Cyanidin attenuates pressure overload-induced cardiac remodeling in mice via mitochondrial fusion enhancement

AIM To investigate the effect of cyanidin （Cyn） on pressure overload-induced cardiac remodeling and the underlying mechanism. METHODS Six-week-old male C57BL/6 mice （n=120） were divided into 4 groups： sham group （n=20）, sham+Cyn group （n=20）, transverse aortic constriction （TAC） group （n=40） and TAC+Cyn group （n=40）. The model of cardiac chronic pressure overload was induced by TAC, and the first day of TAC was defined as day 0. The animals in sham+Cyn group and TAC+Cyn group were treated with Cyn dissolved in DMSO and normal saline （5 mg·kg^-1·d^-1） for 5 d before TAC, while the animals in sham group and TAC group were treated with the same amount of DMSO and normal saline. Four weeks after TAC, the survival rate of the animals in each group was analyzed, the heart function of the mice was measured by ultrasound echocardiography, and the heart weight/body weight and lung weight/body weight were calculated. The cross-sectional area of the cardiomyocytes was measured by wheat germ agglutinin staining and hematoxylin-eosin staining. The degree of cardiac oxidative stress was evaluated by dihydroethidium staining and measurement of superoxide dismutase （SOD） and malondialdehyde （MDA） levels. The cardiomyocyte apoptosis was detected by TUNEL method. The mRNA expression levels of atrial natriuretic peptide （ANP）, brain natriuretic peptide （BNP） and β-myosin heavy chain （β-MHC） were detected by RT-qPCR, and the protein expression levels of Bax, Bcl-2, optic atrophy protein 1 （OPA1） and dynamin-related protein 1 （Drp1） were determined by Western blot. The mitochondrial morphological changes were observed by transmission electron microscopy. RESULTS Compared with TAC group, the survival rate of the mice in TAC+Cyn group was significantly increased （P<0.05）, the myocardial apoptosis, the cross-sectional area of myocardial cells, the heart weight/body weight, the lung weight/body weight, the level of reactive oxygen species and the MDA content were decreased （P<0.05）, and the SOD was activated （P<0.05）. M-mode ultrasound tests showed that Cyn treatment significantly increased left ventricular ejection fraction and left ventricular fractional shortening in the mice after TAC （P<0.05）, while left ventricular end-diastolic diameter and left ventricular posterior wall thickness in diastole were reduced （P<0.05）. Transmission electron microscopic observation showed that the number of myocardial mitochondria was increased and the mitochondrial area was decreased after TAC （P<0.05）, while treatment with Cyn increased the area of myocardial mitochondria and decreased the mitochondrial number （P<0.05）. Compared with sham group, the protein level of OPA1 in TAC group was significantly reduced （P<0.05）, while treatment with Cyn significantly increased the protein level of OPA1. CONCLUSION Cyanidin significantly increases the survival rate, improves the cardiac function and attenuates the cardiac remodeling of the mice after TAC. The mechanism may be related to the inhibition of myocardial mitochondrial OPA1 cleavage and the promotion of mitochondrial fusion.     

Scutellarin affects LPS-induced oxidative stress and apoptosis of glomerular epithelial cells by up-regulating miR-7-5p

AIM To investigate the effect of scutellarin （SCU） on oxidative stress and apoptosis induced by lipopolysaccharide （LPS） in human glomerular epithelial cells and its mechanism. METHODS Human glomerular epithelial cells were cultured in vitro， and were treated with LPS （1.0 mg/L） to establish a cell injury model. The cells were divided into normal control （NC） group， LPS group， NC+SCU group， LPS+SCU group， LPS+miR-NC group， LPS+microRNA-7-5p （miR-7-5p） group， LPS+SCU+anti-miR-NC group and LPS+SCU+anti-miR-7-5p group. Cell viability was detected by CCK-8 assay. Apoptosis was detected by flow cytometry. The intracellular malondialdehyde （MDA） content and superoxide dismutase （SOD） activity， and lactate dehydrogenase （LDH） activity in the cell culture supernatant were determined by kit. RT-qPCR was used to detect the expression level of miR-7-5p. RESULTS Compared with NC group， the cell viability， miR-7-5p expression and SOD activity in LPS group were significantly reduced， and the apoptotic rate， MDA content and LDH activity were significantly increased （P<0.05）. Compared with LPS group， the cell viability， miR-7-5p expression and SOD activity in LPS+SCU group were significantly increased， and the apoptotic rate， MDA content and LDH activity were significantly reduced （P<0.05）. Compared with LPS+miR-NC group， the cell viability and SOD activity in LPS+miR-7-5p group were significantly increased， and the apoptotic rate， MDA content and LDH activity were significantly reduced （P<0.05）. Compared with LPS+SCU+anti-miR-NC group， the cell viability and SOD activity in LPS+SCU+anti-miR-7-5p group were significantly reduced， and the apoptotic rate， MDA content and LDH activity were significantly increased （P<0.05）. CONCLUSION Scutellarin inhibits LPS-induced oxidative stress damage and apoptosis in glomerular epithelial cells via up-regulating miR-7-5p expression.     

