Interleukin 12 p40 production by barrier epithelial cells during airway inflammation.

Human airway epithelial cells appear specially programmed for expression of immune response genes implicated in immunity and inflammation. To better determine how this epithelial system operates in vivo, we analyzed its behavior in mouse models that allow for in vitro versus in vivo comparison and genetic modification. Initial comparisons indicated that tumor necrosis factor alpha induction of epithelial intercellular adhesion molecule 1 required sequential induction of interleukin (IL)-12 (p70) and interferon gamma, and unexpectedly localized IL-12 production to airway epithelial cells. Epithelial IL-12 was also inducible during paramyxoviral bronchitis, but in this case, initial IL-12 p70 expression was followed by 75-fold greater expression of IL-12 p40 (as monomer and homodimer). Induction of IL-12 p40 was even further increased in IL-12 p35-deficient mice, and in this case, was associated with increased mortality and epithelial macrophage accumulation. The results placed epithelial cell overgeneration of IL-12 p40 as a key intermediate for virus-inducible inflammation and a candidate for epithelial immune response genes that are abnormally programmed in inflammatory disease. This possibility was further supported when we observed IL-12 p40 overexpression selectively in airway epithelial cells in subjects with asthma and concomitant increases in airway levels of IL-12 p40 (as homodimer) and airway macrophages. Taken together, these results suggest a novel role for epithelial-derived IL-12 p40 in modifying the level of airway inflammation during mucosal defense and disease.

Identification and characterization of aldose reductase in cultured rat mesangial cells.

Although the enhanced activity of the polyol pathway has been detected in diabetic glomeruli, the intraglomerular localization of this pathway has not yet been well defined. In this study, we attempted to identify aldose reductase, a key enzyme of the polyol pathway, in cultured rat mesangial cells and to characterize the properties of this enzyme using enzymological and immunological methods. When the aldose reductase (DL-glyceraldehyde-reducing) activity was analyzed in mesangial cell extract, the Lineweaver-Burk plot showed concave downward curvature, and the Michaelis constant was 0.83 mM DL-glyceraldehyde, and this activity was noncompetitively inhibited by an aldose reductase inhibitor, ICI-128,436. The enzyme activity was enhanced by the addition of sulfate ion and partially suppressed by barbital. The enzyme cross-reacted with the antisera against rat lens and testis aldose reductases on Ouchterlony plate, and migrated to the region of molecular weight of about 36,500 Da on Western blotting. The presence of aldose reductase mRNA was also confirmed by Northern analysis using cDNA for rat aldose reductase, 10Q. From these results, it was concluded that the aldose reductase may exist in rat glomerular mesangial cells and may play a role in the development of diabetic glomerulopathy, though the coexistence of aldehyde reductase(s) may not be fully ruled out.

Role of sympathetic activity in blood pressure reduction with low calorie regimen.

To investigate the effects of a low calorie regimen on sympathetic function and its relation to blood pressure response, 22 untreated obese essential hypertensive patients (50 +/- 2 years, body mass index 29 +/- 1 kg/m2) were hospitalized and a diet was prescribed of 2,000 kcal/day for 5 days (control period) followed by 800 kcal/day for 21 days without changing salt intake (8-10 g/day). The dose of intravenous phenylephrine infusion needed to elevate systolic blood pressure 20 mm Hg (CD20) and the 24-hour urinary excretion of norepinephrine (UNE) were measured. During the low calorie period, blood pressure normalized in 14 patients (responder group, 124 +/- 3/79 +/- 4 mm Hg) and eight remained hypertensive (poor responder group, 158 +/- 6/103 +/- 3 mm Hg). At the control period, blood pressure and body mass index were similar, but the responder group had higher UNE (134 +/- 15 micrograms/day) and CD20 (127 +/- 11 micrograms) than the poor responder group (89 +/- 6 micrograms/day and 79 +/- 13 micrograms, respectively). During the low calorie period, both UNE (87 +/- 15 micrograms/day) and CD20 (74 +/- 10 micrograms) decreased in the responder group; no change was seen in the poor responder group. Changes in UNE and systolic blood pressure were correlated (r = 0.6, p less than 0.05). In conclusion, suppression of sympathetic activity plays a role in blood pressure reduction during moderate caloric restriction.

