Intestinal fuels: glutamine, short-chain fatty acids, and dietary fiber. In recent years, considerable research has focused on the physiologic effects and clinical uses of three dietary constituents thought to be trophic to the intestinal tract in human beings: glutamine, short-chain fatty acids (SCFAs), and dietary fiber. Glutamine is an important nitrogen-carrying amino acid that may be "conditionally essential" in certain disease states to support the gut barrier and immune function and overall protein use. Colonic irrigations with SCFA preparations have demonstrated enhanced healing of bowel tissue in animals and human beings. Dietary fiber supports bacterial SCFA production, normal stool output, and the gut barrier and immune function. However, optimal fiber doses for various medical conditions are not known, and the risk for gastrointestinal (GI) obstruction, diarrhea, gas, and bloating necessitates careful selection of patients and daily monitoring of fiber tolerance. A review of the current literature indicates that widespread use of glutamine and SCFA additives parenterally and enterally awaits further evidence of safety and efficacy in human beings, establishment of appropriate doses, and advances in formulation technology. Administration of dietary fiber to enhance bowel motility should be considered in long-term tube-fed patients with intact GI function and sufficient fluid tolerance to permit hydration of fiber. Industrywide agreement on fiber analysis methods and labeling standards (eg, fiber fermentability vs solubility) would facilitate selection of enteral products. To streamline studies and optimize research efforts in future clinical trials, standard criteria for evaluating GI function, diarrheagenic factors, and intestinal outcome variables should be established.


The relationship between electromyographically documented peripheral neuropathy and falls. To determine if the presence of an electromyographically demonstrated peripheral polyneuropathy involving the lower extremities is associated with falls. Case control study. A University-based referral center. Twenty-five patients with an axonal peripheral polyneuropathy affecting the lower extremities, demonstrated by electromyography more than 8 and less than 16 months before, and 25 age- and sex-matched controls who had normal electromyographic studies of the lower extremities during the same time period, were identified from a computer record of all electromyography results during the year prior to the study, using diagnostic codes and demographic data. Twenty-eight pairs were identified, 25 of whom completed the study. Number of falls and resultant injuries during the year before the interview, and number of previously identified risk factors for falling, based on interview. Peripheral neuropathy was found to be significantly associated with falling (14/25, 56% vs 2/25, 8%--odds ratio 23.0, 95% confidence interval 2.5, > 100) and repetitive falling (10/25, 40% vs 1/25, 4%--odds ratio, 95% confidence interval 1.4, > 100). There was a trend toward increased injury requiring medical attention (5/25, 20% vs 0/25, 0%), but it did not reach statistical significance. Previously identified risk factors were not significantly more common in the peripheral neuropathy group. An electromyographically identified peripheral neuropathy is strongly associated with and may be a risk factor for falls.


Alpha-adrenoreceptor modulation of neurally evoked circular muscle responses of the guinea pig stomach. The effects of alpha-adrenergic agonists on transmural-evoked motor responses were investigated in guinea pig gastric corpus in vitro, using preparations stripped of mucosa and orientated to record changes in circular muscle tension. Three tetrodotoxin-sensitive components to a 10 s burst of transmural stimulation could be distinguished: an initial 'on' contraction, an 'off' contraction and a transient relaxation. The 'on' response was blocked by atropine (0.1 microM), while the 'off' response and relaxation were unaffected at this dose. A submaximal dose of acetylcholine was used to assess the sensitivity of the preparation. The alpha 1 agonist L-phenylephrine decreased the amplitude of the 'off' response while simultaneously increasing both the 'on' response and the relaxation, although the response to acetylcholine was unchanged. These effects were dose-dependent and reversed by pretreatment with prazosin. In marked contrast, the alpha 2 agonist clonidine inhibited the 'on' response in a dose-dependent manner without affecting the 'off' response, the relaxation or the response to acetylcholine. Yohimbine reversed the effect of clonidine. We conclude that the inhibitory action of alpha-agonists involves both cholinergic and non-cholinergic pathways, with alpha 1 and alpha 2 adrenoceptors modulating different circuits within the enteric nervous system.


HLADR5 and C4BQO high frequency and antinuclear antibodies positivity in patients with 21 hydroxylase deficiency from Campania region. HLA haplotypes, complement C4 factor and factor B immunochemical concentrations and autoantibodies titer have been studied in six patients with mild congenital adrenal hyperplasia (MC-AH), in two patients with classical congenital adrenal hyperplasia (CCAH) and in their parents. A high frequency of DR5 and C4BQO alleles have been found in MCAH patients. Moreover, C4BQO allele is carried out in three out of four cases associated with DR5. In the two CCAH patients we found a B51 and a B14 allele, the last one usually described in the non classical form of the disease in population of different ethnic origin. Signs of autoimmunity in some patients and parents have been found. C4 null alleles were several-fold more frequent among our patients with respect to the same ethnic control group and the autoantibody positivity could be the result of an altered immune regulation. The presence of a positive correlation between cortisol basal levels and C4 and Bf concentrations in the six MC-AH patients suggests an interrelationship between hormonal factors and immunological findings in this disease. Our finding about HLA antigens not previously described in this syndrome may stimulate more profound studies by genomic and cDNA probes.


Acute and chronic effects of high glucocorticoid levels on hypothalamic-pituitary-thyroid axis in man. It is known that glucocorticoids can influence anterior pituitary hormones other than ACTH. Their effects on the hypothalamic-pituitary-thyroid axis are controversial. To further investigate this issue, the acute and chronic effects of high plasma cortisol levels on TSH secretion were evaluated in 20 normal subjects and in 14 patients with Cushing's syndrome. In normals, high plasma cortisol levels were obtained by giving ACTH 250 micrograms or CRH 100 micrograms iv as a bolus or by an hydrocortisone 500 mg infusion over 1 h. Acute cortisol increase produced no effect both on basal and TRH-stimulated TSH secretion. In patients with Cushing's syndrome, basal TSH levels, low or suppressed, showed an impaired response to TRH, inversely correlated with urinary cortisol values. After successful surgery, TSH and its response to TRH became normal in concomitance with the normalization of plasma and urinary cortisol levels. Our data show the lack of an acute inhibitory effect of high plasma cortisol levels on TRH-TSH axis. However, after long-term exposure to high plasma cortisol levels, i.e. Cushing's syndrome, inhibition of both basal and TRH-stimulated TSH secretion was demonstrated. These findings indicate that only prolonged hypercortisolism does interfere with pituitary TSH secretion. The underlying mechanisms are still unclear.


The silent corticotropinoma: is clinical diagnosis possible? Up to now, the diagnosis of silent corticotroph cell pituitary adenomas has been made only on histopathological basis. In this paper we describe 6 women affected with pituitary adenomas, without evident clinical features of hypercortisolism, in whom retrospective data suggested the possibility of clinically diagnosing silent corticotropinomas in vivo. In all patients basal ACTH and cortisol levels were normal, and the low-dose dexamethasone test constantly suppressed serum cortisol and urinary 17-hydroxycorticosteroid levels. The CRH and/or lysine-vasopressin tests, performed in five patients, always induced exaggerated ACTH/cortisol rises. In three cases the response to the opiate agonist loperamide was assessed and no inhibition of ACTH/cortisol levels was found. All patients underwent pituitary surgery. In five cases evidence of corticotropinoma was obtained by immunohistochemistry or immunofluorescence studies; moreover, in one adenoma ACTH was secreted into the culture medium, and in another one CRH and arginine-vasopressin induced a marked intracellular [Ca++] rise. Electron microscopy study of the adenoma, removed from three patients, showed the presence of adenomatous corticotroph cells. Finally, in another woman no hormonal abnormalities were initially observed and she was operated for a "nonfunctioning" pituitary adenoma, but four years later an overt Cushing's disease appeared, suggesting that a silent corticotropinoma subsequently became functional, although the formation of a different adenoma cannot be excluded. In conclusion, the occurrence of ACTH/cortisol hyperresponsiveness to CRH and/or lysine-vasopressin and the lack of suppression of ACTH/cortisol secretion to opioid agonists in patients with apparently "nonfunctioning" pituitary tumors might allow the in vivo recognition of silent corticotropinomas.


Inhibitory effect of dexamethasone on the oxytocin response to insulin-induced hypoglycemia in normal men. Glucocorticoids are known to reduce both ACTH and arginine vasopressin responses to insulin-induced hypoglycemia in normal men. The present study was undertaken in order to establish whether glucocorticoids are capable of modifying the oxytocin (OT) response to hypoglycemia. For this purpose, 8 normal men (28-33 yr) were tested with insulin (0.15 IU/kg in an iv bolus) [insulin tolerance test (ITT)] with and without pretreatment with dexamethasone (2 or 4 mg in an iv bolus 10 min before insulin). Eight different subjects (29-35 yr) were tested with dexamethasone alone. The administration of dexamethasone (2 or 4 mg) alone changed neither ACTH nor OT concentrations in the plasma during the next hour. Insulin produced similar hypoglycemic responses, regardless of dexamethasone treatment. ACTH levels rose significantly in response to insulin-induced hypoglycemia, with a mean peak response at 45 min (p less than 0.01 vs baseline). Two and four mg dexamethasone produced similar significant reductions of the ACTH response to hypoglycemia (p less than 0.02 at 45 min, p less than 0.05 at 30 and 60 min vs ITT). In the ITT, OT levels rose significantly in response to hypoglycemia, with a mean peak response at 45 min (p less than 0.01 vs basal value). The pretreatment with 2 or 4 mg dexamethasone reduced in a similar manner the hypoglycemia-induced OT rise (p less than 0.05 at 30 and 45 min vs ITT). These findings show a partial inhibition by dexamethasone of the OT response to hypoglycemia.(ABSTRACT TRUNCATED AT 250 WORDS)


Effect of awakening on aldosterone.

Effect of awakening on aldosterone. Overnight studies conducted in 10 male volunteers showed that the effect of awakening on aldosterone and its main regulatory hormonal systems depends on the time of its occurrence. Waking periods in the latter half of the night induced concomitant increases in plasma levels of aldosterone and ACTH, whereas plasma renin activity tended to decrease. No effect was apparent, when subjects were awoken during the first hours of sleep, which is known to be a quiescent period of adrenocorticotropic activity. These results provide evidence of the dominant influence of ACTH during the second half of the night insuring the regulation of aldosterone during waking periods.


[Luteinized cystic hyperplasia of the ovaries during pregnancy. A case report]. Multiple lutein cyst (MLC) and luteoma are two aspects of ovarian hyperplasia induced by high levels of human chorionic gonadotrophin (HCG). Using a case report of MLC, the authors compare it with luteoma (the anatomoclinical and pathogenic data). Luteoma are more frequent with multipara, whereas MLC are more often seen with paucipara. Maternal virilization can occur in both cases but MLC does not seem to be responsible for fetal virilization. The fetus could be protected by the placenta, thanks to increased aromatase activity and sex binding globulin production. MLC and luteoma are usually found by chance. The pathology must be known, because of their spontaneous regression after delivery. So, the attitude will be conservative under the control of biopsies.


