Objective:To study the positive expression rate of M2 subtype of macrophage cell surface molecules and the inflammatory factors of PPS in IL-4-induced M2 macrophage.Methods:The experiment was divided into 5 groups:blank control group, Model group,PPS groups(50 μg/ml,100 μg/ml and 200 μg/ml).The expression of CD206 and CD23 was used as bio-maker to confirm IL-4 induced macrophages by treating RAW264.7 with 20ng/ml of IL-4.IL-4 induced RAW264.7 cells were treated with PPS of 50μg/ml,100μg/ml and 200μg/ml for 24 h.Then the expression of CD206,CD16/32 and CD40 were analyzed by flow cytometry, and the mRNA expression of IL-1β,TNF-α,IL-10 and iNOS were detect by qRT-PCR.Results: After treated with IL-4,the positive rate of CD206 of RAW264.7 were high.After treated with PPS ,the rate of CD16/32 and CD40 in IL-4 induced RAW264.7 cells were high ,the expression of CD206 decreased,and the mRNA level of IL-1βand TNF-αincreased.Conclusion:RAW264.7 cells can be polarlized to M2 subtype macrophage by using 20 ng/ml IL-4.PPS enhances the mRNA of IL-1β,TNF-αand the expression of CD40, CD16/32 in IL-4-induced RAW264.7 cells .These results indicate that PPS can induce the M2 subtype to become M1 macrophages, can improve immune function of macrophages.

Objective To examine the stability of high-density lipoprotein cholesterol (HDL-C)during serum incubation at different temperature and time periods.Methods Ten healthy volunteers (4 males and 6 females,aged 24 to 59 years)from Beijing Hospital were recruited in May 2015.Fasting venous blood samples were collected and centrifuged to separate the sera.Serum samples were incubated at 4℃ for 24 h,25℃for 0,1,8 and 24 h (with or without an LCAT inhibitor).Serum to-tal cholesterol (TC),total free cholesterol (TFC)HDL-C and HDL-FC were measured by the HPLC Method.Results HDL-FC and HDL-C changed -6.91% and -2.17% during serum incubation at 4℃for 24h.TFC,HDL-FC and HDL-C changed significantly (averaged -13.70%,-25.88% and -1.53% respectively)during serum incubation at 25℃ for 24 h,in which the decrease of TFC and HDL-FC were inhibited by the addition of the LCAT inhibitor.The decrease of HDL-C was even higher in the presence of the LCAT inhibitor.Conclusion Serum TFC,HDL-C and HDL-FC levels changed during serum incubations,which were caused by the LCAT and CETP activities and the transfer of cholesterol among lipoproteins. For accurate measurement of serum HDL-C,prolonged serum storage should be avoided in clinical laboratories.

Background Proliferative vitreoretinopathy (PVR):no blood vessels,fibrous membrane cell proliferation occurs retinal surface.The development of specific mechanisms has not been completely clarified,but the retinal pigment epithelium(RPE) and platelet-derived growth factor(PDGF) for the past few years are the research hotspot.Objective To explore the effects of hypoxia on the production of PDGF-BB and proliferation in cultured human RPE (hRPE) cells.Methods In the experimental group,10,15,20,30,40 μmol/L CoCl2 were used to mimic hypoxia environment of hRPE cells,the control group did not add CoCl2 on cultured hRPE cells.By using reverse transcriptive PCR (RT-PCR) and enzyme linked immunosorbent assay(ELISA),the expression of PDGF-BB mRNA and protein was detected,and detected proliferation of hRPE cells by MTT.The cells were divided into 6 groups,different siRNAs were transfected into PDGF-BB siRNA1 group,PDGF-BB siRNA2 group,PDGF-BB siRNA3 group (because PDGF-BB has three different siRNAs,one of which is a valid siRNA),β-actin siRNA group,unrelated sequence siRNA group,only Lip2000 was plused in the blank control group.After transfection of 4-6 hours,except for blank control group,the other five groups were added to PDGF-BB mRNA and protein and its effect on the proliferation of hRPE cells obviously CoCl2 concentrations (15 μmol/L) simulated hypoxia environment of 24 hours,the detection of PDGF-BB mRNA and protein and hRPE cell proliferation rate.Results Neither PDGF-BB mRNA or protein was found in the blank control group.The production of PDGF-BB mRNA and protein in hRPE cells cultured by different concentrations were different,with significant differences among them (F=43.737,P＜0.01;F=54.612,P＜0.05),and the expressions of PDGF-BB mRNA and protein of 15 μmol/L CoCl2 group were more than other groups,with significant differences between the two groups (all at P＜0.05).MTT showed that PDGF-BB protein and hRPE cell proliferation rate in different concentions of CoCl2 groups were different,with significant differences among them(F=95.379,P＜0.01 ; F =63.375,P＜0.05),the expression of PDGF-BB protein was more and hRPE cell proliferation rate was higher in 15 μmol/L CoCl2group than other groups,with significant differences between the two groups (all at P＜0.05),postive correlation was found between PDGF-BB protein and hRPE cell proliferation rate (r =0.994,P＜0.05).The production of PDGF-BB mRNA and protein in the different siRNA transfected groups were different,with significant differences among them (F =156.330,125.650,both at P＜0.01),and the expressions of PDGF-BB mRNA and protein of PDGF-BB siRNA2 group were more than other groups,with significant differences between them(all at P＜0.05).MTT showed that PDGF-BB protein and hRPE cell proliferation rate in different siRNA transfectedgroups were different,with significant differences among them (F =73.131,98.564,both at P＜ 0.01).The expression of PDGF-BB protein was less and hRPE cell proliferation rate was lower in PDGF-BB siRNA2 group than other groups,with significant differences between them (all at P ＜ 0.05),postive correlation was found between PDGF-BB protein and hRPE cell proliferation rate (r =0.996,P＜0.05).Conclusions Proper hypoxia can induce the expression of PDGF-BB,PDGF-BB expression can significantly promote the proliferation of hRPE cells.In the transfection targeting PDGF-BB siRNA,PDGF-BB expression is suppressed,can effectively reduce the value of hRPE cell hyperplasia.

