Objective To investigate the cellular receptor pathway and the intracellular signaling of advanced glycation end products(AGE)-induced inflammatory reaction in monocytes. Methods Human peripheral monocytes were isolated from healthy volunteers. Cells were incubated with AGE modified by the addition of human serum albumin (AGE-HSA) either with pretreatment or no pretreatment of anti-AGE receptor (RAGE) IgG, NADPH oxidase inhibitor (apocynin)or a specific inhibitor of p38(SB 203580). The levels of interleukin-1?(IL-1?)and tumor necrosis factor-?(TNF-?) in the supernatants were assayed with enzyme-linked immunoadsorbent assay (ELISA). Reactive oxygen species (ROS) production was determined by MCLA chemiluminescence. Nuclear factor-?B translocation was assayed by immunochemical staining with anti-NF-?B/p65 and electrophoretic mobility shift assay(EMSA). Results AGE-HSA was found to induce activation of NF-?B, increase levels of IL-1? and TNF-? in the supernatants, and enhance production of ROS by monocytes. Pre-treatment of cells with anti-RAGE IgG or apocynin inhibited AGE-HSA to induce NF-?B translocation and IL-1? or TNF-? production. AGE stimulated ROS production could also be blocked by pre-treatment of cultured cells with anti-RAGE IgG or apocynin. Pre-treatment of cultured cells with SB 203580 inhibited both NF-?B activation and cytokines production, but showed no significant effect the cells to produce ROS. Conclusion AGE-HSA could induce IL-1? and TNF-? release as well as ROS production in human monocytes via a pathway mediated by RAGE. Activation of NADPH oxidase may be the upstream of the intracellular pathway. AGE-induced cytokines production was p38 pathway-dependent.

Enhanced expression of adhesion molecules on human endothelial cells of synovial tissue has been demonstrated in patients with dialysis related amyloidosis(DRA).The study was conducted to elucidate the mechanism by which the expression of adhesion molecules on human endothelial cells was up regulated. Human endothelial cells derived from umbilical veins(HUVEC) were coincubated in vitro with native human serum albumin(HSA) or HSA modified with advanced glycation end products(AGE HSA).The expression of adhesion molecule intercellular adhesion molecule 1(ICAM 1),vascular cell adhesion molecule 1(VCAM 1) and E selectin was determined respectively by immunofluorescence staining and flow cytometer analysis.The results showed that ICAM 1 and VCAM 1, but not E selectin,were constitutively expressed on HUVEC. AGE HSA enhanced the expression of ICAM 1, VCAM 1 and E selectin on HUVEC in a time and dose dependent manner. HSA had no effect on the expression of adhesion molecules. The results led to the conclusion that AGE HSA up regulated the expression of adhesion molecules on human endothelial cells and therefore promoted the infiltration of monocytes/macrophages around ? 2 microglobulin amyloid.

The aim of this study was to investigate the effect of patients isolation,paying careful attention to hygienic measure and strict sterilization on prevention of hepatitis C infection in hemodialysis(HD) unit. Patients in our HD unit from May 1994 to May 1998 were isolated and strict sterilization of dialysis apparatuses were performed, while patients from April 1990 to April 1994 were not isolated. The rate of HCV infection in these two groups of patients were 18 20% and 26 72%, respectively ( P

The study was performed to investigate the prevalence of nasal carriage of Staphylococcus aureus (NCSA) in patients with chronic renal failure(CRF) and its relationship with incidence of bacteremia in patients on hemodialysis(HD). The preventive effect of external application of mupirocin ointment on HD patients with venous catheters was also observed. Nasal swabs were taken from 114 CRF patients hospitalized from Nov. 2000 to April 2002. Samples from 42 patients in CCU with normal renal function and 48 staffs working in HD centre were also analyzed. External application of mupirocin ointment near the exit sites of catheters was performed as prophylaxis in HD patients with venous catheters. The prevalence of SA bacteremia was compared with that of the historical control group from Jan.1999 to Oct.2000. The results showed that the prevalence of NCSA in CRF patients was 14% (16/114). Nasal swab cultures were all negative in CCU patients, as well as staffs working in the HD centre. The mean frequency of SA bacteremia in HD patients with NCSA was 0 12/catheter month, compared to 0 in HD patients with no NCSA( P

