Comparison between the thyrotropin response to thyrotropin-releasing hormone in summer and that in winter in normal subjects. A comparison was made between the thyrotropin (TSH) response to 500 microgram thyrotropin-releasing hormone (TRH) in summer and that in winter in ten healthy normal adults living in Supporo. The serum resin triiodothyronine (T3) uptake (RT3U), thyroxine (T4) and T3 levels were also measured. While the TSH response to TRH in summer was similar to that in winter, serum T3 concentration and free T3 index were significantly higher in winter than in summer, associated with the similar values in RT3U and T4 levels in serum. Independently measured 86 specimens (43 in summer and 43 in winter) from normal adults living in the same district also showed a significant increase in serum free T3 index as well as a slight elevation of serum T3 concentration in winter but not in serum T4 level. These results indicate that the primary change in cold winter would be the stimulation of peripheral conversion of T4 to T3 rather than the activation of hypothalamo-pituitary-thyroid axis. The relevance of this interpretation was discussed.


Effect of active and passive immunization with TRH on plasma TSH response to propylthiouracil. The role of thyrotropin-releasing hormone (TRH) in the secretion of TSH from the anterior pituitary was investigated in rats by active and passive immunization with TRH. The plasma TSH response to propylthiouracil (PTU) in TRH-bovine serum albumin (BSA)-immunized rats was significantly lower than that of BSA-immunized or non-immunized rats. Similarly, the increased plasma TSH level following PTU treatment was significantly suppressed after iv injection of antiserum to TRH. However, the decline in plasma TSH levels was not complete. The results of the present study indicate, at least in part, the physiological significance of endogenous TRH in the regulation of pituitary TSH secretion.


Cerebellar stimulation in alumina-gel monkey model: inverse relationship between clinical seizures and EEG interictal bursts. The efficacy of cerebellar stimulation was addressed in a chronic monkey model (N = 12) of spontaneous focal motor and secondarily generalized seizures using 24 hr seizure frequency monitoring and all-night EEG recording. The anterior cerebellar vermis was stimulated employing parameters similar to those used in man, 10 Hz, 1 msec pulses, 10 min on, 10 off, at an average current of 2.0 mA. Six weeks pre- and post-base-line periods were compared to a stimulation period of the same length. The results contribute to a clarification of conflicting findings of previous researchers by revealing an inverse relationship between seizure frequency and interictal EEG bursts during the weeks of stimulation. Seizure frequency increased significantly and interictal bursts decreased. Both of these effects (especially the former) were evident in the post-stimulation period, but for different reasons than hypothesized for the period of stimulation. Whereas the therapeutic value of cerebellar stimulation on seizures may be in question, its utilization in the study of mechanisms of epilepsy may be warranted.


Pharmacokinetic properties of thiopental in two patients treated for uncontrollable seizures. Thiopental was administered for seizure control in 2 patients with uncontrollable seizures. Serum samples were collected from each patient and assayed for thiopental, and the resulting serum concentration--time data were analyzed pharmacokinetically. The biologic half-life in both patients was significantly longer than previously reported values. Based on the limited number of patients studied, it would appear that half-life and volume of distribution increase with the degree of obesity, while clearance remains unchanged. These pharmacokinetic characteristics would be worthy of consideration in cases where there may be prolonged use of thiopental, eg., for the control of uncontrollable seizures.


Behavioral toxicology of carbon disulfide and toluene. Organic solvents are pervasive in the communal and industrial environments. Although many are potent central nervous system agents, clearly delineated behavioral effects have played only a minor role in the formation of exposure standards. A comprehensive behavioral pharmacology and toxicology of these compounds is one aim of US/USSR collaboration. The current report describes some actions of carbon disulfide and toulene. Earlier data about the actions of carbon disulfide on pigeon operant performance indicated disruption of schedule-controlled key-pecking. Primate data are now described from a situation designed to determine aversive thresholds to electrical stimulation. Effective concentrations of carbon disulfide produced both a rise in the amount of electric shock tolerated and a diminution of the response force exerted by the monkeys. In experiments with toluene, pigeons were shown to elevate key-pecking rate in an operant situation at certain concentrations. Toluene also was studied for its capacity to maintain self-administration in the same way as drugs of abuse. Monkeys worked to gain access to toulene vapor just as they work for opiates or amphetamines. The current experiments demonstrate how comprehensive the range of behavioral toxicology needs to be to deal with environmental health issues.


Urethral diverticulum with lithiasis.

Urethral diverticulum with lithiasis. Urethral diverticulum, either congenital or acquired, is very rare and becomes symptomatic when complicated by infection and lithiasis. This paper reports on such a case that was cured by radical excision of the lesion.


Abnormal immunoglobulins in severe combined immunodeficiency: analysis by immunoelectrophoresis and SDS-polyacrylamide gel electrophoresis. The plasma immunoglobulins of patients with severe combined immunodeficiency were studied by immunoelectrophoresis and, following isolation by affinity chromatography, by SDS-polyacrylamide gel electrophoresis. Immunoglobulins in plasma from the eight patients studied were immunoelectrophoretically abnormal. Although certain of the immunoglobulins in plasma from five patients could not be identified antigenically, all possessed two mu determinant-bearing proteins with abnormally fast electrophoretic mobilities. Molecular analysis of immunoglobulins of three of these patients revealed two mu heavy chains of abnormally low molecular weight which lacked the ability to polymerize into the pentameric structure of IgM. The failure of concanavalin A to precipitate these molecules suggests that they lack the carbohydrate moiety of normal IgM. Using these techniques, we documented the acquisition of normal IgM synthesis by a patient grafted with maternal leukocytes and the partial immunologic development of a child maintained under gnotobiotic conditions. In the latter patient, between the age of 1 and 4 years, an abnormal mu component disappeared from plasma and normal IgM appeared.


Comparison of rat, mouse, guinea-pig and human slow reacting substance of anaphylaxis (SRS-A). Slow reacting substance of anaphylaxis obtained from rat, mouse, guinea-pig and human tissues have exhibited similar biological activity and have reacted in the same way to chemical and enzymatic treatments. It is concluded that they appear to be the same substance or a similar class of compounds.


Ethanol metabolism in Drosophila melanogaster. A quantitative study of the transformation of ethanol into acetaldehyde shows that, in Drosophila melanogaster, the mitochondrial ethanol oxidizing system is not very active but that the part played by catalase appears more important than expected. For a strain without alcoholdehydrogenase, ethanol is highly toxic. The presence of acetaldehyde in the culture medium is toxic for all the strains studied. But, since even a strain without any aldehydeoxidase lives normally, the metabolic production of acetaldehyde does not seem dangerous.


alpha-Amanitin: inactivation by bovine lactoperoxidase. The principle amatoxin, alpha-amanitin, is found to be extremely sensitive toward lactoperoxidase catalyzed degradation, rather than iodination, of the indole nucleus. Extensive attenuation of inhibitor potency against eukaryotic DNA-dependent RNA polymerase II accompanies the treatment of alpha-amanitin with lactoperoxidase, iodide and hydrogen peroxide.


Position effect influencing alcohol dehydrogenase activity in Drosophila melanogaster. A small decrease in alcohol dehydrogenase activity was found in translocation stocks of Drosophila melanogaster in which a section of the heterochromatic Y-chromosome is inserted proximal to Adh. This small position effect is consistent with our growing knowledge of the control and transcription of the Adh locus.


The distribution of carcinogens, 4-nitroquinoline-1-oxide and 4-hydroxyaminoquinoline-1-oxide, in the nervous system and its possible neurotoxicological significance. 4-NQO-14C can enter the grey matter parenchyma of the central nervous system of mice after i.v. injection. The level of its uptake by the central grey is higher than that taken up by the central white and by the trigeminal and spinal dorsal root ganglia. This pattern of distribution is strikingly different from that obtained after i.v. injection of 4-HAQO-14C, suggesting the possible occurrence of 4-NQO encephalomyelopathy having entirely different sites of lesions from those of 4-HAQO neuropathy.


Biochemical changes in avian tissues during infection. Major sources of error in studies of diet--infectious disease interactions relate to failure 1) to define the stage of a disease cycle; and 2) to determine the extent of disease involvement at time of sampling. The former can be determined from clinical and biochemical observations from time of inoculation to recovery or mortality; the latter can be calculated from indexes obtained from histological preparations, changes in body temperatures, and other clinical symptoms, including weight loss and efficiencies of feed utilization. Other significant errors are derived from the normal 24-hour dynamics of particular tissue constituents, which include oscillations and circadian rhythms. Experimental designs and accuracy of data are always helped by prior knowledge of 24-hour patterns of these normal fluctuations. For example, such patterns will reveal that enzyme and hormonal changes are far more dynamic than body water changes. Studies of nutrition--disease interactions are extremely complex and the researcher must be aware of and eliminate as many sources of experimental error as possible to avoid confounding the data.


The incidence of complications during pregnancy after treatment of hyperprolactinemia with bromocriptine in patients with radiologically evident pituitary tumors. The course of pregnancy achieved after bromocriptine therapy is described in nine patients with radiologically evident prolactin-secreting pituitary tumors. In six patients no complications occurred. No changes in sellar size or secondary endocrine deficiencies developed. In three patients, however, complications developed between the 22nd and 24th weeks of pregnancy. Despite prior external pituitary irradiation, one patient developed transient bitemporal hemianopsia and one patient had apoplexy of the pituitary tumor with transient paresis of the left abducens nerve. A third patient developed parasellar expansion of the pituitary tumor with bone destruction and paresis of the right abducens and oculomotor nerves. After transsphenoidal surgery the paresis of both nerves disappeared. Microscopically, the tissue removed at surgery was a chromophobe adenoma with focal fibrosis and calcifications without recent hemorrhages. In the course of more than 100 pregnancies achieved in The Netherlands after bromocriptine therapy, five patients reportedly developed complications of the pituitary tumor. At present, patients in whom complications can be expected cannot be predicted by the size or configuration of the sella turicica or the magnitude of elevation of the plasma prolactin level. In two patients external pituitary irradiation did not prevent complications during pregnancy.


Representation of heavy but not light chain Ig idiotypes on T cell receptors for alloantigens. We have analyzed idiotypic determinants on B and T lymphocytes reactive against the same antigenic determinants by the use of different anti-idiotypic antibodies. Such antisera were produced in (Lewis X DA) F1 rats against Lewis anti-DA alloantibodies (= B cell product) and Lewis T lymphocyte receptors with the same specificity. We found that B lymphocytes bear unique idiotypic determinants which are not present on the corresponding T lymphocytes. T cell unique (not shared by B lymphocytes) idiotypes were so far not detected. T cells idiotypic determinants which are present on heavy but not light chains of the corresponding alloantibodies.


