Cytochemical detection of superoxide in cerebral inflammation and ischemia in vivo. We used a cytochemical technique for the detection of superoxide in cerebral inflammation and ischemia-reperfusion in anesthetized cats. The technique is based on the oxidation of Mn2+ to Mn3+ by superoxide; Mn3+, in turn, oxidizes diaminobenzidine. The oxidized diaminobenzidine forms an osmiophilic electron-dense product that is detected by electron microscopy. The reagents, manganese chloride (2 mM) and diaminobenzidine (2 mg/ml), were placed topically on the brain surface of anesthetized cats equipped with cranial windows. Inflammation was induced by topical carrageenan with or without phorbol 12-myristate 13-acetate to activate leukocytes. In inflammation, superoxide was detected in the plasma membrane and in the phagocytic vacuoles of leukocytes. In ischemia-reperfusion, superoxide was identified in the meninges in association with blood vessels. It was located primarily in the extracellular space and occasionally in endothelial and vascular smooth muscle cells. In both inflammation and ischemia, the reaction product was eliminated by superoxide dismutase or by the omission of either manganese or diaminobenzidine. It was unaffected by sodium azide, which inhibits peroxidases. No superoxide was detected in the brain parenchyma. The findings confirm the generation of superoxide is cerebral ischemia-reperfusion and show that it is produced in cerebral vessels.


Stretch-induced increases in intracellular calcium of isolated vascular smooth muscle cells. Vascular smooth muscle responds to stretch with an increase in active force development. To investigate the role of Ca2+ in this response, we used the fluorescent dye fura-2 to quantitate changes in cytosolic Ca2+ in single, vascular smooth muscle cells during rapid stretch. Cells were enzymatically dispersed from pig coronary arteries, loaded with fura-2/AM, and studied using a digital-imaging microscope. Stretch of individual cells was accomplished by attachment with suction to two patch-type micropipettes to apply force to the ends of the cell. Stretch induced the release of Ca2+ from intracellular stores as well Ca2+ influx across the plasma membrane. In physiological saline solution containing 1.5 mM Ca2+, intracellular calcium increased with cell stretch in a sigmoidal fashion. This relationship was shifted upward in 10 mM Ca2+ bath solution and abolished after several minutes in Ca(2+)-free solution. The dihydropyridine Ca2+ channel blocker nifedipine, in doses sufficient to completely block inward Ca2+ current, produced only a partial block of the sustained stretch-induced intracellular Ca2+ response. It is concluded that in isolated pig coronary arterial smooth muscle cells, stretch-induced Ca2+ influx occurs in part via a nifedipine-resistant pathway, which may be a stretch-activated cation channel.


Vasopressin and angiotensin II in reflex regulation of ACTH, glucocorticoids, and renin: effect of water deprivation. Angiotensin II (ANG II) and vasopressin participate in baroreflex regulation of adrenocorticotropic hormone (ACTH), glucocorticoid, and renin secretion. The purpose of this study was to determine whether this participation is enhanced in water-deprived dogs, with chronically elevated plasma ANG II and vasopressin levels, compared with water-replete dogs. The baroreflex was assessed by infusing increasing doses of nitroprusside (0.3, 0.6, 1.5, and 3.0 micrograms.kg-1.min-1) in both groups of animals. To quantitate the participation of ANG II and vasopressin, the dogs were untreated or pretreated with the competitive ANG II antagonist saralasin, a V1-vasopressin antagonist, or combined V1/V2-vasopressin antagonist, either alone or in combination. The findings were as follows. 1) Larger reflex increases in ANG II, vasopressin, and glucocorticoids, but not ACTH, were produced in water-deprived dogs compared with water-replete dogs. 2) ANG II blockade blunted the glucocorticoid and ACTH responses to hypotension in water-deprived dogs, but not water-replete dogs. In contrast, vasopressin blockade reduced the ACTH response only in water-replete dogs. 3) Vasopressin or combined vasopressin and ANG II blockade reduced the plasma level of glucocorticoids related either to the fall in arterial pressure or to the increase in plasma ACTH concentration in water-replete dogs, and this effect was enhanced in water-deprived dogs. 4) In both water-deprived and water-replete animals, saralasin and/or a V1-antagonist increased the renin response to hypotension, but a combined V1/V2-antagonist did not. These results reemphasize the importance of endogenous ANG II and vasopressin in the regulation of ACTH, glucocorticoid, and renin secretion.(ABSTRACT TRUNCATED AT 250 WORDS)


Interleukin-1 stimulates aldosterone secretion: involvement of renin, ACTH, and prostaglandins. Interleukin-1 (IL-1), a cytokine produced during infection and inflammation, mediates some of the endocrinological alterations that parallel these processes. The purpose of this study was to determine whether human recombinant IL-1 (hrIL-1) affects aldosterone output as well as renin and adrenocorticotropic hormone (ACTH) release, two key factors in the regulation of mineralocorticoid secretion. We observed that intravenous administration of hrIL-1 into conscious unrestrained rats elicited a marked and rapid rise in aldosterone plasma levels in a dose-dependent manner. The hrIL-1-induced increase in aldosterone levels was associated with enhanced renin activity and increased ACTH levels in plasma. Furthermore, aldosterone levels of IL-1-injected rats were positively correlated with plasma renin activity (PRA), suggesting that the renin-angiotensin system contributes to the changes observed in the levels of the mineralocorticoid hormone. ACTH seems also to be implicated in the aldosterone response to hrIL-1 because the profile of the kinetic curves of changes in the levels of the pituitary hormone and aldosterone was similar. Pretreatment with the cyclooxygenase inhibitor indomethacin markedly reduced the increase in aldosterone plasma levels and PRA induced by IL-1, indicating that prostaglandins are involved in these effects of the cytokine. These results suggest that IL-1 may play an important role in the control of homeostasis during infectious and inflammatory diseases.


Control of adipose tissue lipolysis in ectotherm vertebrates. Lipolytic activity of fish (Hoplias malabaricus), toad (Bufo paracnemis), and snake (Philodryas patagoniensis) adipose tissue was investigated in vivo and in vitro. Catecholamines or glucagon did not affect the release of free fatty acids (FFA) by incubated fish and toad adipose tissue. Catecholamines also failed to activate snake adipose tissue lipolysis, which even decreased in the presence of epinephrine. However, glucagon stimulated both the lipolytic activity of reptilian tissue in vitro and the mobilization of FFA to plasma when administered to snakes in vivo. The release of FFA from incubated fish, amphibian, and reptilian adipose tissue increased markedly in the presence of cAMP or xanthine derivatives, inhibitors of phosphodiesterase. Forskolin or fluoride, activators of specific components of the adenylate cyclase system, strongly stimulated toad adipose tissue lipolysis. The data suggest that adipocyte triacylglycerol lipase of ectotherm vertebrates is activated by a cAMP-mediated phosphorylation and that the organization of the membrane-bound adenylate cyclase system is similar to that of mammals.


Adrenal corticosteroid secretion in fetal sheep: pulsatile pattern at rest. This study was undertaken to define the resting pattern of fetal pituitary-adrenocortical function. Experiments were performed at 127-145 days gestation in fetal sheep with chronic peripheral and adrenal cannulas inserted under halothane anesthesia. With the fetus in a baseline state, over 6 h, at 30-min intervals, maternal and fetal peripheral samples were collected for blood gases and cortisol (F), corticosterone (B), and adrenocorticotropic hormone (ACTH) concentrations, and three successive, 2-min adrenal samples were collected for determination of F and B secretion rates. We observed high-frequency, episodic bursts of F secretion. A lower frequency oscillation of F secretion, with a period of approximately 90 min, was defined by cosinor analysis. The mean amplitude of the oscillation increased from 45 to 507 ng/min with advancing gestation. The pattern of B secretion was similar to that for F but was quantitatively lower. An oscillatory period of approximately 90 min for plasma F was present in a majority of experiments. Pulsatile rhythms for ACTH were defined in 10 of 14 experiments, with periods ranging from 1.64 h in the least mature group to 2.37 h in the oldest fetus. Mean data revealed exponential increases in both F secretion and plasma ACTH from 129 to 145 days gestation.


Paneth cell-like change and small cell carcinoma of the prostate. Two divergent forms of prostatic neuroendocrine differentiation. Paneth cell-like change of the prostate refers to collections of prostatic cells with eosinophilic cytoplasmic granules that bear a striking histological resemblance to normal intestinal Paneth cells. Paneth cell-like change in malignant prostatic epithelium usually represent neuroendocrine differentiation, with neuroendocrine granules confirmed by immunohistochemical and ultrastructural studies. We report the histopathological, immunohistochemical, and electron microscopic findings in a mixed adenocarcinoma with Paneth cell-like change and small cell undifferentiated carcinoma. This case illustrates two divergent forms of neuroendocrine differentiation occurring in a single prostatic neoplasm. The spectrum of neuroendocrine differentiation in the prostate should be expanded to include tumors with Paneth cell-like change in addition to carcinoid tumors and small cell undifferentiated carcinoma. These three distinct forms of prostatic neuroendocrine neoplasia appear to correlate with three size ranges of neuroendocrine granules seen by electron microscopy.


Reduction of the extent of ischemic skeletal muscle necrosis by perfusion with oxygenated perfluorocarbon. Oxygenated perfluorocarbon emulsion has been shown to preserve feline cerebral function after ischemia. The postulated protective effects of perfluorocarbons include improvement of blood rheology and prevention of neutrophil adherence by nonchemical inhibition of surface receptors. In this study, we used a well-described gracilis muscle model to investigate whether oxygenated perfluorocarbon can minimize skeletal muscle necrosis by mitigating the degree of leuko-sequestration. In eight adult mongrel dogs, both gracilis muscles were weighed and then subjected to 6 hours of normothermic ischemia followed by 48 hours of normothermic reperfusion. However, one randomly selected side (experimental side) was infused with oxygen (O2) Fluosol-DA 20% (4.4 +/- 0.2 mL O2/100 mL) intra-arterially at 12 mL/min for 40 minutes immediately after ischemia. Muscle biopsy specimens were obtained before ischemia and after 1 hour and 48 hours of reperfusion to estimate myeloperoxidase (MPO) activity, a marker of neutrophil infiltration. After 48 hours, both gracilis muscles were harvested and weighed in all animals. Muscle necrosis was measured by serial transections, nitroblue tetrazolium staining, and computerized planimetry. The transmuscular oxygen tension (pO2) of the gracilis muscle on the experimental side increased from 2 to 4 mm Hg during ischemia to 315 +/- 50 mm Hg during O2 Fluosol-DA 20% infusion. The percentage of muscle necrosis on the control side was 48.08% +/- 8.46%, compared with 27.62% +/- 6.96% on the experimental side (p less than 0.001). MPO activity was significantly higher at 48 hours of reperfusion compared with pre-ischemic and 1-hour reperfusion values (5.46 +/- 1.52 U/mg tissue protein versus 0.06 +/- 0.01 U/mg tissue protein and 0.16 +/- 0.06 U/mg tissue protein, respectively, in the control group; 1.78 +/- 0.60 U/mg tissue protein versus 0.16 +/- 0.08 U/mg tissue protein and 0.27 +/- 0.10 U/mg tissue protein, respectively, in the experimental group, p less than 0.05). However, MPO activity at 48 hours of reperfusion in the experimental group was significantly lower than in the control group (p less than 0.05). There was no difference in the percentage of weight gain between the control and the experimental groups (38.31% +/- 9.36% and 28.34% +/- 7.35%, respectively, p greater than 0.05). These data show that perfluorocarbons minimize the extent of skeletal muscle necrosis in this canine model. Based on our data on MPO activity, we believe t hat the protective effect of perfluorocarbons is in part due to th e decreased leuko-sequestration in the muscle during the the periods of ischemia and reperfusion.(ABSTRACT TRUNCATED AT 400 WORDS)


Verruciform xanthoma of the ear with coexisting epidermal dysplasia. We report a case of verruciform xanthoma of the ear in a 79-year-old man. The case is unique in that it contained an area of solar keratosis. In situ hybridization using biotin-labeled probe cocktails for human papillomavirus types 6/11, 16/18, and 31/33/35 yielded negative results.


