Tissue Levels Of TNF-α Research Articles

Abstract 384: Tumor Necrosis Factor-alpha Antagonism Attenuates Hypertension and Renal Injury Induced by Chronic Blockade of Nitric Oxide and High Salt Intake in Mice

Earlier, we demonstrated that systemic blockade of nitric oxide (NO) generation by acute infusion of nitro-L-arginine methyl ester (L-NAME) in mice increases plasma and renal levels of tumor necrosis factor-alpha (TNF-α). In the present study, we examined the role of TNF-α in mediating hypertension and renal injury responses to chronic NO blockade by evaluating the responses to chronic administration of L-NAME (0.05 μg/g/min; by osmotic mini-pump) for 3 wks in mice (C57BL6) treated with or without TNF-α receptor antagonist, etanercept (ETN; 5 mg, ip, every alternate day). These mice were given either normal (NS; 0.03% NaCl) or high salt (HS; 4% NaCl) containing diets. Systemic blood pressure (BP) was measured by tail-cuff plethysmography. At the end of the experimental period, mice were sacrificed to collect plasma and renal tissue samples to measure TNF-α level by ELISA and to determine the extent of tissue injury such as glomerulosclerosis (GS; PAS staining) and renal interstitial fibrosis (RIF; Trichrome staining). L-NAME treatment caused increase in BP which was greater in HS (104±3 to 139±3 mmHg; n=6) than in NS group (96±6 to 124±6 mmHg; n=6). ETN treatment significantly attenuated this increase in BP in both L-NAME+HS (119±11 mmHg; n=6) and in L-NAME+NS (115±8 mmHg, n=6) groups. Renal tissue level of TNF-α was higher in L-NAME+NS (664±100 vs 325±73 pg/mg protein) and in L-NAME+HS (391±38 vs 114±17 pg/mg protein) groups compared to the levels in groups given NS (n=5) or HS (n=5) diet alone. However, the changes in plasma level of TNF-α due to chronic L-NAME treatment were minimal. ETN treatment lowered renal TNF-α level in L-NAME+NS (356±60 pg/mg protein) and in L-NAME+HS (259±65 pg/mg protein) groups. L-NAME treatment significantly increased GS (7.1±1.0 vs 17.9±2.3 % sclerosis area) and RIF, (1.5±0.1 vs 5.8±0.6 % fibrotic area) in HS group. However, these changes were minimal in NS group (GS, 10.3±1.0 vs 13.2±1.5 % and RIF, 1.3±0.1 vs 2.4±0.3 %). ETN treatment markedly attenuated the GS (11.8±0.4% and 11.9±0.6 %) and RIF (2.6±0.3% and 1.9±0.3%) both in L-NAME+HS and in L-NAME+NS groups. These data demonstrate that an enhancement in TNF-α activity is involved in mediating hypertension and renal injury responses to chronic NO blockade and HS intake.

The Role of Tumor Necrosis Factor-α in the Pathogenesis of Vitiligo

Vitiligo is an acquired immune disorder of the skin characterized by the presence of white depigmented macules. Its immunopathogenesis is not completely understood, but inflammatory alterations in the skin microenvironment, and particularly increased expression of the cytokine tumor necrosis factor-α (TNFα), are thought to be essential regulators of melanocyte dysfunction and death. In this article we review the evidence that implicates TNFα in the pathogenesis of vitiligo, including studies on serum and tissue levels of TNFα, TNFα gene polymorphisms, in vitro studies, and therapeutic trials using TNFα inhibitors. TNFα emerges as a complex mediator with apparently conflicting roles in vitiligo.