Isolation and characterization of specific rat epididymal proteins. The partial purification and characterization of specific rat epididymal proteins (SEP) is reported. Starting from the cytosol fraction obtained from epididymal homogenates, protein C was purified 15-fold and proteins D--E were purified 19-fold. The molecular weight, determined by molecular sieving, of protein C was 22,400 while that of D--E was 37,000. These proteins stained as glycoproteins with periodic acid--Schiff reagent. The isoelectric point of protein D was 5.13 while that of protein E was 4.95. Protein C separated into 3 bands during isoelectric focussing. The major component focussed at 5.56 and the two minor components at pH 5.38 And 5.79. Using a specific antiserum we could confirm the organ specificity of SEP and their androgen-dependence.


Hormonal regulation in two rat mammary cancer cell lines: glucocorticoid and androgen receptors. We have studied the hormonal control of two continuous cell lines derived from rat-mammary tumors induced by two different carcinogens (dimethylbenzanthracene and N-nitrosomethylurea). The steroid receptors were assayed using charcoal or hydroxyapatite and the effect of different hormones on cell growth was evaluated by measuring total DNA and following the growth of transplanted cells in nude mice. Similar results were found for the two cell lines; they were unresponsive to estrogen since they did not contain appreciable amounts of estradiol receptor. Conversely, these two cell lines contained high concentrations of androgen and glucocorticoid receptors, which were both transferable to the nucleus. While dexamethasone stimulated, directly or indirectly, the growth of RBA and NMU cells, the effect of androgens on cell growth is still questioned. These two cell lines offer a potential model to study the mechanism of action of androgens and glucocorticoids in mammary cancer.


Radioimmunoassay of glibenclamide.

Radioimmunoassay of glibenclamide. A specific, sensitive, and simple radioimmunoassay was developed for the oral hypoglycemic drug glibenclamide, N-4-[2-(5-chloro-2-methoxy-benzamide)-ethyl]-benzenesulfonyl-N'-cyclohexylurea. Antiserum against glibenclamide was obtained from rabbits immunized with an antigen prepared by conjugating the diazonium salt of N(p-amino-benzamidoethyl)-benzenesulfonyl-N'-cyclohexylurea to bovine serum albumin through the diazocoupling. [3H]glibenclamide was used as a tracer. Dextran-coated charcoal was used to separate bound and free [3H]glibenclamide in the reaction mixture. The radioimmunoassay is able to determine as little as 25 pg of glibenclamide directly in plasma without the need for extraction. The antiserum used for the assay was highly specific for glibenclamide, and did not cross-react with two known major metabolites of glibenclamide. Comparable values of glibenclamide in dog plasma were obtained by radioimmunoassay and liquid chromatography. Plasma concentrations of glibenclamide in diabetic patients on glibenclamide treatment can be determined by radioimmunoassay, and the method has been applied to the routine assay of clinical samples. This radioimmunoassay seems to be useful for monitoring plasma glibenclamide concentrations.


Ultracytochemical calcium distribution in B cells in relation to biphasic glucose-stimulated insulin release by the perfused rat pancreas. Calcium distribution in B cells of the isolated perfused rat pancreas was examined by the pyroantimonate precipitation technique in relation to the insulin secretory pattern of the perfused pancreas in response to 3 mM or 20 mM D-glucose or 20 mM D-glucose in calcium-depleted ethylene glycol tetra-acetic acid (EGTA) medium. Perfusion fixation after various time intervals from 3 to 30 min allowed appropriate relation to secretory phases. Qualitative and quantitative evaluation of the precipitation patterns revealed a significant increase in cell membrane associated percipitates after 3--5 min of perfusion with 20 mM glucose compared with the results after perfusion with 3 mM glucose. After 10--30 min of perfusion with 20 mM glucose there was an additional significant increase in precipitates located in the cytoplasm and the halos of the secretory granules. Perfusion with 20 mM glucose in calcium-deprived EGTA medium strongly reduced the number of precipitates within the B cells. The results suggest that cell membrane associated calcium may be involved in exocytosis, and by its sudden increase may trigger the first phase of insulin secretion. The calcium stores in the cytoplasm and the granules may be of importance for long-term regulation of insulin release.


Insulin resistance caused by massive degradation of subcutaneous insulin. Severe resistance to subcutaneous insulin but sensitivity to intravenous insulin persisted for 15 months in a 17-year-old diabetic girl. Heat-labile insulin-degrading activity was present in the patient's ketotic sera and in the 100,000 g fraction (soluble fraction) of adipose tissue. Serum-degrading activity was not inhibited by N-ethylmaleimide. The soluble fraction also degraded glucagon and B chain but not growth hormone or myoglobin. It was inhibited by incubation with the patient's nonketotic sera, normal sera, or Trasylol. Glutathione-insulin-transhydrogenase (GIT) activity was 66% of normal. The biopsy of adipose tissue at remission showed a normal level of insulin- and glucagon-degrading activity. The activity was eluted from Sephadex G200 as a single peak and had properties consistent with those of the insulin-specific protease (ISP). The increased degrading activity present during insulin resistance had properties not shared with ISP, suggesting the presence of an uncharacterized protease.


Prophylactic effect of BCG cell-wall skeleton on the tumor induction by 7,12-dimethylbenz[a]anthracene in mice: strain difference. Prophylactic effect of repeated intravenous administrations of oil-attached BCG cell-wall skeleton (BCG-CWS) on the induction of tumor by 7,12-dimethylbenz[a]anthracene (DMBA) was investigated in various strains of mice. The subcutaneous injection of DMBA emulsified in oil induced squamous cell carcinoma in almost all of the strains of mice. Treatment of C57BL/6, BALB/c, and ddO strains with BCG-CWS with appropriate route and timing resulted in the retardation of DMBA-induced tumor development manifested by a prolonged latent period of tumor outgrowth. In contrast, the same BCG-CWS treatment of C3H/He and BTK mice was incapable in preventing such DMBA-induced carcinogenesis. Thus, the treatment with BCG-CWS was effective for preventing the DMBA-induced carcinogenesis in certain strains of mice, but the effectiveness varied depending on the strain. The implication of such a strain variationof the BCG-CWS effect on the prophylaxis of chemical carcinogenesis was discussed in the context of differences in the magnitude of immunopotentiation of the host by BCG-CWS.


Detection of HBe antigen in sera from HBs antigen asymptomatic carrier and hepatitis patients using polyethylene glycol (PEG). Serum and plasma samples concentrated 8 to 10 times with polyethylene glycol (PEG) having a molecular weight of 6,000 were examined by micro-Ouchterlony (MO) analysis with a view to increasing the detection sensitivity for HBe antigen (HBeAg), one of the hepatitis B virus associated antigens, and HBe antibody (HBeAb). The subjects of this investigation consisted of 82 symptom-free HBsAg carriers and 59 patients with B hepatitis. HBeAg was detected in 22 (26.8%) and HBeAb in 43 (52.4%) of 82 asymptomatic HBsAg carriers, and 17 (20.7%) were negative for both HBeAg and HBeAb. The corresponding values for the liver disease patients were 7 (11.9%), 16 (22.1%) and 36 (61.0%). Histologically, the rate of detection for HBeAg was higher in the cases of a mild disturbance.


Effect of chronic alcohol use on hepatic testosterone 5-alpha-A-ring reductase in the baboon and in the human being. Hepatic testosterone 5-alpha-A-ring reductase (HTAR) activity was measured in open liver biopsies in eight alcohol-fed baboons and eight pair-fed controls. The animals were studied after at least 1 yr of alcohol feeding. In the alcholol-fed animals, a significant fall in enzyme activity was noted. This occurred whether the enzyme levels were related to soluble protein, to DNA, or to wet tissue weight, showing that the change was due to a decrease in the specific activity of the enzyme. In addition, aspiration liver biopsy specimens were obtained from 14 men and women with alcoholic liver disease. Again, there was a significant decrease in HTAH activity in these patients compared with a normal population. No relationship was found between hepatic histology and HTAR levels in either the baboon or human population with alcoholic liver disease, suggesting that the changes in enzyme activity were related to an alcohol effect rather than to liver disease per se. This study demonstrates that chronic alcohol use decreases the function of the enzyme which controls an important rate-limiting step in the metabolism of testosterone in the liver and that this effect may be due primarily to alcohol.


Effect of dietary fiber on intraluminal pressure and myoelectrical activity of left colon in monkeys. The effects of varying intake of dietary fiber content on left colon intraluminal pressure, electrical spike discharge activity, and contractions of circular muscle were measured in stump-tailed monkeys. Animals were preconditioned to sit in restraint chairs and then had the sigmoid colon instrumented with an extraluminal strain gage force transducer, a bipolar serosal electrode and a Thomas cannula to provide access for determining luminal pressure with a perfused, open-tip catheter. Diets containing 0, 5, 10, 15, and 20 g of dietary fiber daily were fed in varying sequences for periods of 3 wk. Observations were made twice during the last week of each diet. Fecal weight increased in animals on a high fiber diet, but changing dietary fiber content did not alter the duration of spike discharge activity and produced only a minimal change in the contraciton frequency of colon circular muscle. Colon intraluminal pressure was increased significantly by feeding low-fiber diets, but intraluminal pressure responses did not always correlate with the myoelectrical activity of the colon circular muscle.


Effect of fasting on metabolite-mediated hepatotoxicity in the rat. Acetaminophen and bromobenzene are transformed in the liver into chemically reactive metabolites that may either bind to glutathione and be detoxified or bind to hepatic proteins and produce liver cell necrosis. Fasting for 42 hr (a) decreased hepatic glutathione concentration, (b) increased the amount of chemically reactive metabolite irreversibly bound to hepatic proteins after administration of 3H-acetaminophen or 14C-bromobenzene, and (c) increased the hepatotoxicity of acetaminophen or bromobenzene. In rats fasted for various lengths of time, there was an inverse relationship between the concentration of glutathione in the liver and the activity of serum glutamic pyruvic transaminases after administration of acetaminophen or bromobenzene. In vitro, there was an inverse relationship between the concentration of glutathione in the incubate and the amount of chemically reactive metabolite bound to microsomal proteins after incubation of 3H-acetaminophen or 14C-bromobenzene with hepatic microsomes. It is concluded that fasting may decrease the inactivation of chemically reactive metabolites by glutathione, increase their binding to hepatic proteins, and enhance the hepatotoxicity of drugs transformed into chemically reactive metabolites that are detoxified by binding to glutathione.