[Failure of relaxometry in diabetic polyneuropathy]. Disturbances of the peripheral nervous system, summarised under the term "diabetic polyneuropathy", are a well-known complication of long-term diabetes mellitus. However, there is little information about the fact that these conditions may lead to difficulties in monitoring neuromuscular blockade by peripheral nerve stimulation during general anaesthesia. We report two diabetic patients (40 resp. 62 years of age) in whom a total of four ophthalmological operations were performed under general anaesthesia. In all cases monitoring of neuromuscular transmission was attempted by stimulation of the left ulnar nerve slightly proximal to the wrist, but not even the usual calibration could be obtained in either patient. Checks confirmed that the nerve stimulator was functional and the electrodes--in all cases self-adhesive, pregelled surface electrodes--were placed correctly. In order to evaluate the hypothesis that disturbances of peripheral nerve function might be responsible, the patients were examined neurophysiologically in the postoperative period. Both patients showed severe alterations in peripheral nerve conduction: a sensory response could not be evoked in any of the extremities and the ulnar distal motor latency time was pathologically prolonged (5.4 ms in patient 1 and 4.9 ms in patient 2; normal: 1.4-4.0 ms). Therefore, in accordance with the clinical symptoms, the diagnosis of diabetic polyneuropathy was established. These cases demonstrate that severe disturbances of the peripheral nervous system may render neuromuscular monitoring impossible.


Poly(ADP-ribose) synthesis and degradation in mammalian nuclei. Poly(ADP-ribose) built from NAD+ on histones and other nuclear proteins by poly(ADP-ribose) polymerase is involved in repair, replication, gene expression, recombination, and chromatin remodeling in embryogenesis. Such nuclear processes are believed to be facilitated by opening up of condensed chromatin structures and by removal of histones from DNA at damaged sites as well as at origins of replication and transcription initiation sites. In addition, poly(ADP-ribosyl)ation might be involved in the up or down regulation of the activity of key nuclear enzymes. Poly(ADP-ribose) is rapidly synthesized at sites containing DNA strand breaks and is then rapidly degraded (half-life 0.5-5 min) by poly(ADP-ribose)glycohydrolase. High-resolution polyacrylamide gel electrophoresis is used in this study to analyze the rate of consumption of [32P]NAD+, the rate of formation of poly(ADP-ribose) molecules, and the rate of appearance of ADP-ribose, AMP, and phosphoribosyl-AMP, the catabolites of poly(ADP-ribose) in isolated nuclei from mouse cells in culture. Our method permits direct loading of aliquots of nuclei at time intervals on the polyacrylamide gel. The action of poly(ADP-ribose) glycohydrolase that degrades the polymer starts at less than 2 min from polymer formation. A poly(ADP-ribose) phosphodiesterase present in mammalian cell nuclei begins degrading poly(ADP-ribose) or unincorporated NAD+ and free ADP-ribose at 10 min. Mammalian phosphodiesterase is identified as an enzyme more important than previously thought which might degrade poly(ADP-ribosyl)ated proteins but also recycle the ADP-ribose produced from di- to poly(ADP-ribosyl)ated proteins by glycohydrolase into utilizable AMP units.


Adrenocorticotropic cell distribution in adult and embryonic pituitaries of the little (lit) mutant mouse. Immunofluorescence and colloidal gold immunocytochemistry were used to analyze the regional distribution of adrenocorticotropic (ACTH) cells and their ultrastructural relationship with growth hormone (GH) cells in adult and 17-day embryonic little (lit/lit) mice that exhibit GH cell defects. Adult lit/lit pituitaries lack the distinctive regionalization of ACTH cells that characterizes normal pituitaries, although typical ultrastructural relationships occur between some ACTH and GH cells. In 17-day embryos, normal and lit/lit pituitaries show similar distributions of ACTH cells. However, in lit/lit pituitaries the pars distalis cells are more loosely arranged, with poorly defined clusters and cords, than in normal glands. The results indicate that whereas the lack of ACTH cell regionalization in adult glands may be a secondary effect of the GH cell defect, the differences in overall integrity of the 17-day embryonic glands suggest the possibility of a developmental disturbance during early stages of gestation in this mutant.


Tubule formation and elemental detection in developing opossum enamel. Most marsupials and some placental mammals possess enamel characterized by the presence of tubules, and the cellular origin of these structures has been the subject of a number of previous studies (See, for example, Lester, 1970; Azevedo and Goldberg, 1987). In the present report, tooth germs of the American opossum were examined to determine the structure and composition of enamel tubules during development and to analyze the enamel matrix relative to that of placental mammals with atubular enamel. For this purpose, tissues prepared by aqueous (decalcified and undecalcified) and anhydrous (undecalcified) methods were investigated by conventional transmission (TEM) and high voltage electron microscopy (HVEM), as well as by electron probe x-ray microanalysis (EPMA), selected-area electron diffraction (SAED), and electron spectroscopic imaging (ESI). Results indicate that most enamel tubules in the opossum begin as cytoplasmic remnants of Tomes' processes of ameloblasts. During development of the matrix, some of the tubules do not appear to be continuous throughout the prismatic layer. Sulfur is detectable around the lumen of the tubule in decalcified sections by EPMA and in and around the tubule by ESI. Calcium/phosphorus (Ca/P) molar ratios of the mineralizing matrix are generally higher than those found in enamel of other mammals and appear to decrease rather than increase with enamel maturation. The summary of data indicates the presence of sulfated glycoproteins or proteoglycans in this tissue, specifically around enamel tubules. Calcium and phosphorus are also present within the tubules, with the sulfated groups possibly binding calcium to prevent mineralization of the enamel tubules themselves.


Localization and activity of Na+,K(+)-ATPase in the ductuli efferentes of the rat. The Na+,K(+)-ATPase enzyme through its p-nitrophenyl phosphatase activity was localized in the ductuli efferentes of rats. Enzymatic activity was demonstrated along the cytoplasmic side of the plasmalemma of the ductular epithelial cells. The most intense deposition of reaction products was found on the plasmalemma delimiting the lower lateral and basal regions of the cells. The plasma membranes forming the microvilli, apical junctional complexes were devoid of reaction product while the midlateral membranes showed a weak reaction. The enzyme reaction was potassium-dependent and was abolished by addition of 10 mM ouabain to the incubation media. Enzyme activity decreased significantly from proximal to distal regions of the ductules (8,101.47 +/- 274.53, 6,658.95 +/- 269.53 and 4,668.10 +/- 575.41 pmoles p-nitrophenol/mm/h, respectively in initial, conus vasculosus and terminal zones). A unified model for water absorption is proposed in the efferent ductules based upon this data and that of others.


Stress hormone response during and after cardiopulmonary resuscitation. The purpose of this study was to assess whether plasma adrenocorticotropin, cortisol, vasopressin, and renin concentrations are higher in resuscitated than in nonresuscitated patients during cardiopulmonary resuscitation, and whether there are possible correlations between these hormones and blood pressure or heart rate in the immediate postresuscitation phase. Of 34 consecutive patients (36-85 yr of age) with out-of-hospital cardiac arrest, 20 could be successfully resuscitated and admitted to hospital, whereas in the remaining 14 patients restoration of spontaneous circulation could not be achieved. During cardiopulmonary resuscitation, median adrenocorticotropin, cortisol, vasopressin, and renin concentrations in the external jugular vein were 237 pg/ml, 32.6 micrograms/dl, 122 pg/ml, and 46.5 ng/l, respectively, in resuscitated patients, and 45 pg/ml (P = 0.018), 18.4 micrograms/dl (P = 0.481), 88 pg/ml (P = 0.049), and 11 ng/l (P = 0.017), respectively, in nonresuscitated patients. Median adrenocorticotropin, cortisol, vasopressin, and renin concentrations were 101 pg/ml, 34.6 micrograms/dl, 22 pg/ml, and 25 ng/l, respectively, 60 min after successful resuscitation. No significant correlations were found between hormone levels and blood pressure or heart rate, but there was a significant negative correlation between the interval from collapse to the start of cardiopulmonary resuscitation and plasma cortisol concentrations during cardiopulmonary resuscitation (Spearman rank correlation coefficient = -0.967, P less than 0.001), indicating an impaired cortisol release from the adrenal cortex. The lower hormone concentrations of the nonresuscitated patients measured during cardiopulmonary resuscitation might indicate an impairment in neuroendocrine response.


Intrathecal pertussis toxin treatment attenuates opioid antinociception and reduces high-affinity state of opioid receptors. The effect of pertussis toxin on opioid antinociception was studied in rats. Intrathecal injection of a single dose of pertussis toxin reduced the antinociceptive effect of PL017, a highly selective mu-opioid agonist, in a dose- and time-dependent manner. The maximal effective dose of pertussis toxin was 1 microgram, and the maximal effect was seen at day 3. The effect of the toxin lasted for 2 weeks, and the antinociceptive response recovered partially at the third week. The dose-response curves of the antinociceptive effect of PL017 were shifted to the right with increasing doses of pertussis toxin. Three days after pertussis injection, receptor-binding activity of membranes in the lumbosacral segment of spinal cords decreased to 70% of control as assayed by 125I-FK33824, a highly selective mu-receptor agonist. In experiments using [3H]naloxone as the radiolabeled ligand, displacement curves of FK33824 were shifted to the right after pertussis toxin treatment. The shift also was dose and time dependent. Scatchard analysis of binding data showed that, after pertussis toxin treatment, there was no significant change in the total number of binding sites, but a class of low-affinity binding sites appeared in addition to the high-affinity sites. When spinal membranes were washed in Na+ (100 mM) and guanosine diphosphate (100 microM) and binding was assayed in the presence of Mg2+ (5 mM), all the mu-receptors were in the high-affinity state in control membranes. After the pertussis toxin treatment, the ratio of low-affinity sites to high-affinity sites increased.(ABSTRACT TRUNCATED AT 250 WORDS)


DNA sequence of the skeletal muscle calcium release channel cDNA and verification of the Arg615----Cys615 mutation, associated with porcine malignant hyperthermia, in Norwegian landrace pigs. Porcine calcium release channel (CRC) cDNA from skeletal muscle has been cloned and sequenced. The deduced amino acid sequence showed 97% identity to the corresponding rabbit and human sequences. Using oligonucleotide primers based on the nucleotide sequence, CRC cDNA fragments from seven pigs representing HALNN, HALNn and HALnn genotypes have been amplified. Sequencing and restriction digestion of the amplified cDNA confirm that the reported C----T mutation, which gives rise to Arg615----Cys615 change in the calcium release channel, is associated with the halothane sensitive allele in Norwegian Landrace pigs. The mutation may alter the reactivity of a neighbouring serine residue which is potentially phosphorylated.