Role of retinoic acid X receptor in regulation of autophagy in rat alveolar type Ⅱ epithelial cells induced by hypoxia/reoxygenation

AIM To investigate the regulatory effect of retinoic acid X receptor （RXR） on autophagy induced by hypoxia/reoxygenation （H/R） in rat alveolar type Ⅱ epithelial cells （AEC Ⅱ） and its molecular mechanism. METHODS AEC Ⅱ were cultured in normoxia. The cells growing to logarithmic growth phase were randomly divided into 5 groups： （1） control （Con） group： cells were cultured for 30 h under normal operation； （2） H/R group： cells were cultured in hypoxia condition for 6 h and then in reoxygenation condition for 24 h； （3） DMSO group： cells were pretreated 1.5 h with medium containing less than 0.1% DMSO before modeling， and the rest were treated the same as the H/R group； （4） 9-cis-retinoic acid （9-RA） group： cells were pretreated for 1 h with 9-RA （100 nmol/L） before hypoxia； （5） HX531 group： cells were treated with 9-RA （100 nmol/L） for 0.5 h， then treatment with HX531 （2.5 μmol/L） for 1 h. CCK-8 assay was used to detect the cell viability. Immunofluorescence staining was used to observe the expression of RXRα. Transmission electron microscope was used to observe the changes of intracellular ultrastructure， and the mRNA expression of adenosine AMP-activated protein kinase （AMPK）， beclin 1， LC3， mammalian target of rapamycin （mTOR） and P62 was detected by RT-PCR. Western blot was used to detected the protein levels of p-AMPK， beclin 1， LC3-Ⅱ， p-mTOR and P62. RESULTS Compared with Con group， the cell viability in H/R， DMSO， 9-RA and HX531 groups were significantly decreased. The mRNA expression of AMPK， beclin 1 and LC3 was significantly increased， and the protein levels of p-AMPK， beclin 1 and LC3-Ⅱ were also increased. The mRNA expression of mTOR and P62 was decreased， and the protein levels of p-mTOR and P62 were also decreased （P<0.05）. The cell injury in 9-RA group was alleviated and autophagy level was significantly lower than that in H/R， DMSO and HX531 groups （P<0.05）， and no significant difference among H/R， DMSO and HX531 groups was observed （P>0.05）. CONCLUSION H/R induces autophagy of AEC Ⅱ. Activating RXR reduce the damage of AEC Ⅱ cells induced by H/R， and its mechanism may be related to the inhibition of autophagy.     

Quercetin activates mitochondrial autophagy via PINK1/parkin pathway to alleviate cerebral ischemia-reperfusion injury in rats

AIM To analyze the regulatory effect of quercetin （QUE） on PTEN-induced putative kinase 1 （PINK1）/parkin mitochondrial autophagy pathway, and to explore the mechanism of quercetin in relieving cerebral ischemia/reperfusion （I/R） injury. METHODS Sixty SD male rats were randomly divided into sham operation group, model group （I/R group）, QUE group,3-methyladenine （3-MA） group and QUE+3-MA group. Administration started in each group 3 days before modeling, once a day, at 30 min after the last administration,except sham group, the other groups used 4-vessel blockage method to establish the whole brain I/R model. On the day after modeling, the neural function was evaluated by neuropathy disability score （NDS）. The volume of cerebral infarction was measured by 2,3,5-triphenyltetrazolium chloride （TTC） staining. The morphological changes of mitochondria in hippocampus were observed by transmission electron microscopy. The contents of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α） in hippocampus were measured by ELISA. The activity of superoxide dismutase （SOD） and contents of malondialdehyde （MDA） in hippocampus were detected by xanthine oxidase method, thiobarbituric acid condensation method. Western blot was used to detect the proteinex pression of PINK1, parkin and LC3-II in brain tissue. RESULTS Compared with sham group, the hippocampus of the rats in I/R group and QUE+3-MA group showed swelling of mitochondria, destruction or disappearance of internal crista and other pathological damage,also the volume of cerebral infarction, the contents of IL-6, TNF-α and MDA, the protein expression levels of PINK1, parkin and LC3-II were increased （P<0.05）, while NDS score and activity of SOD were decreased （P<0.05）. Compared with I/R group and QUE+3-MA group, the pathological damage degree of hippocampus in QUE group was reduced, the volume of cerebral infarction, the contents of IL-6, TNF-α and MDA were decreased （P<0.05）, the proteinexpression levels of PINK1, parkin and LC3-II, and NDS score and activity of SOD were increased （P<0.05）.The above indexes in 3-MA group were opposite to QUE group. No significant difference in the above indexes between I/R group and QUE+3-MA group was observed （P>0.05）. CONCLUSION Quercetin activates mitochondrial autophagy and reduces cerebral I/R by regulating the expression of PINK1/parkin pathway proteins.     