Home parenteral nutrition in children with Crohn's disease: an effective management alternative. Seventeen pediatric patients, ages 9.25--20.5 yr, were placed on a program of home parenteral nutrition (HPN) for severe, symptomatic Crohn's disease. Prior therapy with sulfasalazine in 14, adrenocorticosteroids in 12, inpatient total parenteral nutrition in 7, and/or surgical resections in 6 failed to suppress disease activity. Remission was attained in 12 of the 17 after one course of HPN alone. Four patients had surgical procedures and 1 required steroids in addition to HPN. Remissions have been maintained in 4 of those 12 for a mean duration of 315 days after discontinuation of HPN. Of the 8 who relapsed after a mean duration of 68 days, second courses of HPN were undertaken in 7 and third courses in 2. All 17 had a marked improvement in disease symptoms while receiving HPN in addition to gaining weight sufficient to place them at a higher percentile on standard growth charts. Ten patients demonstrated "catch-up" growth and 4 others increased their height appropriately. A chromium 51-labeled albumin stool collection of greater than 1% in 5 of 7 patients at the completion of a HPN course correlated with relapse within 4 mo. Serial radiographic contrast studies and erythrocyte sedimentation rates were not predictive of prolonged remissions. Home parenteral nutrition complications were minimal, with only one episode of sepsis per 5.8 catheter experience years. It is concluded that HPN is a safe and effective means of inducing remissions and providing optimal nutritional support in pediatric patients with severe Crohn's disease. Patients with less than adequate response to standard medical management should be considered candidates for this therapeutic modality.


Lack of spontaneous sister chromatid exchanges in somatic cells of Drosophila melanogaster. Neural ganglia of wild type third-instar larvae of Drosophila melanogaster were incubated for 13 hours at various concentrations of BUdR (1, 3, 9, 27 micrograms/ml). Metaphases were collected with colchicine, stained with Hoechst 33258, and scored under a fluorescence microscope. Metaphases in which the sister chromatids were clearly differentiated were scored for the presence of sister-chromatid exchanges (SCEs). At the lowest concentration of BUdR (1 microgram/ml), no SCEs were observed in either male or female neuroblasts. The SCEs were found at the higher concentrations of BUdR (3, 9, And 27 micrograms/ml) and with a greater frequency in females than in males. Therefore SCEs are not a spontaneous phenomenon in D. melanogaster, but are induced by BUdR incorporated in the DNA. A striking nonrandomness was found in the distribution of SCEs along the chromosomes. More than a third of the SCEs were clustered in the junctions between euchromatin and heterochromatin. The remaining SCEs were preferentially localized within the heterochromatic regions of the X chromosome and the autosomes and primarily on the entirely heterochromatic Y chromosome.--In order to find an alternative way of measuring the frequency of SCEs in the Drosophila neuroblasts, the occurrence of double dicentric rings was studied in two stocks carrying monocentric ring-X chromosomes. One ring chromosome, C(1)TR94--2, shows a rate of dicentric ring formation corresponding to the frequency of SCEs observed in the BUdR-labelled rod chromosomes. The other ring studied, R(1)2, exhibits a frequency of SCEs higher than that observed with both C(1) TR94--2 and rod chromosomes.


Organization of the rosy locus in Drosophila melanogaster: further evidence in support of a cis-acting control element adjacent to the xanthine dehydrogenase structural element. The present report summarizes our recent progress in the genetic dissection of an elementary genetic unit in a higher organism, the rosy locus (ry:3--52.0) in Drosophila melanogaster. Pursuing the hypothesis that the rosy locus includes a noncoding control region, as well as a structural element coding for the xanthine dehydrogenase (XDH) peptide, experiments are described that characterize and map a rosy locus variant associated with much lower than normal levels of XDH activity. Experiments are described that fail to relate this phenotype to alteration in the structure of the XDH peptide, but clearly associate this character with variation in number of molecules of XDH per fly. Large-scale fine-structure recombination experiments locate the genetic basis for this variation in the number of molecules of XDH per fly to a site immediately to the left of the XDH structural element within a region previously designated as the XDH control element. Moreover, experiments clearly separate this "underproducer" variant site from a previously described "overproducer" site within the control region. Examination of enzyme activity in electrophoretic gels of appropriate heterozygous genotypes demonstrates the cis-acting nature of this variation in the number of molecules of XDH. A revision of the map of the rosy locus, structural and control elements is presented in the light of the additional mapping data now available.


A method for construction of specialized transducing phage rho 11 of Bacillus subtilis. DNA from a temperate phage rho 11 and chromosomal DNA of Bacillus subtilis 168 were digested with endonuclease EcoRI and then ligated with T4 polynucleotide ligase. The ligated DNA fragments were used to transform a lysogenic strain, B. subtilis spoA12 lys21 hisA1 leuA8 p11, and Lys+, His+ or Leu+ transformants were selected. The cells of each type were then mixed, grown and treated with mitomycin C; the induced phages were tested for abilities abilities to form plaques and to tranduce the auxotrophic marker. Various types of plaque-forming or defective phages which transduce hisA or lys marker at considerably high frequencies were thus obtained.


Studies on the organization of the alpha-satellite DNA from African green monkey cells using restriction nucleases and molecular cloning. alpha-Satellite DNA from African green monkey cells was analysed with restriction nucleases in some detail confirming and complementing our earlier results. With EcoRI and HaeIII (or BsuRI isoschizomer), about 25 and 10%, respectively, of the satellite DNA were cleaved into a series of fragments of the 172 bp repeat length and multiples thereof. To allow studies with fragments of homogeneous sequence unit length, HindIII fragments were covalently joined with the plasmid pBR 313. After transformation 19 clones were obtained, containing up to three monomer fragments. Nine of the clones were characterized by digestion with EcoRI. Three of these had cleavage sites for this nuclease in the satellite DNA portion. In the six clones tested with HaeIII no cleavage site was detected in the cloned DNA. The results are discussed in relation to the nucleotide sequence data recently published by Rosenberg et al. (1978) and in the context of random and nonrandom processes in satellite DNA evolution


Distribution of HLA histocompatibility antigens, ABO blood groups and Rh antigens in alcoholic liver disease. The distribution of 16 antigens of the HLA-A and 15 antigens of the HLA-B series of HLA system, the blood groups ABO, and Rh antigens were studied in 40 alcoholics with cirrhosis, 18 alcoholics without cirrhosis, and in normal control subjects. The group of alcoholics with cirrhosis showed a significantly high frequency of HLA-B13 (corrected P less than 0.01) when compared with normal subjects, while the frequency of HLA-B13 was similar to normal in alcoholics without cirrhosis. On the basis of these findings, its seems that the carriers of HLA-B13 are more susceptible to liver damage caused by alcohol. Both groups of alcoholics and the normal controls had a similar distribution of ABO blood groups and Rh antigens.


Encephalitozoonosis in laboratory animals--a serological survey. Serum samples of rabbits, guinea pigs, Syrian golden hamsters and mice from various breeding stations and laboratory colonies were examined for the presence of antibodies to Encephalitozoon cuniculi by the indirect fluorescent antibody test (IFAT). Of more than 500 rabbits, only those from commercial farms and laboratory colonies were found positive. The number of positive animals fluctuated in different colonies from 0% to 95%. 140 guinea pigs were examined. Two colonies were antibody-free, four colonies contained 29-85% of positively reacting animals. 110 Syrian golden hamsters from five colonies showed that in four colonies 14-80% of animals reacted positively and only one colony was negative. Of 126 rats examined, 15% and 30% of animals from two colonies contained significant antibody levels. Two colonies were negative. 200 mice from a number of colonies did not show any significant positivity. The significance of these results is discussed.


[Drug therapy of chronic arterial occlusive diseases of the lower extremities with butalamine-HCl]. In a multicentre study a total of 113 patients suffering from disturbed blood flow conditions of the lower extremities (type II Fontaine) were treated with butalamine hydrochloride tablets, each containing 80 mg of the active principle. In 96% of the cases given a daily dose of 3 x 2 tablets and in 82% of those treated with a daily dose of 3 x 1 tablet both subjective and objective clinical symptoms, such as walking distance and Ratschow's testing parameters, were found to be improved. In view of the findings that metabolic functions as well as systemic blood flow conditions were not affected and that the drug was furthermore well tolerated by the gastrointestinal system, butalamine hydrochloride proved to be an effective therapeutic in long-term treatment of patients suffering from blood flow disorders of the lower extremities.


Islet transplantation in experimental diabetes of the rat. VI. Rate of regression in diabetic kidney lesions after isogeneic islet transplantation: quantitative measurements. Intraportal transplantation of isogeneic adult islets in diabetic rats resulted in long-lasting amelioration of the metabolic disorder. The effect upon diabetes-induced kidney changes (enlargement of mesangial space, capillary changes, cell-proliferation) was examined quantitatively by morphometric studies. The major effect was a remarkable reduction of the mesangial space and re-widening of the capillary lumina. The number of endothelial cells was lowered. Regarding epithelial and mesangila cells no difference was observed between normal, diabetic and transplanted animals of age-matched groups.


Formation and metabolism of 3',5'-diiodothyronine and 3,5-diiodothyronine by cultured monkey hepatocarcinoma cells. Cultured monkey hepatocarcinoma cells were incubated with [3',5'-125I] diiodo-L--thyronine and with [3,5-125I] diiodo-L-thyronine. In both instances monodeiodination as well as sulfoconjugation took place. [3.-125I] iodothyronine and [3',5'-125I] diiodothyronine were identified as metabolites of [3'-5'-125I]-L-thyroxine in the cells, but neither [3-125I]-iodothyronine nor [3,5-125I] diiodothyronine was detected after incubation of the cells with ]3,5-125I]-L-thyroxine. No products of diphenyl ether splitting were observed in the medium after incubation of the cells with either [3,5-125I] diiodo-L-thyronine or [3,5-125I]-L-thyroxine.


Should hospitals use zero-base planning and budgeting? Since most hospital programs are not discretionary, they cannot be successfully evaluated by the "decision package" methodology of ZBPD. For carefully selected hospital services, however, ZBPD may be an effective tool.


Immunological nature of genetic resistance of mice to herpes simplex virus type 1 infection. Treatment of mice genetically resistant to HSV-1 with agents which impair T-cell or macrophage function markedly diminished resistance. Bone marrow transplantation of genetically susceptible mice with marrow from resistant F1 mice produced chimeras resistant to HSV-1. Resistance thus appears to be immunological in nature. Striking similarities were found between resistance to HSV-1 and allogeneic resistance.


Genetic control of resistance to Marek's disease. Various MD-resistant and MD-susceptibel lines of chickens were challenged with the RPL-1 cell line, an MD tumor cell line which is capable of growth both in vivo and invitro. Birds which possessed the B21 allele, an allele shown to be associated with MD resistance, were also resistant to the growth of the RPL-1 tumor. Lines of chickens which possessed B alleles associated with susceptibility to MD were also very susceptible to the growth of the RPL-1 tumour. The growth of RPL-1 cells can therefore be used as a marker of B21-associated resistance to MD. In contrast, lines of chickens which differ in susceptibility to MD, due to allelic differences at a non-B genetic locus (or loci), did not differ with respect to their capacity to reject RPL-1 cells. We suggest that this might indicate that different mechanisms might be operative in B-associated versus non-B-associated resistance to MD. In addition, females were more susceptible than males to RPL-1 challenge and the RPL-1 tumour demonstrated a predilection for growth in the ovary.