Overexpression of alpha 2-adrenergic receptors in fetal rat heart: receptors in search of a function. alpha 2-Adrenergic receptors are transiently overexpressed by many types of developing cells. In the current study, the developmental profile and cellular function of these receptors were examined in the fetal and neonatal rat heart. alpha 2-Receptors, assessed with [3H]rauwolscine binding, were extremely high in fetal hearts on gestational day 19, 30-fold higher than values seen in adults. Receptor binding decreased by two-thirds by gestational day 21 and dropped by half again by postnatal day 3. To assess potential cellular functions controlled by the alpha 2-receptors, the capability of an alpha 2-agonist (clonidine) to inhibit membrane-associated adenylate cyclase activity was measured in three different settings: basal enzyme activity, the enzymatic response to isoproterenol (dependent upon beta-receptor linkages to the regulatory protein, Gs), and the response to forskolin (independent of receptor-Gs interactions). Despite the high number of alpha 2-receptors in fetal hearts, clonidine failed to alter any of the adenylate cyclase activity measures. In light of the postulated role of alpha 2-receptors in the maintenance of fetal/neonatal atrioventricular conduction, the excess alpha 2-receptors are probably linked to other cellular events, such as movement of calcium into the cell.


Large-cell carcinoma of the lung. With major bleeding and intussusception. We describe an unusual case of major small-bowel bleeding and intussusception due to jejunal metastases from a large-cell type carcinoma of the lung. Review of the surprisingly frequent case reports of small bowel metastases from primary lung tumors indicates that there is only one prior case report of intussusception and that ours is the first description of major intestinal hemorrhage due to metastatic pulmonary carcinoma.


Does hormonal therapy have any benefit for bleeding angiodysplasia? Sixty-four patients took part in a cohort study measuring the efficacy of daily hormonal therapy in diminishing intestinal bleeding from small bowel angiodysplasia. Thirty patients received 5-10 mg norethynodrel either with mestranol, 0.075-0.15 mg (24 patients) or with conjugated estrogens, 0.625 mg (six patients). The cohort group consisted of 34 patients who either refused hormonal therapy (six patients) or were diagnosed early in our experience, before the resurgence of hormonal therapy (28 patients). Mean follow-up was 15.6 months (range 2-31 months) for the treated group and 13.4 months (range 1-23 months) for the untreated group. In addition, the change in monthly transfusion requirement with therapy was analyzed ("within group" analysis). Fifty percent (15 of 30) of the treated group required no further transfusion during therapy, while 44% (15 of 34) of the untreated group required no further therapy (not significant). During therapy, the mean monthly transfusion requirement of packed red blood cells in the treated group was not significantly different from that found before therapy (1.5 vs. 2.2 units, NS) or from that of the control group (1.5 vs. 1.6 units, NS). The findings do not support the use of hormonal therapy for bleeding from small intestinal angiodysplasia.


Norepinephrine-induced plasma dopamine decrease in man: pharmacological evidence of the involvement of alpha 2-adrenoceptors. The purposes of this study were: (1) to test whether intravenous infusion of norepinephrine can affect plasma dopamine levels; and (2) to explore to what extent dopamine-2 or alpha 2-receptors play a role in this response. Norepinephrine infusion in man was performed to test whether the increase in norepinephrine during sympathetic stimulation can affect dopamine release. Specific antagonists of presynaptic dopamine-2 and alpha 2-receptors were administered to test the receptor(s) involved in this possible regulatory phenomenon. Plasma catecholamine levels were investigated in seven normal subjects before and after administration of domperidone (dopamine-2 antagonist), yohimbine (alpha 2-antagonist) and norepinephrine. Both oral domperidone and yohimbine induced a significant increase in both plasma norepinephrine and plasma dopamine. Norepinephrine infusion induced a significant decrease in plasma dopamine. Pretreatment with domperidone only partially counteracted this inhibitory effect of norepinephrine infusion, whereas yohimbine fully counteracted it. Our data show that norepinephrine may act as a hormone at plasma concentrations as low as 450 pg/ml. The norepinephrine-induced plasma dopamine decrease seems to be alpha 2-adrenoceptor-mediated. This norepinephrine effect may be involved in the physiologic decrease in plasma dopamine that we demonstrated in the upright position in normal subjects.


Effect of local intra-arterial NG-monomethyl-L-arginine in patients with hypertension: the nitric oxide dilator mechanism appears abnormal. There is indirect evidence that the nitric oxide system may be impaired in hypertensive patients. The objective of this study was to examine basal nitric oxide-mediated dilation in hypertensive patients. The forearm blood flow (FBF) response to noradrenaline and NG-monomethyl-L-arginine (L-NMMA), a stereospecific inhibitor of nitric oxide synthesis, was compared in seven untreated hypertensive patients and 17 normotensive controls. Drugs were infused locally into the brachial artery and FBF measured using venous occlusion plethysmography. In normotensives noradrenaline (60, 120 and 240 pmol/min) and L-NMMA (1,2 and 4 mumol/min) produced similar reductions in resting FBF. In the hypertensives L-NMMA was significantly less effective than noradrenaline, such that the threshold dose for L-NMMA vasoconstriction was increased and the overall response to L-NMMA reduced. Furthermore, when noradrenaline was used as an internal control there was a significant negative relationship between the response to L-NMMA and blood pressure. When the responses to L-NMMA and noradrenaline were compared between groups, the response to L-NMMA was significantly less in hypertensives compared with normotensives, whereas there was no statistical difference in the response to noradrenaline between the two groups. The results suggest an abnormality of basal nitric oxide-mediated dilation in the forearm arteriolar bed of patients with untreated essential hypertension.


Glomerular and tubular antinatriuretic actions of low-dose angiotensin II infusion in man. The aim was to study the physiological effects of angiotensin II upon the glomerular and tubular handling of sodium. Healthy volunteers were examined before and during infusion with either low-dose angiotensin II (n = 11) or placebo (n = 13). Lithium clearance was used to estimate the segmental tubular reabsorption of sodium. During infusion with angiotensin II a sustained and marked fall in renal plasma flow was observed. The glomerular filtration rate (GFR) decreased to a minor extent so that the filtration fraction increased during angiotensin II infusion. Angiotensin II caused an extensive and instantaneous fall in both urinary flow and urinary sodium excretion. Proximal absolute reabsorption of sodium was unchanged despite the fall in GFR, showing that proximal fractional reabsorption was enhanced by angiotensin II. Distal absolute reabsorption was decreased during the entire period of angiotensin II infusion. However, when the distal reabsorption was related to the delivery of sodium from the proximal tubules, distal fractional reabsorption in fact increased after 30 min angiotensin II infusion. None of the measured parameters changed during infusion with placebo. A significant increase in plasma aldosterone was observed 30 min after the start of the angiotensin II infusion. Plasma atrial natriuretic peptide did not change during infusion with either angiotensin II or placebo. We conclude that physiological increments in angiotensin II affect glomerular haemodynamics and cause a marked antinatriuresis in man. The antinatriuretic effect of angiotensin II is caused initially by a combination of a decrease in the GFR and an increase in proximal fractional sodium reabsorption, and later by the enhanced distal fractional reabsorption of sodium.


The role of atrial natriuretic peptide in natriuresis of fasting. To examine the relation between plasma atrial natriuretic peptide (ANP) and the natriuresis of fasting. ANP, aldosterone and renin were examined during natriuresis of fasting in 25 obese essential hypertensive patients and nine overweight normotensive subjects placed on a supervised 500-KCal diet composed of 50% carbohydrates, 30% fat and 20% protein, and unlimited salt. Twenty-four-hour urinary electrolytes were measured on days 0, 4, 7 and 10 of the diet. Urinary sodium concentration nearly doubled in the patients on day 4, and increased 1.4-fold in the normotensive controls. Plasma ANP rose nearly threefold in the hypertensives on day 4 and nearly doubled in the normotensives. Patients and controls showed similar patterns of natriuresis and ANP secretion during the diet. We conclude that there is a clear association between ANP levels and natriuresis of fasting.


Unmasking of the hypotensive effect of nifedipine in normotensives by addition of the angiotensin converting enzyme inhibitor benazepril. To study the pharmacological interaction between a dihydropyridine derivative (nifedipine slow release 20 mg) and inhibition of the renin-angiotensin system (benazepril 10 mg). Single application at intervals of 2 weeks in an open-label three-way cross-over design. Institutional pharmacological unit. Nine healthy male volunteers. Blood pressure and heart rate were recorded in the supine position for 24 h as well as plasma drug levels, plasma angiotensin converting enzyme activity and active plasma renin concentration. Nifedipine increased active plasma renin two-fold and benazepril increased it five-fold. The combination of the two drugs accelerated the increase of active renin during the first 2 h after drug intake. Whereas no hypertensive effect could be detected after nifedipine or benazepril alone, a significant fall in systolic and diastolic blood pressure was observed for up to 9 to 12 h after the combination. The increase in heart rate induced by nifedipine was minimized by the addition of benazepril. There was no interaction between the pharmacokinetics of benazeprilate and nifedipine which would explain these pharmacodynamic effects. These results demonstrate that, in normotensive volunteers, the renin-angiotensin system contributes to mask the hypotensive effect of a single oral dose of dihydropiridine. The concomitant administration of a converting enzyme inhibitor discloses the hypotensive effect and limits the baroreflex-mediated increase in heart rate secondary to vasodilation.


Weight reduction versus antihypertensive drug therapy in obese men with high blood pressure: effects upon plasma insulin levels and association with changes in blood pressure and serum lipids. First, to compare dietary and antihypertensive drug treatment in obese men with mild hypertension with respect to effects upon insulin, glucose, lipid metabolism and blood pressure. Second, to test the hypothesis that in the diet group changes in blood pressure and serum lipid concentration were associated with changes in plasma insulin concentration. A 6-week run-in period followed by random assignment to either diet or drug treatment, lasting for 1 year. Blood pressure measurements were performed blind after 5 and 45 min rest, and during isometric exercise. Plasma insulin and blood glucose concentrations were measured before and after an oral glucose load. Outpatient clinic in a city hospital. Sixty-four men aged 40-69 years with a body mass index > or = 26 kg/m2 and with a diastolic blood pressure of 90-104 mmHg when untreated were recruited (screening after an advertisement in a newspaper). Exclusion criteria were diabetes mellitus, organ damage secondary to hypertension and diseases that may have interfered with compliance and the interpretation of results. Sixty-one patients completed the study. Dietary treatment was based upon weight reduction and sodium restriction. Drug treatment used a stepped-care approach with atenolol as first choice drug. Absolute reductions in blood pressure, plasma insulin, blood glucose, serum lipid concentration and the waist:hip circumference ratio. Mean body weight decreased in the diet group and increased in the drug-treatment group. Plasma insulin concentrations, the waist:hip circumference ratio and serum lipid profile improved in the diet group compared with the drug group. Blood pressure control was significantly better in the drug group. In the diet group the changes in mean arterial pressure after 5 min rest and serum triglyceride levels correlated with changes in plasma insulin concentrations independent of changes in body mass index or body weight. Diet treatment was inferior to drug treatment in controlling hypertension, but superior in lowering plasma insulin concentrations and improving the serum lipid profile. The hypothesis of a relation between changes in blood pressure, serum triglycerides and plasma insulin was supported.


