Abstract:Objective:　To investigate the effects and mechanisms of ghrelin on angiogenesis in vulnerable plaque and myocardial infarction. Methods:　The ApoE^-/-mice were divided into vulnerable plaque group, vulnerable plaque+myocardial infarction group and ghrelin-intervened group. All ApoE^-/-mice were fed with a high-fat diet for 12 weeks to induce atherosclerotic vulnerable plaques. The induced-atherosclerotic vulnerable plaque model was established in vulnerable plaque group. The C57BL/6J mice were fed for 12 weeks on a normal diet and divided into control group and myocardial infarction group. At the 8th week of this study, myocardial infarction group, vulnerable plaque+myocardial infarction group and ghrelin group were subjected to acute myocardial infarction model. After this model, the ghrelin group was administered ghrelin (100 μg/kg, bid) until the end of the 12th week. The left ventricular ejection fraction (LVEF) and left ventricular fraction shortening (LVFS) were detected by echocardiography and the percentage of atherosclerotic plaque area in aortic sinus was observed by Oil red O staining. The microvascular density (MVD) in vulnerable plaque and surrounding area of myocardial infarction were measured by CD31 immunohistochemistry and Masson staining, respectively. The mRNA and protein expression of vascular endothelial growth factor (VEGF), angiopoietins (Ang) -1/2 and tyrosine-protein kinase receptor (Tie) -2 were respectively examined by real-time PCR and Western blot. Results:　①Compared with myocardial infarction group, the myocardial infarct size in vulnerable plaque+myocardial infarction group [(65.518±4.160) %, P=0.000]was increased, whereas the LVEF [(58.560±11.900) %, P=0.012]and LVFS [(27.182±7.807) %, P=0.013], the MVD (5.800±0.837, P=0.000) in the peripheral area of myocardial infarction, and the expression of VEGF (1.870±0.122, P=0.001), Ang-1 (1.830±0.056, P=0.007), Ang-2 (1.660±0.217, P=0.006) and Tie-2 (1.660±0.115, P=0.020) were decreased, significantly. Additionally, the ghrelin intervention had significantly reduced myocardial infarct size [(39.751±3.039) %, P=0.000], and improved LVEF [(74.679±6.535) %, P=0.013], LVFS [(37.507±5.210) %, P=0.020], the MVD (9.857±1.345, P=0.000) in the peripheral area of myocardial infarction, and the expression of VEGF (3.503±0.384, P=0.004), Ang-1 (3.277±0.186, P=0.017), Ang-2 (3.450±0.187, P=0.000) and Tie-2 (3.133±0.139, P=0.009) of the vulnerable plaque+myocardial infarction group.②There were no significant differences in the percentage of aortic sinus plaque area, the MVD in vulnerable plaque, and the expression of VEGF, Ang-1, Ang-2 and Tie-2 among the vulnerable plaque group, the vulnerable plaque+myocardial infarction group and the ghrelin-intervened group. Regrettably, ghrelin had no effect on angiogenesis in vulnerable plaque+myocardial infarction mice. Conclusion: Ghrelin can promote angiogenesis in ischemic myocardium, reduce myocardial infarct size, and stabilize vulnerable plaque by increasing the mRNA and protein expression of VEGF, Ang-1, Ang-2 and Tie-2.Ghrelin had no effect on angiogenesis in vulnerable plaque of vulnerable plaque+myocardial infarction group.

Abstract:Objective:　To explore the protective effect of neuropilin-1 overexpression on cardiac function and against myocardial tissue injury in rats with isoproterenol (ISO) induced heart failure (HF). Methods:　The adenovirus vector of neuropilin-1 overexpression (Ad-neuropilin-1) was constructed. The overexpression efficiency was detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Sixty SD rats were enrolled and randomly divided into control group, ISO group, ISO+Ad group (negative control group) and ISO+Ad-neuropilin-1 group. Except control group, subcutaneous injection of ISO was conducted to prepare HF rats models. The cardiac function in each group was detected. The levels of myocardial damage markers and inflammatory factors in each group were detected by RT-PCR and enzyme-linked immunosorbent assay (ELISA). The apoptosis of myocardial cells was observed by HE staining and TdT-medated dUTP nick end labeling (TUNEL) staining. The expression of proliferation cells related antigen Ki-67 and cysteine aspartase-3 (Cas3) was detected by Western blot. Results:　Compared with control group, neuropilin-1 mRNA and protein in ISO+Ad-neuropilin-1 group were increased (P<0.05). Compared with control group, heart rate (HR), left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS), and expression levels of neuropilin-1 and Ki-67 proteins were significantly decreased in ISO group, while levels of serum cardiac troponin (cTnI), creatine kinase (CK), creatine kinase-MB (CK-MB), inducible nitric oxide synthase (iNOS) and interleukin-10 (IL-10) mRNA, apoptosis rate of myocardial cells and level of cleaved caspase-3/Cas3 were significantly increased (P<0.05). Compared with ISO group, HR, LVFS, LVEF, and relative expression of neuropilin-1 and Ki-67 proteins in ISO+Ad-neuropilin-1 group were significantly increased, while cTnI, CK, CK-Mb, iNOS, content of peripheral blood IL-10 mRNA, apoptosis rate of myocardial cells, and level of cleaved caspase-3/Cas3 were significantly decreased (P<0.05). Conclusion: Neuropilin-1 overexpression can inhibit cardiomyocytes apoptosis and inflammation response in rats with ISO-induced HF, and improve their cardiac function.