Ascaris-induced respiratory responses in the conscious rhesus monkey. Reproducible immediate-type respiratory responses were evoked in conscious monkeys, sensitive to inhaled Ascaris suum, for periods up to 18 months. These responses were characterized by decreases in tidal volume and increases in breathing rate that persisted for about 40 min. Maximum changes were seen 3--5 min after exposure to the aerosolized antigen and were often accompanied by coughing and increased movement of the animals within the plethysmograph used for monitoring their ventilatory changes. Significant inhibition of the Ascaris-induced respiratory changes were seen in animals treated with either isoproterenol or cromolyn sodium. However, the latter agent was not effective in reversing histamine-induced ventilatory changes. The conscious monkey appears to be a suitable animal for evaluating potential antiallergic drugs.


Effects of disodium chromoglycate on systemic anaphylaxis in guinea pigs, rats and mice. Disodium chromoglycate (DSCG) exerted a protective action in passively-sensitized guinea pigs undergoing anaphylaxis but no such effect was found in actively-sensitized animals. The degree of protection depended on the route and the time of administration. DSCG also protected actively-sensitized rats undergoing anaplylaxis but no effect was found in mice.


Cross-reactivity of the O antigens among Haemophilus influenzae type b strains. The double-immunodiffusion technique was used to compare the O antigens of 18 meningeal Haemophilus influenzae type b strains with each other and with H. influenzae strains representing the capsular types a-f. The O antigens of the meningeal strains and the reference type b strain (strain Rab) cross-reacted to a large extent, while only a few cross-reactions were demonstrable with the other serotypes.


Immunodiffusion studies of ribosomes in classification of mycobacteria and related taxa. Ribosomal preparations consisting of crude ribosomes (CR), 30S subunits (30S) and 16S core particles (16S) from four strains of the species Mycobacterium bovis (BCG), Mycobacterium fortuitum, Mycobacterium phlei and Mycobacterium smegmatis were analyzed by immunodiffusion technique for taxonomical purposes. The ribosomal preparations tested contained several interspecies cross-reacting precipitinogens. The number of precipitinogens demonstrated at the homologous reactions was generally larger than the number of precipitinogens shown at the heterologous reactions indicating a probable presence of species-specific antigens in ribosomes. The largest number of possible species-specific precipitinogens was demonstrated when crude ribosomal preparations were studied. However, such precipitinogens were also shown in the 30S subunits and they were individually analyzed. The 16S core particles were dominated by cross-reacting precipitinogens. The number of ribosomal precipitinogens shared by M. phlei and M. smegmatis was large indicating a close taxonomical relationship between these two species. Apart from the four mycobacterial strains studied, 15 other strains representing the genera Mycobacterium, Arthrobacterium, Corynebacterium, Kurthia, Nocardia and Rhodococcus were included in the study. The presence of intergenerically cross-reacting precipitinogens in the ribosomal preparations was demonstrated.


Immunodiffusion studies of various structural preparations from mycobacterial cells. Various structures and other preparations from mycobacterial cells were analyzed by immunodiffusion. The preparations were obtained from four strains referred to the species Mycobacterium bovis (BCG), Mycobacterium fortuitum, Mycobacterium phlei and Mycobacterium smegmatis. They represented cell walls (CW), culture filtrates (CF), artificially disintegrated cell material (XP), protoplasms (PP), crude ribosomes (CR), ribosomal 50S subunits (50S), ribosomal 30S subunits (30S), ribosomal 16S core particles (16S) and ribosomal-free fractions (UCS). The preparations were analyzed by reference precipitation systems based on CF, CR and 30S preparations. Serological comparisons were made with material from the same species and cross-testing between material from different species was not performed. It was shown that from the precipitinogenic point of view the CF and XP materials were very similar and that they both contained protoplasmic material to a large extent. Furthermore, it was found that ribosomal precipitinogens constitute an important part of the precipitinogens in PP as well as in the CF and the XP preparations. The ribosomal precipitinogens were found to represent the two subunits as well as the 16S core particle. One ribosomal precipitinogen, designated beta, was shown in all preparations except the CW and the UCS.


Interaction among IgE-mediated hypersensitivity reaction, PCA reaction and delayed hypersensitivity reaction (at local skin sites of monkeys). The effect of the IgE-mediated reaction on the passive cutaneous anaphylaxis (PCA) reaction was studied at local skin sites of monkeys, and we found that the IgE-mediated reaction appeared to enhance the PCA reaction. Interactions among IgE-mediated reaction, PCA reaction and delayed hypersensitivity reaction were also determined. Contact dermatitis induced with DNCB was utilized as the delayed hypersensitivity reaction. The IgE-mediated reactior or PCA reaction, as well as simple serum irritation, enhanced the delayed hypersensitivity reaction. It is thus assumed that the IgE-mediated reaction enhances the PCA reaction and that this in turn accelerates the delayed hypersensitivity reaction.


Immunoglobulin E, mast cells, and eosinophils in the skin of rhesus monkeys immunized with x-irradiated cercariae of Schistosoma japonicum. Immunoglobulin E (IgE), mast cells, and eosinophils in the skin of rhesus monkeys immunized with highly X-irradiated cercariae of Schistosoma japonicum were studied. IgE was stained by the unlabeled antibody enzyme method and was found on mast cells. Before challenge, mast cells were found only in the dermis. Immediately after the challenge, mast cells were found in both the dermis and epidermis and many of them were degranulated. Soon after, margination and emigration of granulocytes, predominantly eosinophils, occurred along the blood capillaries in the dermis. Gradulally, preivascular infiltration of eosinophils was seen in the dermis and migration of eosinophils from the dermis into the epidermis appeared, resulting in the formation of minute eosinophilic abscesses in the epidermis. In addition, the IgE was found as a thin coat on the integument of the schistosomula. Deteriorated schistosomula were seen amid the eosinophilic abscesses in the epidermis and in eosinophilic infiltrations in the dermis. The present findings suggest the possibility that the reaction of IgE, mast cells, and eosinophils were integrated into one immunological effect, namely the schistosomulicidal action.


Effects of ascorbic acid deficiency and of erythorbic acid on blood components in the Cynomolgus monkey. Eight male Cynomolgus monkeys were fed an ascorbic acid-free total liquid diet until plasma levels decreased from a mean of 1.1 mg/dl to 0.04 mg/dl at 8 weeks. They showed no visible signs of scurvy. The animals were then given a daily oral dose of 10 mg ascorbic acid/kg body weight for 4 weeks, when the experiment was ended. Four of the animals were given, in addition, 200 mg erythorbic acid/kg body weight orally each day. In all animals repletion was accomplished in two to three weeks using return to initial plasma ascorbic acid levels as the criterion. During deficiency, blood cellular elements were found to be more resistant to depletion than plasma. For erythrocytes, this may be explained at least partially by the observation that in vitro uptake of ascorbic acid tended to be related inversely to blood ascorbic acid levels. However, no such relationship was seen in leucocytes or platelets. Other measurements made on blood did not vary in response to changing ascorbic acid levels. These include serum cholesterol; erythrocyte, leucocyte, or platelet counts; leucocyte differential; hemoglobin concentration; and hematocrit. Urinary hydroxyproline/creatinine ratios were also unchanged. Erythorbic acid, a stereoisomer of ascorbic acid and a common food additive, has been cited as a possible interferent in the determination of whole blood or plasma ascorbic acid, since in the guinea pig it is absorbed from the gut and no commonly used ascorbic acid analysis can distinguish between the isomers. Under conditions of the present experiment, however, no elevation of apparent whole blood or plasma ascorbic acid was produced by inclusion of high levels of erythorbic acid in the diet. Animals given erythorbic acid in addition to ascorbic acid during repletion did not differ from those given ascorbic acid alone in any aspect mentioned above.


Application of optical transform techniques to laser irradiation of retina. Novel retinal irradiation techniques for laser damage studies are described; these are straightforward applications of the Fourier transform properties of an optical system. Interposing an appropriate diffracting screen between the laser source and the test eye enables one to simultaneously induce a number of discrete and precisely located retinal exposures with a known gradation of energy. Various kinds of diffracting objects are considered, and it is seen that transparent sinusoidal phase gratings offer the most promise for damage threshold studies. The method provides an objective measure of the ocular focal length at the time of the exposure. The production of arbitrary retinal irradiation patterns for more general research or clinical applications is possible with the extension of the technique by the use of simple holographic methods for forming the appropriate diffracting screens.


Dysgenetic lens (dyl)--a new gene in the mouse. A new autosomal recessive gene, dysgenetic lens (dyl), in the mouse is described. Homozygotes are fully viable and exhibit smaller eye, corneal opacity, adhesion of the iris, cataractous degeneration, and extrusion of the lens nucleus and persistent lens-epithelium attachment. Developmental failure of lens vesicle--ectoderm separation is recognized as the earliest expression of the genetic defect. Possible significance of this mutant in the understanding of Peter's anomaly is indicated.


Regional differences in the fine structure of the ciliary epithelium related to accommodation. The ciliary bodies of five monkey eyes and one human eye were subdivided into five zones. The ciliary epithelium with its bordering stroma was investigated electron microscopically. The number of cell organelles of the nonpigmented (NPE) and pigmented (PE) epithelium (mitochondria, rough endoplasmic reticulum, Golgi complexes); intercellular junctions between NPE and NPE, PE and PE, and NPE and PE (desmosomes, puncta adhaerentia, gap junctions, tight junctions); and fenestrations of the capillary endothelium were quantitatively evaluated. All these types of cell organelles, fenestrations of the capillary endothelium, and gap junctions in the NPE were found in greater numbers at the crests of the ciliary processes than in the valleys between processes. On the other hand, the number of puncta adhaerentia is significantly higher in the valleys than at the crests. In the valleys, the internal limiting membrane performs an elaborate network of electron-dense strands in which many fine zonular fibers terminate. These fibers are believed to belong to the "tension fiber system." Their firm attachment to the ciliary epithelium and the great number of intercellular junctions known as mechanical structures lend further support to our concept that these structures function as a fulcrum in the process of accommodation.