Responses of sodium balance, blood pressure, and other variables to sodium loading in Papua New Guinea highlanders.

For determination of the responses of sodium balance, blood pressure, and other relevant variables to Na loading in people with a low intake of Na, 10 male Papua New Guinea highland subjects were given additional Na at two levels (128 and 256 mmol/d) for 10 d after a 3-d control period of low-Na diet. Na loading caused a marked positive balance of Na, decreases of aldosterone concentration and renin activity in the plasma, and a decrease of urinary aldosterone excretion. The blood pressure, particularly that measured at noon, increased in the latter half of the Na-loading period, the increase being significant in the group given 256 mmol of sodium daily: the systolic and diastolic blood pressure increased from 92 +/- 8 over 56 +/- 7 mm Hg in the control period to 102 +/- 7 over 60 +/- 4 mm Hg in the latter half of the test period (p less than 0.05).

Prospective studies on left ventricular geometric patterns and exercise tolerance in unmedicated men with borderline and mild hypertension.

OBJECTIVE: This study was designed and conducted to assess the clinical significance of left ventricular geometric patterns and physical fitness in subjects with untreated borderline and mild hypertension. METHODS: Symptom-limited maximal treadmill stress testings and echocardiographic examinations were administered to 192 previously unmedicated men. Left ventricular geometric patterns were determined by the combined criteria of left ventricular mass index and relative wall thickness. Subjects whose left ventricular mass index was < 125 g/m2 were followed up for more than 3 years. RESULTS: Normalized treadmill time was lower and pressure rate products at peak exercise were higher in patients with concentric hypertrophy than in those with normal geometry. Of the 77 patients who revealed left ventricular mass index at baseline < 125 g/m2 and who were successfully followed without medication for more than 3 years, 18 demonstrated concentric hypertrophy at the final follow-up examination. During the follow-up period, these 18 patients had significant further augmentation of concentric geometric features, significant decreases in both cardiac output and normalized treadmill time, and significant increases in casual blood pressure and total peripheral resistance compared with those at baseline. CONCLUSION: Patients with concentric hypertrophy exhibited slightly but significantly impaired levels of physical fitness and cardiac work efficiency, and the progression of concentric hypertrophy demonstrated further impairments of these conditions. Therefore, not only lowering blood pressure, but also improving left ventricular hypertrophy, cardiovascular hemodynamics, and physical fitness might be required in patients with concentric hypertrophy.

The relationship of hyperinsulinemic state to left ventricular hypertrophy, microalbuminuria, and physical fitness in borderline and mild hypertension.

The relationship of the hyperinsulinemic state to left ventricular hypertrophy, left ventricular geometric patterns, microalbuminuria, and physical fitness were studied in 52 middle-aged unmedicated men with borderline and mild hypertension. Left ventricular mass index and relative wall thickness were assessed by echocardiography. Physical fitness was determined by symptom-limited maximal treadmill stress testings. The urinary concentration of microalbumin and C-peptide was measured in 24-h urine samples by radioimmunoassey. The 24-h urinary C-peptide excretion rate was correlated with left ventricular mass index (r = 0.46), relative wall thickness (r = 0.41), treadmill time (r = -0.35), normalized treadmill time (r = -0.52), systolic blood pressure at peak exercise (r = 0.29), and 24-h urinary microalbumin excretion (r = 0.48). Stepwise multiple regression analysis identified the left ventricular mass index, the 24-h urinary albumin excretion, and the normalized treadmill time as variables in the equation for the 24-h urinary C-peptide excretion. Thus, the hyperinsulinemic state is related to left ventricular hypertrophy, microalbuminuria, and impaired physical fitness in patients with borderline and mild hypertension.

The efficacy of monatepil, a new calcium antagonist, in the treatment of essential hypertension.