Abstract:Objective:　To investigate the effect of melatonin (Mel) on the JAK2/STAT3 signaling pathway of hepatic stellate cell-T6 (HSC-T6) induced by platelet-derived growth factor (PDGF). Methods:　HSC-T6 cells were divided into 6 groups: control group, model group, experimental groups (Mel 1 nmol/L, Mel 1 μmol/L, Mel 0.1 mmol/L) and inhibitor group (AG490 JAK2/STAT3 pathway inhibitor). Real-time fluorescent polymerase chain reaction (RT-PCR) was used to detect the mRNA expression of α-smooth muscle actin (α-SMA) and collagen typeⅠ (collagen-Ⅰ) in HSC-T6, and the protein expression detection of JAK-2, P-JAK2 and STAT3, P-STAT3 proteins in HSC-T6 cells was made by immunohistochemistry. Results:　Compared with the control group, PDGF significantly activated the proliferation of HSC-T6, and significantly up-regulated the expression of α-SMA, collagen-ⅠmRNA and JAK-2, P-JAK2, STAT3 and P-STAT3 in HSC-T6.Compared with model group, Mel and JAK2/STAT3 pathway inhibitor inhibited PDGF induced HSC-T6 proliferation, and significantly down-regulated the expressions of α-SMA, collagen-ⅠmRNA and JAK-2, P-JAK2, STAT3 and P-STAT3.Conclusion: Mel can inhibit the activation and proliferation of HSC-T6 induced by PDGF, and the mechanism may be related to the inhibition of JAK2/STAT3 signaling pathway, which may be one of the mechanisms of melatonin improving liver fibrosis.

Abstract:Objective:　To investigate the effect of glucagon-like peptide 1 (GLP-1) analog liraglutide on the expression of PARP and NF-κB in diabetic peripheral neuropathy (DPN) rats by establishing a rat model of DPN. Methods:　A total of 84 SD rats were randomly selected as control group, and the other rats were induced by streptozotocin to establish DPN model. After successful modeling, 72 rats were randomly divided into model group, insulin group, mecobalamin group, and liraglutide low, medium and high dose groups. Normal group and model group were subcutaneously injected with equal volume of normal saline. Insulin group was treated with insulin, mecobalamin group was treated with mecobalamin, and drug groups were treated with different doses of liraglutide for 8 weeks. The body weight and blood glucose of rats in each group were recorded, nerve reaction speed was measured, inflammatory factors were detected by ELISA, apoptosis was detected by flow cytometry, and PARP-1 and NF-κB protein expression were detected by Western blot. RSC96 cells were selected to establish cell damage model in high glucose environment. PARP target gene was predicted by gene chip and tested by transfection of PARP-1. Results:　Compared with the model group, liraglutide significantly increased the body weight, significantly decreased blood glucose, and increased the speed of nerve reaction (P<0.05); rat and cell experiments showed that the content of inflammatory factors and the expression of PARP and NF-κB protein were significantly decreased after liraglutide treatment, and the reduction was more obvious in the high-dose group (P<0.05); the apoptosis rate of nerve cells in rats was significantly decreased than that in the model group (P<0.05). Liraglutide inhibited the transcriptional activity of NF-κB. The levels of inflammatory factors, PARP-1 and NF-κB protein in PARP-1 transfection group were significantly higher than those in middle dose group (P<0.05). PARP-1/NF-κB is an important target of GLP-1 in anti-inflammatory effect of diabetic neuropathy rats. Conclusion: Liraglutide can reduce the expression levels of PARP-1 and NF-κB in sciatic nerve of rats with DPN, thereby reducing the sciatic nerve injury of rats and preventing the damage of DPN.