Hyperopia and loss of accommodation following ciliary muscle disinsertion in the cynomolgus monkey: physiologic and scanning electron microscopic studies. Twenty-three cynomolgus monkeys underwent 360-degree disinsertion and retrodisplacement of the ciliary muscle in one eye. Ten to 12 weeks after unilateral disinsertion, resting refraction in the "disinserted" eyes was more hyperopic than in the opposite eyes by 1.12 +/- 0.21 (mean +/- S.E.M.) diopters (p less than 0.001). Accomodative responses to intramuscular pilocarpine (2 or 3 mg/kg) were 0.90 +/- 0.14 (mean +/- S.E.M.) diopters in the disinserted eyes and 13.88 +/- 0.79 diopters in the opposite eyes. The induced hyperopia and loss of accommodation in the disinserted eyes seemed permanent, persisting for at least 14 months in one monkey and 29 months in three monkeys tested periodically after disinsertion. By light microscopy, the ciliary muscle in the disinserted eyes appeared normal and was contracted by pilocarpine. Scanning electron microscopy of the accommodative apparatus revealed retrodisplacement of the ciliary muscle, ciliary processes, and zonular plexus in the disinserted eyes. Structural alterations in the zonular apparatus seemed insufficient to account for the physiological findings. Hyperopia and loss of accommodation following ciliary muscle retrodisplacement are consistent with a new theory of zonular action during accommodation.


Further investigation of the role of calcium in human lens protein aggregation. High-molecular-weight (HMW) protein from human cataractous lenses, isolated by differential centrifugation, was deaggregated in 7M urea and then reaggregated in either the presence or absence of 10 mM CaCl2. Over 90% of the material reaggregated in the presence of calcium appears to have a size greater than 50 X 10(6) daltons. By contrast, only 20% to 25% of the material reaggregated in the absence of calcium has molecular weight greater than 50 X 10(6) daltons. Disulfide formation during reaggregation is unlikely in the latter experiment, since the addition of 50 mM mercaptoethanol caused no change in results. About 60% to 70% of the low-molecular-weight (LMW) protein fraction deaggregated in 7M urea buffer can be converted to HMW species in the presence of 10 mM CaCl2, when the deaggregating agent is removed. However, only 5% to 10% of this protein is converted to HMW species if the deaggregation step is eliminated. Experiments with 45 Ca indicate that whereas calcium is necessary for the formation of the HMW aggregates, only one calcium per approximately 5 X 10(5) daltons remains bound in the reaggregated material. The data suggest that although calcium may be required to induce aggregation to HMW species, it is not required to stabilize such macromolecules. SDS-polyacrylamide gel electrophoresis of the HMW species formed upon reaggregation of the dissociated HMW species with calcium indicates the presence of all the major polypeptide subunits of the original HMW species present in the lens; however, reaggregation in the absence of calcium yields HMW species lacking in the 9600 dalton component.


Umbilical vein as ureteral replacement.

Umbilical vein as ureteral replacement. In 13 dogs the middle one-third of the ureter and in four dogs the entire ureter was replaced by fresh Rhesus monkey or human umbilical cord vein. The overall success rate was 60 per cent (10/17); one dog revealed an excellent intravenous pyelogram; the typical roentgenographic appearance of the kidney was a slight hydronephrosis. Animals surviving for more than 1 year were nephrectomized on the contralateral side. One dog is still alive at 36 months. Histologically, the typical structures of the umbilical vein remained unchanged after 2 years, but endothelium was replaced by urothelium. The umbilical vein seems to be an embryonic tissue of extremely low antigenicity and therefore suitable for transplantation.


The prostatic carcinoma in laboratory animals: a bibliographic survey from 1900 to 1977. This survey was performed in relation to a study of the role of endocrine glands in rodents that develop prostatic cancer. It is our belief that the data synthesized here can save much tedious work and time, and provide accurate information for investigators in the field of experimental and clinical prostatic cancer.


Intravesical thio-tepa: a study of 3H-thymidine uptake in normal urothelium and FANFT-induced tumors in rats. Treatment of superficial low-grade bladder cancer often incorporates intravesical instillations of the alkylating agent thio-TEPA. Current dosages and administration schedules are empiric inasmuch as cytokinetic data are sparse. The FANFT-induced bladder rat tumor model closely approximates human bladder cancer. This study measuring 3H-thymidine uptake identified peaks of maximal synthetic activity at 3 and 9 days after thio-TEPA exposure in both normal and neoplastic urothelium. Thus, repeat instillation at these intervals may improve cytotoxicity and overall response rate.


Gastrointestinal pathology in South America. Non-neoplastic gastrointestinal disease in South America is largely related to environmental conditions. Parasitic disorders, including Chagas' disease with megacolon, predominate in endemic regions. Common enteritides of various etiologie are frequent, whereas appendicits, diverticulosis of the colon, ulcerative colitis and Crohn's disease are extremely rare. There was no appendicitis in native Indians of Paraguay.


Primate model of sprue-like syndrome.

Primate model of sprue-like syndrome. An experimental model resembling so-called tropical sprue has been produced in young, growing rhesus monkeys kept on a protein-deficient diet. At a 2% level of protein intake, the animals became frankly diseased after two months, but at a 5% protein intake the symptoms appeared after five months. The animals lost weight, their skin became brittle, the fur lost luster, and facial edema appeared. The hematocrit, and serum folate, protein and albumin were significantly decreased, and intestinal absorption of fat, D-xylose, radioactive vitamin B12 and folic acid showed marked depression. The jejunal mucosa showed moderate villous atrophy. Histochemical and electron microscopy changes were consistent with those seen in the human tropical sprue syndrome. It appears, therefore, that protein malnutrition plays an important role in the experimental tropical-sprue-like syndrome in monkeys.


Infantile jejunal mucosa in infection and malnutrition. Sixteen children with refractory diarrhea and three malnourished children who had frequent episodes of acute gastroenteritis but little diarrhea at the time of hospital admission, were studied by peroral upper small intestinal biopsy. Six children were adequately nourished; five children weighed 62 to 79% of expected weight and eight weighed less than 60% of expected weight. Two of the malnourished children had giardiasis. Pathogenic bacteria were found in only one case. Varying degrees of mucosal atrophy with reduction of mean villous height were seen in 18 cases. The concentration of mononuclear inflammatory cells and plasma cells was about half that seen in well-nourished children with severe nongastrointestinal infections. The concentration of mononuclear cells in the lamina propria was about twice that seen in normal adults. The proportions of IgA-producing cells and cells that stained for secretory component were significantly reduced, as compared with normal adult control values. This reduction was most striking in children with malnutrition complicated by giardiasis. Enzyme histochemical studies were performed for leucine aminopeptidase, alkaline phosphatase and acid phosphatase. There was a tendency for considerably reduced acid phosphatase activity in all clinical groups (kwashiorkor, marasmic kwashiorkor and marasmus) of growth-retarded infants.


Primary intestinal lymphoma in South Africa. All adult cases of primary intestinal lymphoma seen during the years 1953--77 at Groote Schuur Hospital, South Africa, were reviewed. Seventy percent of patients with solitary lymphoma and 80% with immunoproliferative small intestinal disease were mulatto. Patients in the latter group presented with malabsorption and those with a solitary lymphoma presented with intestinal obstruction. Four of 15 patients tested for the presence of alpha-heavy-chains were found to have alpha-heavy-chain disease. Family studies of potential genetic and immunologic factors showed some significant differences in IgA and IgG levels in the families of two patients with alpha-heavy-chain disease. Elevated alkaline phosphatase of intestinal origin was found in four of six patients with immunoproliferative small intestinal disease and in a high proportion of relatives. Fifty percent of the six patients were of blood group B. Minor blood groups, ABH secretor state and Pi phenotype distribution were similar to those of the control subjects. HLA gene frequency was particularly increased in the HLA-A9 antigen. These studies suggest that genetic factors may be relevant to the pathogenesis of immunoproliferative small intestinal disease.


Gastrointestinal mucosa and primary gastrointestinal lymphoma. The primary gastrointestinal lymphomas in Shiraz, Iran, and Richmond, Virginia, USA were compared. Upper duodenojejunal lymphoma is always associated with atrophy of the surrounding nonlymphomatous mucosa, plasma cell infiltration and formation of lymph follicles. This is frequently linked to repeated gastroenteritis leading to mucosal atrophy, mutation of plasma cell precursors and secretion of alpha-heavy-chain. Gastrointestinal lymphoma in the USA and other industrialized countries is found in the stomach, where it is accompanied by superficial perifoveolar plasma cell gastritis of the surrounding mucosa, or in the performed lymphoid tissue of the ileocolon, surrounded usually by normal mucosa. A hypothesis for the pathogenesis of the different types of primary gastrointestinal lymphoma, considering the geographic distribution, and mucosal and immunologic antecedents, is presented.


Isolation of Brucella abortus and Brucella abortus, strain 19, from cattle. Tissues from 104 cows in herd were examined for brucellae. Brucella abortus, strain 19, was isolated from 22 cows, a field strain of B abortus, biotype 1, was isolated from 9 cows, and both strains were isolated from 2 cows.


Membrane-bound D-gluconate dehydrogenase from Pseudomonas aeruginosa. Purification and structure of cytochrome-binding form. A membrane-bound D-gluconate dehydrogenase [EC 1.1.99.3] was solubilized from membranes of Pseudomonas aeruginosa and purified to a homogeneous state with the aid of detergents. The solubilized enzyme was a monomer in the presence of at least 0.1% Triton X-100, having a molecular weight of 138,000 on polyacrylamide gel electrophoresis or 124,000--131,000 on sucrose density gradient centrifugation. In the absence of Triton X-100, the enzyme became dimeric, having a molecular weight of 240,000--260,000 on sucrose density gradient centrifugation. Removal of Triton X-100 caused a decrease in enzyme activity. Enzyme activity was stimulated by addition of phospholipid, particularly cardiolipin, in the presence of Triton X-100. The enzyme had a cytochrome c1, c-554(551), which might be a diheme cytochrome, and it also contained a covalently bound flavin but not ubiquinone. In the presence of sodium dodecyl sulfate, the enzyme was dissociated into three components with molecular weights of 66,000, 50,000, and 22,000. The components of 66,000 and 50,000 daltons corresponded to a flavoprotein and cytochrome c1, respectively, but that of 22,000 dalton remained unclear as to its function.


Purification and characterization of testosterone-binding globulin of canine serum. Testosterone-binding globulin (TeBG) of canine serum was purified to apparent homogeneity by affinity chromatography on testosterone-17 alpha-ethynylcarboxyaminoethyl-Sepharose 4B followed by hydroxyapatite column chromatography. Canine TeBG was a glycoprotein containing 5.5% carbohydrates. Equilibrium sedimentation analysis in the presence and absence of 6 M guanidine hydrochloride gave molecular weights of 40,000 and 76,000, respectively, suggesting that native TeBG consists of two subunits. Equilibrium dissociation constants at 0 degrees C for testosterone and dihydrotestosterone were estimated to be 5.58 x 10(-8) M and 1.43 x 10(-8) M, respectively, and the number of binding site per native molecule was approximately unity for both androgens. Canine TeBG had virtually no affinity for estradiol, progesterone, or cortisol. Canine TeBG did not cross-react with a rabbit antiserum raised against bovine TeBG.