Gastric antral polyps: a manifestation of portal hypertensive gastropathy

Editor, Gastric mucosal lesions are common in portal hypertension, occur in up to 65 % of cirrhotics, and represent an important cause of gastrointestinal (GI) blood loss. They include portal hypertensive gastropathy (PHG) and gastric vascular ectasia (GVE) [1]. The diagnosis of PHG is based on the presence of a characteristic mosaic-like pattern of the gastric mucosa on endoscopic examination. Superimposed on this mosaiclike pattern may be red spot lesions that usually are greater than 2 mm in diameter [1]. Polypoidal lesions in portal gastropathy on endoscopy have been described in two studies [2, 3]. We present eight cases of cirrhosis and portal hypertension with severe PHG and antral polyps. We prospectively evaluated patients admitted to our department, with a diagnosis of cirrhosis, between January 2011 and March 2012 and performed upper GI endoscopy. Clinical, laboratory, endoscopy, and histology data of these patients were analyzed. During the study period, 786 patients with cirrhosis underwent upper GI endoscopy. Of these, eight (1.01 %, seven male) patients, ranging in age from 43 to 70 years, had portal hypertensive polyps. Seven patients had alcoholic cirrhosis, and one patient had hepatitis B-related cirrhosis, all of Child’s C class. The CTP score ranged from 8 to 13 and model for end-stage liver disease ranged from 9 to 21. None had hepatocellular carcinoma or portal vein thrombosis. Six patients had history of upper GI bleed and had undergone esophageal variceal ligation previously. Endoscopy in all eight patients showed multiple antral polyps two to five in number and with size ranging from 0.5 to 1 cm. All patients had associated severe PHG. Two of the six patients with upper GI bleed continued to have drop in hemoglobin and stool occult blood positivity despite eradication of varices, and one had an overt bleeding polyp on endoscopy shown in Fig. 1d. These patients underwent endoscopic polypectomy once the platelet count was corrected to more than 50,000/mm and prothrombin time was corrected with plasma. On follow up, none of the patients had a drop in hemoglobin or rebleed. Small-bowel evaluation was not done in any of these patients as they did not have a drop in hemoglobin after polypectomy. Biopsy of polyps showed elongation, tortuosity, and dilatation of gastric foveolae with the surrounding stroma showing edema, mild inflammation, and scattered smooth muscle bundles. Numerous proliferating capillaries were present in the subepithelial tissue differentiating these polyps from usual hyperplastic polyps. There were no areas of dysplasia. Rapid urease test was negative, and histology and special stains did not reveal Helicobacter pylori in any of the patients. Cirrhotics with portal hypertensive polyps did not differ from cirrhotics without portal hypertensive polyps with regard to etiology or severity of cirrhosis, duration of illness, or past history of variceal eradication. Gastroscopy in PHG shows a mosaic-like pattern of the gastric mucosa with mild and superimposed discrete cherry red spots, fine pink speckling, or scarlatina-type rash when severe [1]. Biopsies show vascular ectasia and congestion in the mucosal layer without a significant degree of inflammatory infiltrate [4, 5]. Perturbations in the tissue levels of TNF-α, prostaglandins, endothelin, and nitric oxide/peroxynitrite may participate in the vascular congestion and mucosal damage characteristic of PHG [4, 5]. Some patients with cirrhosis and portal hypertension have GVE [5]. The entity is characterized by aggregates of ectatic vessels that C. Panackel (*) :H. Joshy :B. Sebastian : S. K. Mathai Department of Gastroenterology, Medical Trust Hospital, Kochi 682 016, India e-mail: charlespanackel@hotmail.com

Preventive Effects of a Natural Anti-Inflammatory Agent, Astragaloside IV, on Ischemic Acute Kidney Injury in Rats

This study investigated the anti-inflammatory effects of astragaloside IV(AS-IV) on ischemia/reperfusion (IR) induced acute kidney injury (AKI) in rats. Experimental model of ischemic AKI was induced in rats by bilateral renal artery clamp for 45 min followed by reperfusion of 12 h and 24 h, respectively. AS-IV was orally administered once a day to rats at 10 and 20 mg·kg−1·d−1 for 7 days prior to ischemia. AS-IV pretreatment significantly decreased serum urea, creatinine, and cystatin C levels at 12 h and 24 h of reperfusion in AKI rats. AS-IV pretreatment also ameliorated tubular damage and suppressed the phosphorylation of p65 subunit of NF-κB in AKI rats. Moreover, NF-κB and MPO activity as well as serum and tissue levels of TNF-α, MCP-1, and ICAM-1 were elevated in AKI rats. All of these abnormalities were prevented by AS-IV. Furthermore, AS-IV downregulated the mRNA expression of NF-κB, TNF-α, MCP-1, and ICAM-1 in AKI rats. These results suggest that AS-IV might be developed as a novel therapeutic approach to prevent ischemic AKI through inhibition of NF-κB mediated inflammatory genes expression.

279 Antioxidant Effects of N-Acetylcysteine in a Neonatal Rat Model of Necrotizing Enterocolitis

Background and AimHypoxia and ischemia appear to play an important role in the pathogenesis of necrotizing enterocolitis (NEC) which is related to oxygen-derived free radical formation. This study was designed...