A health-economic comparison of diet and drug treatment in obese men with mild hypertension. To compare dietary and antihypertensive drug treatment in obese men with mild hypertension in economic terms. A 6-week run-in period followed by randomization to either diet or drug treatment, lasting for 1 year. Blood pressure was measured blindly and serum lipid concentrations assessed at run-in and after 1 year. A computer-based model was used in five cost-effectiveness simulations with different assumptions as to the effect upon coronary heart disease risk from the changes in diastolic blood pressure and cholesterol, both total and high-density lipoprotein. A cost-benefit analysis was also performed, calculated as willingness to pay for treatment, as assessed by questionnaire, minus total cost. Outpatient clinic in city hospital. Sixty-four men aged 40-69 years with body mass index > or = 26 kg/m2 and a diastolic blood pressure 90-104 mmHg when untreated were recruited (screening after advertisement in newspaper). Exclusion criteria were diabetes mellitus, organ damage secondary to hypertension, and diseases that might have interfered with compliance and the interpretation of results. Sixty-one patients completed the study. Dietary treatment was based upon weight reduction and sodium restriction. Drug treatment used a stepped-care approach, with atenolol as the drug of first choice. Life years gained and willingness to pay. Drug treatment was the preferred option in three of the five cost-effectiveness simulations. The cost-benefit analysis did not show any difference between the two groups. Non-pharmacological treatment seemed to be less cost-effective than drug treatment. However, more studies and further methodological development are needed to verify this finding.


Life style changes improve insulin resistance in hyperinsulinaemic subjects: a one-year intervention study of hypertensives and normotensives in Dalby. Insulin resistance and hyperinsulinaemia are, in some prospective studies, linked to an increased cardiovascular risk, at least in men. We tested the hypothesis that hyperinsulinaemia may be reduced by non-pharmacological methods independently of other cardiovascular risk factors. In a non-pharmacological intervention study for 1 year three groups of subjects (hypertensives as well as normotensives) were selected after stratification for insulin level at baseline. Half of the hyperinsulinaemic subjects were randomly assigned to active intervention with physical exercise and dietary regulation (HI-A group), the other half were followed passively during the study period (HI-P group). Normo-insulinaemics and hypo(low)-insulinaemics also underwent active intervention (NI-A and LI-A groups, respectively). Primary health care in Sweden. During the 1-year follow-up subjects in the HI-A group reduced their weight, waist:hip ratio and systolic and diastolic blood pressure, as well as their low:high-density lipoprotein (LDL:HDL)-cholesterol ratio. Glucose levels before and during an oral glucose tolerance test did not change. However, plasma insulin and plasma-C-peptide decreased both in the fasting state and after 1 and 2 h of oral glucose tolerance testing. This decrease was independent of the previously mentioned reduction in weight, waist:hip ratio, blood pressure and LDL:HDL-cholesterol ratio. No reduction in insulin levels was seen in the HI-P, NI-A or LI-A groups, but in the HI-P group there was a slight decrease in fasting plasma-C-peptide levels. In the HI-A group dietary improvements were observed during the study period, with a reduction in energy intake, fat consumption and cholesterol intake. Fibre intake was increased. No major changes were seen in the HI-P group. We conclude that in hypertensive and normotensive subjects with hyperinsulinaemia insulin levels can be reduced by active non-pharmacological treatment for 1 year without altering glucose tolerance. This shows that insulin resistance may be lowered by non-pharmacological treatment, which may be of considerable importance, and not only for hypertensives.


Emotional coping and tonic blood pressure as determinants of cardiovascular responses to mental stress. The aim was to assess the combined influence of biological risk for hypertension and patterns of emotional control upon cardiovascular responses to mental stress tests. The study involved the administration of mental stress tests in the laboratory, designed to elicit substantial blood pressure and heart rate responses accompanied by suppression of cardiac baroreflex sensitivity. Thirty-seven young men were selected as being at relatively high or low risk through having high or low normal blood pressure. Blood pressure, recorded continuously using the Finapres, heart rate, cardiac baroreflex sensitivity, skin conductance and respiration rate were monitored at rest and during mental arithmetic and mirror drawing tasks. Hypertension risk category had no overall effect upon cardiovascular reactions to mental stress. Two dimensions of emotional coping were identified through factor analysis of psychological questionnaires--anxious emotional inhibition (ratings of trait anxiety, anger in and self-concealment), and anger experience and expression (ratings of trait anger and anger out). Subjects with high and low scores on these dimensions were equally represented in the two blood pressure risk categories. Hypertensive risk interacted with anxious emotional inhibition, with the greatest systolic blood pressure and heart rate responses (accompanied by cardiac baroreflex inhibition) being recorded in subjects at high risk coupled with high anxious emotional inhibition. Anger experience and expression did not interact with hypertension risk, but had a direct effect upon cardiovascular responses to mental stress. No differences were seen in skin conductance or respiratory responses, suggesting specific disturbances of cardiovascular regulation. The results suggest that normotensives at risk for future hypertension are likely to show heightened stress-related cardiovascular responses if they also tend to inhibit the expression of negative emotions. This pattern may be relevant to the postulated links between hypertension and emotional inhibition.


Familial influence upon NaCl sensitivity in patients with essential hypertension. The role of genetic factors in the pathogenesis of essential hypertension has not been fully clarified. In order to characterize the relation between NaCl sensitivity and genetic features of hypertension, the responses to dietary Na manipulations were examined in essential hypertensive patients with and without a family history of hypertension. Fifteen essential hypertensive patients, both of whose parents were hypertensive, were compared with 25 hypertensive patients in whom a family history of hypertension was known to be absent in first- and second-degree relatives. There were no differences in gender distribution, age, blood pressure or duration of hypertension between the two groups. Subjects were studied as inpatients on a daily diet of 170 mmol NaCl for 1 week, followed by 1 week on a low-NaCl diet (50 mmol/day) and then by 1 week on a high-NaCl diet (340 mmol/day). Plasma renin activity on a low-NaCl diet was lower, and the erythrocyte Na+ concentration on both diets higher, in hypertensive patients with than in those without a family history. The elevations in mean blood pressure and in erythrocyte Na+ concentration with a high-NaCl diet were greater in patients with a family history. A significant correlation between the change in mean blood pressure and the change in erythrocyte Na+ concentration was found in patients with a positive family history, but not in those with a negative family history. Essential hypertensive patients with an apparent hereditary component of hypertension can be characterized as the low-renin and Na-sensitive subgroup with intracellular Na+ accumulation. Genetic features may underlie, at least in part, the heterogeneity of essential hypertension.


Arterial hypertension in urban Africa: an epidemiological study on a representative sample of Dakar inhabitants in Senegal. To assess the prevalence and risk markers of arterial hypertension in the context of a rapidly growing urbanization in Africa. A cross-sectional study was performed on a representative sample of inhabitants of Pikine, an urban suburb of Dakar, Senegal. Blood pressure was measured by properly trained field workers at subject homes. A total of 2580 individuals were included, aged > or = to 15 years and with a duration of residence > or = 2 months. Complete data including risk markers were obtained for 2300 subjects. The prevalence of hypertension (World Health Organization criteria) was found to be 10.4%. Due to the young age of the population, nearly half of the cases of moderate-to-severe hypertension were observed in middle-aged subjects. Risk markers of hypertension were age and obesity in both sexes: illiterate and multiparous women also had a higher prevalence of hypertension. Awareness of hypertension was infrequent among the hypertensive subjects: of the 189 subjects who had been labelled hypertensive, 10 were treated with antihypertensive therapy but 111 had normal blood pressure levels without any treatment, suggesting poor-quality prior diagnosis. Hypertension was found to be frequent in this urban African population. These findings emphasize the need for improvement in the management of hypertension, focusing upon the high-risk groups.


Resistance vessel gene expression of nerve growth factor is elevated in young spontaneously hypertensive rats. In this study we determined whether the enhanced production of nerve growth factor (NGF) and the associated hypernoradrenergic innervation of the vasculature of the spontaneously hypertensive rat (SHR) was associated with an increased gene expression of messenger (m)RNA encoding for nerve growth factor. It has been shown previously that the hypernoradrenergic innervation of the SHR occurs early, as does the enhanced expression for NGF. In this study we analysed the content of NGF mRNA in blood vessels from young SHR and normotensive Wistar-Kyoto (WKY) rats. Total RNA was isolated from mesenteric arteries from 2-, 10- and 43-day-old SHR and WKY rats and RNA was also isolated from caudal arteries from 43-day-old rats. The RNA was subjected to Northern transfer or slot blots and the content of NGF mRNA measured after hybridization with a 32P-labelled complementary (c)DNA probe for NGF. Slot blot analysis indicated a larger concentration of NGF mRNA in mesenteric and caudal arteries from SHR than for tissues from WKY rats. In this genetic model of hypertension the results indicate an association between an enhanced level of NGF mRNA and the appearance of vascular hypernoradrenergic hypertension.


Effects of enalapril and hydralazine treatment and withdrawal upon cardiovascular hypertrophy in stroke-prone spontaneously hypertensive rats. To test the hypothesis that effects of angiotensin converting enzyme (ACE) inhibitors upon resistance vessel structure are responsible for their ability to cause long-term reduction in blood pressure. Stroke-prone spontaneously hypertensive (SHRSP) and Wistar-Kyoto (WKY) rats were treated with enalapril or hydralazine from 4 to 15 weeks of age. Effects upon tail-cuff blood pressure, left ventricular hypertrophy and structural indices of the superior mesenteric artery (SMA) and its resistance vessels were assessed at 11 weeks of treatment and up to 11 weeks post-treatment. Left ventricular hypertrophy was assessed by left ventricular weight:body weight ratios. Evidence of vascular structural change was obtained from tissue weight:body weight ratios, levels of RNA, DNA and expression of alpha-actin and elastin messenger (m)RNA. The effects of enalapril and hydralazine upon left ventricular hypertrophy in SHRSP were consistent with their respective effects upon blood pressure. Both drugs prevented the development of medial hypertrophy in SMA and resistance vessels. This was accompanied by substantial reductions in RNA:DNA ratios. Alpha-actin mRNA levels were not affected by either drug but elastin mRNA levels were reduced by both drugs. During the first 12 days post-treatment there was evidence of structural change in SMA accompanying the increases in blood pressure but importantly not in the resistance vessels. The effects of enalapril upon left ventricular hypertrophy and mesenteric arterial hypertrophy are totally consistent with responses to blood pressure and the persistence of structural changes post-treatment does not underlie the ability of the ACE inhibitors to persistently suppress hypertension.