Improved procedure for the conjugation of rabbit IgG and Fab' antibodies with beta-D-galactosidase from Escherichia coli using N,N'-o-phenylenedimaleimide. The procedures for the conjugation of rabbit IgG and Fab' antibodies with beta-D-galactosidase from Escherichia coli using N,N'-o-phenylenedimaleimide were improved in several respects as compared with the previous methods (Eur. J. Biochem. 62, 285--292, 1976; J. Immunol. 116, 1554--1560, 1976). Maleimide residues were efficiently introduced into antibodies under an atmosphere of nitrogen; the average number of maleimide residues introduced into IgG and Fab' antibodies were 0.78 (0.65--0.86) and 0.86 (0.80--0.95) per molecule, respectively. The conjugation with the enzyme was performed at 4 degrees C at pH 6.5 for 15 or more hours. The conjugates were almost completely separated from unreacted IgG and Fab' by gel filtration. When the recoveries of IgG, Fab', and beta-D-galactosidase in the conjugates were 23-29, 35-44, and 99%, respectively, the average numbers of IgG and Fab' molecules conjugated with the enzyme were 1.5-1.7 and 2.1-2.8 per molecule, respectively. There was no significant impairment of beta-D-galactosidase activity or the activity of anti-human IgG antibody to bind to human IgG upon conjugation. However, the conjugate preparation was heterogeneous, and one-third of each preparation consisted of aggregated conjugates less useful in sandwich enzymoimmunoassay than the remaining material. The conjugate with Fab' antibody gave lower control values in sandwich enzymoimmunoassay with silicone rubber as a solid phase than that with IgG antibody.


Inhibition of fatty acid synthesis prevents the incorporation of aldosterone-induced proteins into membranes. 2-Methyl-2-[p-(1,2,3,4-tetrahydro-1-naphthyl)phenoxy]propionic acid (TPIA), an acetyl coenzyme A carboxylase inhibitor, blocks the aldosterone-induced increase in transepithelial sodium transport. To examine the requirement for ongoing fatty acid synthesis and/or elongation in the aldosterone-induced alteration of cellular protein metabolism in the toad's urinary bladder, the effect of TPIA has been examined in double-labeled amino acid incorporation experiments. TPIA itself has no effect on the pattern of protein labeling in either the "soluble" or a plasma membrane-enriched fraction. However, inhibition of fatty acid synthesis selectively inhibits the aldosterone-induced incorporation of membrane proteins without altering the labeling of soluble cell protein. These results indicate that ongoing fatty acid synthesis is required for the hormone-induced changes in plasma membrane protein metabolism.


Purification and properties of prostaglandin D synthetase from rat brain. The prostaglandin D synthetase system was isolated from rat brain. Prostaglandin endoperoxide synthetase solubilized from a microsomal fraction catalyzed the conversion of arachidonic acid to prostaglandin H2 in the presence of heme and tryptophan. Prostaglandin D synthetase (prostaglandin endoperoxidase-D isomerase) catalyzing the isomerization of prostaglandin H2 to prostaglandin D2 was found predominantly in a cytosol fraction and was purified to apparent homogeneity with a specific activity of 1.7 mumol/min/mg of protein at 24 degrees C. The enzyme also acted upon prostaglandin G2 and produced a compound presumed to be 15-hydroperoxy-prostaglandin D2. Glutathione was not required for the enzyme reaction, but the enzyme was stabilized by thiol compounds including glutathione. The enzyme was inhibited by p-chloromercuribenzoic acid in a reversible manner. The purified enzyme was essentially free of the glutathione S-transferase activity which was found in the cytosol of brain.


Arrangement of a highly repeated DNA sequence in the genome and chromatin of the African green monkey. The DNA of the African green monkey contains three components that are distinguishable by the kinetics of reassociation. The rapidly reassociating component represents about 20% of the total DNA and is composed almost entirely of a sequence (AGMr(HindIII)-1) which is repeated 6.8 x 10(6) times. The majority of the AGMr(HindIII)-1 sequences are organized in long tandem repeats of a segment of 172 base pairs in length. However, a fraction of the AGMr (HindIII)-1 sequences is interspersed with another 37% of the genome. The structure of the chromatin containing the AGMr-(HindIII)-1 sequence is indistinguishable from that containing total DNA. Furthermore, there is nothing inherent in the nucleotide sequence of AGMr(HindIII)-1 which specifies a unique location for nucleosomes.


The purification and sequence of a temperature-sensitive tryptophan tRNA. Escherichia coli can be temperature-sensitive due to a lesion in the gene for tRNATrp (Yanofsky, C., and Soll, L. (1977) J. Mol. Biol. 113, 663-677). Purification of tRNATrp from this strain (temperature-sensitive tRNATrp) was achieved by one of two methods. Either a combination of benzoylated DEAE-cellulose column chromatography and two-dimensional polyacrylamide gel electrophoresis, or hybridization to plasmid DNA covalently bound to cellulose (this is a recombinant plasmid carrying the gene for tRNATrp) and electrophoresis of the eluted material on a 10% polyacrylamide gel, produced isotopically pure tRNA. The sequence of the temperature-sensitive tRNATrp was determined by standard methods. We find that the sequence differs from that of wild type tRNATrp by a single residue; G in position 7 (G7) in wild type tRNATrp is A7 in temperature-sensitive tRNATrp. This base difference results in one less base pair in the CCA stem of the temperature-sensitive species. The effect of this base change in the in vitro and in vivo properties of tRNATrp (presented elsewhere (S.P. Eisenberg and M. Yarus, manuscript in preparation.)) are discussed.


Purification and characterization of a carboxypeptidase-transpeptidase of Bacillus megaterium acting on the tetrapeptide moiety of the peptidoglycan. The enzyme carboxypeptidase-IIW of Bacillus megaterium incorporates free diaminopimelate into purified bacterial walls. This enzyme can be solubilized from toluene-treated cells by LiCl extraction and has now been purified 106-fold to one major band on polyacrylamide gel electrophoresis. The enzyme has an apparent molecular weight of approximately 60,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by Sephadex G-100 gel filtration. Carboxypeptidase-IIW requires divalent cations and thiol group(s) for optimal activity. Product analysis indicates that the enzyme can hydrolyze the terminal D-alanine from the tetrapeptide of the peptidoglycan or replace it with a variety of amino acids with D-asymmetric centers for transpeptidation. Substrate specificity studies reveal that the enzymatic activity depends on the presence of N-acetyl-D-glucosamine of the GlcNAc-MurNAc-tetrapeptide. This specificity of carboxypeptidase-IIW for the N-acetyl-D-glucosamine explains in part the affinity of the enzyme for the cell wall of B. megaterium. The enzyme is compared to the carboxypeptidases-transpeptidases of other organisms with the similarities and differences discussed.


Formation of intermolecular disulfide bonds on nascent immunoglobulin polypeptides. The initial step of intermolecular covalent assembly of immunoglobulins molecules involves formation of heavy chain-light chain or heavy chain-heavy chain disulfide bonds. Using QAE-Sephadex chromatography to isolate microsomal nascent polypeptides, we have shown that this initial step of intermolecular covalent assembly occurs, to a substantial extent, on nascent heavy chains, as well as on completed heavy chains as previously demonstrated by others. In MPC 11 mouse myeloma cells, completed light chains are assembled covalently to nascent heavy chains, whereas in MOPC 21 mouse myeloma cells, completed heavy chains are assembled covalently to nascent heavy chains. These results are consisted with the heavy-light half-molecule being the major initial intermediate in the assembly of MPC 11 IgG2b and heavy-heavy dimer being the major initial intermediate formed in assembly of MOPC 21 IgG1. The nascent MPC 11 heavy chain must be at least 38,000 daltons in size before assembly with the light chain occurs, even though the heavy chain cysteine involved in this disulfide bond is 131 residues (approximately 15,000 daltons) from the NH2 terminus. In addition, pulse-chase labeling studies of MPC 11 cells have shown that the assembly of completed light chains with the nascent heavy chain must occur within a few minutes of the synthesis of the light chain even though a large excess of unassembled MPC 11 light chains remain inside the cell for an average time of 2 h before being secreted.


Studies on the association of cytochrome P-450 and NADPH-cytochrome c reductase during catalysis in a reconstituted hydroxylating system. The interaction between cytochrome P-450 and NADPH-cytochrome c reductase during catalysis has been investigated with a reconstituted monooxygenase system composed of the two purified enzyme components and synthetic phospholipid. Steady state kinetic data are consistent with a scheme in which the formation of a binary complex between the two proteins precedes catalysis. The formation of this binary complex is described by a simple mass action equation. In agreement with this equation, the observed Vmax for benzphetamine N-demethylation was found to be directly proportional to the calculated concentration of the cytochrome P-450 . reductase complex. Furthermore, with appropriate reductase/cytochrome P-450 mole ratios, the Vmax could be shown to be linearly dependent on either the reductase or the cytochrome P-450 concentration alone. In contrast, the Km parameter is independent of the complex concentration, indicating that no change in the rate-limiting step has occurred. Thus a distinction should be made between a rate-limiting enzyme component and the rate-limiting step in this multienzyme system.


Total synthesis of a tyrosine suppressor transfer RNA gene. XVII. Transcription, in vitro, of the synthetic gene and processing of the primary transcript to transfer RNA. Primer- and promoter-dependent transcription of the synthesis gene had been studied. Primer-dependent transcription gave, as a major product, an end-to-end transcript which was strand-specific. The transcript was characterized rigorously by two-dimensional separation and analysis of the oligonucleotides formed on digestion with T1-RNase and pancreatic RNase and by nearest neighbor analyses of the oligonucleotides obtained when different alpha-32P-labeled ribonucleoside triphosphates were used as substrates. Minor products accompanying the major transcript were characterized similarly. The major transcript, when treated with an Escherichia coli S-100 extract, was processed to the tRNATyr with correct 5'- and 3'-ends. The nucleolytic cleavages occurring at the 3'-end were characterized. In promoter-dependent transcription, transcription of a restriction fragment containing phi80psu+III gene and the synthetic gene with and without the promoter were compared. Transcription of the synthetic gene was promoter-dependent and strand-specific, the initiation of transcription occurring at the same point as previously found in vivo. Although the synthetic gene contains only 16 base pairs corresponding to the natural sequence following the C-C-A end, processing of the transcript at the 3'-end occurred normally, the endonucleolytic cleavage being followed by exonucleolytic cleavages. The products of promoter-dependent transcription were completely characterized. An examination of the base modifications of the primary transcript during treatment of the latter with E. coli S-100 extract showed couplete modification of uridine to pseudouridine and partial methylation of uridine to ribosylthymine in TpsiCG sequence and partial formation of pseudouridine in the anticodon loop. However, hardly any formation of 2'-O-methylguanosine or of 2-methylthio-6-isopentenyl adenosine could be detected.


