Phosphatidylinositol kinase activity has been reported to be present in a variety of cellular membranes, and multiple isozymes of phosphatidylinositol 4-kinase are present within the cell, suggesting that the product of this reaction may have more than one biological function. The activity of phosphatidylinositol 4-kinase is regulated by growth factors, further underscoring the importance of this enzyme in cellular regulation. Recent data suggest that in addition to serving as substrates for phospholipase C, the polyphosphoinositides may themselves function as intracellular mediators of hormone action. For example, polyphosphoinositides have marked effects on the activity of certain actin binding proteins that may allow these lipids to participate in the regulation of actin polymerization. This review focuses on the properties of the phosphatidylinositol 4-kinases and the potential role of polyphosphoinositides in the regulation of cellular processes.


Purification and characterization of a mitochondrial endonuclease from Drosophila melanogaster embryos. A mitochondrial endonuclease from Drosophila melanogaster embryos was purified to near homogeneity by successive fractionation with DEAE-cellulose and heparin--avidgel-F, followed by FPLC chromatography on mono S, Superose 12 and a second mono S column. This enzyme digests double-stranded DNA more efficiently than heat-denatured DNA. The endonuclease activity has a molecular mass of 44 kDa, as determined under native conditions using a gel-filtration Superose 12 column. The prominent peptide detected by SDS/polyacrylamide gel electrophoresis likewise has a molecular mass of 44 kDa, suggesting a monomeric protein. The enzyme has an absolute requirement for divalent cations, preferring Mg2+ over Mn2+. No activity could be detected when these cations were replaced by Ca2+ or Zn2+. The pH optimum for this enzyme activity is 6.5-7.4 and its isoelectric point is 4.9. Both single-strand and double-strand breaks are introduced simultaneously into a supercoiled substrate in the presence of MgCl2 or MnCl2. Endonuclease-treated DNA serves as a substrate for DNA polymerase I from Escherichia coli, suggesting that 3'-OH termini are generated during cleavage. The enzyme is free from any detectable DNA exonuclease activity but not from RNase activity. Partial inhibition by antibodies raised against mitochondrial endonucleases derived from bovine heart and Saccharomyces cerevisiae have revealed a potential structural homology between these nucleases.


Affinity labelling of the active site of brain phosphatidylinositol 4-kinase with 5'-fluorosulphonylbenzoyl-adenosine. 5'-p-Fluorosulphonylbenzoyl-adenosine (FSO2BzAdo), an affinity labelling analogue of ATP, was used to label the active site of sheep brain phosphatidylinositol 4-kinase (PtdIns 4-kinase). The incubation of PtdIns 4-kinase with concentrations of FSO2BzAdo as low as 50 microM resulted in considerate inactivation of the enzyme. (e.g. 55% less after 60 min with 50 microM FSO2BzAdo). The kinetics of inactivation of PtdIns 4-kinase by FSO2BzAdo suggest a two-step mechanism, in which a rapid reversible binding of FSO2BzAdo to the enzyme is followed by a covalent sulphonation step. The first-order rate constant (k2) for the inactivation of PtdIns 4-kinase was calculated to be 0.063 min-1, and the steady-state constant of inactivation (Ki) to be 200 microM. Preincubation of the enzyme with either ATP plus Mg2+, or PtdIns alone, prior to addition of FSO2BzAdo reduced the degree of inactivation of the enzyme; suggesting that FSO2BzAdo binds within the active site PtdIns 4-kinase. Moreover, since ATP plus Mg2+ provided the greatest protection against inactivation, it is concluded that the main site of labelling of PtdIns 4-kinase by FSO2BzAdo is within the ATP-binding site of the enzyme. Results obtained from chemical modification experiments, which employed pyridoxal 5'-phosphate and tetranitromethane, are consistent with a catalytically-essential lysine being present within the ATP-binding site of PtdIns 4-kinase. Therefore, it is hypothesised that the inactivation of PtdIns 4-kinase by FSO2BzAdo may be due to the labelling of this lysine residue.