Effects of nedocromil sodium and WEB 2086 on chemoattractant-stimulated neutrophil migration through cellular and noncellular barriers. Nedocromil sodium (Tilade) is an effective therapeutic agent against asthma and has been shown to exhibit antiinflammatory activity in vitro; however, its mode of action is yet to be described fully. Using an in vitro assay designed to mimic the extravasation of neutrophils from the peripheral circulation through cellular barriers to sites of inflammation, the effect of nedocromil sodium on chemoattractant-stimulated neutrophil migration was examined. We also examined the effects of WEB 2086, a platelet-activating factor (PAF) receptor antagonist, in parallel. Neutrophils and the cellular barrier were pretreated and/or co-incubated with nedocromil or WEB 2086 and the effects on neutrophil chemotaxis measured. In all treatments, nedocromil did not significantly affect chemotaxis through cellular or noncellular barriers to N-formyl-methionyl-leucyl-phenylalanine (FMLP), leukotriene B4 (LTB4), or PAF. In contrast, WEB 2086 inhibited PAF-induced neutrophil migration through both naked filters and endothelial and epithelial monolayers cultured on these filters. We conclude that while nedocromil has been shown to have inhibitory effects on neutrophils and is an effective therapeutic agent for asthma and inflammatory conditions, its activity is not primarily mediated by inhibition of neutrophil chemotaxis. Platelet-activating factor antagonists may partially be effective in asthma through inhibitory effects on neutrophil chemotaxis.


Allergen-induced changes in adenosine 5'-monophosphate bronchial responsiveness: effect of nedocromil sodium. Bronchial hyperresponsiveness to adenosine 5'-monophosphate (AMP) was studied after allergen challenge in allergic asthmatic patients. Measurements were made with and without nedocromil sodium pretreatment. Nedocromil sodium inhibited both the early and late asthmatic reactions (P < .01). After allergen challenge a significant decrease in PC20 AMP from 12.2 mg/mL to 4.47 mg/mL (P < .05) at three hours was found, returning almost to baseline values: 10.85 mg/mL (P > .05) at 24 hours. Nedocromil sodium, 6 mg, given before allergen challenge prevented the increased responsiveness to AMP at three hours [PC20 10.12 mg/mL (P < .05)], but caused a decrease in PC20 AMP at 24 hours to 6.32 mg/mL (P < .05). Desensitization of the adenosine receptor during the late asthmatic reaction, which is prevented by nedocromil sodium, may explain the lack of increased responsiveness at 24 hours. AMP may play a physiologic role in allergen-induced late phase reactions.


Health care worker exposure to HIV-1 and HTLV I-II in critically ill, resuscitated emergency department patients. Exposure to HIV-1 is of profound concern to health care workers. HTLV-I and HTLV-II, retroviruses with similar modes of transmission as HIV-1, also cause disease in human beings. Emergency department resuscitations are high-risk situations for such exposure. The purpose was to determine the seroprevalence of HIV-1 and HTLV I-II in patients undergoing ED resuscitations, the magnitude of health care worker exposure, and risk factors associated with infection. Prospective identity-unlinked seroepidemiologic study. ED of a 950-bed private inner-city teaching hospital. Participants included 370 patients undergoing ED resuscitations. Serum was tested for antibodies to HIV-1 and HTLV I-II. Questionnaires were completed by the physician in charge of the ED resuscitations. Fifteen (4.1%) (95% confidence interval [CI], 2.1% to 6.1%) patients were HIV-1 seropositive, and seven (1.9%) (95% CI, 0.7% to 3.1%) were HTLV I-II positive. Eleven (5.6%) (95% CI, 2.4% to 8.8%) of 197 trauma patients and 11 (6.4%) (95% CI, 2.8% to 10.0%) of 173 medical patients were infected with one of these viruses. Health care workers had direct cutaneous contact with patient blood during 114 (31%) ED resuscitations and with infected patient blood during 11 (3%) ED resuscitations. An additional 11 ED resuscitations involved parenteral exposures, one to HIV-1-infected blood. No factors could be identified that would quickly and reliably predict infection. Health care workers must protect themselves in such high-risk situations by strict compliance to mandatory universal precautions.


Search for bovine papilloma virus DNA in bovine ocular squamous cell carcinomas (BOSCC) and BOSCC-derived cell lines. Although the cause of bovine ocular squamous cell carcinoma (BOSCC) is attributed to viruses in addition to cofactors (eg, UV light), to our knowledge, the final causative agent has not been described. Bovine papilloma virus (BPV)-like particles were detected in approximately 33% of various putative precursor lesions of BOSCC. In contrast, it was reported that, using BPV-specific antibodies, it was not possible to detect viral antigens in BOSCC. Fourteen established BOSCC and 9 BOSCC-derived cell lines were examined for BPV DNA. Probes of all 6 known BPV types were used in various hybridization assays. Neither Southern blot analysis, under high and low stringency conditions, nor in situ hybridization resulted in detection of BPV DNA. Papilloma viruses were not observed in electron microscopic studies. Results exclude direct association between BOSCC and BPV types 1 to 6, or as yet unknown closely related BPV types. However, BPV may contribute to induction of precursor lesions or events leading to carcinogenic transformation, without being relevant for maintenance of the tumor.


Induction and enhancement of feline infectious peritonitis by canine coronavirus. Preexisting antibody to feline infectious peritonitis virus (FIPV) causes acceleration and enhancement of disease on subsequent infection of cats with FIPV. Other workers have shown that canine coronavirus (CCV) can infect cats subclinically, but have found no evidence of enhancement of, or protection against, subsequent FIPV infection. With various isolates of CCV, we determined that 1 strain of CCV can induce transient mild diarrhea in cats and, furthermore, that previous infection with CCV causes acceleration and enhancement of subsequent infection with FIPV. In addition, sequential inoculation of cats with another strain of CCV caused lesions indistinguishable from those of FIP, without exposure at any time to FIPV.


Influence of arachidonic acid metabolites and steroids on function of bovine polymorphonuclear neutrophils. Polymorphonuclear neutrophils (PMN) from 4 ovariectomized healthy cows were incubated with 0 (control), 10(-8), 10(-7), and 10(-6) M arachidonic acid metabolites of the cyclo- and lipoxygenase pathways for 30 minutes, and with steroids for 2 hours. Immediately after incubation, PMN were subjected to the following function assays: chemotaxis against zymosan-activated serum, chemotaxis against arachidonic acid metabolite or steroid at the doses given (only control PMN were tested), random migration, ingestion of 125I-iododeoxyuridine-labeled Staphylococcus aureus (125I-IdUR-S aureus), iodination of proteins, cytochrome C reduction, antibody-independent and -dependent cell-mediated cytotoxicity (AICC and ADCC). Prostaglandin F2 alpha was chemoattractant and stimulated ingestion of 125I-IdUR-S aureus. Prostaglandin E2 stimulated cytochrome C reduction, whereas prostacyclin inhibited iodination of proteins. Thromboxane B2 stimulated ADCC. Leukotriene B4 was chemoattractant for bovine PMN and stimulated random migration and AICC. 5-Hydroxyeicosatetraenoic acid was also chemoattractant, but inhibited ingestion of 125I-IdUR-S aureus. 15-Hydroxyeicosatetraenoic acid was chemoattractant and decreased ADCC. Lipoxin A4 stimulated random migration, whereas lipoxin B4 inhibited chemotaxis against zymosan-activated serum, but was chemoattractant and stimulated cytochrome C reduction. 12-Hydroxyhepadecatrienoic acid and 12-hydroxyeicosatetraenoic acid did not influence any of the PMN functions tested. Of the steroids tested, cortisol increased ADCC, and progesterone stimulated cytochrome C reduction, but decreased ADCC. 17 beta-Estradiol and estrone were chemoattractant and stimulated cytochrome C reduction. In addition, estrone also stimulated random migration.(ABSTRACT TRUNCATED AT 250 WORDS)


Maintenance energy requirement of llamas. Five castrated male llamas (mean body weight, 94 kg) were studied in an energy balance trial to determine maintenance energy requirement of llamas. Llamas were fed a 50% oat hay-50% pelleted concentrate diet (2.43 Mcal of metabolizable energy/kg of diet dry matter) at approximately 1.6% of body weight (BW). An 8-day total collection digestion trial was used to determine fecal and urine energy losses. Heat production and methane emissions were determined via indirect respiration calorimetry measurements on each llama fed at the same level of intake as during the digestion trial and subsequently on days 3 and 4 of a period of nonfeeding. Fecal, urine, and methane energy losses of the llamas fed near-maintenance intake were 32.5, 3.5, and 7.1% of gross energy intake, respectively. The postabsorptive metabolic rate, commonly called nonfed (fasting) heat production, was 59.3 kcal/BW0.75. Using a linear relation between postabsorptive and maintenance energy requirement and efficiency of energy use below maintenance of 0.702, metabolizable energy requirement at maintenance was determined to be 84.5 kcal/BW0.75.


In search of how people change. Applications to addictive behaviors. How people intentionally change addictive behaviors with and without treatment is not well understood by behavioral scientists. This article summarizes research on self-initiated and professionally facilitated change of addictive behaviors using the key trans-theoretical constructs of stages and processes of change. Modification of addictive behaviors involves progression through five stages--pre-contemplation, contemplation, preparation, action, and maintenance--and individuals typically recycle through these stages several times before termination of the addiction. Multiple studies provide strong support for these stages as well as for a finite and common set of change processes used to progress through the stages. Research to date supports a trans-theoretical model of change that systematically integrates the stages with processes of change from diverse theories of psychotherapy.


Dynamics of the site visit. Laboratory for the small group study of scientific administration. Site visits are conducted by National Institutes of Health review groups when obstacles arise in peer appraisal of research projects. Because such visits provide a window into the state of a scientific community and present highly charged group dynamics that revolve about significant scientific issues, examples from the 1970s and 1980s were subjected to behavioral analysis. Selye's general adaptation syndrome was used to model the forms of coping behavior manifested by investigators targeted for a site visit. A Lewinian approach was taken to analyze the course and outcome of site visit team deliberations. The perspective was that of the NIH official responsible for the operation of the review process.


Abnormal lung function in polyurethane foam producers. Weak relationship to toluene diisocyanate exposures. Exposures to toluene diisocyanate (TDI) were studied for effects on respiratory health of workers in two plants manufacturing polyurethane foams. Intensive personal monitoring was used to characterize job exposures. Of 4,845 12-min personal samples, 9% exceeded 5 ppb and 1% exceeded 20 ppb. Initial questionnaire and spirometry were obtained in 386 workers (88.7% of target population). Current smoking was associated with lower mean FEV1 and FEF25-75, but percent predicted (% pred) means were normal in all smoking categories. Multiple regression showed significant adverse effects of cumulative TDI exposure on initial level of FVC and FEV1 of current smokers, and an effect at borderline significance (p less than 0.063) on FEF25-75 over all smoking categories. Logistic regression showed that chronic bronchitis was more prevalent among those with higher cumulative exposures, after controlling for smoking, age, and sex. Methacholine (MCh) reactivity was associated with reduced airway function, -8.5% pred for FEV1 and -20.0% pred for FEF25-75. In 227 with adequate follow-up, the slopes of annual change were abnormal, for example, FEV1 of -67 ml/yr in current and -53 ml/yr in never smokers. Men had worse FEV1 declines than did women, -71 ml/yr versus -43 ml/yr. TDI exposure, lifetime or concurrent, had no significant effect on slopes, despite its demonstrated effects on initial level of lung function and on prevalence of chronic bronchitis.


