Protective Effects of Nafamostat Mesilate on Liver Injury Induced by Lipopolysaccharide in Rats: Possible Involvement of CD14 and TLR-4 Downregulation on Kupffer Cells

Nafamostat mesilate (NM) is a synthetic protease inhibitor with various biological effects. To determine its effect on liver injury related to sepsis, we investigated the effects of NM on lipopolysaccharide (LPS)-induced liver injury. Wistar rats were allocated into two groups; the NM group underwent intraperitoneal NM administration 30 min before LPS administration, and the control group underwent PBS administration. Serum AST and ALT levels were significantly decreased in NM-treated rats. Reduced levels of TNF-α, IL-1β, and IFN-γ were observed after LPS administration in NM-treated rats. No significant differences were observed in IL-6 levels between the NM and the control group. In contrast, HGF levels were significantly increased only in control rats. NM treatment decreased protein and mRNA levels of TLR-4 and CD14. Our data suggest that NM treatment has protective effects against LPS-induced hepatotoxicity through downregulation of TLR4 and CD14 in liver, which decreased TNF-α, IL-1β, and IFN-γproduction in liver.     

LPS-stimulated production of TNF-α by peripheral blood monocytes in patients with Behcet’s Disease

Tumor necrosis factor alpha (TNF-α) is believed to play a significant role in disease pathogenesis of Behcet’s disease (BD). High serum levels of TNF-α were repeatedly reported in patients with active BD and anti-TNF agents are effective in its treatment. The pathophysiology of TNF-α in BD is still unknown and conflicting results regarding TNF-α overproduction by peripheral blood monocytes (PBM) from patients with BD were reported. The aim of the study is to compare stimulated production of TNF-α by PBM of BD patients with that of healthy volunteers (HV) and to examine correlations between the ability of PBM to produce TNF-α and organ/system involvement in patients with BD. Eighteen patients with BD (mean age 38.4±12.4 years, 12 males) gave informed consent and completed the European BD Current Activity Form. The PBM were separated and treated with lipopolysaccharide (LPS) overnight. TNF-α levels in the supernatants were assayed by ELISA and values were expressed in terms of cell protein contents. The control group included 15 HV (mean age 34.2±9.9 years, seven males). The mean production of TNF-α/cell protein (ng/mg) and in-group dispersion were similar in both groups (p=0.98). In the subgroup analysis, TNF-α production by PBM in BD patients who reported “bad” or “very bad” global well-being over the last month (n=4) was higher compared to other patients with better self-rating (p=0.03). PBM of BD patients in the present study did not overproduce TNF-α upon stimulation with LPS. However, BD patients with a higher TNF-α-producing capacity had worse sense of well-being.     

Inhibitory effect of troglitazone on diabetic neuropathy in streptozotocin-induced diabetic rats

Summary Free-radical scavengers and inhibitors of tumour necrosis factor-α (TNF-α) such as N-acetylcysteine and pentoxifylline have been shown to inhibit the development of peripheral neuropathy in streptozotocin(STZ)-induced diabetic rats. In this study we examined the effect of troglitazone, an anti-diabetic thiazolidinedione, on diabetic neuropathy, since it also is a free-radical scavenger and a TNF-α inhibitor. Rats were fed powder chow mixed with troglitazone at 0.5 % and 0.125 % ad libitum. Although blood glucose concentrations were remarkably higher and body weight lower in diabetic than in nondiabetic rats, troglitazone had no effect on these throughout the 24-week experiment. Serum lipoperoxide concentrations, tibial nerve lipoperoxide content and serum TNF-α activity induced by lipopolysaccharide was increased in diabetic rats, but inhibited in troglitazone-treated rats. Motor nerve conduction velocity (MNCV) of the tibial nerve slowed in diabetic rats, compared with that in nondiabetic rats. On the other hand, the slowed MNCV was (p < 0.05–0.01) inhibited after weeks 12 and 16 of the experiment in diabetic rats treated with high and low doses of troglitazone, respectively. Morphometric analysis showed that troglitazone suppressed the decrease of the myelinated fibre area (p < 0.05), axon/myelin ratio (p < 0.01) and fascicular area (p < 0.05) and suppressed the increase of myelinated fibre density (p < 0.001) in diabetic rats. These results indicate that troglitazone has a beneficial effect on peripheral neuropathy in STZ-induced diabetic rats irrespective of blood glucose concentrations. [Diabetologia (1998) 41: 1321–1326] Received: 16 March 1998 and in revised form: 8 June 1998     

