Objective To investigate the expression of OX40 on CD4+T cells in patients with ulcerative colitis (UC)and the role of OX40/OX40L interaction for the cytokine production of lamina propria(LP)-CD4+T cells from UC.Methotis LP-CD4+T cells were purified.The expression of OX40 molecule was measured with FACS.LP-CD4+T cells were cultured with different stimuli and proliferation was assessed.The cytokines concentrations of the culture supernatant were detected.Results No difference of the OX40 expression was observed among the CD4+T cells from peripheral blood(PB)of UC patients,LP of non-inflammatory colonic tissue in UC patients and control PB.However.the expression of OX40 was significantly higher on LP-CD4+T cells from inflammatory colonic tissue in UC patients.In vitro culture with antigen presenting cells,the levels of IFNγ and TNFα secreted by LP-CD4+T cells from the inflammatory colonic tissue were significantly higher than those from the non-inflammatory colonic tissue(both P＜0.01).The levels of IFNγ and TNFα secreted by LP-CD4+T cells from the inflammatory colonic tissue were further increased by anti-OX40 MoAb stimulation.but suppressed significantly by adding anti-OX40L MoAb (compared with non stimulation,P＜0.01,respectively).The IFNγ and TNFα secretion of the LP-CD4+T cells from the non-inflammatory colonic tissue were not significantly different with and without anti-OX40 or anti-OX40L MoAbs stimulation.IL-4 and IL-10 produced by LP-CD4+T cells from the inflammatory or non-inflammatory colonic tissue were not significantly changed when adding different stimuli.Conclusions OX40 is highly expressed on LP-CD4+T cells from inflammatory colonic tissue in patients with UC.AntiOX40L MoAb can inhibit the proinflammatory cytokines secreted by these cells.It is indicated that OX40+T cells are involved in the immunopathological process in UC and blockage of the interaction of OX40 and OX40Lis a new strategy to be considered for the treatment of the disease.

Objective To investigate the levels of CD4 + CD25 + regulatory T cells and the Foxp3 in the peripheral blood of patients with ulcerative colitis ( UC), and analyze its role in the pathogenesis of UC. Methods From February 2007 to December 2008, 40 UC patients (23 active and 17 remissive), 33 irritable bowel syndrome (IBS) patients, and 32 normal controls entered our study. CD4 + CD25 + T cells were detected with flow cytometric assay. The expression of Foxp3 mRNA in peripheral blood mononuclear cell (PBMC) was detected by RTPCR. Interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) in the serum from the peripheral blood of UC patients, IBS patients, and normal controls were determined with ELISA. Results There were no significant differences of the peripheral CD4 +T cell numbers among the active, remissive UC and IBS patients, and normal control groups (P=0. 126). The positive rate of CD4+ CD25+ T cells in patients with active and remissive UC were significantly lower than those in IBS patients and normal controls ( both P ＜ 0. 01 ) it was more lower in the active UC patients than the remissive UC patients ( P ＜ 0. 001 ). However, the positive rate of CD4 + CD25 + T cells showed no significant difference between IBS patients and normal control groups ( P = 0. 343 ). The percentage of CD4 + CD25 + T cells was negatively correlated with UC disease activity index ( r = - 0. 660, P ＜ 0. 001 ) and with erythrocyte sedimentation rate (r = -0. 572, P =0. 001 ). The expressions of Foxp3 mRNA in PBMC from active and remissive UC patients were significantly lower than those in both IBS patients and normal controls ( both P ＜0. 001 ). There was no significant difference of the plasma levels of IL-10 or TGF-β among the active/remissive UC patients, IBS patients, and normal controls ( all P ＞ 0. 05 ). Conclusions CD4+ CD25 + regulatory T cells remarkably decline in the active UC and show certain increase in remissive UC, indicating that these cells may be involved in the pathogenesis of UC. The decreased expression of Foxp3 mRNA may be an important factor of the aberrant developmental disorder of CD4 + CD25 + regulatory T cells.