Enhanced production of oxygen radicals in nocturnal asthma. Although the mechanisms of nocturnal worsening of pulmonary function in asthmatics have not been entirely established, airway inflammation is felt to be a major factor in disease severity. Consequently, to determine whether changes in bronchoalveolar lavage (BAL) fluid cellular components and their functions are related to nocturnal airway obstruction, we performed BAL at 4:00 A.M. and at 4:00 P.M. in asthma subjects with (n = 5) and without (n = 10) nocturnal asthma. No significant changes were observed from 4:00 P.M. to 4:00 A.M. in the concentration of total cells or the percentage or concentration of eosinophils or neutrophils in BAL fluid from subjects with or without nocturnal asthma. However, superoxide anion generation by air-space cells from subjects with nocturnal asthma was significantly greater at 4:00 A.M. than at 4:00 P.M. (6.9 +/- 1.7 versus 1.8 +/- 0.5 nmol/500K cells/h, p less than 0.05). Moreover, superoxide production at 4:00 A.M. was greater in subjects with than in those without nocturnal asthma (6.9 +/- 1.7 versus 2.2 +/- 0.6, p less than 0.02). Furthermore, in our group of asthmatics, the change in generation of superoxide anion from 4:00 P.M. to 4:00 A.M. was significantly correlated with the change in FEV1 (r = -0.71, p less than 0.01). We conclude that the development of nocturnal airway obstruction in asthma is associated with enhanced production of oxygen radicals by air-space cells. Because oxygen radicals can cause airway injury and thus enhance bronchial obstruction, it is possible that the release of these reactive compounds is causally associated with nocturnal asthma.


Preoperative radiological evaluation of the liver by computerized tomographic portography in patients with hepatic tumors. The main objective of preoperative imaging studies is to define as accurately as possible the number, size, location, and relationship of tumor masses in the liver to pertinent portal and hepatic venous vasculature. Computerized tomographic portography images hepatic veins and segmental portal vein branches and identifies the anatomical location of tumor nodules with excellent sensitivity and a low false-positive rate. The intraoperative correlation of computerized tomographic portography on 30 patients in the last 20 months at this institution shows a sensitivity of 88 per cent with a low rate of false-positivity. The ability to detect metastatic lesions in the liver by computerized tomographic portography diminishes when the lesions are noted to be less than 1 cm. The authors conclude that the preoperative interpretation of the computerized tomographic portogram provides valuable information not previously available to the surgeon operating on the liver.


The J-pouch Swenson procedure for ulcerative colitis and familial polyposis. Total colectomy with mucosal proctectomy and ileal pouch-anal anastomosis has proven to be a favorable option in the treatment of ulcerative colitis and familial polyposis coli. The main advantages of this procedure are that it obviates the need for a permanent stoma, it preserves anal continence, and it removes all disease-prone mucosa. As an alternative to this procedure, the authors have found success with the Swenson pull-through following proctocolectomy in children. This technique involves resection of the rectum at the dentate line, thus, eliminating the need for mucosal proctectomy. This may be particularly advantageous in patients with severely diseased rectal mucosa. Ileoanal anastomosis is performed after creation of an ileal J-pouch using the terminal ileum. The procedure has been used in two children with familial polyposis coli and in three with ulcerative colitis. Median follow-up after closure of the diverting ileostomy is 13 months (5-33 months). Continence has been preserved in all five patients. There have been no complications involving bladder or sexual dysfunction. This technique provides a reliable alternative for the definitive treatment of ulcerative colitis and familial polyposis coli.


Spiral modiolar vein: its importance in viral load of the inner ear. Guinea pig-specific cytomegalovirus and Sendai virus were inoculated into the cochleas of seronegative guinea pigs to study the route of entry of cells participating in inner ear inflammation. Inflammatory cells accumulated around the spiral modiolar vein and appeared to be streaming from this vein into the scala tympani via a collecting venule. Inactivated virus inoculated into the cochlea and normal control cochlea failed to show inflammatory cell infiltrates. The spiral modiolar vein appears to play an important role in the movement of cells from the systemic circulation into the inner ear as part of the host's normal defense against invading pathogens such as viruses.


Prevention of bacterial infection and sepsis in acute severe pancreatitis. Between 1984 and 1986 six patients with acute respiratory failure (requiring ventilation for at least 3 days) complicating acute pancreatitis were managed on the intensive care unit (median ventilation period 6 days; range 3-41 days). Between 1987 and 1989 nine similar patients were managed (median ventilation period 35 days, range 4-69 days), and a regimen of enteral tobramycin, polymyxin and amphotericin to selectively decontaminate the digestive tract (SDD) was introduced. Five of six patients treated before 1987 had serious infections (three Gram-negative, one fungal), compared with only one of nine patients treated with SDD (P < 0.05). Clinical signs of sepsis were evident for 62% of the pre-SDD period, compared with 39% of the period during SDD therapy (P < 0.001). Systemic antibiotic prescribing was reduced in the SDD group; however, mortality remained unaffected with only two patients surviving pre-SDD and three during SDD treatment. SDD reduces infection rates and sepsis in patients with acute pancreatitis and may help to improve the prognosis of this life-threatening condition.


[Induction of hypertrophy of a small left hepatic lobe by preoperative right portal embolization, preceding extended right hepatectomy]. The aim of this study was to evaluate the compensatory hypertrophy of the left lobe of the liver, induced by a preoperative right portal embolization (PORPE), and then the feasibility of a right extended hepatectomy. The small size of the left lobe did not initially permit such a resection. Eight patients (mean age: 62 years) underwent PORPE for cancer between September 1987 and December 1991. They represented 4% of the 187 patients undergoing hepatectomy for liver cancer during the same period. The PORPE was conducted by percutaneous access and puncture of the left portal vein (Rex's recessus). The clinical and laboratory safety were good, with fewer adverse effects than with arterial chemo-embolization. The mean increased volume of the left lobe, four weeks after PORPE, was 54% (range: 32-100%) allowing hepatectomy to be performed. The post-operative course of these right extended hepatectomies was uneventful. In conclusion, we think that PORPE needs a careful technique but that it is well tolerated and effective to induce hypertrophy of the future remnant left lobe. It allows resection of some initially unresectable tumors. This technique warrants further development.


Malignant mixed tumour arising from minor salivary gland tissue of the right parapharyngeal space with metastasis to the scalp. A case of malignant mixed tumour arising from minor salivary gland tissue in the right parapharyngeal space with metastasis to the scalp is described. Both the clinical presentation and the histological picture were unusual. The simultaneous discovery of the primary parapharyngeal tumour and its scalp metastasis, the relatively young age of the patient (43 years-old), the origin of the tumour in minor salivary gland tissue, and the presence of a benign stromal component in the metastasis are features not commonly described in the three entities covered by the term "malignant mixed tumour". We believe this case represents a distinct variant, whose behaviour and progression have not been previously well documented.


Embryonal carcinoma arising in Turner's syndrome. Risk of malignant transformation in a dysgenetic gonad is high. This is a report of a 23 year-old female who presented with symptoms and signs of ovarian neoplasm which was histologically confirmed as embryonal carcinoma. She had features of Turner's syndrome and was found to have XO chromosomal constitution. Embryonal carcinoma arising in a dysgenetic gonad is uncommon. It is even rarer when it arises in a patient with pure 45 XO. The following is a case report which highlights the role of chemotherapy and surgery in the management. It includes a literature review on the clinical features, genetic variants and malignant transformation in dysgenetic gonad. The role of prophylactic removal of dysgenetic gonad is discussed.


Abnormalities of somatosensory evoked potentials in the quinolinic acid model of Huntington's disease: evidence that basal ganglia modulate sensory cortical input. Intrastriatal injection of quinolinic acid (QA) in rats provides an animal model that mimics some of the neuropathological and neurochemical alterations observed in the striatum of patients with Huntington's disease (HD). One of the very early neurophysiological signs in HD is a diminution of amplitude of early somatosensory evoked potentials (SEPs) recorded over the parietal cortex. The present study investigated whether the QA model exhibits similar neurophysiological abnormalities. Two weeks after unilateral intrastriatal injection of QA (240 nmol) or of the solvent, early SEPs were recorded with chronically implanted electrodes from the somatosensory cortex or from the ventrobasal nucleus of the thalamus of lightly pentobarbital-anesthetized rats, in response to single-shock electrical stimulation of the contralateral forepaw. Whereas intrastriatal injection of solvent did not influence SEPs, the striatal QA lesion significantly reduced the amplitude of early cortical SEPs by about 40% without affecting the latency. SEPs recorded from the ventrobasal nucleus were unchanged after QA lesion. Histological examination and glial fibrillary acid protein staining after intrastriatal injection of QA revealed no evidence for damage in the somatosensory system. It is concluded that (1) the QA animal model of HD mimics some of the SEP abnormalities of patients, and (2) a striatal lesion modulates somatosensory transmission to the cortex in rats.


CI-960, a new fluoroquinolone, for therapy of experimental ciprofloxacin-susceptible and -resistant Staphylococcus aureus endocarditis. CI-960 is a new fluoroquinolone with enhanced in vitro activity against gram-positive pathogens. The efficacy of the drug was compared with that of vancomycin by using the rabbit model of nafcillin- and ciprofloxacin-susceptible and -resistant Staphylococcus aureus endocarditis. Animals received intravenous therapy with CI-960, 20 mg/kg of body weight every 8 h, or vancomycin, 17.5 mg/kg every 6 h, for 4 days. In a comparison with the effects on untreated controls, both antimicrobial agents effectively cleared bacteremia and significantly reduced bacterial counts in vegetations and tissues of animals infected with any of the test strains. In some cases, the efficacy of CI-960 was superior to that of vancomycin. The therapeutic activity of CI-960 was reduced, but still very good, against ciprofloxacin-resistant strains. One rabbit infected with such a strain and treated with CI-960 was found to harbor a small number of vegetation-associated organisms resistant to the drug at fivefold its original MIC; this was associated with a microbiological, but not a clinical, failure of therapy. We conclude that CI-960 is as effective as vancomycin is in this model of a serious systemic S. aureus infection, including that caused by strains resistant to ciprofloxacin. Increases in CI-960 MICs may develop during therapy of infections caused by strains highly resistant to ciprofloxacin, but they appear unlikely to occur in ciprofloxacin-susceptible strains.


Identical genes confer high-level resistance to gentamicin upon Enterococcus faecalis, Enterococcus faecium, and Streptococcus agalactiae. The structural gene coding for the bifunctional aminoglycoside-modifying 6'-acetyltransferase-2''-phosphotransferase (6'AAC-2''APH) enzyme was specifically amplified by the polymerase chain reaction using template DNA of clinical isolates of enterococci (both Enterococcus faecalis and Enterococcus faecium) and the single high-level gentamicin-resistant Streptococcus agalactiae strain identified thus far. The results of the present study demonstrated that the genes encoding this antibiotic resistance trait are highly homologous in these species. In dot blot hybridization assays using nonradioactively labeled oligonucleotide probes, strains with and without high-level gentamicin resistance could be discerned unequivocally. The gene segment encoding 6'-acetylating activity and the gene segment encoding 2''-phosphorylating activity were simultaneously present in all isolates exhibiting high-level gentamicin resistance.