AT<Subscript><Emphasis Type="Italic">1</Emphasis></Subscript> Receptor Blockade Prevents Cardiac Dysfunction after Myocardial Infarction in Rats

Summary Myocardial infarction (MI) can induce severe alterations of contractile function that can, in turn, lead to heart failure. In a previous study, we have demonstrated that TNF-α was involved in cardiac contractile dysfunction 7 days after coronary artery ligation in rats. Since Angiotensin II type 1 (AT₁) receptor can be involved in TNF-α production, we have investigated whether early short-term treatment with irbesartan, an AT₁ receptor blocker, is able to limit TNF-α production within the heart and to improve cardiac function and geometry following MI in rats. Male Wistar rats were subjected to permanent coronary artery ligation and received either a placebo or irbesartan (50 mg/kg/day) per os daily from day 3 to day 6 after surgery. On day 7, cardiac TNF-α was significantly reduced in MI rats receiving irbesartan (p < 0.05). Moreover, irbesartan improved residual LV end-diastolic pressure under both basal conditions and after volume overload (p < 0.01). In addition, a significant leftward shift of the pressure-volume curve in the irbesartan-treated group was found versus placebo. Finally, infarct expansion index was also significantly improved by irbesartan (p < 0.01). In conclusion, early, short-term AT₁ receptor blockade limits post-infarct cardiac TNF-α production and diminishes myocardial alterations observed 7 days after MI in the rat.     

Melatonin Modulates the Severity of Taurocholate-Induced Acute Pancreatitis in the Rat

The aim of this study was to investigate the effects of melatonin on serum amylase, tumor necrosis factor-alpha (TNF-α) and histological changes in rats with taurocholate-induced acute pancreatitis. Thirty male Wistar rats were randomly divided into three groups; group 1, group 2 and group 3 were enrolled as melatonin, control and sham groups, respectively (n = 10 per group). Acute pancreatitis was induced by 1 ml/kg body weight using 5% taurocholate injection into the biliopancreatic duct in groups 1 and 2 after clamping the hepatic duct. Those in group 1 received 50 mg/kg body weight melatonin by intraperitoneal (i.p.) injection. Group 2 received physiological saline i.p. at the same dose. Group 3 solely underwent laparotomy with cannulation of the biliopancreatic duct. Twenty-four hours after the intervention, the rats were killed, and serum samples were collected to measure amylase and TNF-α levels. Simultaneously, pancreatic tissues were removed, stained with hematoxylin-eosin and examined under a light microscope. Serum amylase and TNF-α levels were significantly lower in the melatonin group compared to the controls (P < 0.001). The total histological score, including edema, inflammation, perivascular infiltrate, acinar necrosis, fat necrosis and hemorrhage, was also significantly lower in the melatonin group as compared to the control (P < 0.0001). In conclusion, melatonin is potentially capable of reducing pancreatic damage by decreasing serum TNF-α levels in taurocholate-induced acute pancreatitis in rats. This result supports the idea that melatonin might be beneficial in ameliorating the severity of acute pancreatitis.     