Abstract:Objective:　To investigate the effects of artemisinin and its derivatives on blood glucose, body weight and expression of inflammatory factors in db/db mice. Methods:　Five healthy db/m mice aged 4-6 weeks were collected as the normal control group with responsive feeding. Twenty db/db mice were randomized into four groups: the placebo group, the artemisinin group, the artemether group and the artesunate group, with five mice in each group. The placebo group was administrated with hydroxymethylcellulose daily, and the intervention group was administrated with artemisinin[200 mg/ (kg·d)], artemether[200 mg/ (kg·d)] and artesunate[200 mg/ (kg·d)] for 10 weeks. The body weight and fasting blood glucose of mice were measured once a week, and the random blood glucose was measured every two weeks. The changes of inflammatory factors in serum were detected by ELISA, including intercellular adhesion molecule-1 (ICAM-1/CD54), interleukin-6 (IL-6), interleukin-8 (IL-8), leptin (LEP), nuclear factor kappa B (NF-κB) and interleukin-1β (IL-1β). Results:　Firstly, compared with db/m group, fasting blood glucose and random blood glucose were all significantly higher in db/db group. Compared with placebo group, the fasting and random blood glucose in artemisinin, artemether and artesunate group decreased significantly. Secondly, compared with db/m group, the body weight of db/db group increased significantly. Compared with placebo, artemisinin could significantly reduce the weight of diabetic mice, while artemether and artesunate had no significant weight loss effect on diabetic mice. Thirdly, compared with db/m group, the levels of ICAM-1/CD54, IL-6, IL-8, LEP, NF-κB and IL-1β in db/db group were all increased. Compared with placebo, artemisinin, artemetherandartesunatesignificantly reduced the levels of ICAM-1/CD54, IL-6, IL-8, LEP, NF-κB and IL-1β in diabetic mice. In comparison, artesunate had the strongest anti-inflammatory ability, and the level of inflammatory factors was reduced significantly, followed by artemisinin, and artemether had relatively weak anti-inflammatory ability. Conclusion: Artemisinin and its derivatives, artemether and artesunate, could significantly reduce the expression of blood glucose and inflammatory factors in db/db mice. Artemisinin can reduce the body weight of db/db mice, but artemether and artesunate have no obvious effect on body weight. In conclusion, artemisinin has the best therapeutic effect on diabetic mice.

Abstract:Objective:　To study the effect of resveratrol (RES) on left ventricular remodeling in spontaneous hypertension rat (SHR) and its role in silent information regulator 1/nuclear transcription factor-κB (SIRT1/NF-κB) signaling pathway. Methods:　Eight male Wistar-Kyoto (WKY) rats aged 8 weeks were taken as the normal group (WKY group), and 40 male SHR aged 8 weeks were randomly divided into SHR group, RES group, EX527 group, RES+EX527 group, and RES+pyrrolidine dithiocarbamate (PDTC) group, with 8 rats in each group, given corresponding drug intervention for 12 weeks. The rats’weight, systolic pressure, echocardiography, left ventricular mass, left ventricular mass index (LVMI), myocardial collagen volume fraction (CVF), oxidative stress index, SIRT1 and NF-κB protein expression levels, and inflammatory factors protein expression levels were measured. And the morphological and pathological changes of the heart under HE and Masson staining were observed. Results:　Compared with WKY group[LVMI (2.12±0.10) mg/g, CVF (0.18±0.09) %, SIRT1 protein expression (1.40±0.05), NF-κB protein expression (0.46±0.06)], SHR group[LVMI (3.07±0.07) mg/g, CVF (2.60±0.56) %, SIRT1 protein expression (0.78±0.08), NF-κB protein expression (1.41±0.08)] showed severe left ventricular remodeling, significantly increased levels of oxidative stress and inflammatory factors as well as NF-κB expression (P<0.05), and decreased SIRT1 expression (P<0.05). In RES group[LVMI (2.72±0.09) mg/g, CVF (1.10±0.30) %, SIRT1 protein expression (0.99±0.07), NF-κB protein expression (0.98±0.12)], compared with SHR group, left ventricular remodeling degree was reduced, oxidative stress and inflammatory factors as well as NF-κB expression levels were decreased (P<0.05), and SIRT1 expression was increased (P<0.05). The results of EX527 group[LVMI (3.18±0.08) mg/g, CVF (2.83±0.98) %, SIRT1 protein expression (0.60±0.02), NF-κB protein expression (1.68±0.07)] were opposite to those of RES group. Compared with RES group, RES+EX527 group showed a trend of reversing RES to ameliorate left ventricular remodeling. RES+PDTC group showed a tendency to further ameliorate left ventricular remodeling. Conclusion: RES can ameliorate left ventricular remodeling of SHR, and its mechanism may be related to regulation of SIRT1/NF-κB pathway to reduce oxidative stress and inflammatory response damage.

