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.     

CYP450 epoxygenase/EET pathway attenuates adipose inflammation of obese mice through HIF-1α

AIM To investigate the effects of cytochrome P450 （CYP450） epoxygenase/epoxyeicosatrienoic acid （EET） pathway on insulin resistance in obese mice, and to explore the possible mechanisms. METHODS High-fat diet-induced obesity model was established in C57BL/6Cnc mice, and the obese mice were randomly divided into 3 groups, including obesity group （treated with saline; n=10）, EET group （treated with 11,12-EET; n=10） and EET inhibitor 14,15-epoxyeicosa-5（Z）-enoic acid （EEZE） group （n=10）. Normal C57BL/6Cnc mice （n=10） treated with saline served as control. Protein expression of CYP2J2 （one of CYP450 epoxygenases） and hypoxia-inducible factor-1α （HIF-1α） was measured by Western blot. Vessel-like structure was detected by immunofluorescence staining. The serum levels of insulin, tumor necrosis factor-α （TNF-α）, interleukin （IL）-1β, IL-6 and monocyte chemoattractant protein-1 （MCP-1） were measured by ELISA. RESULTS In obese mice, homeostasis model assessment of insulin resistance （HOMA-IR） values were increased, the protein level of CYP2J2 was reduced, and the protein level of HIF-1α was increased in adipose tissues as compared with the controls （P<0.05）. The serum levels of MCP-1, IL-1β, IL-6 and TNF-α were also significantly increased in obese mice （P<0.05）. After treatment with 11, 12-EET, the HOMA-IR values were decreased compared with vehicle-treated obese mice, HIF-1α expression levels were decreased in the adipose tissue, and the serum levels of MCP-1, IL-1β, IL-6 and TNF-α were reduced （P<0.05）. Immunohistochemical results of adipose tissue from vehicle-treated obese mice showed a marked decrease in vessel-like structures （CD31-positive） compared with normal control mice （P<0.05）. EET treatment significantly increased the newly formed vessel-like structures in the visceral adipose tissues of obese mice as compared with vehicle-treated obese mice （P<0.05）. CONCLUSION High-fat diet-induced obesity and insulin resistance are closely related to the CYP450 pathway. Exogenous EETs effectively decrease obesity-induced insulin resistance possibly through pro-angiogenesis and attenuation of hypoxia and inflammation.     

Correlation between progression of atherosclerosis accelerated by emotional stimulation and imbalance of SDF-1/CXCR4 in mice

AIM To observe effects of emotional stimulation on expression of stromal cell-derived factor-1（SDF-1） and CXC chemokine receptor 4 （CXCR4） in plasma, platelets, aortas, hippocampus and bone marrow of apolipoprotein E gene knockout （ApoE^-/-） mice, and to reveal the possible mechanism of the aggravated atherosclerotic plaque vulnerability by emotional stimulation. METHODS Thirty 8-week-old male ApoE^-/- mice were randomly divided into normal control group, high fat group, and emotional stimulation group. Ten 8-week-old inbred C57BL/6J mice served as blank control group. After 12 weeks of intervention, the serum levels of SDF-1 and CXCR4 were investigated by ELISA. The protein levels of SDF-1 and CXCR4 in platelets, aortas, hippocampus and bone marrow were determined by Western blot. The pathological damage of aortas was observed by oil red O staining. RESULTS Compared with blank control group, normal control group and high fat group, the mice subjected to emotional stimulation showed more serious atherosclerosis in aortas detected by oil red O staining, and increased levels of SDF-1 and CXCR4 in the plasma and aortas were also observed （P<0.05）. The results of Western blot showed that the protein levels of SDF-1 and CXCR4 in platelets, aortas and hippocampus were increased in the mice subjected emotional stimulation, but the expression of SDF-1 and CXCR4 in the bone marrow was decreased （P<0.05）. CONCLUSION Imbalance of SDF-1/CXCR4 may be the key target by which emotional stimulation accelerates the progression of atherosclerosis.     