BACKGROUND: Currently, the materials used in clinical practice to repair cruciate ligament of knee joint contain auto-graft bone- mid 1/3 patella tendon-bone (B-T-B), auto-semitendinous muscle, gracilis muscle and allogenic tissue graft. All of them are limited to a certain degree in clinical application. Therefore, people hope to consistently develop artificial ligaments to take the place of auto- and allografts. OBJECTIVE: To investigate the feasibility to construct artificial biological ligament (ABL) by applying a novel biochemical technique using porcine tendon as the raw material. DESIGN: Research of new biological material. SETTING: Department of Orthopedics, Third Affiliated Hospital of Sun Yat-sen University. MATERIALS: Adult pigs of either gender were provided by the Animal Center of Sun Yat-sen University. Scanning electron microscope (SEM, S-520) was provided by Hitachi, Japan, and micro-controlled electron tension-testing device (Model LWK-10B) by Guangzhou Experimental Devices Factory. METHODS: The experiment was performed at the Animal Center of Sun Yat-sen University from January 2004 to June 2005. ABL was established by means of treating porcine tendon with epoxy cross-linking fixation, diversified antigen minimization process, mechanic enhancement modification and surface activating process. Under aseptic condition, a 6-month-old goat's bone marrow was abstracted, and then the bone marrow matrix stem cells were cultured in ABL stent for 3 weeks. Scanning electron microscope was used to observe structure and compatibility of artificial ligament, and mechanics test was used to analyze biomechanics characteristics of ABL. MAIN OUTCOME MEASURES: Structural features, cell compatibility and biomechanics characteristics of ABL.RESULTS: ① Structural features of ABL: The appearance of ABL was similar to that of the normal human ligament. Histological examination showed that the ABL was collagen fibers with no cells. Electron microscope examination revealed that the ABL was composed of hair-looking and fiber-like objects running uniformly in a certain direction and closely parallel-arranged. ② Cell compatibility: Three weeks after xenogenic marrow matrix cells were cultured on the surface of the ABL, it was noted that cells adhered and the matrix secreted by the cells precipitated around the cells. There were no cells found inside the ABL. ③ Mechanical strength of the ligament: The average diameter of ABL was 5 mm and the mechanical test at a speed of 100 mm/min showed that its averaged tensile limit was 927.19 N and the tension-resistant strength was 47.22 N/mm those were close to the corresponding parameters of the normal goat's ACL. The normal goat's ACL was 5 mm. The greatest tensile load was 807.50 N and the tension-resistant strength was 41.13 N/mm.CONCLUSION:As we used the unique biochemical technique and minimized the xenogenic protein immunogenicity of the porcine tendon, ABL has acceptable biomechanical properties and superior biocompatibility. As a substitute of the ligament in the reconstruction of the ACL, ABL has a promising prospect in clinical applications.