Differential effects of high-fat-diet rich in lard oil or soybean oil on osteopontin expression and inflammation of adipose tissue in diet-induced obese rats

To examine the effect of different dietary fat types on osteopontin (OPN) expressions and inflammation of adipose tissues in diet-induced obese rats. Male Sprague-Dawley rats were randomly assigned to one control group fed standard diet (LF, n = 10) and two high-fat diet groups fed isoenergy diet rich in lard or soybean oil (HL or HS, n = 45 each). Diet-induced obese rats in HL and HS group were then subdivided into two groups either continuously fed high-fat diet or switched to low-fat diet for 8 more weeks. Fasting serum glucose, insulin, and OPN concentrations were assayed and QUICKI was calculated; the expression of OPN, IL-6, IL-10, TNF-α, NF-κB, and F4/80 in adipose tissue was determined. Both high-fat diets lead to comparable development of obesity characterized by insulin resistance and adipose tissue inflammation. Obese rats continuously fed high-fat diet rich in lard oil exhibited the highest fasting serum insulin level and adipose tissue OPN, F4/80, TNF-α, and NF-κB expression level. In both high-fat diet groups, switching to low-fat diet resulted in less intra-abdominal fat mass, decreased expression of F4/80, TNF-α, and NF-κB, while decreased OPN expression was only observed in lard oil fed rats after switching to low-fat diet. Reducing diet fat or replacing lard oil with soybean oil in high-fat diet alleviates obesity-related inflammation and insulin resistance by attenuating the upregulation of OPN and macrophage infiltration into adipose tissue induced by high-fat diet.

Temporary rapid bowel ligation as a damage control adjunct improves survival in a hypothermic traumatic shock swine model with multiple bowel perforations

BackgroundPrimary intestinal anastomosis is not the right choice for multiple bowel perforations under hemodynamically stable conditions. Our group has employed temporary rapid bowel ligation as a damage control procedure in a hypothermic traumatic shock swine model with multiple bowel perforations and hypothesized that damage control treatment would improve survival in the setting of a damage control surgery. Materials and methodsThe abdomen was shot one time with an experimental modified gun while pigs were hemorrhaged to a mean arterial pressure of 40 mm Hg and maintained in shock for 40 min. Cold lactated Ringer solution was gradually infused to induce hypothermia. Animals were randomized to control (no resuscitation), primary anastomosis (PA), or temporary rapid bowel ligation (damage control group, DC). Animals were resuscitated for 12 h with the shed blood and lactated Ringer solution. Delayed anastomosis was performed in DC animals after resuscitation. Surviving animals were humanely killed 24 h after operation. Systemic hemodynamic parameters were recorded and blood samples were obtained for biochemical assays. The lung and ileum was harvested at the end of the experiment for pathologic evaluation and test of wet/dry weight ratio, TNF-α level, and nuclear factor-κB activations. ResultsAll animals suffered extreme physiologic conditions: hypothermia, severe acidosis, hypotension, and depressed cardiac output. Control animals suffered 100% mortality. Compared with the PA group, DC animals required less resuscitation fluid, normalized lactate levels faster, had lower serum creatine kinase, aspartate amino transferase levels and tissue TNF-α level and nuclear factor-κB activations, suffered less severe histopathology, had greater early survival. ConclusionsMultiple bowel perforations under hemodynamically stable conditions seem better managed with DC than with PA. Temporary rapid bowel ligation as a damage control adjunct is important to rapid control of multiple bowel perforations instead of a prolonged operative time.

Ischaemic postconditioning reduces serum and tubular TNF-α expression in ischaemic-reperfused kidney in healthy rats

We studied the protective effects of postconditioning (PS) in healthy and hypercholesterolemic rats after renal ischaemia-reperfusion (IR) injury. We aimed to examine cytokine expression and apoptosis in tissue damage after revascularisation (TNF-α levels in serum and tissue). Male Wistar rats (n = 32) were divided into four groups. The animals of normal feed groups (NF) were fed with normal rat chow and the cholesterol feed groups (CF) were fed with 1.5% cholesterol containing diet for 8 weeks. Anaesthetized rats underwent a 45-min cross-clamping in both kidney pedicles. Ischaemia was followed by 120-min reperfusion with or without PS protocol (group PS vs. IR). Postconditioning was induced by four intermittent periods of ischaemia-reperfusion of 15-s duration each. Serum cholesterol, triglyceride, urea and creatinine levels were determined. Proinflammation was characterized by the measurement of serum TNF-α. Tissue injury in kidney was determined by formaline-fixed, paraffin-embedded tissue sections. Tissue TNF-α levels were determined by immunohistochemistry. Significant elevation was observed in serum TNF-α level after IR injury in normal feed groups, which was reduced by PS. In CF group neither the elevation nor the postconditioning induced reduction were as significant as in the NF groups. In normal feed group PS caused a significant reduction in tissue TNF-α level which was significantly higher in CF. Ischaemic postconditioning proved to be an effective defense against IR in NF groups, but it was ineffective in CF groups in kidney tissue.