Effect of antacid on absorption of the quinolone lomefloxacin. The effect of antacid on the absorption of lomefloxacin (LFLX) in humans was studied. When LFLX was orally administered concomitantly with aluminum- and magnesium-containing antacids under fasting conditions, its level in plasma decreased by one-half and its area under the concentration-time curve was reduced by 40% compared with the levels observed after treatment with LFLX alone. The urinary recovery value also decreased by 40%. No such effects were noted after coadministration of LFLX and a nonmetallic antacid. This study confirmed the existence of chelate complexes of LFLX with Al3+ and Mg2+ and examined the chelating strength. The stability constants of LFLX with Al3+ and Mg2+ were measured and compared with those of ofloxacin and norfloxacin; little difference was observed among them. LFLX was found to bind more strongly with Al3+ than with Mg2+. Further, the existence of chelate formation was proven by 13C-nuclear magnetic resonance spectroscopy. The decrease in the LFLX level in plasma in humans could be explained by a reduced absorption of the Al(3+)- and Mg(2+)-LFLX chelate complexes.


Clindamycin, erythromycin, and roxithromycin inhibit the proinflammatory interactions of Pseudomonas aeruginosa pigments with human neutrophils in vitro. The Pseudomonas aeruginosa-derived phenazine pigments pyocyanin and 1-hydroxyphenazine (1-hp) prime human neutrophils for enhanced, stimulus-activated release of superoxide and myeloperoxidase (MPO), respectively. In the present study, the modulatory potentials of the antimicrobial agents clindamycin, erythromycin, and roxithromycin (10 and 20 micrograms/ml) on the prooxidative interactions of pyocyanin and 1-hp (12.5 microM) with human neutrophils have been investigated. Clindamycin, erythromycin, and especially roxithromycin caused dose-related inhibition of the generation of superoxide by both untreated and pyocyanin-treated neutrophils during activation with either the synthetic chemotactic tripeptide N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP) or the calcium ionophore A23187. The antimicrobial agents also inhibited the generation of reactive oxidants by the MPO-H2O2-halide system during activation of both untreated and 1-hp-treated neutrophils by FMLP. These effects appeared to be due to drug-related interference with membrane-associated oxidative metabolism, since none of the antimicrobial agents inhibited the release of MPO by activated neutrophils, nor did they possess oxidant-scavenging properties. These data demonstrate that clindamycin, erythromycin, and especially roxithromycin antagonize the proinflammatory interactions of pyocyanin and 1-hp with neutrophils and indicate a possible therapeutic role for these antimicrobial agents in the prevention of tissue damage in diseases characterized by P. aeruginosa infection.


In vitro activity of azithromycin compared with that of erythromycin against Actinobacillus actinomycetemcomitans. The in vitro susceptibility of Actinobacillus actinomycetemcomitans to azithromycin, a new macrolide antibiotic of a new class known as azalides, was compared with that of erythromycin by the agar dilution method on Mueller-Hinton Haemophilus test medium. Eighty-two A. actinomycetemcomitans strains, 79 recent clinical isolates obtained from 40 periodontally healthy or diseased subjects, and 3 type strains were included in the study. Erythromycin showed poor in vitro activity against A. actinomycetemcomitans. Azithromycin, however, was highly effective against A. actinomycetemcomitans: all strains were inhibited at 2.0 micrograms/ml. Azithromycin exhibited the best in vitro activity against the serotype a subpopulation of A. actinomycetemcomitans: 100% of the strains were inhibited at 1.0 micrograms/ml. The lowest MICs were, however, recorded by serotype b strains. Since azithromycin has favorable pharmacokinetic properties, including excellent distribution into tissues, it could be expected to pass into gingival crevicular fluid at levels sufficient to inhibit A. actinomycetemcomitans in vivo. Therefore, it is a good candidate for future clinical trials in A. actinomycetemcomitans-associated periodontitis.


Multiple-dose pharmacokinetics and safety of rufloxacin in normal volunteers. The pharmacokinetics and safety of rufloxacin were evaluated in a double-blind, placebo-controlled study. Two groups of 16 healthy volunteers were given a single oral loading dose of 400 or 600 mg of rufloxacin on day 1 of the study. A single daily maintenance dose of 200 or 300 mg was then administered for a further 9 days; in addition, four subjects in each group received placebos. Rufloxacin levels in plasma and urine were determined by high-performance liquid chromatography. Following the initial dose, the mean (+/- standard error of the mean) peak concentrations of rufloxacin in plasma were 3.35 +/- 0.12 micrograms/ml in the 400-mg group and 4.54 +/- 0.19 micrograms/ml in the 600-mg group. They were generally reached 2 to 3 h after dosing. At the end of treatment, maximum levels in plasma rose to 4.51 +/- 0.15 and 7.20 +/- 0.25 micrograms/ml in the 400-mg and 600-mg groups, with a mean extent of accumulation (fold) of 3.1 +/- 0.1 and 3.3 +/- 0.1. For the 400-mg and 600-mg groups, the elimination half-lives were 40.0 +/- 1.5 and 44.0 +/- 1.3 h, mean residence times were 57.8 +/- 2.2 and 63.7 +/- 1.8 h, apparent volumes of distribution were 132 +/- 4 and 139 +/- 5 liters, and apparent total body clearance were 39 +/- 1 and 44 +/- 4 ml/min, assuming complete bioavailability. Of the total dose administered, the percentages excreted in urine were 49.6 +/- 1.3 and 51.1 +/-2.1%, with renal clearances of 21 +/- 1 and 22 +/- 2 ml/min, for the 400-mg and 600-mg groups. On the whole, the treatments were well tolerated, but some minor adverse events (mainly headache, insomnia, or abdominal discomfort) were reported for 7 subjects on abnormalities were detected in the laboratory examinations or in ocular function tests. This study shows that a 200-mg daily oral dose of rufloxacin preceded by a loading dose of 400 mg are well tolerated and produce steady-state concentrations in plasma above the MIC for most susceptible pathogens.


In vitro and in vivo intraleukocytic accumulation of azithromycin (CP-62, 993) and its influence on ex vivo leukocyte chemiluminescence. The accumulation of azithromycin in phagocytic cells was studied both in vitro by using a radiolabelled drug and a bioassay and in vivo for 12 volunteers receiving 1.5 g (total dose) orally within 3 days. In vitro, neutrophils and unfractionated blood leukocytes accumulated azithromycin up to 160-fold the extracellular concentration within 1 h at 37 degrees C but less than 3-fold at 4 degrees C. Dead cells accumulated up to 30-fold azithromycin, whereas NaF-treated cells accumulated up to 60-fold arithromycin. The mean efflux from preloaded cells was at most 31.0% +/- 10.6% (standard error of the mean) of the cell-associated concentration within 4 h of incubation at 37 degrees C in drug-free buffer. In vivo, the azithromycin concentration was 45.2 +/- 6.1 mg/liter of intracellular fluid at 2 h after the third dose and 36.6 +/- 8.3 mg/liter at 1 week thereafter. The corresponding concentrations in serum were 0.2 +/- 0.1 (2 h) and less than 0.05 (1 week). The luminol-enhanced chemiluminescence response induced by phorbol myristate acetate, opsonized zymosan, and two opsonized strains of Haemophilus influenzae (a type b capsulated strain and a noncapsulated strain) was also studied ex vivo by using the blood leukocytes from the 12 test volunteers and 4 control volunteers at 2 and 6 h after the third oral dose of azithromycin and at 2, 4, and 7 days thereafter. Azithromycin did not influence this response despite high levels of cellular accumulation.


In vitro activity of OPC-17116.

In vitro activity of OPC-17116. The in vitro activity of OPC-17116, a new C-5 methyl fluoroquinolone, was compared with the activities of other fluoroquinolones. OPC-17116 inhibited 50% of the members of the family Enterobacteriaceae tested and 90% of Haemophilus influenzae, Neisseria species, and Moraxella catarrhalis isolates at less than or equal to 0.25 microgram/ml. At less than or equal to 2 micrograms/ml, 90% of the Enterobacteriaceae were inhibited, which was comparable to or better than the activities of fleroxacin, ofloxacin, and lomefloxacin but less than the activity of ciprofloxacin. OPC-17116 inhibited 90% of the staphylococci tested at less than or equal to 0.25 micrograms/ml, but it did not inhibit methicillin-resistant, ciprofloxacin-resistant Staphylococcus aureus or Staphylococcus epidermidis. Group A, B, C, F, and G streptococci and Streptococcus pneumoniae were inhibited by less than or equal to 0.5 microgram/ml, being four-fold more active than ciprofloxacin and ofloxacin. Tosufloxacin was the most active agent tested against gram-positive cocci. OPC-17116 inhibited Bacteroides fragilis at 4 micrograms/ml. There was a minimal effect of inoculum size on MIC, and the MBCs were within 1 dilution of the MICs. The activity of OPC-17116 was decreased at pH 6 and in the presence of high Mg2+ concentrations, but it was unaffected by human serum. OPC-17116 showed a postantibiotic effect against Pseudomonas aeruginosa and Staphylococcus aureus similar to the postantibiotic effects reported for other fluoroquinolones. The frequency of spontaneous single-step resistance was low (less than 10(-9)), but repeated passage of organisms in the presence of OPC-17116 resulted in the selection of resistant isolates.


Comparative in vitro activity of PD 127391, a new fluoroquinolone agent, against susceptible and resistant clinical isolates of gram-positive cocci. We examined the in vitro activity of PD 127391, an investigational fluoroquinolone antibacterial agent, against staphylococci (including methicillin-resistant Staphylococcus aureus), enterococci (including beta-lactamase-producing and highly gentamicin-resistant isolates), and streptococci. The compound was active against all organisms tested and compared favorably with antimicrobial agents routinely used to treat infections with these organisms. On the basis of MICs for 90% of the strains tested, PD 127391 was 32-fold more active against all staphylococci, 16-fold more active against methicillin-resistant S. aureus, 8-fold more active against all streptococci, and 4-fold more active against all enterococci than ciprofloxacin. PD 127391 was shown to be more active than sparfloxacin, which in turn was shown to be more active than ciprofloxacin, against these gram-positive cocci. PD 127391 shows promise for the treatment of infections with gram-positive cocci, including organisms which are resistant to other commonly used antimicrobial agents.


Activity of azithromycin against Mycobacterium avium infection in beige mice. The comparative activities of azithromycin and clarithromycin and the activities of azithromycin alone and in combination with other antimycobacterial agents were evaluated in the beige mouse model of disseminated Mycobacterium avium complex infection. Azithromycin was similar in activity to clarithromycin. Azithromycin plus clofazimine plus ethambutol reduced the number of splenic organisms more than azithromycin alone, while the combination was less active than azithromycin alone for bacteria in lungs. Rifabutin had activity similar to that of azithromycin for organisms in spleens and lungs. Rifabutin plus azithromycin was more active than either agent alone for organisms in spleens, but the combination's activity was not significantly different from that of rifabutin for organisms in lungs. The activity of azithromycin against several M. avium complex isolates was evaluated. The reduction of viable cell counts in spleens ranged from 1.7 to 0.8 log units. For the three isolates studied, there was little correlation between the in vitro MIC and the in vivo activity.