Dietary sodium restriction and pressor responsiveness to tyramine in spontaneously hypertensive rats. To determine in vivo whether in young spontaneously hypertensive rats (SHR) dietary sodium restriction decreases adrenergic transmitter release from the sympathetic nerve terminal. Dietary sodium restriction was initiated in young and mature SHR and Wistar-Kyoto (WKY) rats, and subsequently changes in pressor responsiveness to norepinephrine and to the indirectly acting sympathomimetic tyramine were determined in relation to their effects upon plasma catecholamines. In young SHR sodium restriction for 3-6 weeks prevented the development of hypertension, whereas in mature SHR sodium restriction did not affect blood pressure. Sodium restriction caused modest decreases in pressor responsiveness to the exogenous alpha-agonist, not different in young and mature SHR compared with WKY rats. In contrast, sodium restriction markedly inhibited pressor-responses to tyramine in young SHR and WKY rats, but not at all in mature rats. Tyramine increased plasma norepinephrine 5-10-fold. However, sodium restriction did not affect this response. The pressor response to tyramine was related to increases in total peripheral resistance, with minimal changes in cardiac output, and could be blocked by alpha 1-receptor blockade in rats on either control or low-sodium diets. These results show that sodium restriction causes only a small decrease in the pressor response to norepinephrine, but a more marked inhibition of the pressor response to tyramine in young SHR and WKY rats without affecting the plasma norepinephrine response to tyramine. These results suggest that dietary sodium can indeed affect presynaptic functions in vivo, but that plasma norepinephrine responses to tyramine may not reflect changes in arterial norepinephrine release, or that sodium restriction affects a co-transmitter rather than norepinephrine release per se.


Systemic and regional hemodynamic effects of 1,25-dihydroxyvitamin D3 administration. To evaluate the cardiovascular effects of 1,25-dihydroxyvitamin D3 (1,25-D). Recent studies suggest that Ca-regulating hormones may contribute to the genesis of hypertension. We determined systemic and regional hemodynamics 24 h after administration of 1,25-D or vehicle to normal conscious Sprague-Dawley rats. In addition, to dissociate the vascular effects of 1,25-D from changes in serum ionized Ca2+, 1,25-D and vehicle were administered to rats maintained for 3 days on a low-Ca diet. To evaluate the effect of the slight rise in serum ionized Ca2+ with 1,25-D administration, we infused CaCl or vehicle over 1 h into normal rats to raise the serum Ca2+ to near that of rats treated with 1,25-D. The radioactive microsphere technique was used. Systemic hemodynamics (blood pressure, heart rate, cardiac output, total peripheral resistance and stroke volume) did not differ between the two groups receiving a normal-Ca diet. In these rats 1,25-D significantly decreased renal blood flow (RBF), increased renal vascular resistance (RVR) and slightly increased serum ionized Ca2+. Similarly, in rats receiving a low-Ca diet, 1,25-D administration decreased renal blood flow, increased renal vascular resistance and caused only a minimal increase in serum ionized Ca2+. A low-Ca diet also increased heart rate, cardiac blood flow and renal blood flow. Although CaCl infusion raised serum ionized Ca2+, blood pressure, renal blood flow and renal vascular resistance did not change significantly. 1,25-D may constrict the renal vasculature directly or indirectly by enhancing the vascular sensitivity to circulating vasoconstrictors.


Angiotensin II receptor antagonist delays renal damage and stroke in salt-loaded Dahl salt-sensitive rats. To study the effects of blockade of the renin-angiotensin system upon the development of hypertension, end-organ damage and mortality in Dahl salt-sensitive (DSS) rats using an angiotensin II receptor antagonist, losartan. DSS rats (n = 186) were fed 8% NaCl from 6 to 16 weeks of age. One group received losartan whilst the control group was untreated. Changes in blood pressure and plasma renin activity (PRA), as well as renal and cerebrovascular damage and survival were assessed during the study. Losartan blunted the blood pressure rise only transiently. Salt loading suppressed PRA in both groups until week 4 and thereafter it rose more markedly in the treated group. With no treatment renal lesions were first detected at 2 weeks, and strokes at 6 weeks. However, losartan transiently decreased the incidence and delayed the progression of renal damage and cerebrovascular lesions (strokes) and increased survival. PRA correlated with renal damage and the incidence of strokes in both groups. Blood pressure only partially affected survival, but did not correlate with stroke incidence. These results indicate that whereas the rise in blood pressure is dependent upon sodium loading, morbidity and mortality in salt-loaded DSS rats are associated with activation of the renin-angiotensin system and are only partially related to blood pressure.


Secretion of medullipin I by the kidney requires oxygen. To test the hypothesis that the secretion of medullipin I by the kidney involves an oxidative step. Medullipin I is secreted by kidney renomedullary interstitial cells and is converted to medullipin II by the liver. Medullipin I can be derived from the kidney in the renal venous effluent by perfusing normal rat kidneys with 95% O2- 5% CO2 at an elevated pressure (180 mmHg). To evaluate whether the secretion of medullipin I involves an oxidative step normal rat kidneys were perfused at an elevated pressure in the presence of O2, in the absence of O2 and after treatment of the kidneys with a powerful antioxidant. Normal rat kidneys were perfused with 5% albumin bubbled with O2-CO2 at 180 mmHg. This was the control procedure for each of the three approaches. In approach (1), the kidneys were perfused with 5% albumin bubbled with N2. In approach (2), the kidneys were perfused with 'blood' treated with carbon monoxide. In approach (3), the kidneys were treated with the antioxidant butylated hydroxytoluene then perfused with 5% albumin bubbled with O2-CO2. Each perfusate was tested for medullipin I activity by rapid intravenous injection into the SHR. All three approaches, which exclude the action of molecular O2, prevented the secretion of medullipin I by the kidneys. The secretion of medullipin I by the kidneys involves an oxidative step.


H+ pump and Na(+)-H+ exchange in isolated single proximal tubules of spontaneously hypertensive rats. To gain insight into the pathogenesis of hypertension in the spontaneously hypertensive rat (SHR), we compared the maturation of the Na-independent H+ efflux and Na(+)-H+ exchange in microdissected superficial proximal cortical tubule (PCT) S1 and S2 segments of SHR and normotensive Wistar-Kyoto (WKY) rats. Isolated superfused PCT segments were loaded with 2'-7'-bis-carboxyethyl-5(6)-carboxyfluorescein and incubated in nominally HCO3-free solution. We assessed Na-independent N-ethylmaleimide (NEM)-sensitive H+ efflux and Na-dependent H+ efflux by measuring the recovery rate of the intracellular pH following acid loading induced by prepulsing with NH4+. In young prehypertensive SHR the Na(+)-H+ exchange recovery rate in S1 at pH(i) 6.8 was significantly higher than in young WKY rats, whereas in adult rats no significant difference between the two strains could be observed. In S2 segments the Na(+)-H+ exchange recovery rate was similar between SHR and WKY rats for both age groups. In the young, no difference in the NEM-sensitive H+ efflux in S2 PCT was observed between the two strains. In contrast, in the adult, although the NEM-sensitive H+ efflux had increased profoundly with age for WKY rats, it remained markedly low in SHR. These studies indicate that apical Na+ reabsorption coupled with H+ efflux in the S1 segment is increased in the PCT of SHR, and demonstrate a marked impairment in the maturation of H+ pump activity in the S2 segment of the SHR compared with the normotensive strain. The impairment of these cell transport systems in the SHR may be relevant to the pathogenesis or maintenance of hypertension in this model.


Effects of graded vasoconstriction upon the measurement of finger arterial pressure. To assess the effects of incremental phenylephrine infusion rates and subsequent graded vasoconstriction upon the performance of the Ohmeda Finapres. Blood pressure in eight hypertensive patients in the finger and the brachial artery was recorded simultaneously. Systolic blood pressure (SBP), diastolic blood pressure (DPB) and mean arterial pressure (MAP) were compared as well as additional waveform characteristics like the pressure at moment of the dicrotic notch and calculation of the pulsatile-systolic areas. Before phenylephrine infusion SBP and DBP were higher in the finger. At maximal infusion (1.6 micrograms/kg/min) the increase in brachial SBP was significantly underestimated by Finapres. Thus, the computed sensitivities of baroreflex control for SBP differed significantly between the two measurements. Under control conditions, the shape of the finger waveform differed from the brachial-artery waveform in terms of: (1) a more peaked appearance; (2) a dicrotic notch (Pnotch) which is located at a lower percentage of pulse pressure; and (3) a larger pulsatile-systolic area. At maximal infusion rates finger Pnotch increased whilst intrabrachial Pnotch did not. In contrast, the brachial and finger pulsatile-systolic areas changed fully in parallel. Phenylephrine infusion caused a significant, and clinically important, underestimation of the increase in brachial SBP when assessed by Finapres, whereas MAP and DBP and pulsatile-systolic area track intra-arterial pressure reliably.


Non-invasive ultrasound assessment of renal artery stenosis by means of the Gosling pulsatility index. To gauge the effectiveness of a new Doppler test for renal artery stenosis (RAS), based on the pulsatility index of the blood flow velocity spectrum within several interlobar arteries of both kidneys. Twenty normotensive volunteers and 49 hypertensive patients were investigated with ultrasound. Patients with angiographic signs of RAS underwent bilateral renal vein catheterization for renin measurement. Significant RAS was assumed if lateralization of renal vein renin to the stenotic side was proven. The pulsatility index was higher in the hypertensives without RAS than in normal volunteers. Side differences between both kidneys were within methodological variations with the exception of one case, in whom side difference was > 0.12. The pulsatility index was lower in kidneys with significant RAS than in kidneys without RAS. In most patients with significant unilateral RAS the side difference was < 0.12. In the other patients with a low pulsatility index and a side difference < 0.12 RAS was found to be bilateral upon angiography. Doppler signals were absent in all kidneys with renal occlusion. A side difference of > or = 0.12 predicts unilateral RAS, whereas the absence of parenchymal Doppler signals indicate occlusive RAS. A low pulsatility index combined with normal side difference may, in hypertensive patients, indicate bilateral RAS. Renovascular hypertension was correctly diagnosed in 84% of the patients and the presence of RAS in 94%.