Effectiveness ofSaccharomyces boulardiiin a rat model of colitis

To investigate the effects of Saccharomyces boulardii (S. boulardii) in an experimental rat model of trinitrobenzene sulfonic acid (TNBS)-induced colitis. Thirty-two Wistar albino female rats were categorized into five groups. On the first day of the study, 50 mg TNBS was administered via a rectal catheter in order to induce colitis in all rats, except those in the control group. For 14 d, the rats were fed a standard diet, without the administration of any additional supplements to either the control or TNBS groups, in addition to 1 mg/kg per day S. boulardii to the S. boulardii group, 1 mg/kg per day methyl prednisolone (MP) to the MP group. The animals in the S. boulardii + MP group were coadministered these doses of S. boulardii and MP. During the study, weight loss, stool consistency, and the presence of obvious blood in the stool were evaluated, and the disease activity index (DAI) for colitis was recorded. The intestines were examined and colitis was macro- and microscopically scored. The serum and tissue levels of tumor necrosis factor-α (TNF-α) and nitric oxide (NO) were determined, and fungemia was evaluated in the blood samples. The mean DAI scores for the MP and S. boulardii + MP groups was significantly lower than the TNBS group (3.69 ± 0.61 vs 4.46 ± 0.34, P = 0.018 and 3.77 ± 0.73 vs 4.46 ± 0.34, P = 0.025, respectively). While no significant differences between the TNBS and the S. boulardii or MP groups could be determined in terms of serum NO levels, the level of serum NO in the S. boulardii + MP group was significantly higher than in the TNBS and S. boulardii groups (8.12 ± 4.25 μmol/L vs 3.18 ± 1.19 μmol/L, P = 0.013; 8.12 ± 4.25 μmol/L vs 3.47 ± 1.66 μmol/L, P = 0.012, respectively). The tissue NO levels in the S. boulardii, MP and S. boulardii + MP groups were significantly lower than the TNBS group (16.62 ± 2.27 μmol/L vs 29.72 ± 6.10 μmol/L, P = 0.002; 14.66 ± 5.18 μmol/L vs 29.72 ± 6.10 μmol/L, P = 0.003; 11.95 ± 2.34 μmol/L vs 29.72 ± 6.10 μmol/L, P = 0.002, respectively). The tissue NO levels in the S. boulardii, MP and S. boulardii + MP groups were similar. The mean serum and tissue TNF-α levels were determined to be 12.97 ± 18.90 pg/mL and 21.75 ± 15.04 pg/mL in the control group, 18.25 ± 15.44 pg/mL and 25.27 ± 11.95 pg/mL in the TNBS group, 20.59 ± 16.15 pg/mL and 24.39 ± 13.06 pg/mL in the S. boulardii group, 9.05 ± 5.13 pg/mL and 24.46 ± 10.85 pg/mL in the MP group, and 13.95 ± 10.17 pg/mL and 24.26 ± 10.37 pg/mL in the S. boulardii + MP group. Significant differences in terms of the levels of serum and tissue TNF-α and the macroscopic and microscopic scores were not found between the groups. S. boulardii fungemia was not observed in any of the rats. However, Candida fungemia was detected in one rat (14%) in the TNBS group, two rats (28%) in the S. boulardii group, three rats (50%) in the MP group, and three rats (42%) in S. boulardii + MP group. S. boulardii does not demonstrate considerable effects on the DAI, pathological scores, or cytokine levels but does decrease the tissue NO levels.

Exogenous carbon monoxide attenuates inflammatory responses in the small intestine of septic mice