Objective To investigate the relationship of plasma homocysteine and the polymorphism of MTHFR gene with ischemic stroke in type 2 diabetes. Methods Serum Hcy ,folic acid and the polymorphism of MTHFR gene were compared among 81 T2DM patients with brain infarction (T2DM+BI) and 325 T2DM patients without brain infarction (T2DM ). Results All the genotypes of T2DM group and T2DM+BI group followed the hardy‐weinberg law. There was no significant difference in the frequency of mutant alleles (T) in site 677 of MTHFR gene and in frequency of TT genotype between the groups of T2DM and T2DM + BI (64.15% vs 60.15% and 42.5% vs 34.5% ,P > 0.05 ). The concentration of Hcy was significantly higher in patients with TT genotype than with CC genotype (14.4 ± 7.86) vs (10.58 ± 3.37)mmol/L(P<0.01). Conclusion There is no correlation between polymorphism of MTHFR gene and stroke in T2DM patients. The mutation of MTHFR C677T is associated with hyperhomocysteinemia.

Objective To investigate the expression an d the role of interleukin 17(IL 17) in patients with ulcerative colitis (UC). Methods IL 17,IL 6 and IL 8 were measured using ELISA technique , and IL 17 mRNA was assayed by RT PCR in 32 patients with UC and compared with 40 controls. The effect of anti IL 17 m onoclonal antibody (MoAb) on production of IL 6 and IL 8 by lamina propria mo nonuclear cells (LPMC) was studied. Results As compared with controls, serum concentrations of IL 17 ,IL 6 and IL 8 from UC patients were significantly higher, but with no statist ic difference. The expression of IL 17 mRNA and secretion of IL 17 by the peripheral blood CD + 4 T cells in UC patients were higher than that in nor mal controls in the presence of stimuli (both P

Background Retinal flatmount of oxygen-induced retinopathy (OIR) animal models is a useful tool in the study of ischemic retinopathy.The retinas of OIR of rat or mouse pups were small and thick and difficult in operating of conventional preparation and quantitative analysis of retinal flatmounts.Objective This study was to explore an easy and stable operating method of retinal flatmount combined with immunofluorescence staining in rodent.Methods Forty ＜6-hour-old SD rat pups were randomly assigned to OIR model group and normal control group.The pups were raised with nursing mothers in hyperoxia environment (80％) and normal oxygen environment (21％) alternately at a 24-hour interval for 14 days in the OIR model group,and the pups were raised in the room air for 14 days in the normal control group.The eyeballs of the rats were extracted to isolate the retinas intactly.The retinas were stained with glutamine synthetase (GS)-isolectin B4 firstly and then expanded into flatmounted and cut into 4 petals radially.Adobe Photoshop CS3 imaging analysis system was used to match the pictures into entire retinal vascular images and analyzed under the fluorescence microscope.The pixel values of retinal avascular areas and the entire retina were quantified by this system.The percentage of avascular areas to the entire retina was calculated to analyze the severity of non-perfusion areas.The use and care of the animals complied with the Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission.Results An intact and smooth retinal flatmount could be obtained by firstly staining method.Ora serrata structure was seen surrounding the whole retina.Strong green fluorescence was exhibited in retinal vessel net with clearly visible vascular branches;while the background fluorescence was weaker.Fully developed blood vessels were displayed in the retinas of the normal control group.Non-capillary areas around the central optic disk and large peripheral avascular areas could be seen in the retinal flatmounts of OIR models.Conclusions The preparation of retinal flatmount is easy and feasible by first immunofluorescence staining for retinal vessels followed by radially cutting of retina.This method of retinal flatmount can ensure the integrity of retinal vascular system and it is available for the observation and evaluation of retinal vascular structure in OIR models.