Ganglioside binding proteins of calf brain with ubiquitin-like N-terminals. Two ganglioside-associated protein components I and II have been isolated from crude ganglioside preparations of calf brain by DEAE-Sephadex ion-exchange chromatography. Both components exhibited binding capacity in aqueous media for gangliosides of the 'ganglio' series but not for neutral glycosphingolipids (polyglycosylceramides) and only a low capacity for sialosylparagloboside. Each protein bound individual gangliosides with different efficiency. Upon prolonged incubation of component I with gangliosides, complexes with high (30:1) and low (6:1) glycolipid/protein molar ratios were formed. The latter but not the former complex was able to penetrate Sephadex G-200 beads. Both components inhibited plating efficiency of cultured mouse N2a neuroblastoma cells. The molecular masses of components I and II were determined by SDS/PAGE to be 11-12 kDa and 28 kDa, respectively. Carbohydrates (fucose, mannose, galactose, N-acetylglucosamine, N-acetylgalactosamine, and some sialic acid) were found only in component II. When examined by reverse-phase HPLC each component separated into two major closely migrating peaks which were subsequently examined by Edman degradation. Amino acid sequences of the N-terminal portions of three of these peaks (one peak from component I and both peaks from component II) showed, as far as the sequences were established, identity with the sequence of ubiquitin. It is hypothesized that the proteins may be instrumental in intracellular trafficking of gangliosides.


Agarose gel pore radii are not dependent on the casting buffer. Previous studies have shown that the apparent pore size of agarose gels is dependent on the buffer in which the gel is cast and run (D.L. Holmes and N.C. Stellwagen, Electrophoresis 1990, 11, 5-15; N.C. Stellwagen and D.L. Holmes, Electrophoresis 1990, 11, 649-652). However, these studies, based on the mobility of DNA restriction fragments, neglected the effect of electroendosmosis. By measuring the mobility of vitamin B12 under various experimental conditions, it is shown here that electroendosmosis is highly buffer-dependent. When the observed mobilities of DNA are corrected for electroendomosis, the apparent pore radii of agarose gels are found to be independent of the casting buffer.


Identification, purification and characterization of a novel phosphatidylinositol-specific phospholipase C, a third member of the beta subfamily. The partial sequence of a novel PtdIns-specific phospholipase C of the beta subfamily (PtdIns-PLC beta 3) is described. Based upon the predicted protein sequence, monospecific antibodies have been raised and used to identify a suitable source for purification of the protein. Fractionation of HeLa S3 cells revealed that immunoreactive PtdIns-PLC beta 3 is membrane associated; purification (approximately 1000-fold) from this fraction yielded a single immunoreactive protein of 158 kDa, with a specific activity of 136 mumol.min-1.mg-1, with PtdIns 4,5-bisphosphate as substrate. Substrate specificity and Ca2+ dependence of this purified PtdIns-PLC are characteristic of the PtdIns-PLC beta subfamily.


Ceruloplasmin in human plasma and its relationships with the copper-albumin complex. The electron paramagnetic spectrum of human plasma is dominated, in the g = 2 region, by resonances from copper atoms bound to ceruloplasmin, and does not reveal the fraction of copper normally associated with albumin, except in a few cases, where a copper-albumin signal increases with time after blood withdrawal. This copper-albumin complex is responsible for a resonance at a g value below g = 2 in the spectrum of human serum, which has been recently attributed to a modified form of type 2 copper bound to ceruloplasmin [Rylkov, V.V., Tarasiev, M.Y. & Moshkov, K.A. (1991) Eur. J. Biochem. 197, 185-189]. In the plasma, copper associated to albumin comes from ceruloplasmin. Purified ceruloplasmin is unable to exchange copper with albumin, either purified or in plasma. It can not be ruled out that some serum components trigger the metal exchange, in a defence mechanism operating when ceruloplasmin leaks, by unknown processes, its copper content before discharging the metal into the various organs.