Effect of lncRNA FEZF1-AS1 on viability and apoptosis of LPS-induced vascular endothelial cells by regulating miR-363-3p expression

AIM To investigate the mechanism of long noncoding RNA （lncRNA） FEZF1-AS1 regulating microRNA-363-3p （miR-363-3p） on the viability and apoptosis of lipopolysaocharide （LPS）-induced vascular endothelial cells. METHODS Human umbilical vein endothelial cells （HUVECs） were cultured in vitro. pcDNA-NC， pcDNA-FEZF1-AS1， anti-miR-NC， anti-miR-363-3p， miR-NC and miR-363-3p mimics were transfected into the HUVECs and LPS stimulation was applied for 24 h. RT-qPCR was used to detect the expression of FEZF1-AS1 and miR-363-3p. The cell viability was measured by MTT assay. The apoptotic rate was analyzed by flow cytometry. The dual-luciferase reporter experiment was used to verify the targeted regulation of FEZF1-AS1 and miR-363-3p. Western blot was used to determined the expression of cyclin D1， Ki67 and cleaved caspase-3. RESULTS Compared with control group， the expression level of FEZF1-AS1 in LPS group was significantly reduced （P<0.05）， and the expression level of miR-363-3p was significantly increased （P<0.05）. Compared with pcDNA-NC+LPS group， the cell viability in pcDNA-FEZF1-AS1+LPS group was significantly increased （P<0.05）， the apoptotic rate was significantly reduced （P<0.05）， the protein levels of cyclin D1 and Ki67 were significantly increased （P<0.05）， and the protein level of cleaved caspase-3 was significantly reduced （P<0.05）. Compared with anti-miR-NC+LPS group， the cell viability in anti-miR-363-3p+LPS group was significantly increased （P<0.05）， the apoptotic rate was significantly reduced （P<0.05）， the protein levels of cyclin D1 and Ki67 were significantly increased （P<0.05）， and the protein level of cleaved caspase-3 was significantly reduced （P<0.05）. Dual-luciferase reporter experiment confirmed that FEZF1-AS1 targeted miR-363-3p. Compared with miR-NC+pcDNA-FEZF1-AS1+LPS group， the cell viability in miR-363-3p+pcDNA-FEZF1-AS1+LPS group was significantly reduced （P<0.05）， the apoptotic rate was significantly increased （P<0.05）， the protein levels of cyclin D1 and Ki67 were significantly reduced （P<0.05）， and the protein level of cleaved caspase-3 was significantly increased （P<0.05）. CONCLUSION Over-expression of FEZF1-AS1 promotes the viability and inhibits apoptosis of LPS induced vascular endothelial cells by inhibiting the expression of miR-363-3p.     

Role and potential mechanism of fecal microbiota transplantation in treatment of chronic hepatitis B

AIM To investigate the effect of fecal microbiota transplantation （FMT） on the treatment of chronic hepatitis B （CHB） and the potential mechanism. METHODS Fifty C57BL/6J mice （6~8 weeks old） were divided into 5 groups： control group， CHB group， entecavir （ETV） group， comprehensive treatment （ETV+FMT， EFMT） group， and blocker （TAK-242+ETV+FMT， EFMT-TAK） group. The mice in each group were given corresponding treatment. The general condition of the mice was observed daily， and fecal specimens were kept every 10 d. The mice were sacrificed after 12 weeks， and the liver tissues and blood samples were collected. HE staining was used for histological scoring. Serum hepatitis B surface antigen （HBsAg） and interleukin-18 （IL-18） levels were measured by ELISA. Toll-like receptor 4 （TLR4） expression was detected by flow cytometry. Intestinal flora diversity was analyzed by high-throughput sequencing. RESULTS （1） Compared with control group， the body weight of the mice in CHB group was significantly reduced （P<0.05）. The body weight loss of the mice in ETV group， EFMT group and EFMT-TAK group was reversed to some extent as compared with CHB group （P<0.05）. （2） The histological score of the mice in CHB group was significantly higher than that in control group （P<0.05）. The score in ETV group was lower than that in CHB group （P<0.05）. The scores in EFMT group and EFMT-TAK group were lower than that in ETV group （P<0.05）， and that in EFMT-TAK group had a further downward trend compared with EFMT group （P<0.05）. （3） Compared with control group， the serum level of HBsAg in the CHB mice was significantly increased （P<0.05） and decreased after ETV treatment （P<0.05）. The HBsAg level in both EFMT group and EFMT-TAK group was significantly lower than that in ETV group （P<0.05）. （4） The IL-18 level in CHB group was significantly higher than that in control group （P<0.05）. After ETV treatment， the IL-18 level was decreased （P<0.05）， and that in both EFMT group and EFMT-TAK group was decreased more than that in ETV group （P<0.05）. （5） TLR4 expression in CHB group was higher than that in control group （P<0.05）， that in ETV group was lower than CHB group （P<0.05）， and that in EFMT group was further decreased （P<0.05）. （6） The heat map analysis at the class level showed that the abundances of Gammaproteobacteria， Deltaproteobacteria and Negativicutes in CHB group were significantly higher than those in control group， and those of Deltaproteobacteria and Negativicutes in EFMT group were close to those in control group. The heat map analysis at the family level indicated that the abundances of Burkholderiaceae， Desulfovibrionaceae and Veillonellaceae in CHB group were significantly higher than those in control group， while those in ETV group and EFMT group gradually approached normal levels. The α diversity index in CHB group was significantly decreased， while the diversity in ETV group was increased， that in EFMT group was further increased， and that in EFMT-TAK group was the highest. CONCLUSION FMT plays an active role in the treatment of CHB. The mechanism may be related to reducing the level of IL-18 and improving the structure and diversity of intestinal flora. The TLR4 signaling pathway is involved.     

