Upregulation of stromal cell-derived factor-1 alpha/CXCR4 axis-induced migration of human neural progenitors by tumor necrosis factor-alpha and interleukin-8

BACKGROUND： Studies of several animal models of central nervous system diseases have shown that neural progenitor cells （NPCs） can migrate to injured tissues. Stromal cell-derived factor 1 alpha （SDF-la）, and its primary physiological receptor CXCR4, have been shown to contribute to this process. OBJECTIVE： To investigate migration efficacy of human NPCs toward a SDF-1α gradient, and the regulatory roles of tumor necrosis factor-α （TNF-α） and interleukin-8 （IL-8） in SDF-1α/CXCR4 axis-induced migration of NPCs. DESIGN, TIME AND SETTING： An in vitro, randomized, controlled, cellular and molecular biology study was performed at the Laboratory of Department of Cell Biology, Medical College of Soochow University between October 2005 and November 2007. MATERIALS： SDF-1α and mouse anti-human CXCR4 fusion antibody were purchased from R＆D Systems, USA. TNF-αwas purchased from Biomyx Technology, USA and IL-8 was kindly provided by the Biotechnology Research Institute of Soochow University. METHODS： NPCs isolated from forebrain tissue of 9 to 10-week-old human fetuses were cultured in vitro. The cells were incubated with 0, 20, and 40 ng/mL TNF-α, or 0, 20, and 40 ng/mL IL-8, for 48 hours prior to migration assay. For antibody-blocking experiments, cells were further pretreated with 0, 20, and 40 μg/mL mouse anti-human CXCR4 fusion antibody for 2 hours. Subsequently, the transwell assay and CXCR4 blockade experiments were performed to evaluate migration of human NPCs toward a SDF-1α gradient. Serum-free culture medium without SDF-1α served as the negative control. MAIN OUTCOME MEASURES： The transwell assay was performed to evaluate migration of human NPCs toward a SDF-1α gradient, which was blocked by fusion antibody against CXCR4. In addition, CXCR4 expression in human NPCs stimulated by TNF-α and IL-8 was measured by flow cytometry. RESULTS： Results from the transwell assay demonstrated that SDF-1α was a strong chemoattractant for human NPCs （P 〈 0.01）, and 20 ng/mL produced the highest levels of migration. Anti-human CXCR4 fusion antibody significantly blocked the chemotactic effect （P 〈 0.05）. Flow cytometry results showed that treatment with TNF-α and IL-8 resulted in increased CXCR4 expression and greater chemotaxis efficiency of NPCs towards SDF-1α（P 〈 0.01）. CONCLUSION： These results demonstrated that SDF-la significantly attracted NPCs in vitro, and neutralizing anti-CXCR4 antibody could block part of this chemotactic function. TNF-α and IL-8 increased chemotaxis efficiency of NPCs towards the SDF-1αgradient by upregulating CXCR4 expression in NPCs.

Clinical significance and prognostic value of tumor necrosis factor-α and dickkopf related protein-1 in ankylosing spondylitis

BACKGROUND Ankylosing spondylitis(AS)frequently occurs in people aged 30-45 years,and its prevalence is generally believed to be between 0.1%and 1.4%globally.At present,the“gold standard”for diagnosis of AS requires the provision of pelvic X-rays,which makes it more difficult to perform in population-based epidemiological studies.Therefore,the identification of serological indicators related to the diagnosis,treatment,and prognosis of AS patients is of great significance.AIM To analyze the therapeutic,diagnostic significance and prognostic value of dickkopf-related protein-1(DKK-1)and tumor necrosis factor-α(TNF-α)in AS.METHODS A total of 113 patients with active AS were selected as the research group,and 100 healthy subjects who underwent physical examination were selected as the control group.The levels of DKK-1 and TNF-α in peripheral blood in the two groups were compared.The diagnostic and predictive values of DKK-1 and TNF-α for AS were analyzed with ROC curves,and the factors influencing AS recurrence were analyzed with COX regression.RESULTS Before treatment,the research group showed lower DKK-1 levels but higher TNF-αlevels than the control group(both aP<0.05).In the research group,DKK-1 was up-regulated and TNF-αwas down-regulated after 12 wk of treatment(aP<0.05).The area under the curve,sensitivity and specificity of DKK-1 combined with TNF-αfor diagnosing AS were 0.934,82.30%and 97.00%,respectively.Before treatment,the area under the curve,cutoff value,sensitivity and specificity of DKK-1 for predicting the curative effect were 0.825,68.42 pg/mL,73.68%and 80.00%,respectively,and those of TNF-αwere 0.863,32.79 ng/L,92.11%and 77.33%,respectively.DKK-1 and TNF-αlevels after treatment were closely related to the curative effect(aP<0.05).C-reactive protein,the Bath Ankylosing Spondylitis Disease Activity Index,DKK-1,and TNF-αwere risk factors for AS recurrence(aP<0.05).CONCLUSION DKK-1 and TNF-αare effective in the diagnosis and treatment of AS and are risk factors for its recurrence.In addition,DKK-1 may be a potential target for the diagnosis of AS.