To determine whether the carbon monoxide (CO)-releasing molecules (CORM)-liberated CO suppress inflammatory responses in the small intestine of septic mice. The C57BL/6 mice (male, n = 36; weight 20 ± 2 g) were assigned to four groups in three respective experiments. Sepsis in mice was induced by cecal ligation and puncture (CLP) (24 h). Tricarbonyldichlororuthenium (II) dimer (CORM-2) (8 mg/kg, i.v.) was administrated immediately after induction of CLP. The levels of inflammatory cytokines [interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α)] in tissue homogenates were measured with enzyme-linked immunosorbent assay. The levels of malondialdehyde (MDA) in the tissues were determined. The levels of nitric oxide (NO) in tissue homogenate were measured and the expression levels of intercellular adhesion molecule 1 (ICAM-1) and inducible nitric oxide synthase (iNOS) in the small intestine were also assessed. NO and IL-8 levels in the supernatants were determined after the human adenocarcinoma cell line Caco-2 was stimulated by lipopolysaccharide (LPS) (10 g/mL) for 4 h in vitro. At 24 h after CLP, histological analysis showed that the ileum and jejunum from CLP mice induced severe edema and sloughing of the villous tips, as well as infiltration of inflammatory cells into the mucosa. Semi-quantitative analysis of histological samples of ileum and jejunum showed that granulocyte infiltration in the septic mice was significantly increased compared to that in the sham group. Administration of CORM-2 significantly decreased granulocyte infiltration. At 24 h after CLP, the tissue MDA levels in the mid-ileum and mid-jejunum significantly increased compared to the sham animals (103.68 ± 23.88 nmol/mL vs 39.66 ± 8.23 nmol/mL, 89.66 ± 9.98 nmol/mL vs 32.32 ± 7.43 nmol/mL, P < 0.01). In vitro administration of CORM-2, tissue MDA levels were significantly decreased (50.65 ± 11.46 nmol/mL, 59.32 ± 6.62 nmol/mL, P < 0.05). Meanwhile, the tissue IL-1β and TNF-α levels in the mid-ileum significantly increased compared to the sham animals (6.66 ± 1.09 pg/mL vs 1.67 ± 0.45 pg/mL, 19.34 ± 3.99 pg/mL vs 3.98 ± 0.87 pg/mL, P < 0.01). In vitro administration of CORM-2, tissue IL-1β and TNF-α levels were significantly decreased (3.87 ± 1.08 pg/mL, 10.45 ± 2.48 pg/mL, P < 0.05). The levels of NO in mid-ileum and mid-jejunum tissue homogenate were also decreased (14.69 ± 2.45 nmol/mL vs 24.36 ± 2.97 nmol/mL, 18.47 ± 2.47 nmol/mL vs 27.33 ± 3.87 nmol/mL, P < 0.05). The expression of iNOS and ICAM-1 in the mid-ileum of septic mice at 24 h after CLP induction significantly increased compared to the sham animals. In vitro administration of CORM-2, expression of iNOS and ICAM-1 were significantly decreased. In parallel, the levels of NO and IL-8 in the supernatants of Caco-2 stimulated by LPS was markedly decreased in CORM-2-treated Caco-2 cells (2.22 ± 0.12 nmol/mL vs 6.25 ± 1.69 nmol/mL, 24.97 ± 3.01 pg/mL vs 49.45 ± 5.11 pg/mL, P < 0.05). CORM-released CO attenuates the inflammatory cytokine production (IL-1β and TNF-α), and suppress the oxidative stress in the small intestine during sepsis by interfering with protein expression of ICAM-1 and iNOS.

Disruption of the Chemokine-Like Receptor-1 (CMKLR1) Gene Is Associated with Reduced Adiposity and Glucose Intolerance

Adipose tissue secretes a variety of bioactive signaling molecules, termed adipokines, which regulate numerous biological functions including appetite, energy balance, glucose homeostasis, and inflammation. Chemerin is a novel adipokine that regulates adipocyte differentiation and metabolism by binding to and activating the G protein-coupled receptor, chemokine like receptor-1 (CMKLR1). In the present study, we investigated the impact of CMKLR1 deficiency on adipose development, glucose homeostasis, and inflammation in vivo. Herein we report that regardless of diet (low or high fat), CMKLR1(-/-) mice had lower food consumption, total body mass, and percent body fat compared with wild-type controls. CMKLR1(-/-) mice also exhibited decreased hepatic and white adipose tissue TNFα and IL-6 mRNA levels coincident with decreased hepatic dendritic cell infiltration, decreased adipose CD3+ T cells, and increased adipose natural killer cells. CMKLR1(-/-) mice were glucose intolerant compared with wild-type mice, and this was associated with decreased glucose stimulated insulin secretion as well as decreased skeletal muscle and white adipose tissue glucose uptake. Collectively these data provide compelling evidence that CMKLR1 influences adipose tissue development, inflammation, and glucose homeostasis and may contribute to the metabolic derangement characteristic of obesity and obesity-related diseases.