Nonradioactive ribonuclease protection analysis using digoxygenine labeling and chemiluminescent detection. A sensitive nonradioactive ribunuclease protection assay is described which we have used to study c-myc gene transcription and promoter usage in GLC4, a human small cell lung carcinoma cell line with amplified gene. For in vitro transcription we used digoxygenine (DIG)-rUTP instead of [alpha-32P]CTP or [alpha-32P]UTP and after polyacrylamide gel electrophoresis the protected probes were transferred to a nylon membrane from Boehringer Mannheim using electroblotting. Subsequently the membrane was analyzed by chemiluminescent detection. Results were obtained after 1 h of exposure and were comparable with those using radioactivity.


Intensive chemotherapy in poor-prognosis nonseminomatous germ cell tumors of the testis. Forty-one patients with poor-prognosis nonseminomatous germ cell tumors (NSGCT) of the testis were treated between 1980 and 1989. This group was defined by the presence of one of the following features: multiple large lung metastases, bone, liver or brain metastases, abdominal mass greater than 10 cm, abdominal mass greater than 5 cm with high serum concentration of the tumor markers [alpha-fetoprotein (alpha FP) greater than 500 kU/l or beta-subunit of human chorionic gonadotropin (beta HCG) greater than 1,000 IU/l) or very high serum tumor marker concentrations (alpha FP greater than 5,000 kU/l or beta HCG greater than 10,000 IU/l). The first 21 patients were treated with cisplatin, vinblastine, bleomycin (PVB) chemotherapy and the following 20 with an intense, alternating 6-drug chemotherapy consisting of cisplatin, bleomycin, vincristine, methotrexate, etoposide and ifosfamide (BOMP/EPI). Surgery of residual masses was performed when tumor markers were negative. Fifteen patients (71.4%) in the PVB group and 18 patients (85%) in the BOMP/EPI group remained disease-free at a median follow-up of 67 and 41 months, respectively. None of the resected masses in the BOMP/EPI group contained malignant disease whereas viable carcinoma was found in 5 of 14 (26.4%) patients in the PVB group. The toxicity of the BOMP/EPI regimen was severe but tolerable. Intensive chemotherapy regimen seems to be useful in this subset of patients, but randomised prospective trials comparing these with standard chemotherapy are necessary.


Male impotence: a possible beta-adrenergic dysfunction in some patients. In some patients nocturnal penile tumescence (NPT) is abnormal, while the others diagnostic evaluations and medical history do not indicate any specific organic cause of impotence. Aim of this study was to evaluate heart rate variability during sleep in impotent patients with any demonstrable organic cause of impotence. We studied 50 patients: 26 were psychogenic and 24 organic on the basis of NPT and nocturnal rigidity. A lower orthosympathetic activity has been found in organic patients. Because beta-adrenergic receptors mediate relaxation of corpus cavernosum, it could be hypothesized a penile beta-adrenergic dysfunction in our organic patients.


Late-onset congenital adrenal hyperplasia in women with hirsutism. Hirsutism in women is a clinical manifestation of excessive production of androgens. The source of the excess androgen may be either the ovaries or the adrenal glands, or increased peripheral conversion of weak androgenic hormones to more potent androgens. Late-onset (non-classic) congenital adrenal hyperplasia is a cause of hirsutism in adult women, but its frequency and the patterns of abnormalities in adrenal hormone secretion are not well understood. The frequency of non-classical adrenal hyperplasia due to deficiencies of 3 beta-hydroxy-delta 5-steroid dehydrogenase, 21-hydroxylase, and 11 beta-hydroxylase among 65 women with hirsutism were determined. All enzyme defects were identified by comparing the patients' hormonal responses to 0.25 mg intravenous bolus of alpha 1-24-ACTH with those of age-matched normal women. The hormones measured in plasma during the ACTH stimulation tests were progesterone, 17-hydroxypregnenolone, 17-hydroxyprogesterone, DHEA-sulfate, androstenedione, testosterone, 11-deoxycortisol, and cortisol. Similarly these hormones were measured after overnight 1 mg oral dexamethasone. Twelve women (18.5%) had 3 beta-hydroxy-delta 5-steroid dehydrogenase deficiency, 24 (37%) 21-hydroxylase deficiency, and 14 (21.5%) 11 beta-hydroxylase deficiency. Women with 21-hydroxylase deficiency also had evidence of a partial deficiency in 11 beta-hydroxylase activity (12 of the 24 patients). Similarly, most (11 of the 14) of the women with 11 beta-hydroxylase deficiency also had evidence of a deficiency in 3 beta-hydroxy-delta 5-steroid dehydrogenase activity. Among the 15 patients with no adrenal biosynthetic defect, eight had high plasma androgen concentrations, and seven had normal concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)