Secoisolariciresinol diglucoside attenuates renal inflammatory injury of mice induced by chronic intermittent hypoxia via inhibiting TXNIP and activating NLRP3 inflammasome

AIM To investigate the effectof flax lignan/secoisolariciresinol diglucoside （SDG） on the inflammatory damage of kidney induced by chronic intermittent hypoxia （CIH）. METHODS C57BL/6N mice were divided into normal （control） group， model （CIH） group and treatment （SDG） group. The changes of the body weight was recorded. Hematoxylin-eosin （HE） staining was used to observe the morphological alterations in the renal tissues. The levels of serum creatinine and blood urea nitrogen were measured by a biochemical analyzer. Hydroxylamine and thiobarbituric acid methods were used to detect the activity of superoxide dismutase （SOD） and the content of malondialdehyde （MDA） in the renal tissues. The protein levels of thioredoxin-interacting protein （TXNIP） and nucleotide-binding oligomerization domain-like receptor protein 3 （NLRP3） were detected by immunohistochemical staining， while those of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6） and IL-1β were measured by ELISA. The protein levels of TXNIP， NLRP3， apoptosis-associated speck-like protein containing a CARD （ASC）， caspase-1， IL-1β and IL-18 in the renal tissues were also determined by Western blot. RESULTS No significant difference in the body weight and kidney index among the 3 groups was observed （P>0.05）. HE staining showed the swollen epithelial cells of renal tubules with vesicular degeneration， and irregular glomerular morphological change in CIH group， while SDG treatment attenuated the above changes. Compared with control group， the levels of serum creatinine， TNF-α， IL-6 and IL-1β were significantly increased in CIH group （P<0.05）. The significantly increased expression levels of NLRP3 and TXNIP in the cytoplasm of renal tubular epithelial cells in CIH group were detected by immunohistochemical staining. Compared with control group， the activity of SOD was decreased， the content of MDA was increased in CIH group， and the protein expression levels of TXNIP， NLRP3， ASC， caspase-1， IL-1β and IL-18 were up-regulated and then decreased after SDG treatment （P<0.05）. CONCLUSION SDG attenuates the renal inflammatory damage of the mice induced by CIH， and its mechanism may be associated with the inhibition of oxidative stress and activation of NLRP3 inflammasome.     

Effects of oxidative stress and inflammatory response on kidney injury in mice with hyperthyroidism

AIM To explore the effects of oxidative stress and inflammatory response on kidney injury induced by hyperthyroidism in mice. METHODS Forty male Kunming mice were randomly divided into control group （n=20） and L-thyroxine （T4） group （n=20）. The mice in T4 group were intraperitoneally injected with T4 diluent at a dose of 1 mg/kg to induce hyperthyroidism， and those in control group were injected with normal saline of the same volume. After 7 weeks， the mice were weighed and dissected， the kidneys were removed and weighed， and the length of tibia was also measured. The activity of superoxide dismutase （SOD） and the content of malondialdehyde （MDA） in the kidney tissues were detected. The pathological changes of the kidney tissues were observed by HE staining. The levels of 4-hydroxynonenal （4-HNE）-modified proteins， interleukin-1 receptor-associated kinase 1 （IRAK1） and tumor necrosis factor receptor-related factor 6 （TRAF6） were determined by Western blot and immunohistochemistry. RESULTS Compared with control group， the body weight of the mice was decreased， while the kidney size and weight were increased significantly in T4 group. In addition， the ratios of kidney weight/body weight and kidney weight/tibia length were also increased （P<0.05）. In T4 group， the renal tubules were enlarged， and the epithelial cells of renal tubules were swollen and exfoliated， with vacuolar degeneration. Furthermore， reduced SOD activity， and increased MDA content and 4-HNE-modified proteins were found in T4 group， all of which were related to oxidative stress （P<0.05）. The levels of inflammation-related proteins IRAK1 and TRAF6 were significantly increased in T4 group （P<0.05）. CONCLUSION Excessive T4 may lead to kidney hypertrophy and injury in mice， and the mechanism may be related to oxidative stress and inflammatory response.     