Influence of hypoxia-inducible factor-1 alpha on lung cancer cell A549 growth in vitro and in vivo

AIM: To study the role of hypoxia-inducible factor-1alpha(HIF-1α) on lung cancer cells A549 growth in vitro and in vivo. METHODS: To observe the growth rate of A549 cells after HIF-1α transfected, A549 cells (1×10⁶/mouse) were inoculated subcutaneously into 20 nude mice, which were randomly divided into two groups: the control group (group A, n=10), the HIF-1α transfected group(group B, n=10). The weights of subcutaneous tumor were detected. The resected specimens were made into paraffin-embedded sections. The proliferating cell nuclear antigen (PCNA) was identified by immunohistochemistry(ISH). The expressions of HIF-1α、 apoptosis-related protein survivin and bcl-2 were analyzed by Western blot. RESULTS: The growth rates of the HIF-1α transfected lung cancer cells A549 were significantly increased, and more importantly, the HIF-1α transfected lung cancer cells A549 was able to enhance lung cancer growth in nude mice(P＜0.05). The PCNA were increased significantly in group B, compared with group A. The expressions of HIF-1α, survivin and bcl-2 in group B were increased significantly than that of group A. CONCLUSIONS: HIF-1α increases lung cancer cells A549 growth in vitro and in vivo and its mechanism may be due to promotion of proliferation and inhibition of apoptosis.     

Effects of different ages on ventricular remodeling after ischemic heart failure in mice

AIM: To observe ventricular remodeling induced by ischemic heart failure in the mice at different ages.METHODS: Three-month-old (young group, n=50) and 18-month-old (old group, n=50) male C57BL/6J mice were selected in the study. Forty mice underwent ligation of left coronary artery with certain infarct size, and 10 were sham-operated for control. Echocardiography was performed after 8 weeks of infarction. All mice were killed and the hearts were collected for examinations. Masson trichrome staining was used to detect myocardial fibrosis. The expression of type I and type Ⅲ collagens was measured by the method of immunohistochemistry.RESULTS: The incidences of cardiac rupture (18% vs 10%, P<0.05) and heart failure (22% vs 10%, P<0.05) were significantly higher in aged mice than those in young mice. The degrees of left ventricular dilation, contractile dysfunction and heart rate were significantly higher in aged mice than those in young mice (P<0.05). The left ventricular mass index, collagen volume fraction, the expression of type I collagen and ratio of type Ⅰ/Ⅲ collagens were significantly increased in aged mice as compared with young mice (P<0.05).CONCLUSION: After heart failure, aged mice show abnormal collagen distribution, and suffer from worse cardiac functions and more serious ventricular remodeling.     

Experimental research on cardiac fibrosis induced by recombinant mouse interleukin-11 in mice

AIM To observe the effect of recombinant mouse interleukin-11 （rmIL-11）injected subcutaneously into mice on heart structure and function and to determine its pro-fibrotic effect. METHODS C57BL/6 mice were randomly divided into experimental group and control group.The mice in experimental group were injected subcutaneously with recombinant mouse IL-11 at the dose of 100 μg·kg^-1·d^-1 for 3 consecutive weeks, while the control group were given equal volume of normal saline in the same way. After the experiment was finished, the parameters of heart function were measured by echocardiography.The heart weight was weighed and the cardiac weight index (CWI) was calculated. HE staining and Masson's trichrome staining were performed to observe the pathological changes and the extent of myocardial fibrosis in mouse myocardia respectively, and the cardiac collagen volume fraction (CVF) was calculated. The expression levels of extracellular matrix proteins in the myocardial tissues of mice, including type Ⅰ collagen, type Ⅲ collagen and fibronectin, were determined by Western blot. RESULTS Left ventricular ejection fraction and left ventricular fraction shortening in experimental group were obviously lower than those in control group (P<0.01), however left ventricular end-diastolic diamension and left ventricular end systolic dimension were significantly higher than those in control group (P<0.05).Compared with control group, the CWI was increased (P<0.01), the myocardial arrangement was disorder, the necrosis of cardiac myocytes was increased, and excessive deposition of collagen was observed in the myocardial tissues in experimental group. Correspondingly, the CVF and protein levels of type Ⅰ collagen, type Ⅲ collagen and fibronectin in the left ventricle in experimental group were increased significantly (P<0.05). CONCLUSION Injection of rmIL-11 into the mice subcutaneously induces fibrogenesis in the heart, which implies that IL-11 is likely a novel pro-fibrotic factor.     