Haemodynamic and renal effects of urodilatin bolus injections in patients with congestive heart failure. Urodilatin (ANF(95-126)) is an analogue of the atrial natriuretic factor (ANF(99-126)), which has been isolated from human urine. Recently we have shown in healthy volunteers, that intravenous bolus injections of synthetic urodilatin produce more pronounced reductions of pulmonary arterial pressure than ANF(99-126). To compare haemodynamic and renal effects of synthetic urodilatin with those of ANF(99-126) in congestive heart failure (CHF), 12 patients (66.3 +/- 1.4 years) received either two high dose intravenous bolus injections of 4 micrograms kg-1 bw Urodilatin (URO) at a 30 min interval (n = 6) or the same doses of ANF(99-126) (n = 6). Prior to i.v. URO, no URO immunoreactivity was found in human plasma (specific RIA, no crossreactivity to ANF). Similar to ANF, the increase in diuresis (1.4 +/- 0.7 to 3.7 +/- 1.6 ml min-1) and natriuresis (169 +/- 114 to 430 +/- 197 mumol min-1) was moderate after URO in CHF. During the 90 min study period, mean plasma cyclic GMP levels increased much more after URO (by 53.4 +/- 15.1 nM) than after ANF (by 13.1 +/- 3.0 nM; P = 0.04). In contrast to ANF, i.v. bolus injections of URO produced sustained haemodynamic effects in CHF lasting up to 90 min: The average (0-90 min) reduction of systemic vascular resistance was more pronounced after URO (-578 +/- 148) than after ANF (-204 +/- 65 dyn*s*cm-5, P = 0.04).(ABSTRACT TRUNCATED AT 250 WORDS)


Transforming growth factor alpha and atrial natriuretic peptide in white adipose tissue depots in rats. To detect the presence in adipose tissue of peptides known to affect tissue growth and to investigate potential regional differences, epididymal and perirenal adipose tissue depots from male Sprague-Dawley rats were separated into adipocyte and stroma-vascular fractions by collagenase digestion, sequential centrifugation and filtration. Identity and integrity of the fractions were demonstrated by light and electron microscopy, while dose-response curves for angiotensin-converting enzyme (ACE) were performed, revealing maintained functional capacity of the stroma-vascular fraction. ACE, atrial natriuretic peptide (ANP), and transforming growth factor-alpha (TGF-alpha) concentrations were significantly greater in epididymal than perirenal stroma-vascular tissue. Adipocyte fractions from both depots contained significant concentrations of ANP and TGF-alpha. There was no detectable ACE in the adipocyte fractions, indicating that no contaminating stromal-vascular cells were present in these fractions. These data show significant concentrations of peptides with effects on growth in subfractions of adipose tissue and demonstrate regional differences in concentrations between fat depots.


Pancreatic and pituitary hormonal responses to insulin-induced hypoglycaemia during muscarinic cholinergic blockade in man. To investigate the role of muscarinic cholinergic mechanisms in mediating the pancreatic and pituitary hormonal responses to hypoglycaemia, six normal subjects were studied during acute insulin-induced hypoglycaemia under control conditions, and during blockade with intravenous atropine. During atropine blockade the response of pancreatic polypeptide was suppressed while the maximum response of plasma glucagon was significantly higher. The increment in plasma vasopressin was also increased significantly during cholinergic blockade. During blockade with atropine the responses of plasma prolactin was reduced, with a slight but significant reduction in the growth hormone response, and although a similar maximum response of plasma ACTH was achieved, this rise was delayed. These results implicate involvement of a cholinergic muscarinic inhibitory and stimulatory mechanisms in regulating the responses of pancreatic and pituitary hormones to hypoglycaemia.