Geniposide attenuates cognitive dysfunction in sleep-deprived rats by inhibiting TLR4/NF-κB signaling pathway

AIM To investigate the effects of geniposide （Gen） on Toll like receptor 4/nuclear factor-κB （TLR4/NF-κB） signaling pathway and cognitive dysfunction in sleep deprived rats. METHODS Wistar rats （n=120） were randomly divided into normal control （NC） group， model （M） group， low-dose （5 g·kg^-1·d^-1） Gen （Gen-L） group， medium-dose （10 g·kg^-1·d^-1） Gen （Gen-M） group， high-dose （20 g·kg^-1·d^-1） Gen （Gen-H） group and Gen-H+LPS （0.4 mg·kg^-1·d^-1， tail vein injection） group. After 7 days of intervention， the sleep deprivation model of rats in M group， Gen-L， Gen-M， Gen-H and Gen-H+LPS group was established by improved small platform water environment. The escape latency of Morris water maze experiment and the behavior correct rate of Y maze experiment were measured. The serum levels of S100B and neuron-specific enolase （NSE）， and the levels of interleukin-1β （IL-1β）， IL-6 and tumor necrosis factor-α （TNF-α） in hippocampus were detected by ELISA. The mRNA levels of TLR4 and NF-κB p65 were detected by RT-qPCR， and the protein levels of TLR4 and NF-κB p65 were determined by Western blot. RESULTS Compared with NC group， the escape latency， the serum levels of S100B and NSE， the hippocampal levels of IL-1β， IL-6 and TNF-α， and the mRNA and protein expression of TLR4 and NF-κB p65 were increased significantly in M group （P<0.01）， and the behavior correct rate was decreased significantly （P<0.01）. Compared with M group， the escape latency， the hippocampal levels of IL-1β， IL-6 and TNF-α， and the expression of TLR4 and NF-κB p65 at mRNA and protein levels were decreased significantly in Gen-L， Gen-M and Gen-H groups （P<0.01）， and the behavior correct rate was increased in turn （P<0.01）. Compared with Gen-H group， the escape latency， the serum levels of S100B and NSE， the hippocampal levels of IL-1β， IL-6 and TNF-α， and the expression of TLR4 and NF-κB p65 at mRNA and protein levels were increased significantly in Gen-H+LPS group （P<0.01）， and the behavior correct rate was decreased significantly （P<0.01）. CONCLUSION Geniposide may inhibit the TLR4/NF-κB p65 signaling pathway to effectively improve cognitive function in sleep-deprived rats and reduce hippocampus inflammation.     

Alleviating effect of exenatide on ectopic lipid accumulation in skeletal muscle of ob/ob mice is independent on reducing body weight

AIM To investigate the alleviating effect of exenatide （Exe）， a glucagon-like peptide-1 （GLP-1） receptor agonist， on the ectopic lipid accumulation in skeletal muscle of ob/ob mice and its mechanism. METHODS Eight-week-old male ob/ob mice and their wild-type （WT） littermates were randomly divided into 3 groups， ob/ob group， ob/ob+Exe group and WT group， and treated with Exe at 24 nmol/kg or the same volume of saline intraperitoneally once daily for 4 weeks. The body weight， fasting blood glucose （FBG） and fat content were measured after the 4-week treatment. The oil red O staining and the quantification of triglyceride （TG） were performed on the skeletal muscle. The serum levels of TG， total cholesterol and free fatty acid （FFA） were also measured by ELISA. The expression levels of AMP-activated protein kinase （AMPK） and lipid metabolism-related proteins were determined by Western blot. Mouse myoblast C2C12 cells were used as an in vitro model to further investigate the effects of Exe. RESULTS As compared with the ob/ob mice treated with saline， 4-week Exe treatment did not reduce body weight， FBG， food intake and fat content in ob/ob mice （P>0.05）. However， serum FFA was decreased （P<0.05）. Oil red O staining and the quantification of TG showed that 4-week Exe treatment significantly attenuated the ectopic lipid accumulation in the skeletal muscle of ob/ob mice （P<0.05）. The results of Western blot showed that the levels of phosphorylated AMPK （p-AMPK） and lipolysis-related proteins were up-regulated， while the lipid synthesis-related proteins were down-regulated by Exe （P<0.05）. Treatment with Exe alleviated the lipid accumulation in the C2C12 cells induced by sodium palmate （P<0.05）， and the effects of Exe on the levels of p-AMPK and lipid metabolism-related proteins in the C2C12 cells were consistent with those in the ob/ob mice （P<0.05）. Treatment with Exe also up-regulated the protein expression of glucose transporter 4 and improved the ability of glucose uptake in the C2C12 cells （P<0.05）. CONCLUSION Short-term Exe treatment attenuates the ectopic lipid accumulation in skeletal muscle of ob/ob mice by up-regulating lipolysis-related proteins and down-regulating lipid synthesis-related proteins， which is independent on body weight loss.     

Effect of continuous renal replacement therapy on mRNA expression of autophagy-related molecules and prognosis in patients with acute kidney injury