Inhibitory effect of Wendan decoction on atherosclerotic plaque formation in ApoE-/- mice by regulating expression of membrane proteins involved in reverse cholesterol transport

AIM To explore the anti-atherosclerotic mechanism of Wendan decoction based on reverse cholesterol transport. METHODS Eight-week-old apolipoprotein E gene knockout （ApoE^-/-） mice with high-fat diet and daily drug gavage were randomly divided into model group, simvastatin group, and low-, middle- and high-dose Wendan decoction groups, with 15 mice in each group. The C57BL/6 mice of the same age served as control group. The mice were weighed once every week. After 10 weeks, the mice were anesthetized with chloral hydrate. The serum were collected for lipid level examination. The atherosclerotic plaque buildup in aortic root and whole aorta was observed by HE staining and oil red O staining, respectively. The levels of proteins related to cholesterol transport, ATP-binding cassette transporter A1 （ABCA1） and caveolin-1 in the aorta, and scavenger receptor class B type I （SR-BI） and CD36 in the liver, were quantified by Western blot. RESULTS Wendan decoction at middle dose inhibited the increase in the body weight of ApoE^-/- mice fed with high-fat diet （P<0.05）. Wendan decoction at different doses significantly reduced the serum levels of triglyceride, total cholesterol and low-density lipoprotein cholesterol in the ApoE^-/- mice （P<0.05 or P<0.01）, but had no effect on serum high-density lipoprotein cholesterol level （P>0.05）. Wendan decoction at different doses inhibited the formation of atherosclerotic plaques in whole aorta of the ApoE^-/- mice （P<0.05 or P<0.01）. Middle- and high-dose Wendan decoction significantly inhibited the formation of atherosclerotic plaques in the aortic root （P<0.05）. Bedsides, Wendan decoction at different doses increased the protein level of ABCA1 and decreased the protein level of caveolin-1 in the aorta of the ApoE^-/- mice （P<0.01）. Middle- and high-dose Wendan decoction increased the liver protein level of SR-BI in the ApoE^-/- mice （P<0.01）. However, Wendan decoction at different doses had no effect on the liver protein level of CD36 in the ApoE^-/- mice （P>0.05）. CONCLUSION Wendan decoction reduces the body weight, serum lipid levels and formation of atherosclerotic plaques in ApoE^-/- mice fed with high-fat diet, and its mechanism is related to up-regulation of ABCA1 protein level in the aorta and SR-BI protein level in the liver as well as down-regulation of caveolin-1 protein level in the aorta.     

Effect of bFGF on promoting angiogenesis in infarct area and improving myocardial fibrosis in mouse myocardial infarction model

AIM: To investigate the protective effect of basic fibroblast growth factor (bFGF) on the heart of mice with myocardial infarction and its mechanism. METHODS: The model of myocardial infarction was established by the ligation of left anterior descending artery of C57/B6 mice (8~12 weeks old) after lateral thoracotomy. The mice were divided into sham operation group, myocardial infarction group and bFGF administration group. bFGF at 0.5 μg was intraperitoneally injected on alternate days after myocardial infarction for 7 d. Cardiac Doppler ultrasonography was used to detect cardiac function after myocardial infarction for 28 d, and left ventricular end-diastolic diameter, left ventricular end-systolic diameter, left ventricular ejection fraction and left ventricular shortening fraction (LVFS) were used to evaluate cardiac function. After myocardial infarction for 28 d, the mice were sacrificed and the hearts were collected for preparing pathological sections. The degrees of myocardial fibrosis and angiogenesis in the myocardial infarction area were observed. Western blot was used to detect the indicators of angiogenesis. RESULTS: The results of Masson staining showed that bFGF administration significantly reduced myocardial fibrosis at 28 d after myocardial infarction. Cardiac ultrasound data showed that cardiac functions in myocardial infarction group were poorer than those in sham group, and bFGF administration significantly improved cardiac functions. Immunofluorescence staining showed that neovascularization in myocardial infarction area of bFGF administration group was more than that in myocardial infarction group. The results of Western blot showed that bFGF activated AKT/HIF-1α/VEGF signaling pathway. CONCLUSION: Intraperitoneal injection of bFGF reduces myocardial fibrosis and improves cardiac function in myocardial infarction mice. bFGF may promote angiogenesis by activating AKT/HIF-1α/VEGF signaling pathway.     