Bromelain Ameliorates the Wound Microenvironment and Improves the Healing of Firearm Wounds

In a previous study, we proposed a new therapy using topical bromelain as a supplement to simple wound-track incision for the debridement of firearm wounds. This enzymatic debridement greatly simplified the management of high-velocity gunshot wounds in a pig model, and bromelain was confirmed to improve wound healing. The purpose of the present study was to investigate the effect of bromelain on the microenvironment of firearm wounds. Sixteen Chinese landrace pigs wounded by high-velocity projectiles were divided randomly into four groups: wound incision (group I), incision + bromelain (group IB), wound excision (group E), and control. Blood perfusion, oxygen partial pressure (pO(2)), and the content of tumor necrosis factor (TNF)-α and transforming growth factor (TGF)-β in wound-track tissue were measured. Wound healing was also noted. The recovery of blood perfusion in tissue and pO(2) in wound tracks was significantly more rapid in group IB and group E than in group I and control. The tissue level of TNF-α was significantly lower in group IB than in group I and control 48 h and 72 h post-wounding, and was lower than in group E 48 h post-wounding. The tissue level of TGF-β in group IB was sustained at a significantly higher level than in the other three groups. Wound healing time was also shorter in group IB. Enzymatic debridement using topical bromelain in incised wound tracks accelerates the recovery of blood perfusion, pO(2) in wound tissue, controls the expression of TNF-α and raises the expression of TGF-β.

Protective Effects of Melatonin and S-Methylisothiourea on Mechlorethamine Induced Nephrotoxicity

In this study, we aimed to investigate the protective effects of melatonin (MEL) and S-methylisothiourea (SMT) on mechlorethamine (MEC) induced nephrotoxicity. A total of 36 male Sprague-Dawley rats were divided into four groups: control, MEC, MEC+MEL, and MEC+SMT. Three groups received single dose of MEC (3.5 mg/kg) via transdermal route. Control animals were given saline only via transdermal route. MEL (100 mg/kg) was administered intraperitoneally 30 min after the application of MEC, and after the same dose of MEL was given every 12 h for a total of six doses. SMT (50 mg/kg) was also given intraperitoneally 30 min after the application of MEC. The tissue TNF-α, IL-1β, and NOx levels were found significantly different for all groups (P < 0.001). MEC application resulted in severe histopathological changes. Melatonin showed meaningful protection against kidney damage. But protection by SMT was weaker. TNF-α and IL-1β levels increased significantly with MEC application, and MEL and SMT ameliorated these increases in kidney tissue. MEC also elevated NOx levels in kidney tissue. Both inflammation and oxidative stress may have an important role in the MEC induced nephrotoxicity. MEL and SMT may also have anti-inflammatory properties, as well as anti-oxidant properties.

Insights antifibrotic mechanism of methyl palmitate: Impact on nuclear factor kappa B and proinflammatory cytokines

Fibrosis accompanies most chronic liver disorders and is a major factor contributing to hepatic failure. Therefore, the need for an effective treatment is evident. The present study was designed to assess the potential antifibrotic effect of MP and whether MP can attenuate the severity of oxidative stress and inflammatory response in chronic liver injury. Male albino rats were treated with either CCl 4 (1 ml/kg, twice a week) and/or MP (300 mg/kg, three times a week) for six weeks. CCl 4-intoxication significantly increased liver weight, serum aminotransferases, total cholesterol and triglycerides while decreased albumin level and these effects were prevented by co-treatment with MP. As indicators of oxidative stress, CCl 4-intoxication caused significant glutathione depletion and lipid peroxidation while MP co-treatment preserved them within normal values. As markers of fibrosis, hydroxyproline content and α-SMA expression increased markedly in the CCl 4 group and MP prevented these alterations. Histopathological examination by both light and electron microscope further confirmed the protective efficacy of MP. To elucidate the antifibrotic mechanisms of MP, the expression of NF-κB, iNOS and COX-2 and the tissue levels of TNF-α and nitric oxide were assessed; CCl 4 increased the expression of NF-κB and all downstream inflammatory cascade while MP co-treatment inhibited them. Collectively these findings indicate that MP possesses a potent antifibrotic effect which may be partly a consequence of its antioxidant and anti-inflammatory properties.