Purification of human liver microsomal epoxide hydrase. Differences in the properties of the human and rat enzymes. Human liver microsomal epoxide hydrase has been highly purified to a specific activity (570 to 620 nmol/min/mg of protein) comparable to that of the rat enzyme using styrene oxide as substrate. Like the purified rat liver microsomal epoxide hydrase, the human enzyme has a minimum molecular weight of 49,000 as determined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate and exhibits broad substrate specificity toward a variety of alkene and arene oxides. Despite these similarities, the human and rat enzymes are different proteins as judged by their immunochemical properties as well as their relative catalytic activities toward certain substrates.


Subunit structure of higher plant glyceraldehyde-3-phosphate dehydrogenases (EC 1.2.1.12 and EC 1.2.1.13). In a previous publication (Cerff, R. (1979) Eur. J. Biochem., 94, 243--247) we demonstrated that chloroplast NADP-linked glyceraldehyde-3-P dehydrogenase (EC 1.2.1.13) from higher plants consists of two separate isoenzymes with apparent subunit compositions A2B2 (isoenzyme 1) and A4 (isoenzyme 2), where Subunits A and B are distinguished by slightly different molecular weights (A smaller than or approximately to B). In the present study we compare isoenzymes 1 and 2 from Sinapis alba and Hordeum vulgare on the basis of antigenic cross-reactivity, tryptic peptides, and amino acid composition. Isoenzymes 1 and 2 show immunochemical identity. They also have very similar tryptic peptide maps and amino acid compositions. This strongly suggests that Subunits A and B of the NADP-linked enzyme are very similar in primary sequence. As opposed to this, cytoplasmic NAD-specific glyceraldehyde-3-P dehydrogenase (EC 1.2.1.12) does not cross-react with antisera raised against the NADP-linked enzyme. Furthermore, tryptic peptide maps of the NAD-specific enzyme show little or no similarity with those of the NADP-linked enzyme. This indicates that the subunits of the NADP-linked enzyme and the subunit of the NAD-specific enzyme are different proteins coded by separate genes. The differences in the amino acid compositions between the two species corresponds to a SdeltaQ value of 21, suggesting some sequence resemblance and a common phylogenetic origin.


Derivatization of cysteine and cystine for fluorescence amino acid analysis with the o-phthaldialdehyde/2-mercaptoethanol reagent. Previous reports (Drescher, D.G., and Lee, K.S. (1978) Anal. Biochem. 84, 559-569; Lee, K.S., and Drescher, D.G. (1978) Int. J. Biochem. 9, 457-467) have shown that high performance liquid chromatographic analysis of amino acids with the o-phthaldialdehyde/2-mercaptoethanol reagent (OPA/2-ME) is one of the most sensitive procedures currently available for micro amino acid analysis. In the present paper, methods are presented for the modification of cysteine and cystine in proteins for micro amino acid analysis using OPA/2-ME. Cysteine and cystine, which both show low fluorescence with OPA/2-ME, are converted to cysteic acid with performic acid directly, or to S-3-sulfopropylcysteine with 1,3-propane sultone after reduction of cystine with tri-n-butylphosphine. Cysteic acid and S-3-sulfopropylcysteine form highly fluorescent adducts with OPA/2-ME. The formation of S-3-sulfopropylcysteine in proteins and the subsequent hydrolysis of the proteins with methanesulfonic acid are particularly useful for complete amino acid analysis at the picomole level using a single sample.


Biochemical analyses of natural and induced null variants of Drosophila enzymes. For the characterization of null mutants identified in Drosophila populations, several Drosophila enzymes including alcohol dehydrogenase, cytoplasmic malate dehydrogenase, alpha-glycerol phosphate dehydrogenase, and phosphoglucose isomerase were co-purified to homogeneity using an 8-(6-aminohexyl)-amino-ATP-Sepharose affinity column followed by DEAE-Sepharose column chromatography. Mitochondrial malate dehydrogenase was purified by the use of CM-Sepharose and the same ATP affinity column. Alcohol dehydrogenase and alpha-glycerol phosphate dehydrogenase were mapped on a two-dimensional gel. Antiserum was raised in rabbits against these Drosophila enzymes. The presence of cross-reacting material in null mutants was characterized by double immunodiffusion, immunoelectrophoresis, and two-dimensional gel electrophoresis. By immunological techniques, two natural null variants of malic enzyme and one of phosphoglucose isomerase were shown to be negative to cross-reacting material. Two low-dose-rate gamma-radiation-induced null mutants of cytoplasmic malate dehydrogenase were shown to be positive to cross-reacting material. Two-dimensional gel analyses enabled the characterization of three natural null variants of alpha-glycerol phosphate dehydrogenase. The viability of some null mutants with homozygous null or null/deficiency genotypes is discussed in terms of the in vivo metabolic roles of the related enzymes.


Serum lipids in hypothyroidism--a re-evaluation. One hundred and forty-two persons referred for assessment of thyroid function were studied in relation to their thyroid function and fasting serum lipids. Forty-five individuals, who had no family history of thyroid disease, were euthyroid, and were negative for thyroid antibodies formed the control group. The remaining 97 patients were fitted into six groups as follows--euthyroid Hashimoto's thyroiditis, thyroiditis with exagggerated TSH response to TRH, and four groups of increasing grades of thyroid failure based on the level of serum TSH. Only in the two most severely hypothyroid groups was cholesterol significantly elevated as compared with control levels. A graded increase in mean serum triglyceride, was observed as thyroid failure increased; however, values were not different (P greater than 0.05) from the control group. However, partial correlation analysis indicated that serum cholesterol was related to increasing levels of serum TSH and that this dependency was not influenced by age or weight. Although serum cholesterol was related to thyroid function in the study population, it was an insensitive indicator of the metabolic effects of thyroid hormone.


Hyperthyroidism without triiodothyronine excess: an effect of severe non-thyroidal illness. Serial changes in thyroid hormone levels are described in two patients in whom hyperthyroidism was associated with transient non-thyroidal illness. In a 74-year-old woman with mild hyperthyroidism, two episodes of cholecystitis were associated with subnormal concentrations of serum T3 and increased concentrations of serum rT3; T3 became elevated during recovery, associated with a simultaneous fall in rT3. The TSH response to TRH was undetectable on three occasions. A cholecystectomy was performed after preparation with Lugol's iodine and subsequent tests showed evolution through T3 toxicosis to classical hyperthyroidism. In the second case, symptoms and signs of classical hyperthyroidism were noted during an undiagnosed illness characterized by severe abdominal pain and fever. Six days after the onset of this illness, an elevated level of serum T4 was associated with a normal total T3 concentration and increased concentration of rT3. After resolution of abdominal symptoms, serum T3 was markedly increased, associated with persistent T4 and rT3 excess. These findings indicate that the changes in T3 and reverse T3 described in non-thyroidal illness also occur in hyperthyroid patients, and suggest that the fall in T3 may be of sufficient magnitude to make T3 measurement diagnostically unreliable in the presence of non-thyroidal illness.


Thyrotropin secretion in patients with central hypothyroidism: evidence for reduced biological activity of immunoreactive thyrotropin. TSH concentration was measured in plasma before and after TRH administration (200 micrograms, iv) in 89 patients with documented hypothyroidism consequent to various hypothalamic-pituitary disorders. Basal plasma TSH was less than 1.0 microI/ml in 34.8%, between 1.0-3.6 microU/ml in 40.5% and slightly elevated (3.7-9.7 microU/ml) in 24.7% of the cases. The plasma TSH response to TRH was absent in 13.5%, impaired in 16.8%, normal in 47.2%, and exaggerated in 22.5% of the cases, with delayed and/or prolonged pattern of response in 65% of the cases. The dilution curves of several plasmas drawn before and after TRH were parallel to those obtained with TSH standard preparation. After gel filtration, the elution pattern of TRH-stimulated plasmas from 4 patients did not show any major difference from that of pooled plasmas from normal subjects given TRH or from that of patients with primary hypothyroidism. Plasma TSH values determined by cytochemical bioassay on both basal and TRH-stimulated samples of 5 patients were markedly lower than those obtained by RIA. The serum T3 response to TRH was absent or low in 40 out of 53 patients in whom it was evaluated. The administration of T3 (100 micrograms/day for 3 days) or dexamethasone (3 mg/day for 5 days) respectively suppressed or reduced both basal and TRH-induced plasma TSH levels. Two patients became hypothyroid shortly after pituitary surgery in spite of basal and TRH-induced plasma TSH levels similar to or higher than those before surgery. Though thyroid atrophy due to chronic understimulation could explain the low T3 response to TRH in secondary hypothyroidism, it is difficult to reconcile thyroid understimulation with normal or increased plasma TSH unless the immunoreactive material has low biological activity. Present data suggest that several patients with hypothyroidism consequent to hypothalamic-pituitary diseases secrete a material which is immunologically similar to pituitary standard TSH and responds to stimulatory and suppressive agents in a manner similar to normal TSH but has low or absent biological activity. Thus, hypothyroidism due to insufficient TSH stimulation can be termed central hypothyroidism and can be due 1) to pituitary insufficiency (secondary hypothyroidism), 2) to a hypothalamic defect (tertiary hypothyroidism), or 3) to the secretion of biologically inactive TSH.


Lack of evidence for thyrotropin-releasing hormone deamidation in normal and hyperthyroid human sera. Two-hour incubations of human serum with 50 ng TRH or pyroglutamyl-histidyl-proline (TRH-OH) were performed under substrate conditions of 0.2 microgram substrate/ml serum. During incubations with normal serum, 46.3 +/- 1.3 (SEM) ng TRH were degraded while only 14.2 +/- 5.1 ng TRH-OH were degraded (P less than 0.001). During incubations with serum from patients with hyperthyroidism, 42.7 +/- 2.6 ng TRH were degraded compared to only 19.6 +/- 2.0 ng TRH-OH (P less than 0.001). Despite the fact that TRH degradation was significantly greater than TRH-OH degradation in both normal and hyperthyroid serum, no formation of TRH-OH (less than 3.1 ng/incubation tube) from TRH was detected. Formation of TRH-OH from TRH was also not noted when normal or hyperthyroid serum was incubated with TRH at substrate concentrations of 62.5 microgram/ml serum. These data confirm other reports that TRH deamidation does not occur in normal serum and extends this observation to hyperthyroid serum.