Objective To investigate the effect of vitamin E supplementation on oxidative stress in hemodialysis patients. Methods Fifty-six uremic patients undergoing hemodialysis (HD) were involved in this self-controlled study. The patients were observed for one month to evaluate the status of oxidative stress and then divided into two groups randomly. Group A patients (n=28) received 200mg/d of vitamin E and Group B patients (n=28) were treated with 400mg/d of vitamin E. Vitamin E was supplied for one month in both groups. The serum levels of advanced oxidation protein products (AOPP), malondialdehyde (MDA), vitamin E and activity of glutathione peroxidase (GSHPx) were measured at the initial and the end of vitamin E supplementation respectively. Results Serum levels of AOPP and MDA were significantly higher in HD patients compared with healthy controls (n=56, P

Objective A close relationship between atherosclerosis and plasma levels of advanced oxidation protein products (AOPP) has been demonstrated in patients with chronic renal failure. The present study was performed to investigate the effect of AOPP on human endothelial cells. Methods Human umbilical vein endothelial cell line ECV304 was incubated with different concentrations of AOPP-BSA, which prepared by combining BSA with HOCl at different molar ratios. Cell viability was measured by MTT assay. Intracellular production of reactive oxygen species (ROS) was evaluated kinetically using VICTOR Wallac 1420 mutilabel counter. Results AOPP-BSA decreased endothelial cell viability, which was dependent on the molar ratio of BSA/HOCl and the concentration of AOPP-BSA. AOPP-BSA also enhanced the intracellular ROS production in ECV304. AOPP-induced ROS production was correlated with the molar ratio of BSA/HOCl and the concentration of AOPP-BSA. Pretreatment of cells with a small molecular glutathione peroxidase mimic (ebselen) reduced AOPP-induced ROS production by 53% with preservation of cell viability. Conclusion AOPP decreased endothelial cell viability via induction of oxidative stress.

Objective To test the hypothesis that advanced glycosylation end products(AGEs) increase cellular inflammation in atherosclerotic plaques. Methods Fifty rabbits were randomly divided into five groups. Hypercholesterolemic (0.5% cholesterol in diet) rabbits received repeated intravenous injection of either AGEs modified rabbit serum albumin (AGEs-RSA ) (group A) or unmodified RSA (group B) for 10 weeks. Rabbits treated with either hypercholesterolemic diet (group C)or with a normal diet(group D) or with a normal diet, and intravenous injection of AGEs-RSA (group E) were served as controls. Aortas were harvested at the 10th week, and lipid deposition was quantitated by oil red 0 staining. Macrophage (RAM-11 positive cells) and T lymphocyte (CD43 positive cells) infiltration, smooth muscle cell(?-actin positive cells) migration and proliferation were determined by using immunohistochemical staining and image-analysis techniques. Results Atherosclerotic plaques could be found in animals fed with hypercholesterolemic diet.Lipid deposition in plaque was significantly higher in group A (71.86%?8.3%) than those in group B (53.76%?3.72%)and group C (56.67%?9.2%). Infiltrations of macrophage[ (23.1?8.5)/0.01 mm2]and T lymphocyte[ (15.1 ? 3.8)/0.01 mm2]as well as migration and proliferation of smooth muscle cell [ (19.2?5.7)/0,01 mm2] in atherosclerotic lesions were significantly increased in animals treated with hypercholesterolemic diet and received injection of AGEs-RSA (group A) when compared with group B [macrophage (14.4? 5.9)70.01 mm2; T lymphocyte (9.1?2.6)/0.01 mm2; smooth muscle cell (12.9?3.8)/0.01 mm2]and group C[macrophage (15.4?4.4)/0.01 mm2; T lymphocyte (10.5?2.2)/0.01 mm2, smooth muscle cell (13.8?3.9)/0.01 mm2]. Neither plaque nor a cellular inflammation was found in animals fed with normal diet (group D)and in those received repeated injections of AGEs-RSA (group E). Conclusion AGEs increase cellular inflammation in atherosclerotic plaques and may accelerate formation of atherosclerosis in AGEs associated diseases.