Sump syndrome is a rare complication of side-to-side choledochoduodenostomy (CDD) and occasionally occurs after spontaneous gallbladder-bile duct-digestive tract fistula or end-to-side choledochojejunostomy.Before the development of minimally invasive surgery,conventional surgical operation used to be the most important treatment method.This article reviews the research advances in sump syndrome in recent years and points out that endoscopic retrograde cholangiopancreatography is the major diagnostic method for this disease,and endoscopic sphincterotomy combined with bile duct debridement is the most simple and effective measure for the treatment of sump syndrome.Meanwhile,this article briefly reviews sump syndrome with reference to related literature and clinical practice,in order to raise the awareness for sump syndrome.

Objective To evaluate the role of mammalian target of rapamycin (mTOR) signaling pathway in dexmedetomidine-induced reduction of renal ischemia-reperfusion (I/R) injury in rats and the relationship with hypoxia-inducible factor 1 (HIF-1α).Methods Seventy-two male Sprague-Dawley rats, aged 10-12 weeks, weighing 220-260 g, were randomly divided into 4 groups (n=18 each) using a random number table: sham operation group (group S), group I/R, dexmedetomidine group (group Dex) ,and rapamicyn + dexmedetomidine group (group Rpm+Dex).Renal I/R was produced by occlusion of bilateral renal pedicles for 35 min follow by reperfusion in anesthetized rats in I/R, Dex and Rpm+Dex groups.Bilateral renal pedicles were only exposed, and then the abdominal cavity was closed in group S.Dexdetomidine 50 μg/kg was injected intraperitoneally at 30 min before I/R in group Dex.In group Rpm+Dex, rapamicyn 1.5 mg/kg and dexdetomidine 50 μg/kg were injected intraperitoneally, and renal I/R model was established 30 min later.Immediately after onset of reperfusion, and at 4 and 24 h of reperfusion (T1-3) , blood samples were collected from the caudal vein for measurement of serum creatinine and blood urea nitrogen (BUN) concentrations.After blood sampling at T1-3, the rats were sacrificed, and the renal specimens were obtained for detection of HIF-1αt, erythropoietin (EPO) and mTOR expression by Western blot.Their kidneys were removed at T3, and cut into sections which were stained with haematoxylin and eosin and examined under microscope.Acute renal tubular necrosis was scored.The cell apoptosis in renal tissues was detected by TUNEL assay, and apoptosis index (AI) was calculated.Results Compared with group S,the concentrations of serum creatinine and BUN, expression of HIF-1α, EPO and mTOR at T2,3 , AI at T3 and acute renal tubular necrosis score were significantly increased in the other three groups (P＜ 0.05).Compared with group I/R, the concentrations of serum creatinine and BUN were significantly decreased, and the expression of HIF-1α, EPO and mTOR was up-regulated at T2,3 , and AI and acute renal tubular necrosis score were decreased in group Dex (P＜0.05) , and no significant change was found in the parameters mentioned above in group Rpm + Dex (P ＞ 0.05).Conclusion The mTOR signaling pathway is involved in dexmedetomidine-induced reduction of renal I/R injury, which may be related to dexmedetomidine-produced up-regulation of HIF-1α expression in renal tissues of rats.

Objective To observe the effects of Toxoplasma lysate antigen(TLA)on growth inhibition and tumor angiogenesis in mice B16 melanoma.Methods Twenty mice bearing B16 melanoma were established by subcutaneous inoculating with B16 cells and were randomly divided into the treatment group and the control group 0.1 ml TLA was administered once every two days for the treatment group starting on the seventh day after inoculation.and an equivalent volume of sterile saline was administered similarly in the control group.The mice were sacrificed on 21 th day after the tumor cells was injected.The anti-tumor effects of TLA were observed by measuring the size and weight of tumor.The microvessel density(MVD)and expression of vascdar endotheilial growth factor(VEGF)were observed by immunohistochemical method.Results TLA Can markedly inhibit the B16 melamoa growth in mice.The volume and weight of the tumors in the experimental group were significantly decreased compared with the control group(P＜0.05),and the turnout inhibitory rate was 49.6%.The MVD of experimental group and control was(44.4000±4.7888)and(31.9000±2.6012)respectively,and with a significant difference between two groups(P＜0.05).The VEGF in the treatment group were significantly lower than those in the control group(P＜0.05).Conclusion TLA can inhibit the growth of the B16 melanoma in mice.suggesting that the anti-tumor effect induced by TLA via a possible antianigionesis mechanism.