Reduced cytosolic free Na+ concentration in intact platelets of essential hypertensives. The role of intracellular Na+ concentration in the pathogenesis of essential hypertension is a point of considerable discussion. Since the novel fluorescent dye technique offers the possibility of measuring cytosolic free Na+ concentration in intact living cells, the role of Na+ was reinvestigated in resting and stimulated human platelets. Cytosolic free Na+ concentration was measured in intact blood platelets of 20 essential hypertensive patients and 21 age- and sex-matched normotensive control subjects using the fluorescent dye Na(+)-binding benzofuran isophthalate. Cytosolic free Na+ concentration was significantly reduced in hypertensives compared with normotensives. Inhibition of Na+,K(+)-adenosine triphosphatase by ouabain elevated cytosolic free Na+ concentration in hypertensives and normotensives in a similar way. Addition of thrombin increased cytosolic free Na+ concentration both in hypertensives and normotensives. Previous concepts concerning the role of Na+ in the pathogenesis of essential hypertension based upon measurements in destructed cells need to be reinvestigarted using new techniques in living cells.


Calcium and platelets in normotensive and hypertensive human pregnancy. The primary objective of this study was to determine the baseline platelet intracellular free Ca2+ concentration in normotensive and hypertensive primigravid women and to note any correlation between variables (ie. platelet intracellular free Ca2+ concentration and arterial blood pressure). A cross-sectional study of 24 normotensive and 36 primiparous women whose pregnancies were complicated by gestational hypertension, in the third trimester. Platelet intracellular free Ca2+, serum Ca2+, and renal clearance of Ca2+ were recorded. Mean platelet intracellular free Ca2+ concentration was significantly increased in those subjects whose pregnancies were complicated by proteinuric gestational hypertension (pre-eclampsia) compared with the normotensive primiparous control sample. Those subjects with non-proteinuric pregnancy-induced hypertension did not show this significant increase in baseline platelet intracellular free Ca2+ concentration despite having a persistent and statistically significantly elevated systemic arterial blood pressure. On pooling these data for both normotensive and hypertensive primigravidae a significant positive correlation was noted between the variables of platelet intracellular free Ca2+ concentration and arterial blood pressure. The renal clearance of free Ca2+ was progressively reduced with increasing severity of the disease but had returned to normal 6 weeks postpartum. Serum Ca2+ concentrations corrected for albumin were however higher in the hypertensive patients. Transmembrane Ca2+ fluxes are altered in hypertensive pregnancy by a specific mechanism, probably of placental origin.


Autocrine growth limitation of human papillomavirus type 16-harboring keratinocytes by constitutively released tumor necrosis factor-alpha. TNF-alpha is known to exert antitumor and antiviral effects and to participate in the regulation of the immune response. In our study we demonstrate that human rTNF-alpha specifically blocks growth of SK-v keratinocyte cell line harboring and expressing human papillomavirus type 16 (HPV16) sequences. This inhibitory effect was shown by [3H]TdR incorporation and cell counting. Binding experiments with 125I-TNF-alpha showed that SK-v cells express about 10,000 single class TNF-alpha R per cell with affinity constant of about 0.7 nM. Binding of 125I-TNF-alpha could be inhibited by htr-9 mAb recognizing a 55/60-kDa type I TNF-alpha R but not by utr-1 mAb recognizing 75/80-kDa type II TNF-alpha R or irrelevant mAb specific for HPV16E7 protein. Addition of anti-TNF-alpha antibodies to SK-v cell culture resulted in significant (p < 0.05), dose-dependent stimulation of their proliferation. SK-v cells constitutively expressed TNF-alpha mRNA, and SK-v CM contained TNF-alpha, as demonstrated by Northern blot analysis, a specific ELISA, Western blot analysis, and a bioassay with TNF-alpha-sensitive L-M cells. HPLC gel filtration of SK-v cell CM showed that the factor cytotoxic for L-M cells coeluted with immunoreactive TNF-alpha. These results demonstrate that HPV16-harboring SK-v cells constitutively express and release immunoreactive and biologically active TNF-alpha that in turn may exert an autocrine growth inhibitory effect. This phenomenon could represent one of the self-limiting mechanisms controling growth of HPV-induced neoplasia.


Regulation of paf-acether receptor expression in human B cells. Paf-acether (paf) is a phospholipid cytokine alloted with potent inflammatory and immunoregulatory properties. Recent reports indicated that in human B cell lines, paf modulated both early and late activation events. In our study, we showed that four of six human B cell lines specifically bound [3H]paf irrespective of the stage of differentiation, the presence of EBV genome or cell surface phenotype. Binding was saturated and fit a one receptor model with a dissociation constant ranging from 1 to 6 nM and a number of sites per cell ranging from approximately equal to 4000 in Rjc13 to approximately equal to 30,000 in Raji or IM9. In addition, our data indicate that 1) maximal expression occurred during the log phase growth; 2) paf itself (10-100 nM) or rIL-4 (100 U/ml) up-regulated by two- to threefold the number of paf binding sites without affecting the affinity. Finally, we found that activated normal B lymphocytes exhibited a higher capacity than resting B cells to incorporate and metabolize [3H]paf at 37 degrees C. Resting B lymphocytes lacked specific binding capacity for paf, yet specific paf receptors were induced upon stimulation via Staphylococcus aureus Cowan I or phorbol 12,13 dibutyrate plus ionomycin. These results suggest that B cell activation is a critical event for paf receptor expression and modulation.


Role of phospholipase D-derived diradylglycerol in the activation of the human neutrophil respiratory burst oxidase. Inhibition by phosphatidic acid phosphohydrolase inhibitors. An agonist-activated phospholipase D/phosphatidic acid phosphohydrolase (PAH) pathway was recently demonstrated in human neutrophils, and evidence suggests that phosphatidic acid (PA) and/or diradylglycerol (DG) generated from this pathway participates in activation of the O2(-)-generating respiratory burst. We have used a series of cationic amphiphilic compounds (sphingosine, propranolol, chlorpromazine, and desipramine) and antibiotics (clindamycin, trimethoprim, and roxithromycin) all of which inhibit the respiratory burst, to investigate the role of the phospholipase D/PAH pathway in neutrophil activation. The phosphatidylcholine (PC) pool in intact cells was first labeled using [3H]-1-O-alkyl-lysoPC; released [3H]-PA and [3H]-DG were then quantified after the addition of either chemo-attractant or PMA. Using either agonist, all compounds showed a dose-dependent inhibition of [3H]-DG generation which correlated with inhibition of O2- generation, but compounds failed to inhibit directly the NADPH oxidase in a cell-free system. For either activator, a plot of the ID50 values for O2- generation vs those for DG generation was linear over four orders of magnitude. In many cases, inhibition of [3H]-DG generation corresponded to an increase in [3H]-PA, implicating PAH as the locus of inhibition. Superoxide generation was inhibited under conditions where PA was either elevated or minimally affected. Neither O2- release nor DG generation showed any selectivity for stereoisomers of propranolol, suggesting that this inhibition does not act via a specific binding site on PAH. No evidence was obtained for an effect of the inhibitors on PA mobility as monitored by electron spin resonance studies of spin-labeled PA in a model membrane system. Data are consistent with an effect of the inhibitors at the level of the interaction of PAH with the membrane and/or its substrate. These data imply that DG produced via the phospholipase D/PAH pathway functions in the activation or maintenance of the respiratory burst.


Four cell-secreted cytokines act synergistically to maintain long term proliferation of human B cell lines in vitro. Autocrine production of growth factors is thought to be an essential element in the development of hemopoietic tumors in vivo. Tumor-derived cell lines frequently show this capability in vitro. It is not understood how autonomous growth in vitro is maintained by lymphoid cell lines that are not of tumorigenic origin. We have previously established human B cell clones that proliferate in serum-free media with unlimited potential. However, the cells need a critical density for continuous growth. Culture supernatant conditioned by these cell lines sustained proliferation even in low density cultures. All B cell clones analyzed were found to secrete the cytokines IL-1 alpha, IL-6, TNF-alpha, and TNF-beta whereas no activity of IL-2, IL-4, low m. w.-B cell growth factor, CSF, or IFN-gamma was recorded. In low density cultures supplemented with rIL-1 alpha, +/- IL-6, +/- TNF-alpha, and +/- TNF-beta together, B cell proliferation is maintained to the same extent as with conditioned medium. Addition of anti-sense oligonucleotides directed to the mRNA of IL-1 alpha, IL-6, and TNF-alpha, respectively, resulted in growth arrest and cell death. This effect could be prevented by supplementation with these cytokines. Scatchard plot analyses and internalization studies revealed that the cells express on their surface high affinity receptors for IL-1 alpha, IL-6, and TNF, respectively, and internalize the cytokines from the supernatant. These results demonstrate that (i) autonomous growth of immortalized B cells is maintained by secretion and reinternalization of IL-1 alpha, IL-6, TNF-alpha, and TNF-beta, (ii) these cytokines act in a synergistic fashion, and (iii) autocrine growth stimulation of human B cells in vitro does not necessarily represent their tumorigenic potential in vivo.


Maturation- and differentiation-dependent responsiveness of human CD4+ T helper subsets. The human CD45R0+ (memory) CD4+ T cell population can be subdivided into a large (82%) CD27+ and a small (18%) CD27- subset. Within the CD45R0+CD27- subset, cells accumulate that have been persistently stimulated by Ag in vivo. As an apparent consequence, TLC with a differentiated functional phenotype, producing either high levels of IFN-gamma (Th1-like), high levels of IL-4 (Th2-like) or high amounts of both these cytokines (here referred to as Thx) can primarily be generated from the CD27- memory CD4+ T cell subset. In this study we examined the requirements for induction of proliferation of distinct CD4+CD45R0+ Th subsets. Immobilized CD3 mAb induced proliferation with comparable magnitude and kinetics in all types of TLC. However, interference with intracellular signaling pathways in this activation system, resulted in a strong inhibition of proliferation in TLC derived from CD27+ cells whereas, in contrast, TLC from CD27- cells were relatively resistant to elevation of [cAMP]i, inhibition of protein kinase C activation and the immunosuppressive effects of cyclosporin A. Stimulation with CD3 mAb in soluble form resulted in Il-4 secretion by Th2-like and Thx-type TLC but did not induce IFN-gamma or Il-2 secretion in any Th subset. Interestingly, Th2-like cells but not Thx-type cells were able to use endogenously produced Il-4 for proliferation. These data demonstrate a differential sensitivity of CD45R0+CD4+ Th subsets for immune activation and suppression, which correlated with their maturation stage, as reflected by CD27 membrane expression, as well as with their effector phenotype.