Increased sensitivity of Candida albicans cells accumulating 14 alpha-methylated sterols to active oxygen: possible relevance to in vivo efficacies of azole antifungal agents. The sensitivity of Candida albicans cells to killing by hydrogen peroxide was found to increase markedly when they were grown in the presence of sub-growth-inhibitory concentrations of the azole drug clotrimazole (CTZ). A superoxide anion-generating system consisting of xanthine and xanthine oxidase also killed such CTZ-treated cells more efficiently than control cells, but this seemed to be accounted for by hydrogen peroxide secondarily formed from superoxide anion as judged by the effect of catalase and superoxide dismutase. The increased sensitivity to hydrogen peroxide was considered to be attributable to the inhibition of 14 alpha-demethylation of ergosterol biosynthesis by CTZ, since a 14 alpha-demethylation-deficient mutant of C. albicans exhibited a similar phenotype. It is suggested that the in vivo efficacy of azole antifungal agents against C. albicans infection is at least partially due to the sensitization of the fungal cells to the oxygen-dependent microbicidal system of the phagocyte.


Hexadecylphosphocholine: oral treatment of visceral leishmaniasis in mice. Hexadecylphosphocholine (He-PC), a novel phospholipid derivative, was tested against Leishmania donovani and Leishmania infantum, the causative agents of visceral leishmaniasis. In vitro, promastigotes were highly susceptible to He-PC; the 50% inhibitory concentrations were between 0.89 and 2.25 micrograms/ml for the different leishmanial strains. In vivo, a marked antileishmanial activity in infected BALB/c mice could be demonstrated after oral administration of He-PC. Whereas parasite suppression and killing in the liver were comparable after 5 days of treatment with He-PC (10 or 20 mg/kg of body weight per day administered orally) and sodium stibogluconate (120 mg of pentavalent antimonal agent per kg/day administered subcutaneously), a superior reduction in the parasite load in the spleen and bone marrow was observed after oral treatment with He-PC. After a 4-week treatment period, parasite suppression in the spleen was better than that observed with standard sodium stibogluconate therapy by a factor of more than 600.


Pharmacokinetic studies of CP-74,667, a new quinolone, in laboratory animals. The pharmacokinetics of CP-74,667 (7-(8'-methyl-3',8'-diazabicyclo[3.2.1]oct-3'-yl)-1-cyclopropyl-6- fluoro-1,4-dihydro-4-oxo-3-quinolinecarboxylic acid) were studied following oral or parenteral administration in mice, rats, rabbits, dogs, and cynomolgus monkeys. The mean peak levels of CP-74,667 in serum following a single oral dose of 20 mg/kg of body weight were similar in all species, with a range of 3.7 micrograms/ml in mice to 5.6 micrograms/ml in dogs. In contrast, elimination half-lives were species dependent, with mean values of 2.1, 1.8, 4.5, 7.8, and 13.1 h in mice, rats, rabbits, dogs, and monkeys, respectively. The oral bioavailability of CP-74,667 was 100% in dogs and monkeys, as determined by intravenous-oral crossover experiments. The maximum concentration of drug in serum and area under the concentration-time curve (AUC) of CP-74,667 in dogs were proportional to dose over the range of 5 to 40 mg/kg. Accumulation of drug in serum was observed following the administration of four once-a-day doses of 7.1 mg/kg in monkeys (mimicking a 500-mg human dose), with significant increases in half-life, maximum and minimum concentrations of drug in serum, and AUC. The good tissue penetration of CP-74,667 suggested by a volume of distribution in excess of 2 liters/kg in dogs and monkeys was confirmed by tissue distribution studies with the same species, which demonstrated tissue concentrations (except for those in brain tissue) greater than 1.45 times higher than corresponding levels in serum. The mean urinary recoveries of unchanged drug were 17.7% in rats, 7.8% in monkeys, and 4.9% in dogs. Metabolism studies in dogs, following intravenous dosing, indicated that renal excretion of CP-74,667-related materials accounted for 41.6% of the administered dose, while biliary recoveries accounted for 6.8%. The CP-74,667 N-oxide metabolite was the primary drug-related material eliminated via renal excretion (37.2% of dose). The pharmacokinetics of CP-74,667 describe a quinolone with complete oral absorption, linear pharmacokinetics, a long elimination half-life, and wide distribution into tissues.


In vitro antibacterial activity of Q-35, a new fluoroquinolone. The in vitro activity of Q-35, an 8-methoxy fluoroquinolone, was compared with those of ofloxacin, ciprofloxacin, tosufloxacin, lomefloxacin, and sparfloxacin. The MICs of Q-35 for 90% of strains tested (MIC90s) of Staphylococcus aureus, methicillin-resistant S. aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, and Streptococcus pyogenes were 0.2, 6.25, 0.2, 0.39, and 0.39 micrograms/ml, respectively. The activity of Q-35 was 4- to 16-fold greater than those of ofloxacin, ciprofloxacin and lomefloxacin but equal to those of tosufloxacin and sparfloxacin against these organisms. For 82 ciprofloxacin-resistant staphylococci (MIC90 = 100 micrograms/ml), Q-35 was the most active of the new quinolones tested (MIC90 = 6.25 micrograms/ml). The MIC90s of Q-35 against Escherichia coli, Enterobacter aerogenes, and Pseudomonas aeruginosa were 0.2, 0.78, and 12.5 micrograms/ml, respectively, and Q-35 was 2- to 16-fold less active than the other quinolones tested. Q-35 showed potent bactericidal activity and inhibited the supercoiling activity of DNA gyrase of S. aureus, E. coli, and P. aeruginosa.


Photostability and biological activity of fluoroquinolones substituted at the 8 position after UV irradiation. Q-35 [1-cyclopropyl-6-fluoro-1,4-dihydro-8-methoxy-7-(3-methylaminopiperid ine-1-yl)-4-oxoquinoline-3-carboxylic acid], a fluoroquinolone, has absorbance peaks at 333 and 286 nm. No spectral change was observed even when this aqueous solution was irradiated with 3 J of long-wavelength UV light (UVA) per cm2. On the other hand, its derivatives, which are unsubstituted (8-H analog) or which are substituted with fluorine at the 8 position (8-F analog), were found to have decreased antibacterial activities with a simultaneous increase in their cytotoxicities when they were degraded in a dose-dependent manner with respect to UVA irradiation. Similar results were observed with the other available fluoroquinolones. Enoxacin and lomefloxacin exposed to 0.3 J of irradiation per cm2 and norfloxacin, ofloxacin, and ciprofloxacin exposed to 1 J of irradiation per cm2 underwent absorption spectrum changes, an accompanying decrease in antibacterial activity, and an increase in cytotoxic activity. These results suggest that the introduction of a methoxy group into the 8 position of quinolones plays an important role in the stability of fluoroquinolones against irradiation by UV light.


Pharmacokinetics of foscarnet after twice-daily administrations for treatment of cytomegalovirus disease in AIDS patients. The pharmacokinetics of foscarnet were evaluated in 11 AIDS patients with cytomegalovirus disease after twice-daily infusion of 90 mg/kg of body weight for 2 weeks. All patients were hydrated during foscarnet infusion. Blood and urine samples were collected on days 1, 7, and 14 of therapy. Foscarnet concentrations were measured by high-pressure liquid chromatography. Despite large interindividual variations, no significant differences were seen between day 1, day 7, and day 14 concentrations in plasma. Mean peak and trough concentrations on day 14 of therapy were 605 +/- 118 and 52 +/- 59 microM, respectively. In all patients, peak concentrations were well above those necessary to inhibit cytomegalovirus. Pharmacokinetic parameters remained stable throughout the study. On day 14, the mean half-life was 3.4 h, total and renal clearances were 118 and 92 ml/min, respectively, and the volume of distribution was 0.6 liter/kg. These data and previous clinical trials demonstrate that this more convenient dosage regimen can be safely used for patients with cytomegalovirus disease. The side effects were comparable to those reported with other dosage regimens, although no renal impairment was seen in this study, probably because of the hydration.


Treatment of murine cytomegalovirus infections in severe combined immunodeficient mice with ganciclovir, (S)-1-[3-hydroxy-2-(phosphonylmethoxy)propyl]cytosine, interferon, and bropirimine. Severe combined immunodeficient (SCID) mice were found to be highly susceptible to murine cytomegalovirus (MCMV) infection. Treatment of infected mice with ganciclovir (12.5, 25, and 50 mg/kg of body weight for 10 days) starting 24 h after virus challenge resulted in delays in death by 2 to 8 days, and no animals survived the infection. (S)-1-[3-Hydroxy-2-(phosphonylmethoxy)propyl]cytosine (HPMPC) was much more potent, with doses of 1, 3.2, and 10 mg/kg/day (for 10 days) increasing the mean survival time by 15 to 30 days. Twenty-day treatments with HPMPC starting 5 days after virus inoculation increased the mean survival time by 24 to 32 days, with once-weekly (50-mg/kg) treatments being equivalent to daily (10-mg/kg) treatments. Delays in the development of liver, lung, and spleen virus titers in ganciclovir- and HPMPC-treated groups correlated with extensions in the mean survival times relative to the survival times of the placebo controls. The two compounds were approximately equally toxic to uninfected BALB/c mice treated for 10 days, causing 80 to 100% mortality after a dose of 150 mg/kg and 0% mortality after a dose of 75 mg/kg. Thus, the relative therapeutic index of HPMPC was 50-fold greater than that of ganciclovir. Recombinant alpha interferon delta 4 alpha 1/alpha 2 (1 x 10(4) and 5 x 10(4) units per mouse per day) and bropirimine (100 and 300 mg/kg/day) provided no protection from the lethal MCMV infection. The severe combined immunodeficient mouse MCMV infection is an important new model that will permit chemotherapy regimens to be studied over several months.


Evidence of incorporation of the chromosomal beta-lactamase gene of Enterococcus faecalis CH19 into a transposon derived from staphylococci. We recently reported the chromosomal location of the staphylococcal beta-lactamase gene in four strains of Enterococcus faecalis. Transfer of this gene from strain CH19 to an enterococcal recipient was accompanied by transfer of numerous other antimicrobial resistance determinants in the absence of detectable plasmid DNA. A restriction map developed by comparing digestions of the regions surrounding the beta-lactamase gene in donor and recipient chromosomes resembles published maps of previously described staphylococcal beta-lactamase transposons, particularly in the area of the structural gene and its downstream region. In addition, DNA sequence analysis of the region immediately downstream of the beta-lactamase gene from both CH19 and its transcipient, CX19, revealed the presence of a 121-bp inverted repeat region found in Tn552 and Tn4002, two previously described staphylococcal beta-lactamase transposons. These results suggest that the chromosomal beta-lactamase gene of E. faecalis CH19 is incorporated into a transposonlike element derived from staphylococci.