BACKGROUND:As the potent, specific immunosuppressants emerge, the survival rate after intestinal transplantation is improved to some extent. However, the adverse effects of immunosuppressants and expensive treatment costs are not tolerable for many patients. Therefore, it is clinical y meaningful to choose traditional Chinese medicine which presents immunosuppressive effects. Artesunate has immune suppression effect, reduces acute rejection fol owing smal intestine transplantation, and improves the success rate of smal intestine transplantation.
OBJECTIVE:To observe the effect and action mechanism of artesunate in acute rejection after smal intestine transplantation in rats.
METHODS:Al ogeneic smal intestine transplantation models were established in the closed group of
Sprague-Dawley rats and Wistar rats, and then were randomly divided into three groups, syngenic transplantation group (SD→SD), al ogeneic transplantation group (Wistar→SD), and artesunate treatment group (Wistar→SD+artesunate 60 mg/kg per day, intraperitoneal injection).
RESULTS AND CONCLUSION:Rats in syngenic transplantation group survived for more than 10 days and they were al kil ed on day 10. The average survival of rats in al ogeneic transplantation group and artesunate treatment group was respectively (6.73±0.58) days and (8.50±0.74) days, with significant differences between the two groups (P<0.01). Histopathological examination showed that, there was no apparent rejection in syngenic transplantation group specimens, but mild, moderate and severe rejections in al ogeneic transplantation group on days 3, 5, 7. In treatment group, some specimens had mild rejection, but appeared relatively late to a low degree. Enzyme linked immunosorbent assay results revealed that, serum interleukin-2 and interferon-gamma expression levels in al ogeneic transplantation group were significantly higher than other two groups after surgery (P<0.01), serum interleukin-2 gene expression level in treatment group was also higher than syngenic transplantation group, but there was no significant difference (P>0.05), serum interferon-gamma expression level in treatment group was higher than syngenic transplantation group (P<0.05). Artesunate can inhibit acute rejection after rat smal intestine transplantation, and its mechanism may be related to inhibition effect on the secretion and expression of interleukin-2, interferon-gamma and other cytokines.

Objective To investigate the protection effect of dexmedetomidine against H2O2 injury in Human renal tubular epithelial cells(HK-2 cells). Methods HK-2 cells cultured in vitro were randomly divided into four groups(n = 24): control group, dexmedetomidine pretreatment group, H2O2 injury group, H2O2 injury +dexmedetomidine pretreatment group. Cell viabilities were measured by MTS assay, cell apoptosis were detected using flow cytometry, and expression of HIF-1α protein was quantified by western blot. HK-2 cells were divided into 8 groups by combining with three treatment factors such as PI3K inhibitor LY294002, dexmedetomidine and H2O2 injury. MTS assay was used to detect cell viability and western blot was used to quantify protein expression of HIF-1α,Bcl-2 and Bax after treatment in each group. Results Dexmedetomidine significantly increased the level of HIF-1α、 Bcl-2 in HK-2 cells after H2O2 injury, thus improved viabilities and reduced apotosis of cells. Moreover, effect on H2O2 injury cells of Dexmedetomidine was reversed by PI3K inhibitor LY294002. Conclusion Dexmedetomidine could protect against H2O2 injury by up-regulating HIF-1α expression through activating PI3K/Akt/mTOR signaling pathway in HK-2 cells.