Role of cathepsin D in antigen presentation of ovalbumin. Modification of protein Ag by proteolysis is one of the principal steps in the presentation of Ag to Th cells. However, little is known about the enzymes participating in these events, their specificity or the characteristics of the natural fragments that they produce. Cathepsin D (CD) is an aspartyl protease identified in endosomes of APC. In this report, the role of CD in the processing of OVA has been investigated. OVA digested in vitro with purified CD was able to stimulate IL-2 secretion by three different OVA-specific I-Ad restricted Th cell hybridomas when it was presented by fixed APC. The digest of OVA was recognized in the context of I-Ad, but not by I-Ak-restricted OVA-specific Th cells. No difference was observed in the ability of OVA digested with CD to stimulate Th cells in the absence of FCS or in the presence of protease inhibitors indicating that extracellular proteases were not likely to contribute to processing of OVA. Taken together, these results suggest that CD is necessary and sufficient for the generation of an antigenic epitope from OVA. A fragment containing the epitope was isolated from the OVA digest by reverse phase HPLC. This fragment, which migrates in SDS-PAGE as a 10-kDa polypeptide, is a potent epitope. Its capacity to activate Th cells is compared to that of the tryptic peptide OVA323-339.


Prostaglandin E2 and cAMP inhibit B lymphocyte activation and simultaneously promote IgE and IgG1 synthesis. Macrophage-secreted prostaglandins of the E series inhibit numerous immunologic events, including IgM secretion by B lymphocytes. In this study, we investigated whether PGE also regulates the activation of normal quiescent murine B cells and subsequent isotype differentiation to IgE and IgG1 production. PGE2 and PGE1 were found to inhibit cellular enlargement induced by IL-4 or bacterial LPS, IL-4 and LPS, or anti-mu and IL-4 by approximately 75%, and completely inhibit enlargement in response to anti-mu antibody. PGE2 also suppresses activation-induced class II MHC up-regulation by 35% and expression of the low affinity IgE receptor, Fc epsilon RII/CD23, by 30%. Interestingly, PGE completely inhibits a fraction of cells from these activation events, while other cells fully respond to activation stimuli, even in the presence of high PGE2 concentrations. Therefore, a PGE-resistant subset of B lymphocytes may exist. A closely related PG, PGF2 alpha, had no immunoregulatory effect in these systems. Because PGE induces production of cAMP in B cells, we determined whether other agents that increase cAMP could inhibit B cell activation. Cholera toxin and dibutyryl cAMP mimicked the ability of PGE2 to inhibit B cell enlargement, and class II MHC and Fc epsilon RII induction, suggesting that PGE2 signaling occurs via cAMP. In addition, cholera toxin and dibutyryl cAMP inhibited B cell activation much more potently (90-100% inhibition) than PGE, indicating that whereas all B cells are cAMP-sensitive only some are PGE-sensitive. Although PGE inhibits activation-associated events, we previously reported that PGE enhances IL-4 and LPS-induced differentiation to IgE and IgG1 synthesis. To investigate the relationship between the cells that are activation-inhibited and those that are differentiation-enhanced by PGE, we sorted B cell subsets by FACS and determined their relative abilities to produce IgM, IgG1, and IgE in response to IL-4 and LPS in the presence of PGE. The population of lymphocytes that was unaffected by PGE in terms of class II hyperexpression was also unaffected by PGE for Ig synthesis, again indicating a PGE-resistant subpopulation of B cells. Furthermore, the PGE activation-inhibited subset of B cells was responsive to PGE enhancement of IL-4-induced class switching, reducing IgM synthesis and inducing a sevenfold increase in IgE and IgG1 synthesis compared with other sort groups. These results are consistent with the hypothesis that the B lymphocytes that are PGE activation-inhibited are the same cells that are PGE differentiation-enhanced.(ABSTRACT TRUNCATED AT 400 WORDS)


Alpha-1-antichymotrypsin inhibits the NADPH oxidase-enzyme complex in phorbol ester-stimulated neutrophil membranes. The generation of superoxide anion and release of granule contents are essential to the bactericidal function of neutrophils, but may also contribute to host tissue damage during inflammation. In previous studies (J. Immunol. 146:2388), we have demonstrated that the acute phase reactant alpha-1-antichymotrypsin (ACT), a potent inhibitor of the serine protease cathepsin G, also suppresses superoxide anion generation. The inhibitory effect of ACT was not directly linked to its antiproteolytic activity and may reflect interaction at a site other than its reactive loop. To further characterize the mechanism of inhibition, we investigated the direct effects of ACT on the NADPH oxidase enzyme complex and the signaling pathways that regulate motivation of the respiratory burst. We present evidence that ACT does not intefer with agonist-stimulated calcium mobilization or translocation and activity of protein kinase C. ACT was an effective inhibitor of superoxide anion generation in membrane preparations isolated from PMA-activated cells. These results support the notion that ACT is acting on a component of the active assembled NADPH oxidase complex. Thus, ACT may have an important role in regulation of specific aspects of the inflammatory processes and the modulation of toxic oxygen-based host tissue damage.


Ligation of CD23 triggers cAMP generation and release of inflammatory mediators in human monocytes. Transduction through the CD23 molecule (Fc epsilon RII) was analyzed in normal human monocytes using monoclonal antibodies to CD23 (MHM6 and 135) and IgE/anti-IgE immune complexes. Monocytes expressing an increased amount of CD23 molecules were obtained by stimulation with IL-4 (30 U/ml). Anti-CD23 mAb as well as IgE/anti-IgE immune complexes were unable to induce any significant calcium mobilization [Ca2+]i in CD23-bearing monocytes whereas they elicited [Ca2+]i increase in B lymphocytes of the same donors. Despite their failure to induce calcium mobilization, the same CD23 ligands triggered a dose-dependent increase of intracellular cAMP, with a maximum 20 to 30 min after the onset of stimulation. This effect is mediated via CD23 inasmuch as: 1) F(ab)'2 fragments are as active as intact anti-CD23 mAb and 2) it is not observed in CD23- monocytes. The increase in cAMP was only partially altered in the presence of 1 microM indomethacin suggesting that it was not due to the release of PG. The possible role of CD23 in the activation of human monocytes was next documented by showing that anti-CD23 mAb and IgE/anti-IgE immune complexes induced the generation of IL-6 and of thromboxane B2 by CD23+ but not by CD23- monocytes. In addition, the IgE/anti-IgE-induced IL-6 production was potentiated in the presence of cAMP inducer such as the beta 2-adrenoceptor agonist salbutamol. These results indicate that ligation of CD23 induces cAMP generation in CD23+ human monocytes and that CD23 may regulate the IgE-dependent functions in normal human monocytes.


Effect of fixation on the detection of transmissible gastroenteritis coronavirus antigens by the fixed-cell immunoperoxidase technique. The effect of various fixatives and detergents on the in vitro detection of the viral determinants which are expressed in swine testis cells infected with the transmissible gastroenteritis coronavirus (TGEV) was studied using a microwell immunoperoxidase technique. When compared with glutaraldehyde and formaldehyde, 0.1% paraformaldehyde was found to be the fixative of choice for the detection of these determinants on the membranes of infected cells. Among dehydrating fixatives, 80% acetone or a mixture of acetone and ethanol or of acetone, methanol and ethanol were found to be the best fixatives for the detection of these viral determinants which are expressed in infected cells. In the case of acetone, the temperature of fixation and its concentration in the fixative preparation were found to be important. The treatment of 0.05% glutaraldehyde-fixed, infected cells with 0.1% saponin or 0.1% paraformaldehyde-fixed, infected cells with 1%NP-40 led to satisfactory detection of viral determinants. Using Triton X-100 to render cells permeable, the quantities of N and M antigen detected in TGEV-infected cells prefixed with either 0.05% glutaraldehyde or 0.1% paraformaldehyde were equal to those of 80% acetone-fixed, TGEV-infected cells while the quantity of S antigen detected was diminished. The effect of other detergents such as zwittergent, empigen BB, Chaps and N-lauroylsarcosine on the detection of viral determinants was also studied.


A rapid isolation procedure for dendritic cells from mouse spleen by centrifugal elutriation. The standard isolation procedure for antigen presenting dendritic cells (DC) takes 2 days and includes selective adherence to tissue culture plates which may lead to the activation of these cells. This report describes the isolation of DC by centrifugal elutriation (CE). Murine spleen cells were separated on the basis of size and density into 7 CE fractions. This method took 90 min. Cells from each CE fraction were characterized by fluorescence activated cell sorter (FACS) analysis and their antigen presenting cell (APC) activity was determined by a secondary Sendai virus specific T cell proliferation assay. CE fraction 5 contained most of the DC with a concentration of 6-10%, representing an approximately 15-fold enrichment compared to unseparated spleen cells (< 1% DC). This CE fraction also exhibited the highest APC activity, which was almost completely abolished after depletion of DC by treatment with monoclonal antibody 33D1 (DC-marker) and complement. Further enrichment of CE fraction 5 by discontinuous density gradient centrifugation resulted in a cell population containing 35-55% 33D1-positive cells with similar characteristics as DC isolated by the standard procedure, such as the capacity to induce a primary viral peptide specific CTL response. Two-color FACS analysis showed an increase in MHC expression on 33D1-positive cells of CE fraction 5 after 18 h culture involving cell adhesion to a similar level as the MHC expression on DC isolated by the standard procedure. During this same period their morphology changed from a round to a dendritic appearance. In conclusion, our results indicate that CE is well suited for isolating DC more rapidly and without activation of these cells by adherence, a process which readily occurs in the standard isolation procedure.


Enzyme immunoassays for the estimation of adenosine 3',5' cyclic monophosphate and guanosine 3',5' cyclic monophosphate in biological fluids. Simple, rapid and sensitive competitive enzyme immunoassays for the estimation of adenosine 3',5' cyclic monophosphate (cAMP) and guanosine 3',5' cyclic monophosphate (cGMP) in human plasma and urine are described. Specific antisera to each nucleotide were raised in rabbits by immunization with succinyl cyclic nucleotide--human serum albumin conjugates. For the assay, specific antibodies were incubated with a mixture of succinyl cyclic nucleotide labelled with horseradish peroxidase together with unlabelled standard or sample. The antibody-bound enzyme conjugate was separated from free hapten by anti-rabbit (IgG) sera immobilized to a microtitre plate. Activity of the bound enzyme conjugate was determined with tetramethylbenzidine. The assays were capable of detecting levels as low as 2 fmol of cAMP and cGMP. Good correlations were obtained between values generated by enzyme immunoassay and radioimmunoassay.