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.     

Effects of ERK1/2 on pressure overload-induced cardiomyopathy and heart failure in mice

AIM: To explore the changes in extracellular regulated protein kinase (ERK1/2) in the hypertrophic myocardium induced by pressure overload at the different time courses and to determine the molecular mechanism in the myocardium from hypertrophy to heart failure. METHODS: C57/BL mice, aged 12 week old, were subjected to sham-operation (SH) or transversing aortic constriction (TAC) to establish left ventricular hypertrophy. Echocardiographic assessments, hemodynamic determination, organ weight measurement, morphological and histological examination were performed at 1, 4, 8, 12 and 16 weeks after surgery. Meanwhile mRNA levels of atrial natriuretic peptide (ANP), α-myosin heavy chain (α-MHC), bcl-2 and bax were measured by RT-PCR, and ERK1/2 levels were detected by Western blotting. The animals in SH group were performed the same tests then sacrificed at 16 weeks. RESULTS: (1) Compared to SH group, LVESd, LVEDd, Awsth, Awdth, Pwsth and Pwdth progressively increased after TAC. Meanwhile, ejection fraction (EF%) significantly decreased at 16th week (P<0.05). LVSP, dp/dtmax and dp/dtmin in TAC group were progressively increased after 4 weeks. From 8-12 weeks these parameters maintained stable and then sharply decreased at 16th week (all P<0.05). However, LVEDP was statistically increased at 8th week. These echocardiographic and hemodynamic changes indicated a development of LVH and eventually progressing towards to heart failure. (2) Histologically, cardiac collagen measured by percentage of Sirius red positive stained area and apoptosis index showed progressive increases from 4 to 16 weeks. (3) Compared to SH group, mRNA levels of ANP was time-dependently increased while α-MHC and Bcl-2 were time-dependently decreased. The ratio of Bcl-2 /Bax was decreased. Phosphorylation of ERK1/2 was increased at 4th week, then decreased with age of TAC (all P<0.05). CONCLUSION: Pressure-overload induced by TAC results in a development of LVH from early concentric hypertrophy to late eccentric hypertrophy, and eventually toward cardiac dysfunction or heart failure. Those changes are associated with increase in cell size and cardiac fibrosis. ERK1/2 signaling pathway may involve in the regulation of myocardial cell apoptosis in hypertrophic and failure heart.     

Effect of hyperbaric oxygen on HIF-1α expression in rat experimental periodontitis with psychological stress

AIM: To investigate the effect of hyperbaric oxygen (HBO) on hypoxia-inducible factor-1α (HIF-1α) expression in rat experimental periodontitis with psychological stress. METHODS: Male special pathogen-free Wistar rats (n=120) were randomly divided into 4 groups: normal control group; psychological stress stimulation group; experimental periodontitis group: the periodontitis model was induced by wrapping 3/0 silk ligature inoculated with Porphyromonas gingivalis around the left maxillary second molar of the rats; periodontitis model with stress stimulation group. Psychological stress was removed at the 9th weeks after ligature, 6 rats from each experiment group were randomly chosen to HBO treatment. The rats were sacrificed at the 2nd, 4th, 8th and 10th weeks after ligature. Gingival index (GI) and attachment loss (AL) were measured before sacrifice. The histological changes of periodontal tissues were observed under microscope with HE staining. The expression of HIF-1α was observed by the method of immunohistochemistry. RESULTS: The sites of gingival attachment were normal in control group and psychological stress stimulation group. Periodontal pocket, and periodontal attachment loss were observed in experimental periodontitis group. The tissue damage was much serious in periodontitis model with stress stimulation group. No significant difference of GI and AL among psychological stress stimulation group and normal control group during the experiment was observed. GI and AL in periodonitis model with stress stimulating group were significantly higher than those in experimental periodontitis group at the 4th and 8th weeks (P < 0.01). The levels of GI and AL were significantly lower at the 10th weeks after HBO treatmnt than those in untreated groups (P < 0.05). No significant difference of HIF-1α expression scores among psychological stress stimulation group and normal control group was found. HIF-1α expression scores in periodonitis model with stress stimulating group was significantly higher than that in experimental periodontitis group at the 4th and 8th weeks (P < 0.01). At the 10th weeks after HBO treatment the levels of HIF-1α were significantly lower than that in untreated groups (P < 0.01). CONCLUSION: Stress stimulation may aggravate periodontitis by decreasing tissue oxygenation in rats. HBO may represent a useful way in psychological stress periodontitis therapy.     