AIM To explore the effect of continuous renal replacement therapy （CRRT） on mRNA expression of autophagy-related molecules and the prognosis in the patients with acute kidney injury （AKI）. METHODS A total of 174 patients from our hospital who were diagnosed to have AKI and underwent CRRT between February 2015 and March 2018 were involved in this study. The plasma levels of interleukin-1β （IL-1β） and interleukin-6 （IL-6）， the serum creatinine （SCr） level， and the mRNA expression levels of autophagy-related molecules， including microtubule-associated protein 1 light chain 3-II （LC3-II）， autophagy-related protein 5 （Atg5） and beclin-1， in the monocytes from peripheral blood were compared before and after CRRT. According to the survival of AKI patients after 4 weeks of CRRT， the enrolled patients were divided into death group （n=43） and survival group （n=131）， and the mRNA expression levels of the above molecules were compared between the 2 groups. RESULTS After CRRT treatment， the plasma levels of IL-1β and IL-6， the level of SCr， and the mRNA expression levels of LC3-II， Atg5 and beclin-1 in the monocytes were significantly lower than those before CRRT treatment （P<0.05）. The mRNA expression levels of LC3-II， Atg5 and beclin-1 in death group were significantly higher than those in survival group （P<0.05）. The positive correlation between SCr and IL-1β， IL-6， LC3-II or beclin-1 was observed （P<0.05）， and no correlation between SCr and Atg5 was found （P>0.05）. CONCLUSION CRRT decreases the mRNA expression levels of autophagy-related molecules in the patients with AKI and reduces the autophagy activity， which is protective for the patients. These autophagy-related molecules may be applied as a potential markers to predict the prognosis of CRRT.     

Inhibitory effect of Wendan decoction on formation of foam cells induced by ox-LDL

AIM To explore the anti-atherosclerotic mechanism of Wendan decoction based on formation of foam cells. METHODS The optimal concentrations of Wendan decoction without cytotoxity to cells were selected by MTT assay. After Wendan decoction treatment， the formation of foam cells was examined by oil red O staining. The cholesterol efflux， cholesterol level， free cholesterol level and cholesterol esterification rate were analyzed using cholesterol efflux assay， total cholesterol assay and free cholesterol assay. The expression levels of macrophage membrane proteins， including CD36， scavenger receptor class A （SR-A）， ATP-binding cassette transporter A1 （ABCA1） and scavenger receptor class B type I （SR-BI）， were quantified by Western blot. RESULTS The optimal concentrations of Wendan decoction without cytotoxity to the cells were 0~6 g/L. Wendan decoction at the concentrations of 1.5， 3 and 6 g/L were selected for the experiments. Wendan decoction at these concentrations inhibited the formation of foam cells induced by oxidized low-density lipoprotein （ox-LDL）， and reduced the accumulation of intracellular lipids in a concentration-dependent manner （P<0.05 or P<0.01）. Wendan decoction also reduced intracellular total cholesterol level， cholesterol ester level and cholesterol esterification rate （P<0.05 or P<0.01）， promoted efflux of intracellular cholesterol （P<0.01）， and decreased the protein level of CD36 in THP-1 cell-derived macrophages （P<0.01） in a concentration-dependent manner. Wendan decoction at the concentration of 6 g/L significantly reduced the protein level of SR-A in THP-1 cell-derived macrophages （P<0.05）. At the concentrations of 3 and 6 g/L， Wendan decoction significantly increased the protein levels of ABCA1 and SR-BI in THP-1 cell-derived macrophages （P<0.05 or P<0.01）. CONCLUSION Wendan decoction significantly inhibits ox-LDL-induced formation of foam cells by reducing cholesterol deposition and promoting cholesterol efflux， and its mechanism may be related to the down-regulation of CD36 and SR-A and the up-regulation of ABCA1 and SR-BI.     

Effects of eosinophils deletion on sPLA2-X in bronchial asthma model mice

AIM To investigate the association between soluble phospholipase A2-X（sPLA2-X） and eosinophils in bronchial asthma， and to provide new insight and strategies for the treatment of bronchial asthma. METHODS Female Babl/c mice （n=48） of SPF grade and 6~8 weeks old were divided into 4 groups （with 12 in each group： healthy control group，asthma control group， eosinophil deletion group， and asthma /eosinophil deletion isotype control group）. The mouse model of bronchial asthma was constructed. The mice in healthy control group were intraperitoneally injected with saline on days 0， 7， and 14. The mice in other groups were intraperitoneally injected with 50 μg OVA and 2 mg aluminum hydroxide gel（soluble in 200 μL saline.On the 21st d and 26 th d， eosinophil deletion antibody （anti-CCR3） and isotype control were intraperitoneally injected and intranasally respectively， and then the lungs function test was conducted within 48 h after the end of nebulization.Half of the mice in each group were subjected to whole lung lavage， the remaining half were used for lung tissue section with HE staining， the whole blood was used to measure serum IgE， the supernatant of broncho alveolar lavage fluid （BALF） was used to measure cytokines， and total number of cells in the bronchoalveolar lavage fluid was analyzed for cell classification and flow cytometry. RESULTS （1）Compared with asthma control group，the airway and alveolar inflammatory responses in asthma/eosinophil deletion group was significantly alleviated.（2） Compared with asthma control group， anti-CCR3 successfully deleted eosinophils， and the percentage of eosinophils in bronchoalveolar lavage fluid in asthma/eosinophil deletion group was significantly reduced （P<0.05）.（3） The airway hyperresponsiveness in asthma/eosinophil deletion group was significantly decreased as compared with asthma control group（P<0.05）.（4） The levels of sPLA2-X in the serum and BALF was significantly reduced in asthma/eosinophil deletion group as compared with asthma control group（P<0.05）.（5）Compared with asthma control group，the levels of IL-4， IL-5， and IL-13 in the BALF of the mice in asthma/eosinophil deletion group were significantly reduced， and the serum level of IgE was also decreased （P<0.05）. CONCLUSION Eosinophils in bronchial asthma are importantly associated with sPLA2-X.     