Postantibiotic sub-MIC effects of vancomycin, roxithromycin, sparfloxacin, and amikacin. The sub-MIC effects (SMEs) and the postantibiotic sub-MIC effects (PA SMEs) of vancomycin, roxithromycin, and sparfloxacin for Streptococcus pyogenes and Streptococcus pneumoniae and of amikacin for Escherichia coli and Pseudomonas aeruginosa were investigated. A postantibiotic effect was induced by exposing strains to 10x the MIC of the antibiotic for 2 h in vitro. After the induction, the exposed cultures were washed to eliminate the antibiotics. Unexposed controls were treated similarly. Thereafter, the exposed cultures (PA SME) and the controls (SME) were exposed to different subinhibitory concentrations (0.1, 0.2, and 0.3x the MIC) of the same drug and growth curves for a period of 24 h were compared. In general, the PA SMEs were much more pronounced than the SMEs. However, for amikacin and E. coli the SME of 0.2 and 0.3x the MIC also had an initial bactericidal effect. The longest PA SMEs were demonstrated for the combinations with the most pronounced killing during the induction and for the combinations which exhibited the longest PAEs.


Comparison of daptomycin, vancomycin, and ampicillin-gentamicin for treatment of experimental endocarditis caused by penicillin-resistant enterococci. Infections with enterococci that are resistant to multiple antibiotics are an emerging clinical problem. We evaluated the antibiotic treatment of experimental enterococcal endocarditis caused by two strains with different mechanisms of penicillin resistance. Enterococcus faecalis HH-22 is resistant to aminoglycosides and penicillin on the basis of plasmid-mediated modifying enzymes; Enterococcus raffinosus SF-195 is susceptible to aminoglycosides but is resistant to penicillin on the basis of low-affinity penicillin-binding proteins. Animals infected with strain HH-22 received 5 days of treatment with the following: no treatment; daptomycin (20 mg/kg of body weight twice daily [b.i.d.], intramuscularly [i.m.]), vancomycin (20 mg/kg b.i.d., intravenously), or ampicillin (100 mg/kg three times daily, i.m.) plus gentamicin (2.5 mg/kg b.i.d. i.m.). Although vancomycin was superior to ampicillin-gentamicin (P less than 0.01), daptomycin was significantly better than all other treatment regimens (P less than 0.01) in reducing intravegetation enterococcal densities, although no vegetations were rendered culture negative by this agent. Animals infected with strain SF-195 received 5 days of no therapy, ampicillin, ampicillin-gentamicin, vancomycin, or daptomycin (all at the dosage regimens described above). Daptomycin, vancomycin, and ampicillin-gentamicin each lowered intravegetation enterococcal densities significantly better than did ampicillin monotherapy or no treatment (P less than 0.01); moreover, these three treatment regimens rendered significantly more vegetations culture negative than did ampicillin monotherapy or no treatment (P less than 0.05). Serum daptomycin levels remained above the MICs and MBCs for both enterococcal strains throughout the 12-h dosing interval used in the study. Daptomycin and vancomycin were both active in vivo in these models of experimental enterococcal endocarditis caused by penicillin-resistant strains, irrespective of the mechanism of resistance. This activity correlated with the unique cell wall sites of action of these agents (binding to lipoteichoic acid and pentapeptide precursor, respectively) compared with the sites of action of beta-lactams (penicillin-binding proteins). Beta-Lactamase production by strain HH-22 precluded in vivo efficacy with ampicillin-gentamicin combinations. In contrast, this combination was active in vivo against strain SF-195, which exhibited intermediate-level penicillin resistance (MIC, 32 micrograms/ml), likely reflecting the ability of high-dose ampicillin to achieve enough binding to low-affinity penicillin-binding proteins to cause augmented aminoglycoside uptake.


Effects of CO2 and pH on inhibition of TEM-1 and other beta-lactamases by penicillanic acid sulfones. Incubation in 5% CO2 reduced the inhibition zones of piperacillin-tazobactam (75/10 micrograms) disks for Escherichia coli strains with TEM-1, TEM-2, and SHV-1 beta-lactamases. Similarly, MICs of piperacillin-tazobactam and other penicillin-sulfone combinations for TEM producers were up to 500-fold higher at pH 6.5 than at pH 8.0. This effect was greatest for organisms with high levels of enzyme activity. CO2 and mild acidity did not affect the susceptibility of beta-lactamase-negative strains to penicillin-sulfone combinations, and the effects of these conditions were variable for organisms with beta-lactamases other than TEM-1, TEM-2, and SHV-1. These last observations discounted acid-mediated inactivation of piperacillin or tazobactam. MICs of amoxicillin or piperacillin alone or with clavulanate for TEM and SHV producers were affected only less than or equal to 16-fold by 5% CO2 or acidity, indicating that the greater effects seen with the penicillin-sulfone combinations depended on the behavior of the sulfones and not on that of the penicillins. This pH effect was studied in detail for TEM-1 enzyme. Inhibition of this enzyme by sulfones but not clavulanate varied grossly with pH, with 50% inhibitory concentrations of tazobactam and sulbactam up to 300-fold higher at pH 6.5 than at 8.0. By contrast, the hydrolytic activity of TEM-1 enzyme for substrates and its level of production varied threefold or less between pH 6.5 and pH 8.0. Increased inhibition at pH 8.0 reflected sequestration of the enzyme into a secondary noncovalent complex rather than increased irreversible inactivation.


Piperacillin, tazobactam, and gentamicin alone or combined in an endocarditis model of infection by a TEM-3-producing strain of Klebsiella pneumoniae or its susceptible variant. The efficacy of tazobactam, a beta-lactamase inhibitor, in combination with piperacillin, was studied in vitro and in rabbit experimental endocarditis due to a Klebsiella pneumoniae strain (KpR) producing an extended-spectrum beta-lactamase, TEM-3, or its nonproducing variant (KpS). In vitro, piperacillin was active against KpS (MIC = 4 micrograms/ml, MBC = 8 micrograms/ml with 10(7)-CFU/ml inoculum) but not against KpR (MIC = MBC = 256 micrograms/ml). Tazobactam (1 microgram/ml) restored the activity of piperacillin against KpR (MIC = 2 micrograms/ml, MBC = 4 micrograms/ml). Gentamicin was active against both strains (MIC = 0.25 and 0.5 micrograms/ml for KpS and KpR, respectively). The piperacillin-tazobactam-gentamicin combination was synergistic in vitro. The piperacillin/tazobactam ratio in plasma and in vegetations was always lower than the 4/1 injected dose ratio. In vivo, piperacillin (300 mg/kg of body weight four times a day [QID]) was active against KpS but not against KpR. Tazobactam (75 mg/kg QID) was able to restore the in vivo effect of piperacillin (300 mg/kg QID) against KpR (-3.0 log10 CFU/g of vegetation versus that of controls). Gentamicin (4 mg/kg twice a day [BID]) was active against both strains. Compared with controls, the combination of gentamicin plus piperacillin against KpS (-5.6 log10 CFU/g of vegetation), and the gentamicin-piperacillin-tazobactam combination against KpR (-4.4 log10 CFU/g of vegetation) achieved the greatest decrease in bacterial counts in vegetations and were the only regimens that significantly increased the proportion of sterile vegetations. It is concluded that (i) tazobactam was able to restore the effect of piperacillin against a TEM-3 extended-spectrum Beta-lactamase-producing strain of K. pneumoniae, both in vitro and in a severe experimental infection with high inoculum, when used in a 4/1 piperacillin/tazobactam dose ratio; (ii) gentamicin alone was effective because of the high peak/MBC ratio in plasma; (iii) piperacillin-tazobactam-gentamicin, probably because of the effect of gentamicin in reducing bacterial inoculum in vivo, as stressed by the results obtained by piperacillin-gentamicin against KpS, may be the most effective regimen against KpR.


Mechanism of action of the antiherpesvirus biflavone ginkgetin. Screening of plant extracts found that a biflavone from Cephalotaxus drupacea, which was found to be ginkgetin, is active against herpes simplex virus type 1 (HSV-1). This compound caused dose-dependent inhibition of virus replication with a 50% cytotoxic activity at 12.8 micrograms/ml and 50% anti-HSV-1 activity at 0.91 micrograms/ml, the therapeutic index being 14.1. Ginkgetin also showed inhibitory effects against HSV type 2 and human cytomegalovirus with therapeutic indices of 13.8 and 11.6, respectively. Ginkgetin had a weak virucidal activity against HSV-1 at more than 5 micrograms/ml. Both adsorption of HSV-1 to host cells and virus penetration into cells were unaffected by this agent. Ginkgetin suppressed viral protein synthesis when added at various steps of HSV-1 replication and exerted strong inhibition of transcription of the immediate-early genes.


Transposition of the gene encoding a TEM-12 extended-spectrum beta-lactamase. An isolate of Klebsiella oxytoca from the blood culture of a child with leukemia was found to produce two beta-lactamases, at least one of which conferred resistance to ceftazidime. Genes encoding both enzymes were located on a single self-transmissible 100-kb plasmid, pOZ201. This plasmid was introduced into Escherichia coli UB5201 (pACYC184), and the gene encoding one beta-lactamase was transposed onto plasmid pACYC184 by exploiting a gene dosage effect. The transposable gene was found to encode a TEM-12 enzyme as determined by nucleotide sequencing. This gene was subsequently transposed onto plasmid pUB307. The transposable element encoding the TEM-12 enzyme has been designated Tn841. Both plasmids pACYC184::Tn841 and pUB307::Tn841 were shown to encode a beta-lactamase with the same isoelectric point and substrate profile as the TEM-12 beta-lactamase. Transposon Tn841, at approximately 7 kb, is larger than TnA (4.8 kb) and transposes at a lower frequency. Although it produced a resolvase which can complement the resolvase of Tn3, its transposase function was not able to complement the transposition of a TnA element which lacked transposase. The occurrence of a gene encoding an extended-spectrum beta-lactamase on a transposable element in a clinically significant bacterium is potentially a cause for concern for the spread of resistance to the extended-spectrum cephalosporins.


Pharmacokinetics and tissue penetration of tazobactam and piperacillin in patients undergoing colorectal surgery. The pharmacokinetics of tazobactam and piperacillin in plasma and different tissues after a 30-min intravenous infusion of 4 g of piperacillin and 0.5 g of tazobactam were investigated in 18 patients who underwent elective colorectal surgery. Serial blood samples were collected for up to 6 h after the initiation of the infusion. The types of tissue collected were fatty tissue, muscle, skin, appendix, and intestinal mucosa (proximal and distal). On the basis of concentrations in plasma, the following pharmacokinetic parameter values were obtained (values are means +/- standard deviations): maximum concentration of drug in serum, tazobactam, 27.9 +/- 7.67 micrograms/ml; piperacillin, 259 +/- 81.8 micrograms/ml; time to maximum concentration of drug in serum, tazobactam, 0.51 +/- 0.03 h; piperacillin, 0.51 +/- 0.03 h; area under the concentration-time curve, tazobactam, 47.6 +/- 13.3 micrograms.h/ml; piperacillin, 361 +/- 80.3 micrograms.h/ml; clearance, tazobactam, 188 +/- 52.3 ml/min; piperacillin, 194 +/- 42.9 ml/min; half-life, tazobactam, 1.42 +/- 0.32 h; piperacillin, 1.27 +/- 0.24 h; apparent volume of distribution, tazobactam, 0.31 +/- 0.07 liter/kg of body weight; piperacillin, 0.29 +/- 0.06 liter/kg; volume of distribution at steady state, tazobactam, 0.28 +/- 0.04 liter/kg; piperacillin, 0.25 +/- 0.05 liter/kg. The concentrations of tazobactam and piperacillin in fatty tissue and muscle tissue were 10 to 13 and 18 to 30% of the levels in plasma, respectively. In skin, the concentrations of piperacillin were 60 to 95% of the levels in plasma, whereas the concentrations of tazobactam in plasma were 49 to 93% of the levels in skin tissue. The mean concentration of tazobactam in the investigated gastrointestinal tissues (appendix, proximal and distal mucosa) exceeded levels in plasma after 1 h, while piperacillin showed a mean penetration into these tissues of 43 and 53%. The mechanisms that can be used to explain the extent of penetration of piperacillin and tazobactam are discussed. Simple diffusion may take place in fatty and muscle tissue, while penetration into skin and gastrointestinal tissue is governed by more complex mechanisms which lead to differences in penetration between piperacillin and tazobactam. For all tissues investigated (except fatty tissue), the time course of the concentrations of both compounds was similar, with a peak in concentration at between 1 and 2 h after the start of infusion followed by a decline of concentrations that were almost parallel to the curves of the drug concentrations in plasma. In plasma and in all investigated tissues, piperacillin as well as tazobactam reached or exceeded the concentrations found to be effective in vitro.