Hydrogen sulfide preconditioning or neutrophil depletion attenuates ischemia-reperfusion-induced mitochondrial dysfunction in rat small intestine

The objectives of this study were to determine whether neutrophil depletion with anti-neutrophil serum (ANS) or preconditioning with the hydrogen sulfide (H(2)S) donor NaHS (NaHS-PC) 24 h prior to ischemia-reperfusion (I/R) would prevent postischemic mitochondrial dysfunction in rat intestinal mucosa and, if so, whether calcium-activated, large conductance potassium (BK(Ca)) channels were involved in this protective effect. I/R was induced by 45-min occlusion of the superior mesenteric artery followed by 60-min reperfusion in rats preconditioned with NaHS (NaHS-PC) or a BK(Ca) channel activator (NS-1619-PC) 24 h earlier or treated with ANS. Mitochondrial function was assessed by measuring mitochondrial membrane potential, mitochondrial dehydrogenase function, and cytochrome c release. Mucosal myeloperoxidase (MPO) and TNF-α levels were also determined, as measures of postischemic inflammation. BK(Ca) expression in intestinal mucosa was detected by immunohistochemistry and Western blotting. I/R induced mitochondrial dysfunction and increased tissue MPO and TNF-α levels. Although mitochondrial dysfunction was attenuated by NaHS-PC or NS-1619-PC, the postischemic increases in mucosal MPO and TNF-α levels were not. The protective effect of NaHS-PC or NS-1619-PC on postischemic mitochondrial function was abolished by coincident treatment with BK(Ca) channel inhibitors. ANS prevented the I/R-induced increase in tissue MPO levels and reversed mitochondrial dysfunction. These data indicate that neutrophils play an essential role in I/R-induced mucosal mitochondrial dysfunction. In addition, NaHS-PC prevents postischemic mitochondrial dysfunction (but not inflammation) by a BK(Ca) channel-dependent mechanism.

Therapeutic Angiogenesis of Human Early Endothelial Progenitor Cells Is Enhanced by Thrombomodulin

We examined the effect of thrombomodulin (TM) domains 2 and 3 (TMD23) on human early endothelial progenitor cells (EPCs). TM was expressed and released by human EPCs cultured from peripheral blood mononuclear cells (PBMCs). Addition of TMD23 (100 ng/mL) to the cultured PBMCs increased the colony-forming units, chemotactic motility, matrix metalloproteinase activity, and interleukin-8 secretion but decreased tumor necrosis factor-α (TNF-α) release. Analysis of the signal pathway showed that TMD23 activated Akt. Inhibition of phosphatidylinositol-3 kinase-Akt blocked the effects of TMD23 on chemotactic motility, matrix metalloproteinase-9, interleukin-8, and TNF-α. In hindlimb ischemia mice, laser Doppler perfusion imaging of the ischemic limb during the 21 days after arterial ligation showed that the perfusion recovered best with intraperitoneal infusion of TMD23 plus local injection of early EPCs, followed by either infusion of TMD23 or injection of the cells. Animals without either treatment had the worst results. Animals treated with TMD23 also had lower circulating and tissue levels of TNF-α. TM is expressed and released by human circulating EPCs. Exogenous TMD23 enhances the angiogenic potential of early EPCs in vitro through activation of phosphatidylinositol-3 kinase-Akt pathway. Coadministration of TMD23 plus early EPCs augments therapeutic angiogenesis of the EPCs in ischemic tissues.

Involvement of galectin-3 in cadmium-induced cardiac toxicity

Accumulation of the wide spread environmental toxin cadmium (Cd) in tissues results in toxicity. Heart is one of the most effected tissues. Cd exposure induces inflammation in effected tissues. The present study was focused to evaluate roles of tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) in Cd toxicity and their relationships with galectin-3 levels. In this experimental study, male Wistar rats were divided randomly to control and experimental groups. Experimental group was exposed to Cd at the dose of 15 ppm for 8 weeks (n=10/group). Inflammatory status in hearts was evaluated with measurement of tissue TNF-α and IL-6 levels. Histopathological examination of heart was carried out by light microscopy. Heart tissue caspase-3 level was used to identify apoptosis. Tissue galectin-3 level was evaluated by ELISA. Statistical difference between groups was evaluated by unpaired Student t-test, correlation was analyzed by Pearson's test. Heart sizes were increased after Cd toxicity. A significant increase in galectin-3 tissue levels was seen after Cd toxicity, this was accompanied with a significant increase in the TNF-α (control: 402±39, Cd: 793±26 pg/g tissue, p<0.001) and IL-6 (control: 150±78, Cd: 325±65 pg/g tissue, p<0.001) levels. Histopathological examination under light microscope suggested a combination of ongoing necrosis and apoptosis. Increased caspase-3 levels were measured after Cd toxicity (control: 12±2, Cd: 18±3 pmol/µg/min, p<0.001). Chronic Cd administration induces inflammation and apoptosis in rat hearts. Cadmium causes increased galectin-3 production from heart tissue. The formation of TNF-α due to Cd exposure may likely trigger this mechanism.