Demonstration of tuberculostearic acid in sputum from patients with pulmonary tuberculosis by selected ion monitoring. Selected ion monitoring was used to detect tuberculostearic acid (10-methyloctadecanoic acid) in sputum from patients with pulmonary tuberculosis. The specimens were autoclaved, lyophilized, extracted, and methanolysed before being subjected to thin-layer chromatography and injected into the gas chromatograph/mass spectrometer. Tuberculostearic acid could be detected in five of six tuberculous sputum specimens containing acid-fast rods detectable by light microscopy of Ziehl-Neelsen stained smears. After the sputum specimens had been cultured for five days on Löwenstein-Jensen medium, when still no colonies could be observed visually, the presence of tuberculostearic acid was demonstrated in all six cases of tuberculosis. In corresponding analyses of sputum from eight patients with non-tuberculous pneumonia, tuberculostearic acid was not found. This fatty acid, the presence of which was also demonstrated in cultures of various mycobacterial and nocardial species, is characteristic of organisms of the order Actinomycetales. The demonstration of tuberculostearic acid in sputum specimens may constitute a rapid and sensitive way of diagnosing pulmonary tuberculosis.


Complex of meningococcal group B polysaccharide and type 2 outer membrane protein immunogenic in man. A noncovalent complex of meningococcal group B polysaccharide and type 2 outer membrane protein has been characterized and its potential as a vaccine against group B meningococcal disease investigated. The polysaccharide component was found to have a partition coefficient, K(d), of 0.34 on Sepharose CL-4B in the presence of sodium deoxycholate. The protein consisted of four to five major proteins including the principal outer membrane protein. Hydrophobic binding between the protein and polysaccharide was demonstrated by gel filtration and isopycnic CsCl density gradient centrifugation and found to involve all of the proteins. After demonstrating safety and immunogenicity in animals, two lots of vaccine were tested in a total of eight volunteers. Two 120-mug doses were given subcutaneously at 0 and 5 wk. Mild local reactions occurred in all eight volunteers, but no systemic reactions were observed. 2 wk after the first dose, six of the volunteers had increased levels of bactericidal antibodies against both the group B polysaccharide and the outer membrane proteins. Antibody rises to the group B polysaccharide (mean 6-fold) were confirmed by passive hemagglutination assays and rises to the proteins (mean 10-fold) by a solid phase radioimmunoassay. The second dose resulted in little or no increase in antibody titers. Antibody titers declined over a period of 14 wk but mostly remained above preimmunization levels. Bactericidal antibodies with specificity for the group B polysaccharide were mostly of the immunoglobulin (Ig)M class, and were directed against a determinant associated only with high molecular weight polysaccharides. We conclude that both the group B polysaccharide and the outer membrane protein are immunogenic in man when presented as a complex and that the complex warrants further testing and development as a vaccine against group B meningococcal disease.


Effects of nitroglycerin and propranolol on the distribution of transmural myocardial blood flow during ischemia in the absence of hemodynamic changes in the unanesthetized dog. Chronically instrumented awake dogs were used to study the effects of nitroglycerin and propranolol on the transmural distribution of myocardial blood flow during transient ischemia. Studies were carried out 7-14 d after implantation of an electromagnetic flowmeter probe and balloon occluder on the left circumflex coronary artery, placement of epicardial minor axis sonar crystals, and implantation of left atrial, left ventricular, and aortic catheters. The occluder was inflated to completely interrupt flow for 10 s followed by partial release to reestablish flow at 60% of the preocclusion level. During this partial release, which served as the control for the study, regional myocardial blood flow was measured with 7- to 10-mum radioactive microspheres. After control measurements, seven dogs were given nitroglycerin (0.4 mg i.v.) and eight dogs propranolol (0.2 mg/kg i.v.). 5 min later the occlusion and partial release sequence was repeated, and regional myocardial blood flow was measured when heart rate, aortic and left ventricular end-diastolic pressure, and minor axis diameter were unchanged from control values.The data values were selected so that total flow to the ischemic region during partial release after nitroglycerin or propranolol administration was not significantly different from flow during the control partial release. After nitroglycerin administration, endocardial flow (endo) in the ischemic region increased from 0.46+/-0.07 to 0.59+/-0.06 ml/min per g (P < 0.006); epicardial flow (epi) decreased from 0.78+/-0.09 to 0.70+/-0.08 ml/min per g (P < 0.04). The endo:epi ratio increased from 0.65+/-0.07 to 0.92+/-0.10 (P < 0.05). In contrast, administration of propranolol produced no significant change in transmural flow (endo, 0.42+/-0.02 and 0.46+/-0.03 ml/min per g; epi, 0.71+/-0.06 and 0.70+/-0.07 ml/min per g) or in the endo:epi ratio (0.60+/-0.03, 0.66+/-0.06) in the ischemic region. Nitroglycerin and propranolol produce different effects on the transmural distribution of blood flow to ischemic myocardium. Nitroglycerin can increase blood flow to the underperfused endocardium in the absence of alterations in heart size, hemodynamic parameters, and total transmural flow to the ischemic region. Under similar conditions, propranolol has no significant effect on the transmural distribution of blood flow to an ischemic region.


Reversible phagocytosis in rabbit polymorphonuclear leukocytes. We have studied the fate of inert phagocytized particles in rabbit neutrophils. Neutrophils release significant quantities of preingested oil emulsion. Roughly 50% of an ingested load is released in 40 min at 37 degrees C. By electron microscopy the process of release appears to be by exocytosis: particles appear extruded through a network of processes often accompanied by membranous vesicles. Exocytosis is temperature and glucose dependent but unlike phagocytosis does not require divalent cations. From Coulter counter measurements virtually the entire cell population appears to undergo the phagocytosis-exocytosis sequence. Neutrophils undergoing exocytosis remain intact as determined by direct counts, electron microscopy, and absence of lactate dehydrogenase release. Moreover, by sequentially feeding differently labeled particles, it is shown that the processes of phagocytosis and exocytosis can occur concurrently. Indeed, it is found that ingestion accelerates release. The implications of these phenomena for membrane recycling, lysosomal enzyme release, and the killing of microorganisms are briefly discussed.


Complement-dependent histaminase release from human granulocytes. The role of particle-bound complement proteins in the induction of noncytotoxic enzyme release from human granulocytes was investigated with the use of sera genetically deficient in complement and highly purified complement components. Release of histaminase, one of two important histamine catabolizing enzymes, and beta-glucuronidase from polymorphonuclear leukocytes was solely dependent on particle-bound C3b (the larger cleavage product of the third component of complement) when fluid-phase complement was excluded. The extent of enzyme release was a function of particle-bound C3b input, was reduced by exposing the particles to C3b inactivator, and was blocked by fluid-phase C3b. Phagocytosis of the C3b-coated particles was not required for enzyme release from neutrophils. In contrast, phagocytosis of "opsonized" particles was required for noncytotoxic release of histaminase and arylsulfatase from eosinophils; other proteins, as well as C3b, were able to opsonize particles for induction of enzyme release from eosinophils. These studies suggest a dual role for complement (particularly C3) in modulating vascular permeability phenomena, i.e., release of vasoactive mediators by the action of C3a and C5a, and release of the corresponding enzymes that inactivate the mediators by C3b.


Regulation of the metabolism of 25-hydroxyvitamin D3 in primary cultures of chick kidney cells. A primary chick kidney cell culture is described, capable of forming 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], 24,25-dihydroxyvitamin D3 [24,25(OH)2D3], and 1,24,25-trihydroxyvitamin D3 [1,24,25(OH)3D3] over several days. The apparent Km values were 0.125 microM for the 1-hydroxylase and 2.1 microM for the 24-hydroxylase. Exogenous 1,25(OH)2D3 decreased 1-hydroxylase and increased 24-hydroxylase within 4 h. 24,25(OH)2D3 produced similar effects, but only in the absence of fetal calf serum. R and S isomers of 1,24,25(OH)3D3 were about fives times less active than 1,25(OH)2D3. Bovine parathyroid hormone stimulated the 1- and reduced the 24-hydroxylase in 6 h, but this only occurred in cultures either previously treated with 1,25(OH)2D3 and EGTA to lower Ca to 0.8 mM or in cultures grown in the presence of 25-hydroxyvitamin D3 (25(OH)D3). Under the latter condition, the sensitivity to bovine parathyroid hormone was enhanced, 0.04 U/ml producing a maximum response. Synthetic aminoterminal tetratriacontapeptide (1-34) human parathyroid hormone was equally effective. In the absence of D metabolites, estradiol for 6 h produced a dose-dependent inhibition of the 1-hydroxylase, but no change in the 24-hydroxylase. Progesterone, testosterone, and corticosterone had no significant effect. In cultures grown in the presence of 25(OH)D3 no reproducible effects were obtained with either 1 microM estradiol or 1 microM testosterone, alone or in combination, but 5 microM corticosterone decreased the 1- and increased the 24-hydroxylase. Changes in Ca and P concentrations of the medium as well as addition of ethane-l-hydroxy-1, 1-diphosphate for 48 h did not affect any of the hydroxylase activities. The modulation of the hydroxylase activities by vitamin D3 metabolites and parathyroid hormone suggests that these factors regulate the renal hydroxylase by direct actions, whereas it would appear that ethane-1-hydroxy-1,1-diphosphate, Ca, P, and steroid may exert their influence indirectly.


Sodium taurocholate modifies the bile acid-independent fraction of canalicular bile flow in the rhesus monkey. Bile acid-independent secretion and the choleretic response to taurocholate were determined in rhesus monkeys fitted with indwelling silastic cannulas in the common bile ducts. Bile acids were infused intravenously in random order at 3.5, 7.0, or 10.5 mumol/min for 1.5 h each. When data were analyzed with a single regression line, bile flow increased in proportion to the level of bile acid secretion, although the y-intercepts (the conventional measurement of bile acid-independent secretion) varied widely (77.9+/-40.9 ml/24 h). The variation in y-intercepts was observed between animals and with repeated studies in the same animal and could not be explained by sex differences or the effects of the indwelling silastic cannulas, but seemed to be related to the order of bile acid infusion. With only two taurocholic acid infusion rates (7.0 and 3.5 mumol/min), [(14)C]erythritol clearance was greater per mole of secreted bile acid when the initial bile acid infusion was at the high level, but approached zero at low bile acid secretion rates, which suggests that so-called bile acid-independent canalicular flow is closely related to bile acid secretion or is small in size. The augmentation in [(14)C]erythritol clearance when the high infusion rate was given first was also associated with an increase in biliary clearance of [(3)H]inulin, which indicates that the premeability to inulin was also enhanced. Identical experiments which substituted equimolar infusions of a nonmicelle-forming bile acid (taurodehydrocholate) for taurocholate failed to demonstrate any difference in choleretic response or biliary clearance of [(3)H]inulin with the order of bile acid infusion. These experiments demonstrate that a micelleforming bile acid, taurocholate, can increase the permeability of the biliary system to large molecular weight solutes and simultaneously modify the y-intercept and the volume of bile secreted in response to the transported bile acid. Taurocholate may, therefore, modify its own choleretic response, perhaps by altering the structure or function of bile secretory membranes, and appears to be a major determinant of so-called bile acid-independent flow in rhesus monkeys.