BACKGROUND:There are reports concerning differentiation of adipose-derived mesenchymal stem cells(ADSCs)into chondrocytes using gene transfection technique.However,the transfection of adenovirus and adeno-associated virus into ADSCs is various.It is controversial whether adeno-associated virus(AAV)can transfect ADSCs.OBJECTIVE:To observe the enhanced green fluorescent protein(EGFP)expression following Ad5-EGFP and rAAV2-EGFP transfection into ADSCs,and investigate the cell proliferation ability following transfection.METHODS:ADSCs were isolated from the adipose tissue,which was from 6-month-old New Zealand albino rabbit back and neck by mechanical digestion and enzyme digestion,then ADSCs were cultured and amplified in vitro.ADSCs were infected with Ad5-EGFP and rAAV2-EGFP,and the EGFP expression was observed.A total of 2 μL sodium butyrate(1 mol/L)was added into the medium after rAAV2-EGFP transfection.MTT assay was used to detect the gene transfection effects on reproductive activity of ADSCs.RESULTS AND CONCLUSION:ADSCs isolated and cultured in vitro were flat,long-spindle and amplified stabry.The cell morphology was uniform.Many green fluorescent cells were observed in Ad5-EGFP and rAAV2-EGFP groups.Transfection efficiencies were about 88% and 83%.Adenovirus and adeno-associated virus vector can be transfected with ADSCs,and transfection efficiency is high.Adeno-associated virus needs sodium butyrate to increase its level of gene expression.

Objective:To prepare the polyclonal antibodies against advanced oxidation protein products (AOPP),and to provide an effective agent for research on the pathogenesis of AOPP and assess exactly the relationship between AOPP and relative diseases.Methods:AOPP-rabbit serum albumin (AOPP-RSA) was prepared by treating RSA with hypochloric acid.The rabbit anti-AOPP-RSA polyclonal antibodies were generated and purified by affinity chromatography. The titers and the specificity of the antibodies were measured by ELISA.The plasma AOPP and the localization of AOPP in nephridial tissues of some patients with chronic kidney disease (CKD) were determined using rabbit anti-AOPP-RSA.Results:Titers of the antibodies were 10-6.Purified antibodies reacted specifically with oxidized albumin from different genus,but could not react with normal albumin and glycosylated albumin.The high level of AOPP in plasma from CKD patients was confirmed by Western blot.The antibodies could be used to immunostain AOPP deposition in different regions of kidney tissues from both CKD patients and CKD rat models.Conclusion:We successfully generate rabbit anti-AOPP polyclonal antibodies with high titers and striking specificity.The presence of plasma AOPP and localizations of AOPP in kidney tissues of CKD patients can be demonstrated using the antibodies.The development of anti-AOPP polyclonal antibodies may provide a new tool to explore the pathogensis of AOPP and assess exactly the relationship between AOPP and relative diseases.

Objective To study the changes in interleukin-32 (IL-32) in rats with Klebsiella bacillus pneumonia and approach its significance. Methods Seventy-two Sprague-Dawley(SD)rats were divide into control group,model group and experimental group by the method of random digits table,then the experimental group was subdivided into 4 hours and 1,3 and 5 days experimental subgroups(each n=6). The rat model of Klebsiella bacillus pneumonia was established by injection of 0.3 mL Klebsiella bacterial suspension into the trachea. Before the establishment of the model in the experimental group,IL-32 inhibitory agent,protease activated receptor-2(PAR2) was injected into the abdominal cavity. After model establishment,at different time points,blood was collected via tail vein to observe the changes in serum levels of IL-32,tumor necrosis factor-α(TNF-α),IL-6 and IL-8 in all the groups. The lungs were removed and stained with hematoxylin-eosin(HE)method to investigate the histopathological changes of the lung tissues under the light microscope. Results Compared to the control group, with the prolongation of time the levels of IL-32,TNF-α,IL-8 and IL-6 were increased gradually in the model group,and reached their peaks at 3 days〔IL-32(ng/L):84.40±28.24 vs. 18.57±3.86,t=5.544,P=0.002;TNF-α(ng/L):79.27±14.64 vs. 17.82±3.86, t=9.994, P=0.000;IL-8(ng/L):55.85±10.90 vs. 16.66±3.76,t=8.544, P=0.000;IL-6(ng/L):56.65±2.57 vs. 28.48±2.11,t=19.693,P=0.000〕;PAR2 could inhibit above indexes significantly,there was statistical difference at 3 days compared with the model group〔IL-32(ng/L):54.13±6.68 vs. 84.40±28.24,t=2.560,P=0.046;TNF-α(ng/L):49.12±3.56 vs. 79.27±14.64,t=4.901,P=0.003;IL-8 (ng/L):22.95±2.52 vs. 55.85±10.90,t=7.204,P=0.000;IL-6(ng/L):36.49±2.63 vs. 56.65±2.57,t=13.443, P=0.000〕. Under the light microscope,the inflammatory changes in the lung tissue in experimental group were milder than those in the model group. Conclusion As a pro-inflammatory cytokine,IL-32 can induce the production of TNF-α,IL-6 and IL-8,and the inhibition of IL-32 production may play a role in suppression of the development of Klebsiella bacillus pneumonia.