Objective To develop a solid phase extraction and Folin-Ciocalteu method for the measurement of total polyphenols in urine samples.Methods From a group of individuals attending an annual physical examination at Beijing hospital, 123 healthy volunteers (52 males and 71 females, ranging in age from 18 to 81 years ) were recruited during the period from December 2013 to April 2014.Urine samples were stored in 0.5%HCl at -80 ℃.For analysis, samples were applied to the Plexa PAX solid phase extraction cartridge, to purity the polyphenols through washing, evaporating and reconstituting.Total polyphenols were measured by Folin-Ciocalteu colorimetric assay, calculated by gallic acid standard curve, and corrected by urine creatinine concentrations.The relationship between total polyphenols and fruits and vegetable intake and cardiovascular disease risk factors were analyzed. Results Gallic acid standard solution and urine samples were stable in 0.5% HCl for 48 h at RT and 7days at 4 ℃, respectively.The PAX cartridge effectively eliminated the possible interfere materials in urine and had better recovery for most of the polyphenol types.The inter assay and total CVs for the measurement of total polyphenols were 2.7%-3.8% and 2.4%-4.6 %, respectively.Total polyphenol concentrations of 123 healthy subjects were 114.13(82.97-146.70) mg GAE/g Crea.Total polyphenol levels positively correlated with both HDL-C (r=0.194, P=0.032) and apoAI (r=0.312,P<0.001), and negatively correlated with serum uric acid levels(r=-0.220,P=0.014).Conclusions We established a measurement of total polyphenols in urine samples using solid phase extraction and Folin-Ciocalteu method.This simple, precise method reliable and may be use to assess dietary polyphenols intake.

Objective To investigate the clinical features,diagnosis and treatment of von Hippel-Lindau (VHL) disease in order to improve the understanding of this disease.Methods The clinical features,imaging features,pathological data and treatments of 7 patients in 3 families admitted to our hospital from December 2005 to August 2017 were retrospectively analyzed.Results Ten oprations were performed under the guidance of neural navigation system for the 7 patients.There were 4 enjoying total resection at once;one achieved primary surgical resection of the primary lesions,and residual lesions grew up and the patient accepted surgical treatment at the 7th and 9th years;one patient achieved primary surgical resection of the primary lesion,the residual lesions grew up and surgical treatment was performed 8 months later;another one underwent surgical resection of the primary lesion,and the residual lesion was treated with gamma knife therapy 3 months after surgery.The pathological results of the 10 operations of the 7 patients were proved to be hemangioblastoma.No operative deaths were noted.The symptoms of these patients when they were discharged from the hospital were significantly better than those when they were on admission.The 10 patients were followed up so far,no symptoms of neurological impairment were noted,and they could work and live a normal life.Conclusions Surgical treatment is still the preferred method of VHL,and radiotherapy is effective for small lesions.

Objective:To investigate the effects of heme oxygenase-1 (HO-1) on hepatic sinusoidal endothelial cells (LSECs) proliferation, migration, and hepatocyte proliferation.Methods:Eighteen male C57BL/6 mouse aged 6-8 weeks old were underwent partial hepatectomy. Cell proliferation and HO-1 expression in residual liver tissue were detected by immunofluorescence histochemistry at 0 d, 2 d and 4 d after operation. In vitro, LSECs were transfected with adenovirus carrying HO-1 gene (HO-1 group), and the cells were transfected with empty vector adenovirus and the non-transfected cells were used as control. In addition, LSECs from different transfection groups were co-cultured with hepatocyte without contact to evaluate the effect of HO-1 expression on promoting hepatocyte proliferation. Western Blotting and RT-PCR were used to detect the protein and mRNA expression of HO-1, inhibitor of DNA binding and or differentiation (Id1), hepatocyte growth factor (HGF) and Wnt2. Cell proliferation was detected by EdU. The ability of cell migration was detected by Transwell migration assay.Results:Compared with 0 d after hepatectomy, LSECs proliferation and HO-1 expression within LSECs were increased significantly at 4 d after surgery. EdU positive rate of LSECs in HO-1 group (27.20±4.80)% was higher than that in empty vector group (12.47±3.30)% and non-transfected group (15.97±2.50)%. The number of LSECs migration in HO-1 group (258.70±36.56) was higher than that in empty vector group (122.00±38.16) and non-transfected group (107.70±30.01). The protein and mRNA expression level of HO-1, Id1, HGF and Wnt2 in HO-1 group were higher than that in empty vector group and non-transfected group. EdU positive rate of hepatocytes that co-cultured with LSECs in HO-1 group (18.33±2.52) % was higher than that in empty vector group (11.33±1.53)% and non-transfected group (11.7±2.08)%. The differences were statistically significant (all P<0.05). Conclusion:Up-regulation of HO-1 promoted LSECs proliferation and migration of, as well as up-regulation of HO-1 in LSECs enhanced the capacity of LSECs to promote hepatocyte proliferation.