Abstract:Objective:　To explore whether overexpression of NR4A1 can alleviate lupus nephritis (LN). Methods:　A mouse model of LN was constructed by intraperitoneal injection of 0.5 mL pristane, divided into saline control group (SC), pristane control group (PC), vector control group (VC) and overexpressed NR4A1 group. At the end of 20^thweek, the monocytes overexpressed NR4A1 were injected into the tail vein. The diameter of ankles was measured at the end of 28^thweek. Renal pathological sections were stained with HE staining and semi-quantitative scores were used to assess renal pathological damage. Serum TNF-α and anti-ds-DNA concentrations were measured by ELISA. The expression of Bcl-2 and NR4A1 in the spleen was detected by real-time PCR. Results:　The results showed that compared with the SC group, LN-related activity indicators in the PC group were significantly increased, but the ankle joint diameter[4.69 (4.26, 5.13) mm, P=0.000], the anti-ds-DNA antibody concentration [(173.11±25.49) ng/mL, P=0.000], renal pathology score[2.78 (2.68, 2.81), P=0.000], and the expression of NR4A1 (0.40±0.09, P=0.008) was significantly reduced. Compared with the PC and VC groups, the NR4A1 group LN activity indicators were significantly improved, the ankle joint diameter[3.80 (3.44, 4.20) mm, P_PC=0.002, P_VC=0.033], the concentration of anti-ds-DNA antibody [(51.92±10.91) ng/mL, P_PC=0.008, P_VC=0.025], renal pathology score[1.39 (1.30, 1.49), P_PC=0.002, P_VC=0.018].ELISA results showed that the concentration of TNF-α in the NR4A1 group [(542.86±108.28) pg/mL]was significantly lower than that of the PC group [(1 779.00±358.40) pg/mL, P=0.008]and the VC group [(1 759.71±280.27) pg/mL, P=0.010].Real-time PCR results showed that the expression of Bcl-2 gene in the NR4A1 group (1.92±0.58) was significantly higher than that of the PC group (0.48±0.20, P=0.000). Conclusion: Overexpression of NR4A1 alleviates the symptoms of LN and reduces LN activity indicators in LN mice, which may involve the interaction among NR4A1, TNF-α and Bcl-2.

Abstract:Objective:　To explore the regulation between the Shh related signal pathway cytokines, cell cycle regulator and primary cilia depolymerization factors as well as the changing relationship among them after the treatment of dexamethasone and its antagonist vitamin B₁₂intervention in the key period of pregnancy. Methods:　C57BL/6J pregnant mice at specific period of pregnancy were intraperitoneally injected with a certain dose of saline (control group), dexamethasone (dexamethasone group), vitamin B₁₂ (vitamin B₁₂ group), vitamin B₁₂+dexamethasone (dexamethasone+vitamin B₁₂group), and they were observed for the fusion of embryo and palatal process in each group on 13.5 and 17.5 days of pregnancy. The expression changes of Smo, Ptch1, Cyclin D1 and Aurora A in the palatal process of mice on 13.5 days of pregnancy in each group were detected by Western blot, and analyzed statistically. Results:　Dexamethasone retarded the development of the embryonic palatal process leading to cleft palate, while vitamin B₁₂intervention partially restored the development of the embryonic palatal process. Western blot results showed that, compared with the control group, the expressions of Ptch 1, Aurora A and Cyclin D1 were significantly decreased in dexamethasone group (all P=0.000), but the expression of Smo had no significant change (P=0.695). Compared with dexamethasone group, the expressions of Ptch 1, Aurora A and Cyclin D1 in dexamethasone+vitamin B₁₂group were upregulated in different degrees (all P=0.000), but the expression of Smo had no significant change (P=0.818). There was no significant difference in the expressions of each target protein between the vitamin B₁₂group and the control group (P=0.879, 0.399, 0.645, 0.168). Conclusion: Dexamethasone influences Shh downstream signal transmission and primary cilia depolymerization in the embryonic palatal process of mice, thereby inhibiting the expression of Cyclin D1 and reducing the proliferation of mouse embryonic palatal process cells. Vitamin B₁₂can antagonize the inhibitory effect by up-regulating the content of related factor proteins.