Helicobacter pylori tumor necrosis factor-α inducing protein promotes cytokine expression via nuclear factor-κB

AIM:To study the effects of Helicobacter pylori(H. pylori)tumor necrosis factor-α(TNF)inducing protein (Tip-α)on cytokine expression and its mechanism. METHODS:We cloned Tip-αfrom the H.pylori strain 26695,transformed Escherichia coli with an expression plasmid,and then confirmed the expression product by Western blotting.Using different concentrations of Tip-αthat affected SGC7901 and GES-1 cells at different times,we assessed cytokine levels using enzyme-linked immunosorbent assay.We blocked SGC7901 cells with pyrrolidine dithiocarbamate(PDTC),a specific inhibitor of nuclear factorκB(NF-κB).We then detected interleukin(IL)-1βand TNF-αlevels in SGC7901 cells. RESULTS:Western blot analysis using an anti-Tip-α antibody revealed a 23-kDa protein,which indicated that recombinant Tip-αprotein was recombined successfully.The levels of IL-1β,IL-8 and TNF-αwere sig-nificantly higher following Tip-αinterference,whether GES-1 cells or SGC-7901 cells were used(P<0.05).However,the levels of cytokines(including IL-1β,IL-8 and TNF-α)secreted by SGC-7901 cells were greater than those secreted by GES-1 cells following treatment with Tip-αat the same concentration and for the same duration(P<0.05).After blocking NF-κB with PDTC, the cells(GES-1 cells and SGC-7901 cells)underwent interference with Tip-α.We found that IL-1βand TNF-αlevels were significantly decreased compared to cells that only underwent Tip-αinterference(P<0.05). CONCLUSION:Tip-αplays an important role in cyto-kine expression through NF-κB.

Splenectomy Attenuates Severe Thermal Trauma-induced Intestinal Barrier Breakdown in Rats

Summary： The severe local thermal trauma activates a number of systemic inflammatory mediators, such as TNF-α, NF-κB, resulting in a disruption of gut barrier. The gastrointestinal tight junction （T J） is highly regulated by membrane-associated proteins including zonula occludens protein-1 （ZO-1） and oc- cludin, which can be modulated by inflammatory cytokines. As splenectomy has been shown to reduce secretion of cytokines, we hypothesized that （1） severe scald injury up-regulates TNF-α and NF-κB, meanwhile down-regulates expression of ZO-1 and occludin, leading to the increased intestinal perme- ability, and （2） splenectomy can prevent the burn-induced decrease in ZO-1 and occludin expression, resulting in improved intestinal barrier. Wistar rats undergoing a 30% total body surface area （TBSA） thermal trauma were randomized to receive an accessorial splenectomy meanwhile or not. Intestinal in- jury was assessed by histological morphological analysis, and serum endotoxin levels, TNF-α, NF-κB, ZO-1 and occludin levels were detected by Western blotting in the terminal ileum mucosal tissue. 30% TBSA bum caused a significant increase in serum endotoxin levels, but NF-κB, and TNF-α, and the av- erage intestinal villus height and mucosal thickness were decreased significantly. Burn injury could also markedly decrease the levels of ZO-1 and occludin in terminal ileum mucosal tissue （all P〈0.01）. Sple- nectomy at 7th day after burn significantly reversed the bum-induced breakdown of ZO-1 and occludin （all P〈0.01）. The results of this study suggest that severe thermal injury damages the intestinal mucosal barrier. Splenectomy may provide a therapeutic benefit in restoring bum-induced intestinal barrier by decreasing the release of inflammatory cytokines and recovering TJ proteins.