A multicenter, open-label trial in Japan examined the efficacy, safety, and optimal dose of monatepil (AJ-2615) as monotherapy and in combination therapy with angiotensin-converting enzyme (ACE) inhibitors or beta-blockers. Patients with essential hypertension who had never been treated or had been refractory to conventional antihypertensive agents were enrolled in the trial. During a 4-week control period patients assigned to monotherapy received placebo and those assigned to combination therapy received an ACE inhibitor or beta-blocker and placebo. Patients with systolic blood pressure (BP) > or = 160 mm Hg and diastolic BP > or = 95 mm Hg at the end of the control period were enrolled in the study. The initial dose of monatepil was 30 mg/day in monotherapy and 15 mg/day in combination therapy; the daily dose was titrated to 60 mg/day according to the antihypertensive response. The treatment period was 8 to 12 weeks. Blood pressure decreased from 168 +/- 8/100 +/- 6 to 142 +/- 9/85 +/- 7 mm Hg (SD) with monatepil monotherapy, from 171 +/- 11/102 +/- 6 to 141 +/- 9/84 +/- 6 mm Hg in combination with ACE inhibitors, and from 175 +/- 13/102 +/- 7 to 153 +/- 21/91 +/- 9 mm Hg in combination with beta-blockers (P < .001). When patients in whom mean BP decreased by > or = 13 mm Hg were defined as responders, the response rate was 80.4%, 78.1%, and 51.6% in the respective groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Left ventricular geometric patterns and QT dispersion in borderline and mild hypertension: their evolution and regression.

To investigate whether QT dispersion increases in borderline and mild hypertension during a longitudinal observation of > 3 years and whether it is improved with medications, left ventricular geometric patterns and QT dispersion were studied with special regard to their longitudinal changes in 85 male borderline and mild hypertensive subjects with left ventricular mass index < 125 g/m2. These subjects were followed for > 3 years without medication. Thirty-two patients with a left ventricular mass index > 125 g/m2 at the end of follow-up period were further observed using antihypertensive drugs for an additional 3 years. Echocardiograms and electrocardiograms were obtained at the beginning and end of the follow-up period. At the end of the follow-up period, subjects were classified into four groups based on ventricular geometric patterns determined by left ventricular mass index and relative wall thickness in diastole. The QT dispersion was greater in patients with concentric hypertrophy (56+/-18 msec) than in patients with normal geometry (41+/-17 msec) (P < .05) and increased significantly in the former group during the follow-up period. After medication, the left ventricular mass index regressed and the QT dispersion decreased (from 55+/-21 to 50+/-26 msec, P < .01) in these patients. Thus, these findings suggest that changes in the QT dispersion reflect both concentric evolution and regression of left ventricular hypertrophy.

Metabolic actions of insulin-like growth factor I in cultured glomerular mesangial cells.

Glomerular mesangial cells in culture have been reported to possess a considerable number of receptors specific to insulin-like growth factor I (IGF-I), with very small number of receptors specific to insulin. To explore acute metabolic effects of IGF-I on mesangial cells, uptake of glucose and amino acid was measured in the presence of IGF-I or insulin. IGF-I stimulated D-[U-14C]glucose incorporation, 2-deoxy[1-3H]glucose uptake and alpha-[methyl-3H]aminoisobutyric acid (AIB) uptake into cultured mesangial cells by 139.8% +/- 2.1%, 116.6% +/- 1.7%, and 214.9% +/- 12.8% (percent of basal), respectively. Similar maximal stimulation was also induced by insulin, while the ED50 of IGF-I to stimulate these uptake systems (9.98 +/- 2.36, 3.45 +/- 1.86, and 3.35 +/- 0.40 ng/mL, respectively) was significantly lower than that of insulin (120.8 +/- 28.5, 61.8 +/- 7.7, and 76.3 +/- 17.5, respectively). These results indicate that, in cultured glomerular mesangial cells, IGF-I induces acute metabolic effects, possibly through its own receptors.