Abstract:Objective:　To investigate the effect of metformin (Met) on islet β-cell function in young rats with type 1 diabetes mellitus (T1DM) and its possible mechanisms of action. Methods:　Healthy young rats were recruited in this study, 12 in Control group, 24 in induced type 1 diabetes model (T1DM model) group, and 24 in Met administration group (T1DM+Met group) after induction of T1DM. Glucose tolerance test, insulin tolerance test and high glucose clamp technique were used to detect islet function. The morphology of islet was observed by hematoxylin-eosin (HE) staining. Insulin and Ki67 immunofluorescence double staining method was used to determine the proliferation rate of β cells. Apoptosis of β cells was detected by terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) assay. Western blot was used to detect the protein levels of pancreatic duodenal homeobox 1 (PDX-1), B-cell lymphoma-2 (Bcl-2) and endoplasmic reticulum stress-related factors in pancreatic tissue. Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was used to detect mRNA levels of inflammation-related factors in pancreatic tissue. Results:　Met could reduce the fasting blood glucose and insulin levels of T1DM young rats, and improve the abnormal glucose tolerance and insulin sensitivity. Met significantly increased the glucose infusion rate (GIR) value, the insulin secretion 1 min after intravenous glucose stimulation, and the maximum secretion capacity of islet β cells in the clamp steady state in the high glucose clamp experiment. Met restored the islet morphology to a certain extent, promoted β cells proliferation and inhibited their apoptosis by promoting the expression of PDX-1 and Bcl-2 protein levels. Met significantly down-regulated the expression of phosphorylated eukaryotic initiation factor 2 alpha (eIF2α), eukaryotic initiation factor 4 (ATF4) and C/EBP-homologous protein (CHOP) (F=81.322, 215.372, 55.059, all P=0.000), and also significantly down-regulated the mRNA levels of nuclear factor κappa-B (NF-κB), interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) (F=570.275, 369.184, 474.006, all P=0.000). Conclusion: Met can improve the secretion function of islet β cells in type 1 diabetic rats by promoting differentiation, anti-apoptosis, relieving stress of endoplasmic reticulum stress and reducing inflammation, thereby promoting its proliferation and restoring islet morphology in young rats with T1DM.

Abstract:Objective:　To explore a method for separation and purification of satellite cells (SCs) from rats levator ani muscle (LAM), and to observe the proliferation and myogenic characteristics of SCs in vitro. Methods:　The SCs from rat LAM were separated by enzyme digestion with collagenase type 1 and trypsin, and purified by differential wall adhesion method. From that, we obtained highly purified SCs from rat LAM. The morphology and growth of primary and passage cells were observed by inverted microscope. CCK-8 method was used to detect the growth of LAM SCs (cultured in vitro) and draw the growth curve. Immunofluorescence was used to identify SCs in different stages. Results:　Microscopically, it was observed that SCs separated from rat LAM grew well. In CCK-8 test, the SCs grew slowly in the first to second days of in vitro culture, after 3 days, the SCs augmented multiply. On the 10th day, the cell growth rate decreased and some cells began to differentiate. Immunofluorescence and DAPI staining showed that the characteristic gene expression of protein was observed different in different stages of SCs. Conclusion: In this study, SCs were successfully separated from rat LAM tissue with good proliferation and differentiation ability in vitro, laying a preliminary foundation for exploring the mechanism of muscle injury and repair by LAM SCs.

Abstract:Objective:　To evaluate the therapeutic potential of thymoquinone in rats with sepsis and explore the underlying molecular mechanism. Methods:　Cecum ligation and puncture (CLP) was used to induce sepsis rats model. Thymoquinone (20 mg/kg) was intraperitoneally injected every 12 h for three days after CLP, with or without intraperitoneal injection of the sirtuin1 (SIRT1) inhibitor EX527 (5mg/kg). The expression of fasting blood glucose and fasting insulin were detected after the designed treatment. The relative expression of aspartate aminotransferase (AST) and lactic alanine transaminase (ALT) in serum were determined by Reit’s analysis method. Levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in liver tissues were analyzed by immunohistochemistry. Levels of p-Akt, t-Akt, SIRT1, p-signal transducer and activator of transcription 3 (STAT3), PEPCK, G6Pase and t-STAT3 were evaluated by Western blot. The expression of reactive oxygen species (ROS) in liver tissues was detected by DCFH-DA fluorescence probe and the level of acetylated STAT3 in liver tissues was determined by immunoprecipitation analysis. Results:　The hepatic SIRT1/STAT3 pathway of septic rats was significantly inhibited, expressions of AST, ALT, IL-6 and TNF-α were up-regulated, and levels of fasting blood glucose and fasting insulin were increased. Thymoquinone administration was able to promote STAT3 phosphorylation and alleviate liver dysfunction and glucose metabolism disorder. EX527 significantly diminished the protective effect of thymoquinone on sepsis induced liver injury, hyperglycaemia and STAT3 inactivation. Conclusion: Thymoquinone is a potential therapeutic agent for sepsis-associated liver injury and glucose metabolism disorder and its mechanism may be realized by activating SIRT1/STAT3 pathway.

