Objective To investigate the role of autophagy in podocyte damage,and the intracellular mechanism of autophagy activation through passive Heymann nephritis (PHN) animal model.Methods Male Sprague-Dawley rats (n=40) were studied on day 0,2,4,7,and 21 after induction of PHN by injection of anti-Fx1A.Podocyte morphology and autophagosomes were observed by transmission electron microscopy.Podocyte numerical density was estimated by Weibel-Gomez =method.Cell apoptosis was detected by TUNEL assay and caspase-3 immunohistochemical staining.Expressions of autophagy markers and endoplasmic reticulum stress (ERS)-associated proteins were analyzed by Western blotting.Results (1) In PHN rats,immunohistochemical staining showed that C5b-9 deposited along glomerular basement membrane on day 4 to day 21.Small subepithelial electron -dense deposits and a part of foot process fusion were detected in the glomerulus of PHN rats on day 4 by transmission electron microscope,and podocyte damage was aggravated on day 21.Furthermore,compared with control,the urinary protein levels of PHN rats began to increase on day 3,and reached the top on day 21 [(50.6±6.0) mg/24 h].(2) The number of podocytes significantly decreased in PHN rats compared with control group on day 21(P ＜ 0.05).(3) In PHN rats,apoptotic podocytes were found by caspase-3 immunohistochemical staining and TUNEL assay on day 21.(4) The expression of autophagy marker LC3 Ⅱ was markedly increased on day 7 and 21,but down-regulated on day 21 compared with day 7.Moreover,accumulated autophagosomes in podocytes were detected on day 7 and 21 by transmission electron microscope.(5) The level of GRP78 was significantly increased on day 2 and 7 but reduced to baseline on day 21.At the same time,the downstream pathways (ATF6α,p-PERK and p-JNK) of unfolded protein response were also up-regulated in the early process of PHN and down-regulated later.Conclusions Autophagy is an important way to protect against immunemediated podocyte injury in membranous nephropathy.Autophagy activation is mainly related to endoplasmic reticulum stress induced by complement attack.This provides an important basis for a thorough understanding of the role of autophagy in the process of podocyte damage and the pathogenesis of membranous nephropathy.

Objective To determine the effect of rapamycin on sublytic C5b-9-induced podocyte adhesion damage,and whether autophagy is involved in this progression.Methods Sublytic complement C5b-9 stimulation was used in vitro.Autophagosomes were viewed using electron microscopy.Western blotting was used to measure the change of autophagy-related markers.Attachment assay was used to assess the adhesion of podocyte.Confocal microscopy was used to explore the expression patterns of cytoskeletal protein F-actin.Flow cytometry was used to measure the level of adhesion-associated protein integrin α3.Results (1) For ensuring sublytic complement injury,the maximal amounts of anti-podocyte antiserum and 160×-diluted normal human serum were used without inducing cell lysis (defined as ＞ 5％ LDH release).(2) Sublytic C5b-9 promoted autophagy in podocyte in vitro.The proautophagic effect of sublytic C5b-9 manifested in the form of accumulated autophagosomes and enhanced expression of LC3-lⅡ.(3) Inhibition of autophagy by 3-methyadenine enhanced the effect of sublytic C5b-9-induced podocyte injury,including serious cytoskeleton damage and markedly reduced adhesion of podocyte.(4) Rapamycin treatment significantly improved the above lesions.(5) Rapamycin enhanced autophagy induced by sublytic C5b-9 in podocyte.Conclusions In summary,rapamycin can improve sublytic CSb-9-induced podocyte adhesion damage by appropriate autophagy activation.These findings provide important information for the development of appropriate protocols for the application of mTOR (mammalian target of rapamycin) inhibitors in podocytopathy.

Objective To study the effect of lipid peroxidation and anti-lipid peroxidation and the pigment in skin of C57BL/6 mice exposed to arsenic.Methods Forty male C57BL/6 mice were divided into four groups via the random number table method,ten mice in each group,and the mice were fed ad libitum drinking water containing arsenic at 0,1,5 and 25 mg/L concentrations for a period of 6 weeks,respectively.Twelve days before the end of the experiment,procedure of depilation was performed to induce anagen of hair cycle.On the 30th day of experiment,the activity of superoxide dismutase (SOD) was determined by nitrite method,the activity of glutathione peroxidase (GSH-Px) was determined by dithiobisobenzoic acid method,the content of malondialdehyde (MDA) in skin of C57BL/6 mice was determined by thiobarbituric acid method.Melanin was measured by NaOH dissolution method.Results No significant difference was found in body weight and water intaken between groups (F =0.119,0.363,P ＞ 0.05).The activity of SOD [(16.00 ± 5.05),(13.96 ± 2.02),(10.46 ± 3.14) U/mg prot] in 1,5 and 25 mg/L arsenic groups were all significantly lower than that in control group [(20.36 ± 4.82) U/mg prot,P ＜ 0.05].GSH-Px activity in 1,5 and 25 mg/L arsenic groups [(98.14 ± 23.92),(87.18 ± 10.87),(53.56 ± 19.97) U/mg prot] were all significantly lower than that in control group [(119.34 ± 33.14) U/mg prot,P ＜ 0.05].While MDA levels in 5 and 25 mg/L arsenic groups [(9.09 ± 2.04),(11.48 ± 2.21) nmol/mg prot] were significantly higher than that of control group [(6.19 ± 0.56) nmol/mg prot,P ＜ 0.05] and that of 1 mg/L arsenic group [(6.52 ± 1.67) nmol/mg prot,P ＜ 0.05).No significant difference was found in melanin levels (P ＞ 0.05).Conclusions Lipid peroxidation and the decreasing of antioxidation may be one of the mechanisms of arsenic-induced skin damage.Arsenic-induced skin melanin content change is not found.

The 6-min walk test(6MWT)is an effective tool to assess the cardiopulmonary function and exercise tolerance,and is widely used in clinical practice to assess the exercise capacity in the patients with car-diopulmonary disease.Pulmonary hypertension(PH)is a serious cardiovascular disease manifested by a con-tinuous increase in pulmonary circulatory pressure,which limits the right heart function and eventually devel-ops into the right heart failure,moreover its clinical manifestations lack specificity,making diagnosis difficult.With the development of PH diagnosis and treatment technology,the medical staffs have paid more and more attention to the application of 6MWT in the diagnosis,treatment and prognosis of the patients with PH.This article reviews the characteristics of 6MWT,its application in PH,and its limitations and future prospects in order to help clinic to better understand the role of 6MWT in the whole disease course of the patients with PH,and encourage the medical care staffs to broadly apply and improve 6MWT to make it to be more reliable clinical evidence.