Effects of long-term antihypertensive therapy on physical fitness of men with mild hypertension.

This study was conducted to investigate the effects of long-term administration of a calcium-channel antagonist (nifedipine) and a beta-blocker (acebutolol) on physical fitness in men with mild hypertension. All subjects underwent symptom-limited treadmill stress testing and routine echocardiographic studies. Twenty-two subjects who had either a causal diastolic blood pressure of more than 105 mmHg or a left ventricular mass index (LVMI) of 125 g/m2 or more during follow-up were assigned to receive medical therapy. The other 31 men who did not meet either criterion were continuously followed-up without medication. Among the 22 treated men, the age-adjusted treadmill time (normalized treadmill time, TMTn) significantly decreased before the initiation of medication, while 31 untreated men showed no change in TMTn throughout the study. The 22 treated subjects were subsequently divided into two groups; 13 were given nifedipine and 9 were given acebutolol. All treated subjects were followed-up for more than 3 years. After treatment, the two groups showed similar reductions in blood pressure and LVMI, but a different outcome for TMTn: TMTn increased from 104 +/- 8% to 115 +/- 16% in subjects given nifedipine (p < 0.05) and decreased from 106 +/- 12% to 99 +/- 10% (p < 0.01) in those given acebutolol. Thus, the physical fitness of subjects who required medication significantly deteriorated without medication; their physical fitness improved after treatment with a calcium-channel antagonist and deteriorated after treatment with a beta-blocker.

Neuron-specific expression of reporter gene in transgenic mice carrying the 5'-upstream region of mouse P/Q-type Ca2+ channel alpha 1A subunit gene fused to E. coli lacZ reporter gene.

To dissect the molecular mechanisms underlying the neuron-specific expression of the P/Q type calcium channel alpha 1A subunit gene, transgenic mice carrying a 0.5-kb, 1.5-kb, 3.0-kb or 6.3-kb 5'-upstream region of the gene fused to Escherichia coli lacZ reporter gene were produced. In transgenic mice carrying the 1.5-kb, 3.0-kb or 6.3-kb 5'-upstream region, the reporter gene was exclusively expressed in the nervous system, although those with the 0.5-kb 5'-upstream region failed to show reporter expression. Histological examinations showed that the three 5'-upstream regions induced distinct expression patterns of the reporter gene in the CNS and adrenal medulla. The 1.5-kb 5'-upstream region drove reporter gene expression in the olfactory bulb, dorsal cortex and hippocampus, while the regulatory element for the expression in the amygdaloid nucleus, septum, habenula medial nucleus, choroid plexus, substantia nigra, inferior colliculus, pontine nucleus and cerebellum was located in the 5'-upstream sequence between 1.5 kb and 6.3 kb. In the cerebellum, the expression of the reporter gene was induced by the 3.0-kb region in granule cells, whereas it was induced by the 6.3-kb region in Purkinje cells. The expression of the reporter gene in chromaffin cells in the adrenal medulla was induced only by the 6.3-kb 5'-upstream region. These results suggest that the expression of the mouse P/Q-type Ca2+ channel alpha 1A subunit gene is regulated in a complex fashion by both positive and negative cis-regulatory elements.

Nipradilol inhibits rat mesangial cell mitogenesis through the activation of soluble guanylate cyclase.

Nipradilol is a beta-adrenoreceptor blocking agent, whose structure contains an NO2 group. Thus, it is possible that it modulates the function of glomerular mesangial cells through the activation of soluble guanylate cyclase. To prove this hypothesis, we examined the effect of nipradilol on soluble guanylate cyclase, intracellular cyclic guanosine monophosphate (cGMP) accumulation, and the mitogenesis of cultured rat glomerular mesangial cells. Nipradilol increased intracellular cGMP accumulation in a dose-dependent manner through the activation of soluble guanylate cyclase. Furthermore, nipradilol inhibited the incorporation of [3H]thymidine into the mesangial cells stimulated by 2.5% fetal bovine serum in a dose-dependent manner. These results indicate that nipradilol may modulate mesangial cell function through an increase in intracellular cGMP resulting from the activation of soluble guanylate cyclase.