Objective To investigate the protective effect of gastrodin (Gas) on the hepatic ischemia-reperfusion injury (HIRI) in mice,and study possible mechanisms.Methods Forty male C57BL/6 mice were randomly divided into sham-operated group (Sham),HIRI group,gastrodin-treated groups with low and high does (Gas-L,Gas-H,respectively),and cobalt protoporphyrin(CoPP) group(n =8).Mice in Gas-L,Gas-H,and CoPP groups were injected intraperitoneally with individual drugs and dose before operation.Except for Sham group,an 70％ volume HIRI model was established by means of 60 minutes ischemia and then 6 hours reperfusion in the other groups.The levels of serum AST and ALT in each group were compared.The levels of superoxide dismutase (SOD) and malondialdehyde (MDA) in the liver tissue were measured;The level of TNF-αt,IL-6,IL-10 and heme oxygenase-I (HO-1) mRNA level were detected by real time quantitative PCR.Liver tissue was stained with H&E.The hepatocyte apoptosis was studied by TUNEL.Results Respectively,the serum ALT level in the HIRI,Gas-L,Gas-H and CoPP groups was (5 057.34±290.80)U/L,(3 917.49±198.10) U/L,(3 645.63± 171.10) U/L,(2 977.78± 179.00) U/L,and the ALT level was (5 871.25 ± 819.01) U/L,(4 660.88 ± 505.96) U/L,(4 182.00 ± 507.51) U/L,(3 788.65±462.14)U/L.The serum level of ALT and AST in Gas-L,Gas-H and CoPP groups was lower than HIRI group (P＜ 0.05).The apoptotic index in Gas-L,Gas-H and CoPP groups respectively was (37.89±4.27)％,(32.59±3.78)％,(20.45-±2.49)％,which was lower than group (58.92±3.32)％ (P＜ 0.05).Compared with HIRI group,the TNF-α and IL-6 mRNA level in Gas-L,Gas-H and CoPP groups were decreased,and the levels of IL-10 and HO-1 mRNA were increased.Simultaneously,the activity of SOD was promoted,whereas the levels of MDA was reduced (P＜0.05).Conclusions Gastrodin shows an protective function aganist liver ischemia-reperfusion injury in mice by inhibiting inflammation,oxidative stress and cell apoptosis.The up-regulation of HO-1 by gastrodin may be the possible mechanism.

Liver sinusoidal endothelial cells (LSECs) play a positive role in maintaining microcirculation of the liver to achieve liver homeostasis, and they also mediate a balance between regeneration and fibrosis in the process repair of liver injury. After acute liver injury, LSECs are the regulators of liver regeneration, while in chronic injury, abnormally activated LSECs promote development of liver fibrosis. In this paper, we summarized the recent progress in research on the balance between LSECs-mediated liver regeneration and fibrosis, with the aim to provide new ideas in treating liver fibrosis and promoting liver regeneration by targeting LSECs.

OBJECTIVE To screen the best compatibility ratio and administration conditions of Panax ginseng -Poria cocos pair against aging ，and investigate its mechanism . METHODS P. ginseng-P. cocos pair extracts with different compatibility ratios （1∶1，1∶2，2∶1，1∶4，4∶1，m/m）were prepared ；taking Saccharomyces cerevisiae as the aging model organism ，the S. cerevisiae growth curve was drawn by MTT method ，the best compatibility ratio ，administration concentration and administration time point of P. ginseng-P. cocos pair were screened out ；the activities of antioxidant related enzymes [superoxide dismutase （SOD），peroxidase （POD），catalase（CAT）]，the levels of reactive oxygen species （ROS）and malondialdehyde （MDA），the content of adenosine triphosphate（ATP），and the mitochondrial membrane potential （MMP）in S. cerevisiae cells were detected ；mRNA expressions of SOD1，CTT1，GSH1，ATP1，MRS1 and CDC 19 were also detected . RESULTS The optimal ratio of P. ginseng-P. cocos pair for anti-aging activity was 1∶4（m/m），the optimal administration concentration was 220 μg/mL，and the optimal administration time point was the 28th hour . The extracts of P. ginseng-P. cocos pair（1∶4，m/m）could significantly increase the activities of SOD ， POD and CAT ，ATP content ，MMP，mRNA expression of CTT 1，GSH1 and MRS 1（P＜0.01），but decrease the levels of MDA and ROS ，mRNA expressions of SOD 1，ATP1 and CDC 19（P＜0.05 or P＜0.01）. CONCLUSIONS P. ginseng-P. cocos pair（1∶4， m/m）has a good anti -aging effect on S. cerevisiae ，its mechanism may be related to the positive regulation of oxidative stress and energy metabolism of S. cerevisiae cell.