A peroxidase-independent method for the quantitation of extracellular hydrogen peroxide generated by activated phagocytes in vitro. A peroxidase-independent method for the determination of phagocyte-derived hydrogen peroxide in vitro is described. The method is based on the catalase-inhibitable oxidation of added cysteine. Human blood neutrophils (1 x 10(6)/ml) were coincubated with the myeloperoxidase (MPO) inhibitor, sodium azide (10 micrograms/ml), for 30 min at 37 degrees C followed by addition of phorbol 12-myristate 13-acetate (PMA, 10 ng/ml), a potent activator of membrane-associated oxidative metabolism. After incubation at 37 degrees C the cells were removed and the supernatants aliquoted and incubated with and without catalase (500 U/ml) for 10 min at room temperature followed by addition of cysteine (50 microM). Thereafter the catalase-inhibitable, H2O2-mediated oxidation of added cysteine was assayed spectrophotometrically following the addition of the thiol-reactive agent 5,5'-dithiobis-(2-nitrobenzoic acid), and the H2O2 concentration determined using a standard curve constructed from difference data based on the oxidation of cysteine by H2O2 (0-200 nmol). The average amount of H2O2 produced by 10(6) PMA-activated neutrophils was 47 +/- 7 nmol/30 min. This sensitive assay procedure is likely to be particularly useful for investigating the effects of pharmacological agents and anti-oxidant nutrients on H2O2 production by activated phagocytes, since these agents are generally unsuited for use in peroxidase-based assays.


Processing of delta-endotoxin from Bacillus thuringiensis subsp. kurstaki HD-1 and HD-73 by gut juices of various insect larvae. Midgut juices were prepared from Adoxophyes sp., smaller tea tortrix (STT); Bombyx mori, silkworm (SW); Spodoptera litura, common cutworm (CCW); Plutella xylostella, diamondback moth (DBM); and Musca domestica, housefly (HF) and immobilized onto Sepharose 4B. delta-Endotoxins (ICPs) from Bacillus thuringiensis subsp. kurstaki HD-1 and HD-73 were digested by these immobilized gut juice proteases. All gut juices tested derived relatively proteolytic resistant cores from ICP. The molecular sizes of these cores, about 55 kDa in SDS-PAGE, were resulted. In the case of CCW, however, digestion was very strong and only 1/20 concentration of core protein remained relative to other digests. The N-terminal amino acid sequencing of the core proteins showed that they were truncated at the very end of the N-terminus of protoxin, CryIA, at different sites. Although housefly larvae were completely insensitive to active toxin, the gut juice produced the core, suggesting that the housefly may lack the binding sites for the core-active toxin.


Alpha-MSH and other melanogenic activators mediate opposite effects on tyrosinase and dopachrome tautomerase in B16/F10 mouse melanoma cells. alpha-MSH was found to decrease the recently characterized dopachrome tautomerase activity in cultures of B16/F10 mouse melanoma cells. Other stimulating agents of melanogenesis, like dibutyryl cyclic AMP, 3-isobutyl-1-methylxanthine, theophylline, retinol, and retinoic acid, caused the same effect. The grade of inhibition depended on the nature of the agent and the time of exposure. In all cases, both melanin production and tyrosinase activity were activated by these treatments, although the grade of tyrosine hydroxylase and dopa oxidase stimulation was different. Moreover, no correlation among the intensities of dopachrome tautomerase inhibition and tyrosinase activation by the tested agents could be obtained. The significance of these results in the regulation of mammalian melanogenesis is discussed.


Gap-junctional protein connexin 43 is expressed in dermis and epidermis of human skin: differential modulation by retinoids. Retinoids are effective modulators of proliferation and differentiation of keratinocytes in vivo and in vitro. In mouse 10T1/2 cells, retinoid action on proliferation and neoplastic transformation is correlated with the upregulation of gap-junctional communication and expression of connexin 43 (Cx43). In the present study we have determined if retinoids induce similar effects on gene expression in human skin. Studies were conducted in intact skin and on cultured keratinocytes and dermal fibroblasts. In a clinical study, 2 weeks of treatment with 0.05% all-trans retinoic acid resulted in increased expression of Cx43 mRNA and protein in epidermis. Expression occurred predominantly in the suprabasal layer. Cultured cells exhibited a differential response to retinoic acid. In keratinocytes, increased expression of Cx43 occurred at low (10(-11) M) concentrations, whereas inhibition occurred at high (10(-7) M) concentrations; however, junctional communication, measured by dye transfer, was not altered over this concentration range. Dermal fibroblasts, in contrast, exhibited a dose-dependent increased expression of Cx43 at concentrations up to 10(-7) M retinoic acid and proportionately increased their junctional communication over this dose range. These data indicate that control of Cx43 gene expression by retinoids in human skin cells is complex. The production of gradients of junctional channels could play a role in the control of growth and differentiation in epidermis.


Human immunodeficiency virus type 1 (HIV-1) inhibitory interactions between protease inhibitor Ro 31-8959 and zidovudine, 2',3'-dideoxycytidine, or recombinant interferon-alpha A against zidovudine-sensitive or -resistant HIV-1 in vitro. Protease inhibitor Ro 31-8959, a compound that interrupts human immunodeficiency virus (HIV)-specific formation of infectious virions, was evaluated in two-drug combined regimens with zidovudine, 2',3'-dideoxycytidine (ddC), or recombinant interferon-alpha A (rIFN-alpha A) against HIV-1 replication in vitro. By using peripheral blood mononuclear cells infected with HIV-1, drug interactions were evaluated by the median-effect principle and the isobologram technique. A zidovudine-sensitive and -resistant HIV-1 isolate pair was studied. Additive to synergistic anti-HIV-1 interactions were seen with 7.5-30 nM Ro 31-8959 and 0.005-0.02 microM zidovudine (for the zidovudine-sensitive HIV-1 isolate), 0.25-1.0 microM zidovudine (for the zidovudine-resistant HIV-1 isolate), 0.025-0.1 microM ddC, and 8-32 units/mL rIFN-alpha A, without additive toxicity. Phase I/II clinical trials of Ro 31-8959 for therapy of HIV-1 infection are in progress. If results are favorable, combined regimens including Ro 31-8959 deserve consideration for future clinical trials.


Detection of varicella-zoster virus DNA in human geniculate ganglia by polymerase chain reaction. Latent varicella-zoster virus (VZV) has been demonstrated in the human trigeminal and thoracic ganglia by means of nucleic acid hybridization. However, the human geniculate ganglia in VZV latency have not been examined. Tissue DNA extracted from the trigeminal and geniculate ganglia of a newborn and 7 adults was examined by polymerase chain reaction with a pair of VZV-specific primers. None had symptoms of recent infection with VZV (chickenpox or shingles). VZV DNA was detected in 11 (79%) of 14 trigeminal ganglia and in 9 (69%) of 13 geniculate ganglia of the adults. VZV DNA was not detected in either type of ganglion from the newborn or from 1 adult who was seronegative for VZV antibodies. These findings indicate that VZV becomes latent in human geniculate ganglia after primary infection and suggest the possibility that reactivation of the virus from the geniculate ganglia may cause Ramsay Hunt syndrome.


Resistance of hepatitis delta virus replication to interferon-alpha treatment in transfected human cells. Interferon-alpha (IFN-alpha) is known to inhibit both DNA and RNA viruses, including hepatitis B virus (HBV). In humans the antiviral effect, if any, of IFN-alpha on hepatitis delta virus (HDV) is complicated by the fact that HDV is spread only to patients already infected with HBV. An in vitro model system was used to assay for an antiviral effect of IFN-alpha on HDV genome replication. Hep G2 cells were transfected with a plasmid containing a trimer of HDV and treated with IFN (20 or 100 units/mL) starting 1-7 days after transfection. RNA extracted from treated and nontreated cells was assayed by both slot blot and Northern analyses. The IFN-alpha treatment as expected increased the 2'-5' oligo A synthetase RNA activity, but it did not affect HDV genome replication. Thus, in the absence of HBV, it appears that HDV is resistant to IFN-alpha.


Detection of replicative form of hepatitis C virus RNA in peripheral blood mononuclear cells. Paired samples of plasma and peripheral blood mononuclear cells (PBMC) from 7 patients with posttransfusion hepatitis C were collected and studied for the presence of replicative forms of hepatitis C virus (HCV). RNA was extracted and tested for both positive and negative strands of HCV RNA by polymerase chain reaction. Positive-strand RNA of HCV was found in both plasma and PBMC of all 7 patients. However, negative-strand RNA was found in PBMC of 3 patients while none was found in the plasma. Levels of negative-strand RNA were about 10-100 times less than those of positive-strand RNA by semiquantification with serial dilution of viral cDNA. The results suggest that active infection and replication of HCV in PBMC is present in patients with posttransfusion hepatitis C and implicate the extrahepatic infection of HCV.


Colposcopic and histologic findings and human papillomavirus (HPV) DNA test variability in young women positive for HPV DNA. The purpose of this study was to determine the prevalence of human papillomavirus (HPV)-related disease in women positive for HPV DNA by using a commercially available DNA detection kit and to compare these results to HPV DNA variability observed in repeated sampling. Young women attending family planning clinics who were positive for HPV DNA on routine screening were asked to return for a repeat HPV DNA test, cytology, colposcopy, and biopsy if indicated. Of the 78 women examined, 35% had biopsy-verified low-grade dysplasia of the cervix, and 64% had evidence of HPV-related disease somewhere in the anogenital area. Fifty percent of the women had a negative test on repeat sampling, but there was no association between DNA persistence at the second visit and colposcopic/histologic findings. In conclusion, the minority of women positive for HPV DNA have latent infection. In addition, variability in detection is common and unrelated to the presence of HPV-related disease.


Synergistic interaction of interferon-beta and interferon-gamma in coxsackievirus B3-infected carrier cultures of human myocardial fibroblasts. The antiviral effects of human interferon-beta (IFN-beta) and human recombinant interferon-gamma (rIFN-gamma) were studied in persistently coxsackievirus B3-infected carrier cultures of human myocardial fibroblasts over a period of 21 days. Synergism was observed with concentrations as low as 30 IU of IFN-beta plus 10 IU of rIFN-gamma/mL, reducing mean viral titers from 6.0 x 10(7) to 1.3 x 10(4) pfu/mL and number of infected cells from 14.4% to 0.1% as determined by quantitative in situ hybridization. Higher concentrations of IFNs (both > or = 30 IU/mL) were associated with transient antagonism followed by antiviral synergism. With 100 IU of IFN-beta plus 30 IU of rIFN-gamma/mL, elimination of infectious virus was consistently achieved and sustained for 6 weeks after cessation of IFN application, whereas at least threefold higher concentrations were required with single drugs. In summary, our data support a concept of low-dose IFN combination schedules that might become useful in the treatment of enteroviral heart disease.