Clinical significance of dynamic detection for serum levels of MCP-1,TNF-α and IL-8 in patients with acute pancreatitis

Objective:To observe dynamic changes of levels of monocyte chemotactic protein-1(MCP-1),tumor necrosis factor-α(TNF-α) and interleukin-8(IL-8) in patients with acute pancreatitis and to investigate its evaluation value on the severity of acute pancreatitis.Methods:A total of 109 patients with acute pancreatitis admitted were divided into mild acute pancreatitis group(MAP group,42 cases),moderately severe acute pancreatitis(MSAP group,35 cases)and severe acute pancreatitis(SAP group,32 cases).ELISA was used to detect the serum levels of MCP-1,TNF-α and IL-8 of patients at day 1,day 4 and day 7 of admission to hospital.Results:The serum levels of MCP-1,TNF-α and IL-8 from MAP group,MSAP group and SAP group at day 1 of admission to hospital all significantly increased.There was a significant difference between MAP group and control group,MSAP group and MAP group,SAP group and MSAP group(P<0.05).The serum concentrations of IL-8 from MASP group and SAP group obviously increased at day 1,and there was significant difference between MASP group and MAP group,SAP group and MSAP group(P<0.05),while the difference between MAP group and control group was not obvious(P>0.05);The serum concentrations of MCP-1,TNF-α and IL-8 from MAP group all reached the highest level at day 4,which were significantly higher than the detection levels at day 1.In MSAP group and SAP group,the serum concentrations of MCP-1,TNF-α and IL-8 were the highest at day 1,which were significantly higher than the detection levels at day 4 and 7.At each detecting timing,the serum concentrations of MCP-1,TNF-α and IL-8 from MSAP group and SAP group were all higher than those of MAP group and MSAP group,respectively.Conclusions:The dynamic changes of serum levels of MCP-1,TNF-α and IL-8 in patients with acute pancreatitis have their rules,and the change rule of MAP group was different with that of MSAP and SAP group,which showed the reference value for the diagnosis and illness severity evaluation of acute pancreatitis.

Effect of triptolide on secretion of inflammatory cellular factors TNF-α and IL-8 in peritoneal macrophages of mice activated by lipopolysaccharide

Research has been carried out to look for safe and effective anti-inflammation drugs from traditional Chinese herbal medicine. As a powerful research technology of life science, molecular biology has entered many areas of traditional Chinese medicine.This study aimed to investigate the effect of triptolide on tumor necrosis factor-α （TNF-α） and interleukin-8 （IL-8） of peritoneal macrophages activated by lipopolysaccharide （LPS） in mice. Peritoneal elicited macrophages were separated, purified and activated by LPS in mice, then cultured in vitro with triptolide at different concentrations. The activity of TNF-a and the level of IL-8 of cellular supernatants were determined by MTT colorimetric assay and ELISA, respectively. The activity of TNF-a in macrophages was significantly inhibited （P〈0.01） by triptolide （10^-1-10^1μg/ml） during 4-24 hours in a time- and dose-dependent manner. The level of IL-8 in macrophages was significantly inhibited （P〈0.01） by triptolide （10^-1-10^1g/ml） in 12 hours in a dose- dependent manner. Triptolide could inhibit the activity of TNF-a and the level of IL-8 in macrophages activated by LPS.

EXPERIMENTAL WORK and RESEARCH Effect of Reduqing (热毒清) on TNF-α,IL-1,IL-6,IL-8 and PAF Levels in Endotoxin-Induced DIC Model of Rabbits

Objective: To study the mechanism of anti-endotoxemic effect of Reduqing Injection (RDQ)and to explore the essence of traditional Chinese 'heat-clearing and detoxifying therapy'. Methods: A disseminated intravascular coagulation (DIC) model was made in rabbits by intravenous injection with E. colt endotoxin. Increased plasma levels of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8) and platelet activating factor (PAF), as well as the ex-vivo LPS-induced interleukin-1 (IL-1 ) production in peripheral blood monocytes were observed in model (M) group. Results: Same parameters were significantly lower in RDQ and dexamethasone (DXM) groups than those in the M group (P < 0. 01, P < 0. 05). Histopathological examination showed severe damage of the liver, lungs and kidneys in the rabbits of M group, in contrast, only mild affects were seen in the RDQ and DXM groups. Conclusions: RDQ exhibits protective effect on rabbits against endotoxin-induced DIC. The suppression of cytokines and inflammatory mediator PAF by RDQ may play a central role in the inhibition of endotoxin-induced DIC cascade.