Assessment of a selective inhibitor of herpes simplex virus thymidine kinase (L-653,180) as therapy for experimental recurrent genital herpes. Herpes simplex virus (HSV)-coded thymidine kinase (TK) is important in efficient reactivation of latent infection. These studies were designed to investigate whether treatment of latently infected animals with a TK inhibitor altered the natural history of recurrent HSV disease. 9-([(Z)-2-(hydroxymethyl)cyclohexyl]methyl) guanine (L-653,180) is a potent and selective nonsubstrate inhibitor of HSV TK which can suppress or delay reactivation of HSV-1 from latently infected cells in vitro without affecting viral replication. In an initial study, six female Hartley guinea pigs were treated with L-653,180 in their diet (25 mg/30 g of food) and water (300 mg/liter) for 7 days. Blood, urine, kidney, liver, spinal cord, and cerebral cortex specimens were collected. L-653,180 was detected in all specimens at concentrations which, although low, were higher than the in vitro 50% inhibitory concentration of the drug against HSV TK. In the second study, 20 female Hartley guinea pigs were randomized into two groups following recovery from primary genital HSV-2 infection. One group received L-653,180 in diet and water for 4 weeks beginning 21 days postinoculation. Animals were examined daily for recurrent lesions for 10 weeks. Treated animals experienced fewer recurrences during the treatment period but the results were not significantly different from results with controls. During the first 2-week posttreatment period, L-653,180-treated animals had significantly fewer recurrences than control animals (P = 0.02). Over the entire 10-week observation period, treated animals experienced fewer recurrences (P = 0.06). These results suggest that inhibitors of viral TK may be useful in limiting reactivation of latent virus and thus recurrent infections. In these experiments, the amount of drug that could be administered to the animals was limited by its poor solubility. Further studies with more potent and soluble inhibitors of HSV TK appear to be warranted.


In vitro antibacterial activities of tosufloxacin against and uptake of tosufloxacin by outer membrane mutants of Escherichia coli, Proteus mirabilis, and Salmonella typhimurium. The antibacterial activities of tosufloxacin and other quinolones against and apparent uptakes of tosufloxacin and other quinolones by outer membrane mutants of Escherichia coli, Proteus mirabilis, and Salmonella typhimurium were studied. The hydrophobicity of tosufloxacin was nearly equal to that of ofloxacin or lower than those of sparfloxacin and nalidixic acid. OmpF- and OmpC-deficient E. coli and 40-kDa porin-deficient P. mirabilis mutants were twofold more susceptible to tosufloxacin and sparfloxacin but two- to fourfold less susceptible to other quinolones than their parent strains. In S. typhimurium lipopolysaccharide-deficient (rough) mutants, the differences in susceptibility to tosufloxacin were similar to those to sparfloxacin and nalidixic acid. The apparent uptake of tosufloxacin by intact cells was increased in porin-deficient mutants compared with that by their parent strain. These results suggest that the permeation route of tosufloxacin across the outer membrane is different from that of other fluoroquinolones and that tosufloxacin may permeate mainly through the nonporin pathway, presumably phospholipid bilayers. However, this characteristic is independent of the hydrophobicity of the molecule.


In vitro activities of penciclovir and acyclovir against herpes simplex virus types 1 and 2. Penciclovir (PCV) and acyclovir are acyclic guanine analogs which inhibit herpes simplex virus (HSV) DNA polymerase. Their 50% infective doses were 0.5 to 0.8 microgram/ml for clinical isolates of HSV-1 and 1.3 to 2.2 micrograms/ml for HSV-2. Furthermore, HSV-infected cultures receiving 2-h pulses of PCV had 2- to 50-fold less HSV than acyclovir-treated cultures, consistent with the prolonged intracellular half-life of PCV triphosphate.


Interplay of impermeability and chromosomal beta-lactamase activity in imipenem-resistant Pseudomonas aeruginosa. Mutational loss of the D2 porin causes imipenem resistance in Pseudomonas aeruginosa. It was found that this mechanism could function only when the chromosomal beta-lactamase was expressed. Mutants lacking both the beta-lactamase and the D2 porin were almost as susceptible as those that lacked the beta-lactamase but retained the porin. Thus, imipenem resistance reflected an interplay of the enzyme and impermeability, not either factor alone. These findings suggest that the activity of a carbapenem more beta-lactamase stable than imipenem should be less affected by the porin loss. Meropenem approached this behavior.


Antimicrobial activities of two investigational fluoroquinolones (CI-960 and E4695) against over 100 Legionella sp. isolates. The antimicrobial activities of two investigational fluoroquinolones (CI-960 and E4695) were compared with those of five similar compounds and four comparison drugs against 103 strains of Legionella pneumophila and five other Legionella species type strains. When concentrations inhibiting 90% of strains tested (MIC90s) for L. pneumophila were determined, CI-960 and temafloxacin emerged as the most active (0.015 microgram/ml) and were followed in potency by E4695 (0.03 microgram/ml). This activity was two- to fourfold greater than that of the reference drug, ciprofloxacin, and approached that of rifampin (MIC90, 0.008 microgram/ml). All fluoroquinolones studied were more active than erythromycin (MIC90, 0.5 microgram/ml). These two investigational fluoroquinolones appear well suited for further in vivo study of legionellosis therapy.


Ultrastructural changes associated with reduced mitochondrial DNA and impaired mitochondrial function in the presence of 2'3'-dideoxycytidine. Incubation of Molt-4 cells in 4 microM 2'3'-dideoxycytidine did not produce a significant change in the mitochondrial ultrastructure after 4 days; however, by 12 days, the mitochondrial ultrastructure was distorted, with condensed cristae or vacuolization, or both. Concentration-dependent decreases in both cell growth (mean 50% inhibitory concentration, 4.70 +/- 0.5 microM) and mitochondrial DNA content (mean 50% inhibitory concentration, 0.46 +/- 0.06 microM) occurred after incubation with 2'3'-dideoxycytidine for 4 days.


HHV-6 inhibition by two polar compounds. Dimethyl sulphoxide and dimethyl formamide, two polar compounds and powerful cell differentiation inducers, inhibit HHV-6 infection when added to HHV-6-infected HSB2 cultures. This was established by a delay in the time-course of infection and in the development of virus-induced cytopathic effects. Furthermore, viral titration of supernatants showed a significant reduction (3 log10) of the number of infectious particles. Electron microscopy confirmed that viable cells and extracellular virions were present in the cultures containing the polar compounds, while in the non-treated cultures all cells were lysed and no extracellular virus was evident. The mode of action of these compounds is still unclear and warrants further investigation.


Structure/function studies of murine interferon-alpha 1 using site-directed mutagenesis followed by in vitro synthesis. Site-directed in vitro mutagenesis followed by in vitro transcription and translation has been used to study structure/function relationships for murine interferon-alpha 1 (MuIFN-alpha 1). The mature form of the MuIFN-alpha 1 protein was expressed as well as analogue forms with amino acid substitutions at positions 33, 71, 72, 123 and 133. These positions were chosen on the basis of known human interferon-alpha structure/function relationships. Biological assays for antiviral activity on murine cells and natural killer cell activation have been performed for each of the proteins produced. The data obtained have been interpreted in the light of previous human and murine interferon-alpha structure/function work and the recently published three-dimensional structure of murine type I interferon.


The acyclic nucleoside analogue penciclovir is a potent inhibitor of equine herpesvirus type 1 (EHV-1) in tissue culture and in a murine model. Equine herpesvirus type 1 (EHV-1) was sensitive to the nucleoside analogue penciclovir (PCV) when tested in tissue culture; the ED50 was 1.6 micrograms/ml. Drug-resistant mutants were selected which were found to be TK-defective and approx. 45-fold less sensitive to PCV compared with the parental strain. PCV was compared with the phosphonyl derivative, HPMPA in mice infected with EHV-1. Both drugs were shown to be effective in vivo, limiting wild-type virus replication in respiratory tissues, and reducing viraemia. The treated mice also showed less clinical signs and reduced histopathology compared with placebo-treated controls. The establishment of latent EHV-1 in the mice, however, was not prevented. The results obtained with mice suggest that antiviral chemotherapy may be practical in the horse and that this possibility is worthy of further investigation in the natural host.


A view from the Common Cold Unit.

A view from the Common Cold Unit. I have been asked to stand back and describe in broad terms the view I have had of common colds--probably the most frequent of acute human diseases and a long-lasting scientific problem--and in particular our recent work on antivirals. I should be able to do this for two reasons. Like everyone else I have suffered from colds, but in addition I have been studying the problem from the virological and clinical point of view for over 35 years--for the last 31 at the Common Cold Unit, Salisbury. As a result I may have problems with perspective--it is not possible to give a personal view and at the same time to describe something from every possible angle, and quite impossible to be comprehensive, but I have done my best and readers will make their own judgements and corrections.


Effect of anionic polymers on fusion of Sendai virus with human erythrocyte ghosts. The effect of anionic polymers (dextran sulfate, heparin and chondroitin sulfate) on fusion of Sendai virus with erythrocyte ghosts was studied. The effect of pH on the activity of these anionic polymers was also investigated. In order to examine the interaction of such polymers with the Sendai virion and erythrocyte ghost surfaces, the binding of virions to erythrocyte ghosts and the aggregation of virions and/or erythrocyte ghosts were also measured with respect to the same parameters. It was found that the anionic polymers suppressed the fusion of Sendai virus with erythrocyte ghosts. The order of effectiveness of the polymers in suppression was dextran sulfate greater than heparin greater than chondroitin sulfate, for the application of a same quantity (weight/ml) of the polymers. The lower the pH of the suspending medium, the more effective were the polymers in suppressing virion-erythrocyte ghost aggregation and fusion. The suppression of fusion was dependent on the concentration of the polymers applied: the higher the concentration of the polymer applied, the more the suppression was observed. Evidence from binding studies, turbidity measurements and electrophoretic mobility measurements indicates that the anionic polymers interact preferentially with the virion surface.