Niflumic acid inhibits the proliferation,migration and invasion of glioma U87 cells

AIM To investigate the effect of niflumic acid （NFA） on human glioma U87 cells and to clarify the potential mechanism. METHODS The U87 cells were cultured in vitro and divided into blank control group， and 50， 100 and 200 μmol/L NFA groups. MTT assay was performed to determine the viability of cells in various groups. Migration and invasion abilities were measured by real-time cell analysis （RTCA）. RESULTS The results of MTT assay showed that compared with blank control group， the viability of U87 cells was increased after treatment with NFA for 12 h （P<0.05 or P<0.01）， while the viability was significantly decreased after treatment with NFA for 24 and 48 h （P<0.05 or P<0.01） in a concentration-dependent manner. The results of RTCA showed that compared with control group， the cell migration and invasion abilities were inhibited in 100 and 200 μmol/L NFA groups （P<0.05 or P<0.01） and the inhibitory effects were more obvious in 200 μmol/L NFA group （P<0.01）. CONCLUSION NFA inhibits the viability， migration and invasion of human glioma U87 cells.     

Effect of Tripterygium wilfordii multiglucoside on intestinal flora and immune function in IgA nephropathy rats based on C1GALT1/Cosmc pathway

AIM To investigate the effects of Triptergium wilfordii multiglucoside （TWM） on intestinal flora and immune function in IgA nephropathy （IgAN） rats based on core 1 β1，3-galactosyltransferase （C1GALT1） and its chaperone protein Cosmc （C1GALT1/Cosmc pathway）. METHODS The rat model of IgAN was established， and the animals were randomly divided into model group （IgAN group）， dexamethasone （Dex） group and TWM group. Normal rats served as normal control （NC） group. The levels of serum creatinine （SCr） and blood urea nitrogen （BUN）， 24-hour urinary total protein （24 h UTP） and the number of urinary red blood cells were measured by automatic biochemical analyzer. The levels of serum IgA1， and plasma tumor necrosis factor-α （TNF-α）， B-cell activating factor （Baff） and interleukin-17 （IL-17） were detected by ELISA. The level of galactose-deficient IgA1 （Gd-IgA1） was detected by Vicia villosa lectin affinity ELISA. The intestinal colony was cultured in selective bacterial medium. The ratio of CD4⁺ CD25⁺ regulatory T cells （Treg） to CD4⁺ T cells （Treg proportion） in peripheral blood mononuclear cells （PBMC） was detected by flow cytometry.Western blot was used to determine the protein expression of C1GALT1 and Cosmc in intestinal mucosa. RESULTS Compared with NC group， 24 h UTP， the number of urinary red blood cells， SCr， BUN， serum IgA1 and Gd-IgA1， the numbers of Enterobacteriaceae， Enterococcus and Bacteroides， and the levels of TNF-α， Baff and IL-17 in plasma in IgAN group were significantly increased （P<0.05）， while the numbers of Bifidobacteria and Lactobacilli， the Treg proportion in PBMC， and the protein expression levels of C1GALT1 and Cosmc in intestinal mucosa were significantly decreased （P<0.05）. Compared with IgAN group， 24 h UTP， the number of urinary red blood cells， SCr， BUN， serum IgA1 and Gd-IgA1， the numbers of Enterobacteriaceae， Enterococcus and Bacteroides， and the levels of TNF-α， Baff and IL-17 in plasma in Dex group and TWM group were significantly reduced （P<0.05）， and those in TWM group were lower than those in Dex group （P<0.05）. Moreover， the numbers of Bifidobacteria and Lactobacilli， the Treg proportion in PBMC， and the protein expression levels of C1GALT1 and Cosmc in intestinal mucosa were significantly elevated （P<0.05）， and those in TWM group were higher than those in Dex group （P<0.05）. CONCLUSION TWM reduces the abnormal glycosylation level of IgA in IgAN rats by promoting the activation of C1GALT1/Cosmc pathway， and attenuates the intestinal flora disorder and immune dysfunction in IgAN rats， thus exerting the therapeutic effect.     

Curcumin attenuates lipopolysaccharide/D-galactosamine-induced acute rat liver injury via activating autophagy of hepatocytes