Expression of cathepsin C and tumor necrosis factor-α in human coronary artery and their relationships with coronary heart disease

AIM To explore the possible mechanism of cathepsin C （CTSC） and tumor necrosis factor-α （TNF-α） in coronary heart disease （CHD） by detecting the protein expression of CTSC and TNF-α in human coronary artery tissue. METHODS The coronary artery tissues from 52 cases of CHD and 25 cases of accidental death without any heart disease in the Forensic Judicial Expertise Center of Guizhou Medical University from October 2018 to December 2019 were collected as CHD group and control group, respectively. The coronary artery stenosis and intimal plaque formation were examined by histopathology, the protein expression of CTSC and TNF-α was determined by Western blot, and the intracellular expression of CTSC and TNF-α was analyzed by immunohistochemical staining. RESULTS The results of HE staining showed that the intima of coronary artery in control group was smooth, and no thickening or stenosis was observed. In CHD group, the intima thickened irregularly, atherosclerotic plaques formed, the intima became thinner and the lumen showed eccentric stenosis in varying degrees （P<0.05）. The results of Western blot showed that the expression of CTSC and TNF-α in CHD group was significantly higher than that in control group （P<0.05）. Immunohistochemical staining showed that both CTSC and TNF-α were expressed in the cytoplasm of foam cells, and their positive expression was significantly higher than that in control group （P<0.05）. The results of Pearson moment correlation analysis showed that there was positive correlation between the expression of CTSC and TNF-α in CHD （r²=0.743, P<0.05）. CONCLUSION The up-regulated expression of CTSC in coronary artery tissue may promote the expression of TNF-α and affect the occurrence and development of CHD.     

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.     

Expression of hypoxia-inducible factor 1 and neuroglobin in piglet cortex during deep hypothermic circulatory arrest

AIM: To observe the expression of hypoxia-inducible factor 1 (HIF-1) and neuroglobin (NGB) in piglet cortex during deep hypothermic circulatory arrest. METHODS: Wuzhishan piglets were randomly assigned to cardiopulmonary bypass group (CPB group), 40 min of circulatory arrest (CA) at 18 ℃ without cerebral perfusion (DHCA group) or with selective antegrade cerebral perfusion (SACP group). After 180 min of reperfusion, cortical tissue was harvested for determining HIF-1α and NGB expression by HE staining, Western blot and real-time PCR. RESULTS: Severer cerebral injury was observed in DHCA group than that in SACP group. After 180 min of reperfusion, HIF-1α protein and mRNA levels were significantly higher in DHCA group than those in CPB group (P<0.05). Accordingly, SACP animal had higher levels of HIF-1α protein and mRNA than those in DHCA group (P<0.05). Simultaneously, higher NGB protein and mRNA levels were found in DHCA group than those in CPB group after 180 min of reperfusion (P<0.05). The SACP animal had higher levels of NGB protein and mRNA than those in DHCA group (P<0.05). CONCLUSION: Up-regulation of HIF-1 and NGB are involved in the mechanism against cerebral injury resulting from DHCA in the cortex and possibly a part of cerebral protective effect of SACP.