Expression analysis of the 5'-upstream region of mouse P/Q-type Ca(2+) channel alpha( lA) subunit gene fused to Escherichia coli lacZ reporter gene in the spinal cord using transgenic mice.

The P/Q-type Ca(2+) channel alpha(1A) subunit is expressed in spinal cord including ventral motor neurons and interneurons and dorsal horn. To identify the transcriptional mechanisms of the mouse alpha(IA) subunit gene in spinal cord, transgenic mice carrying a 0.5, 1.5, 3.0 or 6.3-kb 5'-upstream region fused to the Escherichia coli lacZ reporter gene were examined. Transgenic mice carrying the 3.0-kb region expressed the reporter gene in dorsal horn and interneurons of ventral horn, although those with the 0.5-kb, 1.5-kb or 6.3-kb region did not. No transgenic mice expressed the reporter gene in motor neurons of ventral horn. These results suggest that in spinal cord, the expression mechanisms of the alpha(1A) subunit gene are complex, involving both positive and negative cis-regulatory elements, and the 6.3-kb 5'-upstream region alone is not sufficient for the expression.

Expression analysis of Escherichia coli lacZ reporter gene in transgenic mice.

To define a gene expression mechanism, it is often advantageous to use a reporter gene and transgenic mouse. The lacZ reporter gene is particularly useful for studies of the cis-regulatory element for tissue-specific expression in transgenic mice because of the ease of the enzyme assay and visualization on sections. In this report, we describe our method for examining the cis-regulatory element in transgenic mice, including choice of the lacZ gene, generation of transgenic mice, and analysis of beta-galactosidase activity.

High blood pressure is a risk factor for the development of microalbuminuria in Japanese subjects with non-insulin-dependent diabetes mellitus.

In this study, 52 nonproteinuric Japanese patients with non-insulin-dependent diabetes (NIDDM) were followed from 1985 to 1990 to investigate the rate of development and progression of microalbuminuria and the factors which influence it. In 1985, 34 patients were normoalbuminuric, and 18 patients were microalbuminuric. Five years later, 11 of 34 initially normoalbuminuric patients (32.4%) developed microalbuminuria, and 6 of 18 initially microalbuminuric patients (33.3%) developed overt proteinuria. At the beginning of the study, hypertension existed more frequently in the patients who later developed microalbuminuria (8 of 11, 72.7%) than in the patients who stayed normoalbuminuric (4 of 23, 17.4%). Age-adjusted values of mean blood pressure (+/- SEM) at the beginning of the study in the patients who developed microalbuminuria (98.2 +/- 3.4 mm Hg, n = 11) were significantly higher than those in the patients who stayed normoalbuminuric (87.3 +/- 2.4 mm Hg, n = 23). In six patients who developed overt proteinuria, initial urinary albumin excretion rates (AER) were higher than those in the patients who stayed microalbuminuric, and four patients who presented with initial AER greater than 100 micrograms/min all developed overt proteinuria. These results indicate that, in Japanese patients with NIDDM, the rate of development of microalbuminuria is faster than that reported in Caucasian IDDM, and preexisting hypertension with relatively poor control of blood pressure may be a risk factor for the development of microalbuminuria.

Effect of dietary protein levels on urea utilization in Papua New Guinea highlanders.

Ability to utilize urea nitrogen for body protein synthesis was examined with Papua New Guinea (PNG) highlanders and Japanese (JPN). Eight male PNG highlanders and 8 male JPN were fed on a low protein diet containing 0.55 g protein/kg or an adequate protein diet containing 1.34 g protein/kg for 1 or 2 weeks. The fate of 15N was measured after oral administration of 15N-labelled urea. There was no difference in 15N incorporation into serum protein between PNG highlanders and JPN receiving low protein diets. On the other hand, on the adequate protein diet, 15N incorporation in PNG highlanders was similar to that on the low protein diet, in contrast to that in JPN which was hardly detected in the adequate protein diet. When PNG highlanders take more protein than protein in their usual diet, they effectively incorporate ingested protein into their body protein and urea nitrogen is utilized for synthesis of body protein.