AIMTo investigate the effect of curcumin (CUR) on autophagy of hepatovyte in rats with lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced acute liver injury (AHI). METHODSThe healthy male Sprague-Dawley (SD) rats were randomized into the control group,AHI group,CUR group, 3-methy-ladenine (3-MA) group and 3-MA+CUR group, with 6 rats in each group. AHI was induced with an intraperitoneal injection of LPS and D-GalN. Liver function was tested 12 h after LPS/D-GalN treatment. Pathological changes of liver tissues were analyzed by HE staining.The amount of autophagic bodies were observed by transmission electron microscopy. The protein levels of autophagy related-proteins LC3 and beclin 1 in livers were detected by Western blot. ELISA were used to examine the serum levels of tumor necrosis factor-α (TNF-α). RESULTSCompared with control group, the serum level of alanine aminotransferase (ALT) and asparated aminotransferase (AST) were significantly increased, hepatic pathological damage were aggravated and serum TNF-α level was significantly increased in AHI group, while the autophagic bodies and the protein levels of LC3 and beclin 1 were increased (P<0.01). Compared with AHI group, the serum level of ALT and AST were significantly decreased, hepatic pathological damage were attenuated and serum TNF-α level was significantly reduced (P?<0.05), while the autophagic bodies and the protein levels of LC3 and beclin 1 were significantly increased in CUR group (P<0.01). Compared with CUR group, the serum level of ALT and AST were significantly increased, hepatic pathological damage were aggravated and serum level of TNF-α was significantly increased in 3-MA group and 3-MA+CUR group, while the autophagic bodies and the protein levels of LC3 and beclin 1 were decreased (P<0.01). CONCLUSION Curcumin protects rats against LPS/D-GalN-induced liver injury, partially due to activation of hepatocyte autophagy in livers.     

Effects of EGCG on GLUT2/G6PD/GS expression and blood glucose level in type 2 diabetic rats

AIM To investigate whether epigallocatechin gallate （EGCG） improves blood glucose in type 2 diabetic rats through glucose transporter 2 （GLUT2）-glucose-6-phosphate dehydrogenase （G6PD）-glycogen synthase （GS） pathway. METHODS Type 2 diabetes mellitus （T2DM） model was established in male Sprague-Dawley （SD） rats by feeding with high-fat diet and injection of streptozotocin （STZ）. The rats were divided into 5 groups （n=10）： control （Con） group， T2DM model （M） group， metformin （Met； 200 mg/kg， ig） group， T2DM+low-dose （50 mg/kg， ig） EGCG （EL） group， and T2DM+high-dose （100 mg/kg， ig） EGCG （EH） group. Diabetic rats were given drugs for 8 weeks. After 8 weeks of administration， the rats were killed， and the blood and liver tissues were collected. The levels of fasting blood glucose （FBG）， fasting serum insulin （FINS） and serum glycosylated hemoglobin were measured by biochemical tests. Liver glycogen were test by periodic acid-Schiff （PAS） staining. The mRNA expression of G6PD in the liver was detected by real-time PCR. The protein levels of GS and GLUT2 were determined by Western blot and immunohistochemistry. RESULTS T2DM rat model was established successfully. Compared with Con group， the levels of FBG， FINS and serum glycosylated hemoglobin in M group were increased significantly （P<0.05）， while the insulin sensitivity index （ISI）， the liver glycogen， the G6PD mRNA expression， and the protein levels of GS and GLUT2 were decreased significantly （P<0.05）. Compared with M group， the levels of FBG and serum glycosylated hemoglobin in Met group and EH group were decreased significantly （P<0.05）， while the ISI， the liver glycogen， the G6PD mRNA expression， and the protein levels of GS and GLUT2 were increased significantly （P<0.05）. CONCLUSION EGCG reduces the blood glucose level in T2DM rats， which may be related to the regulation of GLUT2-G6PD-GS signaling pathway.     

Expression of histone chaperone ASF1B in prostate cancer cells and its effect on cell viability in vitro

AIM To investigate the expression of histone chaperone anti-silencing function 1B （ASF1B） in prostate cancer cells and its effect on cell viability in vitro. METHODS Human prostate cancer PC-3 cells were used， and knockdown of ASF1B was conducted by small interfering RNA （siRNA） transfection into the cells. The cells were divided into control group， siRNA negative control vector （mock） group and siRNA-ASF1B group. The viability of the PC-3 cells treated with ASF1B-siRNA for 12， 24 and 48 h was measured by MTT assay. Flow cytometry was used to analyze cell apoptosis and cell cycle distribution. The mRNA expression of apoptosis-related molecules was detected by RT-qPCR， and the expression levels of MAPK/JNK/ERK signaling pathway-related proteins were determined by Western blot. RESULTS The protein level of ASF1B in the normal cells （benign prostatic hyperplasia） was significantly lower than that in the PC-3 cells （P<0.01）. Compared with control group and mock group， the protein expression level of ASF1B in the PC-3 cells transfected with siRNA-ASF1B plasmid and the viability of the PC-3 cells were significantly decreased （P<0.01）. The apoptotic rate of the PC-3 cells transfected with siRNA-ASF1B plasmid was significantly higher than that in control group （P<0.01）， and the cell cycle was arrested at G₁ phase. The mRNA levels of p53， caspase-3， Bax and PARP-1 in the PC-3 cells transfected with siRNA-ASF1B plasmid were up-regulated compared with those in control group and Mack group （P<0.01）. In addition， the protein levels of MAP2K4 and p-JNK in the PC-3 cells in siRNA-ASF1B group were significantly higher than those in mock group （P<0.01）， while the protein level of p-ERK was significantly lower than that in mock group （P<0.01）. CONCLUSION ASF1B silencing induces G₁ arrest and promotes apoptosis of PC-3 cells. Activating MAPK/JNK/ERK signaling pathway may be a possible contributor to the anti-prostate cancer effect of siRNA-ASF1B.