Gastroprotective effects of amygdalin on experimental gastric ulcer: Role of NO and TNF-

Natural plant origin products like amygdalin with anti-inflammatory effects are still a major part of traditional medicine. We conducted the present investigation to elucidate the amygdalin effects on alcohol-induced gastric ulcer together with the possible role of nitric oxide (NO) and tumor necrosis factor-alpha (TNF-α). Thirty-two male Wistar rats were used to study therapeutic and preventive effects of amygdalin on absolute alcohol-induced gastric ulcer. Animals were equally divided into 4 groups (n=8): (I) control, (II) Alcohol; absolute alcohol (1 ml/200 g b.w) used to induce gastric ulcer, (III) Alcohol/Amygda; amygdalin (2 mg/kg/day intramuscular) administrated for three weeks after gastric ulcer induction and (IV) Amygda/Alcohol; amygdalin was given for three weeks before gastric ulcer induction. Ulcer index, gastric acid secretion, gastric tissue’s NO metabolites andTNF-α level measured. Ulcer index was significantly decreased in Alcohol/Amygda than Alcohol group (p<0.05). Gastric tissue’s NO metabolites level and gastric acid secretion decreased in Alcohol and Amygda/Alcohol groups than control (p<0.05). Gastric tissue TNF-α level increased in Alcohol and Amygda/Alcohol rats than control group (p<0.05). The results of this study show that amygdalin protected gastric mucosa from alcohol-induced gastric ulcer. This gastroprotection may mediate via gastric mucosal nitric oxide production and TNF-α suppression. Key words: Amygdalin, gastric ulcer, nitric oxide, TNF-α.

Tumor necrosis factor-α in vitiligo: direct correlation between tissue levels and clinical parameters

Background: Experimental evidences have shown that tumor necrosis factor (TNF)-α may play a role in the pathogenesis of nonsegmental vitiligo, and successful cases of vitiligo treated with TNF-α inhibitors have been recently reported.Materials and methods: Two cases of refractory generalized vitiligo, which showed high tissue levels of TNF-α, were commenced anti-TNF-α antibody etanercept 50 mg weekly. A retrospective study, considering chart review and immunohistochemical staining for TNF-α, was then carried out on eight additional patients affected by untreated vitiligo.Results: Etanercept achieved improvement of vitiligo in two patients at 6-month follow-up. Five out of eight specimens showed a strong cytoplasmic staining for TNF-α. Considering all 10 cases, patients with a strong TNF-α staining were characterized by a higher vitiligo disease activity score than patients with a weak staining.Discussion: These findings, albeit limited in significance by the low number of cases and the retrospective nature of the study, confirm a probable role of TNF-α in the pathogenesis of vitiligo. The intensity of TNF-α staining in vitiligo lesions may be worth to be further studied as a biomarker for potentially successful anti-TNF-α treatment of nonsegmental vitiligo in cases refractory to conventional treatment.

Curcumin immune-mediated and anti-apoptotic mechanisms protect against renal ischemia/reperfusion and distant organ induced injuries

Renal ischemia followed by reperfusion results in kidney injury which in turn produces and releases destructive inflammatory cytokines into the circulation causing subsequent distant organ injury. Little data suggest the immune mechanism of curcumin on protection against ischemia/reperfusion induced injury. We investigated the immunomodulatory and anti-apoptotic effects of curcumin on ischemia/reperfusion (I/R) injury in rats. Thirty-six rats were randomly divided into three experimental groups (sham, I/R and curcumin pretreated I/R, n = 12 each). Curcumin was administered orally to curcumin pretreated I/R group. Curcumin can significantly decrease both systemic as well as blood levels of cytokines (p < 0.05). Treatment with curcumin also resulted in significant reduction in serum and tissue level of TNF-α, IL-1β, IL-12, IL-18 and INF-γ that were increased by renal I/R injury (p < 0.05). Curcumin pretreatment reduce pulmonary apoptotic pathway via significant inhibition of TGF-β and caspase-3 in kidney and lung tissues. Given that pulmonary apoptosis is an important complication of acute renal injury, we identified curcumin protective effect against distant organ I/R induced injury. Based on our results, we concluded that curcumin protects the kidneys and other vital organs against I/R injury via immune-mediated and the new identified anti-apoptotic mechanisms.