Abstract:Objective:　To explore the effect and mechanism of miR-149-5p in apoptosis, epithelial-mesenchymal transition (EMT) and microtubule formation in human oral squamous cell carcinoma (OSCC). Methods:　miR-149-5p mimic (mimic) and pcDNA-FGF5 (FGF5) were singly or co-transfected with CAL-27 cell line, and RT-PCR and Western blot were used to detected the expression FGF5; dual luciferase report test was used to verify the targeting relationship between miR-149-5p and FGF5. The apoptosis rate was detected by flow cytometry, and the expression of Bax/Bcl-2 protein was detected by Western blot. The morphological transformation of EMT cells was observed, and the expression of epithelial and interstitial markers E-cadherin (E-cad) and N-cadherin (N-cad) in CAL-27 cells was detected by Western blot. The microtubule-like formation ability of CAL-27 cells was detected by tube-forming test, and the expression of vascular endothelial growth factor (VEGF) was detected by Western blot. Results:　miR-149-5p mimic could significantly inhibit the expression of FGF5 in CAL-27 cells (P=0.000); miR-149-5p mimic could significantly reduce the luciferase activity of FGF5 wild plasmid (P=0.000); miR-149-5p mimic increased the apoptosis rate of CAL-27 cells and Bax/Bcl-2 ratio significantly (P=0.000), and reversed the increase of apoptosis rate (P=0.014) and Bax/Bcl-2 ratio caused by FGF5 overexpression (P=0.000). miR-149-5p mimic inhibited the EMT transformation on cell morphology from round to spindle shape, and reversely transformed the mesenchymal-like cell morphology caused by FGF5 overexpression. miR-149-5p mimic up-regulated the expression of E-cad in CAL-27 cells (P=0.000), down-regulated the expression of N-cad (P=0.0009), reversed the down-regulated expression of E-cad (P=0.020) and the up-regulated expression of N-cad (P=0.000) caused by FGF5 overexpression. miR-149-5p overexpression significantly inhibited the formation of microtubule nodules in CAL-27 cells (P=0.002), reduced the expression level of VEGF protein (P=0.000), and reversed the upregulation of VEGF expression caused by overexpression of FGF5 (P=0.000). Conclusion: miR-149-5p can inhibit EMT and microtubules formation and apoptosis promotion of OSCC cells. The mechanism of action is related to the targeted inhibition of FGF5 expression.

Abstract:Objective:　To investigate the protective effect of carvacrol on astrocytes injury in spontaneous type 2 diabetic db/db mice. Methods:　Twenty eight-week-old male db/db mice were randomly divided into model group and carvacrol group, and 10 db/m mice were randomized to the control group. The carvacrol group mice were given 20 mg/kg intragastric administration of carvacrol, and the control group and the model group were respectively given the same amount of intragastric administration of saline, once a day, for 6 weeks. The morphological changes of astrocytes in CA1 area of hippocampus in different groups were observed by labeling glial fibrillary acidic protein (GFAP) through immunohistochemistry. The number of expressed positive cells of lipocalin 2 (LCN2) and aquaporin 4 (AQP4) in CA1 region of control group, model group, and carvarol group were determined by immunohistochemistry. TUNEL was used to detect the relationship among LCN2, AQP4 and astrocyte apoptosis in CA1 region. RT-PCR and Western blot were used to detect the expression of LCN2, AQP4, nuclear factor kappa B (NF-κB) and neutrophilic alkaline phosphatase-3 (NALP3) in CA1 area of hippocampus in control group, model group and carvacrol group. Results:　The expression levels of glucose (GLU), high density lipoprotein (HDL), triglyceride (TG) and total cholesterol (TC) in the model group were significantly higher than those in the control group, while the expression levels of GLU, HDL, TG and TC in the carvacrol group were significantly lower than those in the model group (all P<0.05). Immunohistochemistry showed that the number of GFAP labeled astrocytes in the model group increased and the number of branches increased, while the carvacrol group could improve the morphology of astrocytes and reduce the number (all P<0.01). Immunohistochemistry also showed that the number of positive cells of LCN2, NF-κB and NALP3 in the model group increased, while the number of positive cells of AQP4 decreased. However, in the carvacrol group, the number of positive cells of AQP4 increased, and the number of positive cells of LCN2, NF-κB and NALP3 decreased (all P<0.01). TUNEL staining showed that LCN2 and apoptotic cells increased, while AQP4 decreased significantly in the model group, while the number of apoptotic cells and LCN2 positive cells and AQP4 positive cells increased in the carvacrol group compared with the model group (all P<0.01). The results of RT-PCR and Western blot showed that the expression levels of LCN2, NALP3 and NF-κB in hippocampus of carvacrol group were decreased, and the expression level of AQP4 was increased compared with those of model group (all P<0.01). Conclusion: Carvacrol has protective effect on astrocytes injury in spontaneous type 2 diabetic mice, and its mechanism may be related to the decrease of LCN2, NF-κB and NALP3, and the increase of AQP4 expression level.