磷酸二酯酶4抑制剂Hemay005对角质形成细胞炎性因子产生的影响

目的为了探讨磷酸二酯酶4抑制剂Hemay005对体外培养的角质形成细胞株HaCaT细胞炎性因子产生的影响。方法分别用重组人肿瘤坏死因子-α(rhTNF-α)、重组人干扰素-γ(rhIFN-γ)和312 nm窄波UVB诱导,采用MTT法检测Hemay005对HaCaT细胞增殖的影响,用ELISA法检测其对HaCaT细胞白细胞介素(IL)-8、sICAM-1和肿瘤坏死因子(TNF)-α产生的影响。结果 Hemay005对HaCaT细胞增殖及25μg/L rhTNF-α诱导的IL-8产生无明显影响,但能剂量依赖性地抑制1 000 U/mL rhIFN-γ诱导的HaCaT细胞产生sICAM-1和124.2 mJ/cm2窄波UVB照射引起的HaCaT细胞产生TNF-α。结论 Hemay005可通过抑制角质形成细胞炎症因子的表达发挥抗炎作用。

Downregulation of cyclooxygenase-1 is involved in gastric mucosal apoptosis via death signaling in portal hypertensive rats

Portal hypertension （PHT） gastropathy is a frequent complication of fiver cirrhosis and one of the leading causes of death from cirrhosis. Apoptosis is widely considered to be an active energy-dependent mode of cell death and a distinct entity from necrotic cell death. It is unclear whether gastric mucosal apoptosis is involved in PHT gastropa- thy. Prostaglandins （PGs） produced through cyclooxygenase （COX） are thought to play a key role in protection of the gastrointestinal mucosa from injury and apoptosis. However, the role of COX in PHT gastropathy is still not clearly understood. The aims of this study were to investigate whether （1） gastric mucosal apoptosis is involved in PHT gas- tropathy and （2） downregulation of COX contributes to this apoptosis. In this study, we show that gastric mucosal apoptosis was remarkably increased while mucosal proliferation was inhibited in PHT rats. Gastric mucosal COX- 1 was significantly suppressed at both the mRNA and protein levels, and PGE2 was reduced in PHT rats. Further, PGE2 treatment suppressed gastric mucosal apoptosis in PHT rats. However, gastric mucosal COX-2 levels did not differ between sham-operated rats and PHT rats. Gastric mucosal levels of tumor necrosis factor-α （TNF-α） and Fas ligand, but not TNF-related apoptosis-inducing ligand, were increased, and activated caspase-8 and caspase-3 levels were upregulated in PHT rats. The release of cytochrome c from the mitochondria to the cytosol was not observed in PHT rats. Our data indicate that downregulation of COX-1 is involved in gastric mucosal apoptosis via death signal- ing-mediated type-I cell death in PHT rats.

Pathophysiology of insulin resistance and steatosis in patients with chronic viral hepatitis

Chronic hepatitis due to any cause leads to cirrhosis and end-stage liver disease.A growing body of literature has also shown that fatty liver due to overweight or obesity is a leading cause of cirrhosis.Due to the obesity epidemic,fatty liver is now a significant problem in clinical practice.Steatosis has an impact on the acceleration of liver damage in patients with chronic hepatitis due to other causes.An association between hepatitis C virus (HCV) infection,steatosis and the onset of insulin resistance has been reported.Insulin resistance is one of the leading factors for severe fibrosis in chronic HCV infections.Moreover,hyperinsulinemia has a deleterious effect on the management of chronic HCV.Response to therapy is increased by decreasing insulin resistance by weight loss or the use of thiazolidenediones or metformin.The underlying mechanisms of this complex interaction are not fully understood.A direct cytopathic effect of HCV has been suggested.The genomic structure of HCV (suggesting that some viral sequences are involved in the intracellular accumulation of triglycerides),lipid metabolism,the molecular links between the HCV core protein and lipid droplets (the core protein of HCV and its transcriptional regulatory function which induce a triglyceride accumulation in hepatocytes) and increased neolipogenesis and inhibited fatty acid degradation in mitochondria have been investigated.