Experimental variations in renovascular resistance in normal man as detected by means of ultrasound. The pulsatility index (PI) of blood flow velocities has been reported to vary with changes in peripheral vascular resistance. Since blood flow velocities can easily be detected with the Echo-Color-Doppler technique in interlobar arteries of normally positioned kidneys, we tried in six healthy volunteers to estimate pharmacological induced variations in renal plasma flow (RPF) and renovascular resistance (RVR) by means of PI measurements. In this study no significant correlation between the absolute values of PI and RVR was found. In order to correct PI for different blood pressure-inputs to the renal artery, PI was divided by the pulsatility of the systemic arterial pressure, i.e. the 'blood pressure index' [BPI = (SAD-DAP)/MAP)], resulting in the 'velocity blood-pressure index' (VBI = PI/BPI), which was significantly correlated with RVR (r = 0.54, P less than 0.01). The pharmacological induced changes of RPF and RVR (delta RPF, delta RVR) were also correlated to the respective changes of PI and VBI (delta PI, delta VBI), with the highest significance when delta VBI was plotted against delta RVR (r = 0.83, P less than 0.0001). VBI, i.e. the pulsatility index of blood flow velocities as corrected for the pulsatility of the driving force, may be a tool for noninvasive assessment of changes in RVR and thereby of importance for the diagnosis and follow-up of renovascular diseases.


Metabolic and neuroendocrine effects of pro-inflammatory cytokines. Immune-neuroendocrine interactions occur during physiological and pathological situations. Pro-inflammatory cytokines such as IL-1, IL-6 and TNF alpha play a role in mediating these interactions. Although all three cytokines can stimulate ACTH and glucocorticoid output, IL-1 has the highest potency. It is known that increased glucocorticoid levels result in hyperglycemia. However, administration of low doses of lipopolysaccharide (LPS), an inducer of several cytokines including those mentioned above, causes a profound and long lasting hypoglycaemia. This effect seems to be dissociable from that of insulin, since the same effect was observed in insulin-resistant db/db mice. The data reported here show that IL-1 plays a crucial rôle in the mediation of the hypoglycaemia induced by LPS, since other cytokines released following inoculation of endotoxin, such as TNF alpha and IL-6, have only marginal effects or do not induce hypoglycaemia when administered in doses similar to those of IL-1. The effect of IL-1 seems to be integrated at least in part at CNS level since i.c.v. administration produces hypoglycaemia in spite of the concomitant release of corticosterone. The data reported here reinforce the concept that IL-1 and related cytokines participate in the mediation of immune-neuroendocrine interactions.


Beta-adrenergic receptors in human leukocyte subpopulations. beta 2-Adrenergic receptors, which are encoded on chromosome 5q32-34, belong to the group of G-protein-linked hormone receptors with seven transmembrane domains. Upon agonist binding, adenylate cyclase is activated. Although the function of human leukocyte beta-adrenergic receptors is unknown, these cells are often used as a model system to study tissue beta-adrenergic receptors. In intact mononuclear leukocytes or membrane preparations, 1000-3000 beta 2-adrenergic receptors are found, having an antagonist affinity constant (KD) in the range of 25 pM. beta-Adrenergic receptor numbers are different in leukocyte subsets, with receptor density higher in B than in T cells. CD56+ or CD57+ natural killer cells express more receptors than CD8+ or CD4+ cells. KD is higher in CD8+ than in CD4+ cells. Acute sympathetic activation by isoproterenol infusion or short-lasting exercise leads to an increased number of mononuclear beta-adrenergic receptors with a slightly reduced proportion of those with high agonist affinity. Acute sympathetic activation by adrenaline infusion, short-term exercise, or psychological stress also causes a selective increase in circulating CD56+ or CD57+ lymphocytes which are rich in beta-adrenergic receptors. The results of several studies suggest that adrenaline-induced changes in beta-adrenergic receptors and the redistribution of leukocyte subsets may be linked. beta-Adrenergic receptors may mediate immuno-modulatory effects by causing selective cell mobilization.