Abstract:Objective:　To study the interaction between the DH domain of VAV2 protein (VAV2-DH) and CDC42 by isothermal titration calorimetry. Methods:　The coding sequences of VAV2-DH and CDC42 were cloned into pGEX-6p-1 or pET28a vectors to construct recombinant prokaryotic expression plasmids. Purified proteins were obtained by prokaryotic expression, affinity chromatography and gel filtration chromatography. Then GEF assay and isothermal titration calorimetry were employed to detect the affinity and thermodynamic parameters of the interaction between VAV2-DH and CDC42. Results:　Moderate binding affinity was recorded between VAV2-DH and CDC42 with a K_Dvalue of (11.44±2.12) μmol/L, in which the molar binding enthalpy DH= (-224.33±95.48) kJ/mol, -TDS= (195.97±95.95) kJ/mol, and Gibbs free energy DG= (-28.30±0.49) kJ/mol. Conclusion: VAV2-DH and CDC42 protein interact with each other with moderate intensity, which is an enthalpy-driven binding process.

Abstract:Objective:　To explore a new method to measure the diaphragmatic contraction function by comparing the accuracy of the ultrasonic area method with excursion method. Methods:　Seventy-eight healthy volunteers from Shengjing Hospital of China Medical University from May 2019 to April 2020 were selected in this study, and their general information was collected. Bedside ultrasound was used to measure the movement of the diaphragm and the change of the thoracic area during quiet breathing and deep breathing. Taking the deep inspiratory capacity (IC) of lung function as the evaluation standard, Spearman correlation test was used to compare the correlation between the two methods and IC. Results:　There was no statistical difference in age, BMI and gender between the two measurement methods. There were significant differences in the correlation between the change of thoracic area and IC during quiet breathing (r=0.486, P=0.000) and the correlation between the change of diaphragmatic movement and IC during quiet breathing (r=0.245, P=0.031), and there were also statistical differences in the correlation between thoracic area and IC during quiet breathing (r=0.424, P=0.000) and between diaphragmatic movement and IC during deep breathing (r=0.285, P=0.012). The correlation between the change of thoracic area and IC measured by ultrasonic area method was stronger than that measured by excursion method (the quiet breathing area difference>the quiet breathing diaphragmatic movement, the deep breathing area difference>the deep breathing diaphragmatic movement), and the P value was significantly lower than that of the excursion method (the quiet breathing area difference<the quiet breathing diaphragmatic movement, and the deep breathing area difference<the deep breathing diaphragmatic movement). Conclusion: The area method is superior to the excursion method in the evaluation of diaphragmatic contraction function, and it’s more accurate to evaluate diaphragmatic contraction function combined with excursion method and diaphragmatic thickening rate.

Abstract:Objective:　To quantitatively evaluate the multifidi (MF) degeneration by using multi-echo 3.0 T magnetic resonance (MR) chemical shift-encoding water-fat imaging (Dixon) sequence in patients with degenerative scoliosis (DS) and contrast with those in non-DS volunteers. Methods:　Sixty subjects who underwent 3.0 T MR lumbar examination using Dixon sequence from July 2018 to June 2020 in our hospital were included in this study, with thirty DS patients (DS group) and thirty non-DS volunteers (control group). The fat infiltration (FI) and cross sectional area (CSA) at the MF on levels L4 through L5 (L4/5) of all subjects were measured by two radiologists using the double-blind method to compare consistency. The mean percentage of FI (FI%), relative CSA (RCSA), functional CSA (FCSA), asymmetry ratio of area (AAA) and asymmetry ratio of fat infiltration index (AAFI) of bilateral MF at L4/5 were calculated, and the differences were compared between DS and control groups. The statistical significance of Oswestry disability index (ODI), sum of visual analogue scale scores (VAS) at lumbar and legs and Roland-Morris disability questionnaire (RMD) scores between DS patients and control volunteers were also compared. Receiver operating characteristic (ROC) curve of FI%, AAFI, ODI, RMD and VAS were calculated. Results:　Interobserver agreement of FI%and CSA was excellent between the two radiologists (ICC>0.9). The FI%, RCSA, FCSA and AAFI of bilateral MF were statistically significant (P<0.05). ODI, VAS and RMD obtained from DS group showed a significantly higher scores than those from control group. Areas of FI%, AAFI, ODI, RMD and VAS under the ROC curve were respectively 0.818, 0.754, 0.984, 0.947 and 0.933.Conclusion: The FI%and AAFI of bilateral MF at L4/5 level calculated by multi-echo Dixon image will reflect the worse pain and quality of life of DS patients, which has good consistency with clinical factors, with a valuable diagnostic efficacy to identify non-DS volunteers.