Type I interferons inhibit hepatitis B virus replication and induce hepatocellular gene expression in cultured liver cells. A hepatoblastoma cell line transfected with hepatitis B virus (HBV) DNA (Hep G2.2.15) was used to investigate the effects of interferons (IFNs) on HBV replication and hepatocellular gene expression. IFN-alpha 2b or -beta inhibited HBV replication transiently. In parallel, there was a decrease in the amount of HBV mRNA. Hepatitis B surface antigen and early antigen secretion were not influenced; however, their intracellular levels diminished during treatment. The cellular 2',5'-oligoadenylate synthetase activity was increased 9- to 18-fold during treatment of cells with IFN-gamma, -alpha, or -beta. The number of IFN-alpha and -beta receptors was down-regulated, while the number of IFN-gamma receptors remained constant. The expression of major histocompatibility complex class I antigens was stimulated by addition of IFN-alpha or -beta. These data show that both IFN-alpha and -beta can effectively inhibit HBV replication and induce a cellular IFN response in Hep G2.2.15 cells similar to that seen in humans.


An experimental model for dilated cardiomyopathy after rabbit coronavirus infection. A rabbit model for coronavirus-induced dilated cardiomyopathy is described. Acute rabbit coronavirus infection results in virus-induced myocarditis and congestive heart failure. Of the survivors of rabbit coronavirus infection, 41% had increased heart weight and heart weight-to-body weight ratios, biventricular dilation, myocyte hypertrophy, myocardial fibrosis, and myocarditis consistent with the development of dilated cardiomyopathy. These changes were also seen in the remaining 59% of the survivors, except that the degree of myocyte hypertrophy was reduced and only right ventricular dilation was present. In most survivors, myocarditis was usually mild (1-5 foci/transverse section), but in some cases it was severe (> 20 foci/transverse section). Interstitial and replacement fibrosis was more pronounced in the papillary muscles. These data suggest that rabbit coronavirus infection may progress to dilated cardiomyopathy.


Quantitative oropharyngeal Epstein-Barr virus shedding in renal and cardiac transplant recipients: relationship to immunosuppressive therapy, serologic responses, and the risk of posttransplant lymphoproliferative disorder. Epstein-Barr virus (EBV) infection plays a major role in the pathogenesis of posttransplant lymphoproliferative disorder (PTLD). Quantitative oropharyngeal EBV shedding measured by a DNA-DNA dot blot assay and the genotype of isolates determined by a polymerase chain reaction assay were studied in 23 renal and 23 cardiac transplant recipients followed over the first posttransplant year. Five patients developed PTLD and two additional PTLD renal transplant recipients were studied from the time of diagnosis. Significantly higher levels of EBV were observed in primary versus reactivation infection (P < .04) when sequential courses of antilymphocyte globulins or > 4 g of methylprednisolone were used in the first 6 months after transplant and in patients with versus those without PTLD (P < .04), although the former group had a high incidence of primary infection. Patients with the highest EBV shedding had the poorest serologic responses. All PTLD patients shed EBV-1, which was also shed by patients without PTLD.


Genetic variation and evolution of human parainfluenza virus type 1 hemagglutinin neuraminidase: analysis of 12 clinical isolates. The extent of genetic variation and evolution in a population of human parainfluenza virus type 1 was investigated. The hemagglutinin neuraminidase genes of 13 isolates collected over a 26-year period were sequenced and compared. All isolates except the 1957 type strain were from a single geographic location and demonstrated significant consistent genetic change from the type strain (47/7 [nucleotide/amino acid] substitutions). Antigenic subgroup A isolates demonstrated minor intragroup differences (9/1 substitutions). However, 18/7 unique substitutions separated subgroup A from B regardless of geographic location or year of isolation. Multiple strains of both subgroups appeared and reappeared over decades with only minor variation. There may be significant genetic differences between clinical isolates based on geographic location, and progressive mutational change may occur. Previously defined antigenic and now genetic subgroups were stable and at least regional in distribution over the period studied. The biologic implications and extent of this variation need further evaluation.


Composite grafting of the maxillary sinus for placement of endosteal implants. A preliminary report of 48 patients. The maxillary sinuses in 48 patients were grafted with dense, non-resorbable hydroxylapatite (HA) particles combined with autologous, cancellous bone. After 3 months of healing, HA-coated titanium endosteal implants were placed in the maxilla, and following an additional 3-5 month healing period, the dentitions were restored with various prostheses. Of the 267 maxillary implants placed, 18 (6.4%) failed. Thirteen (6.4%) of the 203 implants placed in the grafted floor of the sinus failed, and 5 (7.8%) of the 64 implants placed in the anterior maxilla failed. Simultaneous lateral and anterior onlay grafting of the alveolar process with the same composite graft material was required in 36 (75%) patients because the width of the alveolar process was considered insufficient for placement of endosseous implants. The mean follow-up period was 17 months (range 12-32). Results from this preliminary study indicate that composite grafting of the maxillary sinus with onlay grafting of the alveolar ridge will provide the bony structure necessary for placement of endosseous implants. Further follow-up of these patients is necessary to determine the long-term stability of this technique; however, these results are promising.


Juvenile angiofibroma of the maxillary sinus. A case report. The clinicopathological features of a rare case of juvenile angiofibroma originating from the maxillary sinus of a 13-year-old boy are reported. This tumor was composed of angiomatous and fibrous structures. The analysis of nucleolar organizer regions (NOR) of the various components of this tumor indicated that the fundamental elements were the fibroblastic cells, rather than the vascular endothelial cells.


Restoration of the lateral sinus wall using a collagen type I membrane for guided tissue regeneration. A collagen type I membrane prepared from bovine pericard was used to restore and thicken the lateral sinus wall after resection in 6 Göttingen minipigs. One rehydrated collagen type I membrane was placed on the intact sinus mucosa. A hydroxyapatite block was placed on this membrane and covered with a second membrane. After 6 months there was complete, bony regeneration in the space between the 2 membranes and thickening of the new bone plate compared to the untreated control side. This study shows that a collagen membrane can be applied for guided tissue regeneration.


[Influence of new quinolones on superoxide anion generation of polymorphonuclear leukocytes]. We investigated the influence of the new quinolones for the ability to kill on polymorphonuclear leukocytes (PMNs) in phosphate buffer solution (PBS) and in the urine in order to clarify the in vivo effects of the new quinolones. Four new quinolones, i.e., norfloxacin, enoxacin, ofloxacin and ciprofloxacin were employed. PMNs derived from peripheral blood in PBS and in hypotonic urine were stimulated with phorbol myristate acetate or opsonized zymosan, and superoxide anion generation of PMNs was measured by means of the chemiluminescence method. The superoxide anion generation of PMNs was significantly enhanced in the presence of 10 micrograms/ml of all new quinolones and significantly suppressed in the presence of 100 micrograms/ml of all new quinolones in PBS and in hypotonic urine. From these results, it was suggested that no influence of new quinolones on the ability to kill the PMNs might be seen in human blood, however, a great influence might be expected in the urine in the human urinary tract.


[Studies on hepatitis C virus infection in haemodialysis patients]. To examine the prevalence of hepatitis C virus (HCV) in haemodialysis patients without blood transfusion in Hiroshima Prefecture, antibody to HCV (anti-HCV) was studied by the Ortho ELISA Kit in sera from 393 consecutive haemodialysis patients and in sera from 510 age and sex matched healthy members of the general population (control). An additional confirmatory test was done by a recombinant immunoblot assay. 1) Anti-HCV was detected in 70 of the 393 dialysis patients and 3 of the 510 healthy controls (17.8% vs 0.6%, p less than 0.01). Prevalence of anti-HCV in haemodialysis patients sera was increased by the volume of blood transfusion, and even in dialysis patients who had no blood transfusion, the frequency of anti-HCV positivity (9.2%) was greater than the healthy controls (p less than 0.01). Thus, the major route of HCV transmission in haemodialysis patients without blood transfusion may be via the haemodialysis treatment. 2) The prevalence of anti-HCV increased significantly with the ALT level and abnormal ALT activity of the anti-HCV positive group were significantly greater than that of the negative group. Thus, it is suggested that HCV infection may be an etiologic factor of liver dysfunction in haemodialysis patients.


[Detection of human papillomavirus DNA in cases of sexually transmitted diseases (STD)]. Human papillomavirus (HPV) has been detected on the genitalia without any macroscopic abnormality and the possibility of latent infection of HPV has been suggested. Using Vira Type (Toure Co.), we have detected 7 genotypes of HPV DNA under a high stringent condition on the genitalia of patients with sexually transmitted diseases (STD), who were suspected of having had many sexual partners. In male cases of STDs other than condyloma acuminatum, the HPV-positive rate of the glans and sulcus coronarius was 4.7% (5/106). In healthy men, the HPV-positive rate was 6.1% (2/33), while in chronic prostatitis cases it was 3.4% (7/205) and in benign prostatic hypertrophy cases HPV was not detected. In female cases of STDs other than condyloma acuminatum, the HPV-positive rate of uterine cervix was 5.1% (3/58). In pregnant women, the HPV-positive rate was 4.6% (9/197). With regard to the HPV-positive rate within different age groups of STD and non STD cases, the rate tended to be higher in young people. After several weeks, follow-up studies were conducted on HPV-positive cases. HPV DNA was detected in one case of 10 males STD cases and two of 10 pregnant women, and the HPV DNA was the same type as at the first examination. However, after 3-4 months, all three of these cases had become negative for HPV DNA.


Prevalence of glioblastoma multiforme subjects with prior herbicide exposure. The charts of 100 patients with established diagnoses of glioblastoma multiforme (GBM) provided the data base for a descriptive study of patients' exposure to herbicides. The study focused on place of residence and occupation during the year prior to diagnosis of GBM. Although subjects reported residences in 33 of the 75 counties in Arkansas, more than one-third of the sample came from just 3 counties in which rice, cotton or wood products are produced. These industries were reported as the occupations of almost one-third of the sample for whom occupations involved a risk of herbicide exposure. The findings of this study add to the epidemiological profile of those at risk for GBM and underscore the need for assessment of residence and occupation on a consistent basis when counseling patients and providing health education.


[Continuous distribution of ventilation-perfusion ratios following lung denervation in dogs by means of inert gas elimination method]. Pulmonary branches of the vagus nerve and sympathetic nerve-chain are severed frequently in thoracic surgery. In this investigation, respiratory function, this transection influence on, was studied, experimentally. Then, denervation of the left lung carried out in mongrel dogs. And distribution of ventilation-perfusion ratio actually present in the lung was measured by multiple inert gases elimination method. In dogs with denervated one side lungs, imbalance of distribution of ventilation-perfusion ratio existed, and it mean that there were areas of high VA/Q in distribution of ventilation. However, there was still no shunt and distribution of blood flow was no dispersion. From the it seems reasonable to infer that blood flow in pulmonary vessel diminished by pulmonary denervation. But arterial blood gas analysis showed normal degree. Accordingly pulmonary gas exchange should not be disturbed to high degree by imbalance of VA/Q distribution due to the denervation.