Organochlorine pesticides and polychlorinated biphenyls in California sea lions.

Concentrations of polychlorinated biphenyls (PCBs), DDTs, chlordanes, HCHs, hexachlorobenzene (HCB), dieldrin, heptachlor epoxide, tris(4-chlorophenyl)methane (TCPMe), and tris(4-chlorophenyl)methanol (TCPMOH) were measured in the blubber of California sea lions (Zalophus californianus) collected in 2000. DDTs were the most predominant contaminants, followed by PCBs, chlordanes, TCPMe, HCHs, TCPMOH, dieldrin, and heptachlor epoxide. Concentrations of PCBs and DDTs varied from a few microg/g to several hundreds of microg/g on a lipid weight basis. Concentrations of DDTs have declined by an order of magnitude over the last three decades in California sea lions; nevertheless, the measured concentrations of PCBs and DDTs in California sea lions are still some of the highest values reported for marine mammals in recent years. Concentrations of organochlorines were highly correlated with one another. Concentrations of PCBs and DDTs in the blubber of gray whale, humpback whale, northern elephant seal, and harbor seal, and in the adipose fat of sea otter, were lower than the levels found in California sea lions, and were in the range of a few to several microg/g on a lipid weight basis.

Organochlorine and organotin compounds in Caspian seals (Phoca caspica) collected during an unusual mortality event in the Caspian Sea in 2000.

Polychlorinated biphenyls (PCBs), organochlorine pesticides and organotin compounds were determined in the blubber and liver of Caspian seals (Phoca caspica) found stranded on the coast of the Caspian Sea during an outbreak of canine distemper virus (CDV) in 2000. Among organochlorines analyzed, DDTs were the most dominant contaminants with concentrations ranging from 6.3 to 470 microg/g on a lipid-weight basis. Caspian seals collected in 2000 during the epizootic had higher concentrations of organochlorines than healthy individuals sampled in 1998. However, the blubber layer was generally thinner in the seals collected in 2000 than those in the previous surveys. Although compositions of organochlorine pesticides in seals suggested that the contamination status in the Caspian Sea is improving, the levels found in Caspian seals in 2000 were comparable to those in other marine mammals that have suffered from epizootics. This implies that the present status of contamination found in Caspian seals poses a risk of immunosuppression. Concentrations of butyltin compounds in livers of seals ranged from 0.49 to 17 ng/g on a wet-weight basis and octyltin compounds were below limit of detection in all the samples analyzed, suggesting less contamination by organotin compounds in the Caspian Sea.

Neurotropism of mouse-adapted haemagglutinating encephalomyelitis virus.

The propagation of a mouse-adapted strain (67N) of haemagglutinating encephalomyelitis virus in infected mice and murine cells was examined by viral re-isolation and immunostaining. Viral propagation was strictly limited to the neurons and to an established line of neuroblastoma cells in in-vivo and in-vitro experiments. These results provide adequate evidence that this virus is neurotropic.

Vertical transmission to embryo and fetus in maternal infection with rat virus (RV).

The influence of maternal rat virus (RV) infection on rat embryogenesis and fetus was examined by viral reisolation, immunostaining and PCR analysis. Vertical transmission caused by the UT-1 strain of RV depended on the stage of gestation when maternal infection occurred. When females were infected at the pre-mating point, the number of fetuses was smaller than that normally obtained, possibly due to infection at the stage of the hatched blastocyst, but almost all of the fetuses obtained were free from infection and developed normally. The incidence of transplacental infection was the highest when pregnant females were infected in the middle of the gestation stage, and some of the fetuses died. In pregnant females which were infected late in the gestation stage, all fetuses developed normally. Some of them were infected transplacentally and harbored the infectious virus. Much attention should be paid to performing reliable rederivation of RV-infected rat colonies by hysterectomy and embryo transfer.