A new in ammation marker of chronic obstructive pulmonary disease– adiponectin

BACKGROUND： This study was undertaken to measure the concentration of adiponectin （APN） in serum and induced sputum in patients with chronic obstructive pulmonary disease （COPD during acute exacerbation （AECOPD） and at stable stage and to determine the role of APN as a marker of in？ ammation in the pathogenesis of COPD.METHODS： All the patients in this prospective study were enrolled from October 2008 to October 2009, including 30 male AECOPD patients from the emergency department, 30 male stable COPD patients from the department of respiratory diseases, and 30 healthy non-smoking male controls from the department of medical examination. The serum and induced sputum were collected from each patient. All of the patients had normal weight （BMI range 18.5-24.9 kg/m2）. Patients with severe bronchial asthma, bronchiectasis or autoimmune disease were excluded. Cell count and classi？ cation was performed for the induced sputum. The concentrations of APN, IL-8, IL-6 and TNF-α were measured by ELISA. Pulmonary function was tested among the three groups. Comparisons between the groups were conducted by Student＇s t test, ANOVA analysis or nonparametric test. Correlation analysis was carried out by Pearson＇s product-moment correlation coef？ cient test or Spearman＇s rank-order correlation coef？ cient test.RESULTS： The concentrations of APN in the serum or induced sputum in AECOPD patients were signi？ cantly higher than those in stable COPD patients or healthy non-smoking controls （P〈0.01）. The concentration of APN in stable COPD patients was signi？ cantly higher than that in healthy non-smoking controls （P〈0.01）. For the AECOPD patients, APN was positively correlated with IL-8 and TNF-α in the serum and induced sputum （r=0.739, 0.734, 0.852, 0.857 respectively, P〈0.05）. For the stable COPD patients, APN was also positively correlated with IL-8 and TNF-α in the serum and induced sputum （r=0.751, 0.659, 0.707, 0.867 respectively, P〈0.05）. In addition, for the AECOPD patients, APN was positively correlated with the percentage of neutrophils in the induced sputum （r=0.439, P〈0.05）.CONCLUSIONS： APN is involved in the process of systematic and airway inflammation ofCOPD. This process is related to neutrophils in the airway, IL-8 and TNF-α. APN could be used as a new marker for in？ ammation of COPD.

Effect of Reduqing (热毒清) on Endotoxin-Induced Production of Tissue Factor and Cytokines in Human Whole Blood

Objective: TO study the effect of Reduqing (RDQ) Injection on the lipopolysaccharide (LPS)induced tissue factor and cytokine production in whole blood. Methods: Heparinized human blood was incubated with LPS in the presence or absence of RDQ. The plasma concentrations of TNF-α, IL-1β, IL-6, and IL-8were measured by enzyme-linked immunosorbent assays (ELISA) and the monocyte tissue factor activity wasmeasured by a one-stage tissue factor induced plasma clotting time assay. Results: RDQ was found to diminishthe LPS-induced increase of TNF-α, IL-1β and IL-6 in plasma but did not completely abolish their production.In contrast to the effect on these cytokines, RDQ caused further increase of the plasma level of IL-8 and themonocyte TF activity in the presence of LPS. Conclusions: In the in vztro whole blood assay system used inthis study, the decrease of LPS-induced production of TNF-α, IL-1β, and IL-6 was similar to a previous in vivostudy on the effect of RDQ on the production of these cytokines in response to two-time LPS injection in rabbits, while the increase of IL-8 and TF production was contradictory to the previous in vivo study. Potentialreasons contributing to the differences are discussed.

苏木对糖尿病大血管病变中SOD、TNF-α、MCP-1的影响

目的:探讨苏木对糖尿病大血管病变大鼠血清超氧化物歧化酶(SOD)、肿瘤坏死因子α(TNF-α)、单核细胞趋化因子1(MCP-1)水平的影响。方法:将24只雄性SD大鼠随机分为4组,每组6只,分别为对照组、模型组、α-硫辛酸组和苏木组。除去对照组,余下的18只大鼠单次腹腔注射STZ50 mg/kg制备糖尿病大鼠模型,对照组予以注射等体积的柠檬酸-柠檬酸钠缓冲液。成功造模后,苏木组给予0.15 g/kg/d的苏木,α-硫辛酸组给予20 mg/kg/d的α-硫辛酸,每日1次给药,连续给药8周。8周末,检测和比较两组大鼠血清中SOD、TNF-α、MCP-1水平的变化。结果:治疗后,苏木组及α-硫辛酸组血清SOD含量均高于模型组(P<0.05),但苏木组和α-硫辛酸组组间比较无统计学差异(P>0.05);苏木组及α-硫辛酸组血清TNF-α、MCP-1含量均显著低于模型组(P<0.05),与对照组没有明显差别(P>0.05)。结论:苏木可提高糖尿病大血管病大鼠血清SOD含量,降低血清TNF-α、MCP-1含量。