Effect of TNF-α inhibition on urinary albumin excretion in experimental diabetic rats

The objective is to assess the effect of TNF-α inhibition on urinary albumin excretion in experimental diabetic rats. Male Wistar rats, 8-week-old, were categorized into four groups, which were the control (n = 9), diabetes (n = 9), infliximab-treated diabetes (n = 10), and FR167653-treated diabetes (n = 9) groups. Diabetes was induced by intraperitoneal injection of STZ (40 mg/kg). Thereafter, infliximab was injected intraperitoneally once a month (5.5 mg/kg) and FR167653 was administered orally by mixing with the rat chow (0.08%). The effects of infliximab and FR167653 on urinary albumin excretion were observed for 12 weeks. Body weight, blood sugar, 24-h urinary TNF-α, and 24-h urinary albumin/creatinine ratio (Ualb/Ucr) levels were determined at 1, 4, 8, and 12 weeks after the STZ-injection. Treatment of rats with STZ caused a significant loss of body weight, as well as polyuria and hyperglycemia within 1 week, while the urinary excretions of albumin and TNF-α were increased. Neither infliximab nor FR167653 affected body weight or blood sugar levels, whereas both decreased urinary albumin excretion, together with a modest decrease in the urinary excretion of TNF-α. These results suggest a role of TNF-α in the pathogenesis of diabetic nephropathy and show that TNF-α inhibition is a potential therapeutic strategy.     

Cotton Smoke Inhalation Primes Alveolar Macrophages for Tumor Necrosis Factor-α Production and Suppresses Macrophage Antimicrobial Activities

The present study determined the effects of cotton smoke inhalation on the functioning of alveolar macrophages (mφ). Smoke inhalation led to dose-dependent impairment of respiratory gas exchange by 48 h postexposure and pulmonary edema by 96 h. Maximal effects were observed in animals ventilated with 54 breaths of cotton smoke (3-min exposure, 18 breaths/min). Macrophages were obtained at 48 h postexposure by bronchoalveolar lavage of rabbits subjected to 54 breaths of smoke or room air (control). Phagocytosis of opsonized bacteria and adherence to solid substratum were reduced in smoke-exposed mφ. Smoke inhalation primed mφ for release of tumor necrosis factor-α (TNF-α) induced by lipopolysaccharide (LPS). Smoke-exposed mφ were also primed for TNF-α release induced by phorbol myristate acetate, which suggests that the priming event occurred downstream of protein kinase C activation in the signal transduction pathway. Further, smoke exposure attenuated the inhibitory effects of phosphodiesterase inhibitors on LPS-induced TNF-α release. Thus, the priming event may be mediated through cAMP and/or protein kinase A. The data indicate that cotton smoke inhalation suppresses the antimicrobial activities of alveolar mφ and can lead to excessive mφ production of TNF-α. These mφ effects would be expected to contribute to the pathophysiological abnormalities associated with smoke inhalation injury. Accepted for publication: 19 February 1998     

Inhibitory effects of edaravone on the production of tumor necrosis factor-α in the isolated heart undergoing ischemia and reperfusion

We evaluated the effects of edaravone, a hydroxyl radical scavenging agent, on the production of tumor necrosis factor-α (TNF-α) in myocardium, and the release of TNF-α and P-selectin from myocardium after ischemia–reperfusion injury in isolated Langendorff-perfused rat hearts. Cardiodynamic function at stable points during perfusion and 5, 15, 30, and 60 min after the initiation of reperfusion was evaluated by left ventricular developed pressure, rate of increase in left ventricular pressure and rate of decrease in ventricular pressure, coronary flow, and heart rate. At 60 min after the initiation of reperfusion, myocardial infarct size was estimated microscopically using triphenyltetrazolium chloride staining, and expression of TNF-α in myocardium was detected by Western blot and immunohistochemistry. At the same time points as the measurement of cardiodynamic function, TNF-α and the soluble form of P-selectin in coronary effluent were measured by enzyme immunoassay. At all time points during reperfusion, edaravone markedly improved cardiodynamic function and reduced myocardial infarct size in comparison to the control. In myocardium in the control, TNF-α was detected in the endothelial cells and other cells bearing some resemblance to interstitial cells and monocyte cells. Edaravone suppressed this cytokine expression in the corresponding sites. P-selectin as well as TNF-α was found in the coronary effluent of the control, and edaravone significantly decreased soluble P-selectin levels in comparison to the control (P < 0.01). Edaravone might have protective effects on cardiac function through reduction of infarct size via decrease of production of TNF-α in myocardium induced by ischemia–reperfusion injury and through reduction of the release of adhesion molecules such as P-selectin from vascular endothelial cells.