Beta 2-adrenergic receptors on peripheral blood mononuclear cells in patients with rheumatic diseases. In order to investigate the influence of the autonomic nervous system on the immune response in rheumatic diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), we determined the number and the dissociation constants of beta 2-adrenergic receptors (beta 2R) on peripheral blood mononuclear cells (PBMC) in patients with RA characterized by either low or high disease activity, patients with SLE, and healthy age-matched controls by means of a radioligand binding assay with [125I]iodocyanopindolol. The number of beta 2R was significantly decreased in all three patient groups as compared with the healthy controls. In SLE patients, a significant negative correlation between beta 2R and the anti-ds-DNA antibody titre was observed (r = -0.57, P < 0.01). These data demonstrate the close correlation between the number of beta 2R on PBMC and the extent of disease activity in patients with RA and SLE and suggest an involvement of the autonomic nervous system in the pathogenesis of rheumatic disorders.


Correlation between density of beta 2-adrenergic receptors on peripheral blood mononuclear cells and serum levels of soluble interleukin-2 receptors in patients with chronic inflammatory diseases. In order to study the influence of the autonomic nervous system on the immune response, we evaluated the density and the equilibrium dissociation constants (KD) of beta 2-adrenergic receptors (beta 2R) on peripheral blood mononuclear cells (PBMC) in patients with rheumatoid arthritis (RA), Crohn's disease (CD), and in controls. Results were correlated with the serum concentrations of soluble interleukin-2 receptors (sIL-2R) as a marker for T-cell activation in vivo. The density of beta 2R was significantly decreased in patients with RA (P < 0.05) and CD (P < 0.05) as compared with controls. The number of beta 2R in patients with RA was significantly lower than in CD patients (P < 0.05). KD values of beta 2R were markedly but not significantly decreased in both patient groups as compared with control values. Serum concentrations of sIL-2R were significantly elevated in RA patients as compared with those in CD patients (P < 0.05) and controls (P < 0.05), while there was no difference between the latter two groups. In patients with RA, a significant negative correlation between beta 2R density and serum IL-2R levels (r = -0.66, P < 0.02) was observed. These results demonstrate the close correlation between the modulation of beta 2R on PBMC and the activation of the immune response. However, the role of beta 2R stimulation in the pathogenesis of immunologically mediated diseases remains to be clarified.


Repeated cocaine modifies the neuroendocrine responses to the 5-HT1C/5-HT2 receptor agonist DOI. Endocrine responses to the serotonin (5-HT) 5-HT1C/5-HT2 agonist (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) were utilised to evaluate cocaine-induced alterations in postsynaptic 5-HT receptor function. Rats received cocaine HCl (0, 5 or 15 mg/kg i.p.) twice daily for 7 days. Effects of DOI (0, 0.5, 2 or 10 mg/kg i.p.) on plasma adrenocorticotropic hormone (ACTH), corticosterone, prolactin, oxytocin and renin concentrations were assessed 42 h after the final cocaine injection. DOI dose dependently increased the plasma concentrations of each hormone. Cocaine potentiated the DOI-induced elevations of plasma ACTH, corticosterone and prolactin concentrations. In contrast, the oxytocin response was reduced, and the renin response was unaltered by cocaine exposure. The data suggest that 5-HT2 receptor-mediated responses for ACTH, corticosterone and prolactin secretion become supersensitive following repeated cocaine. In contrast, the 5-HT2 receptor-mediated response for oxytocin secretion is subsensitive. The cocaine-induced changes in postsynaptic 5-HT receptor function are likely a consequence of deficits in the function of 5-HT nerve terminals, that we have documented previously.


Dihydroergotoxine modulation of the GABAA receptor-associated Cl- ionophore in mouse brain. Dihydroergotoxine non-competitively displaced the binding of t-[3H]butylbicycloorthobenzoate ([3H]TBOB) to crude synaptosomal membranes from the mouse brain (cerebrum minus cortex), and gamma-aminobutyric acid (GABA) (10 microM) enhanced the displacement potency of dihydroergotoxine in a bicuculline-sensitive manner. The same ergot compound prolonged pentobarbital-induced sleeping in mice and diminished the convulsive potency of picrotoxin in the same animal species. The results are indicative of the positive coupling between GABA and dihydroergotoxine.