Abstract:Objective:　To investigate the clinical diagnostic value of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in the peripheral blood of patients with oral squamous cell carcinoma (OSCC). Methods:　Fluoresence quantitative RT-qPCR was used to detect the expression of MALAT1 in the preoperative peripheral blood samples and paired postoperative tumor tissues of patients with OSCC. Results:　The mean expression level of MALAT1 in peripheral blood of patients with OSCC was significantly higher than that of healthy volunteers (1.845±0.142 vs.0.642±0.090, P<0.001). The expression level of MALAT1 in peripheral blood was positively correlated with the paired tumor tissues with a correlation coefficient of 0.783 (P<0.001). The expression level of MALAT1 in peripheral blood was correlated with tumor TNM stage (P=0.001) and cell differentiation (P=0.010). The area under the ROC curve of MALAT1 for the diagnosis of OSCC was 0.892.When the cutoff value was 0.788, the diagnostic sensitivity and specificity of MALAT1 were 85%and 81%respectively. Conclusion: MALAT1 has potential clinical value of diagnosing OSCC, and is expected to be a serum molecular marker for the diagnosis and prognosis of OSCC.

Abstract:Objective:　To study the incidence of post-discharge adverse events occurring after dental treatment under general anesthesia in pediatric patients aged 1-6 years and compare their distribution differences in different age groups. Methods:　A total of 110 children, including 72 boys (65.5%) and 38 girls (34.5%), American Society Anesthesiologists physical statusⅠorⅡ, who received oral treatment under general anesthesia in the Stomatological Clinic of The First Affiliated Hospital of Zhengzhou University from January 2019 to February 2020 were collected. According to the age of the children, they were divided into two groups: infant group (1-3 years, 58 cases) and preschool group (4-6 years, 52 cases). The telephone follow-up records of first 24 hours after surgery were inquired to summarize the incidence of adverse events, including crying, toothache, oral discomfort, emotional instability, fever, drowsiness, bleeding gums, red lips, salivation, mouth breathing and whether the children had any memory of the dental experience, etc, and their distribution differences in different age groups were compared. Results:　In this study, 62.7%of children had one or more adverse events reported. The incidence of adverse events in the infant group and preschool group were 55.2%and 71.1%, respectively (P>0.05). Crying (41.3%) and bad memory of dental visits (40.4%) were the highest incidence of adverse events in the infant group and preschool group, respectively. The incidence of memory, emotional instability and irritability and toothache in the infant group were 13.8%, 6.9%and 8.6%, respectively, which were all lower than those in the preschool group (40.4%, 21.2%, 23.1%, respectively), with statistical differences (P<0.05). The number of treated teeth, the number of extracted teeth, and the number of endodontic treatments in the infant group were (11.5±2.9), (1.6±1.4) and (5.3±3.3), respectively, which were significantly lower than those in the preschool group [(13.3±3.5), (3.2±1.1) and (7.5±4.1), respectively] (P<0.05). The number of total coronary treatments in the two groups was (3.5±2.3) and (4.6±3.7), respectively, without significant difference (P>0.05). Conclusion: In oral treatment under general anesthesia, the effect of intraoperative sedation and analgesia is positive. However, the incidence of discomforts occurring after general anesthesia, such as crying, bad memory of medical experience and toothache, is relatively high, and necessary attention should be given. For infants and young children, the comfort of their crying should be strengthened. Due to the large number of teeth and complex dental conditions in preschool children, the probability of adverse events is higher, and the effect of perioperative forgetting and analgesia should be strengthened.

Abstract:Objective:　To investigate the characteristics and clinical outcomes of prenatal ultrasound diagnosis of fetal cardiac rhabdomyoma. Methods:　The ultrasonographic characteristics and clinical outcomes of 15 fetuses with prenatal diagnosis of cardiac rhabdomyoma in our hospital from August 2016 to August 2019 were retrospectively analyzed. Results:　In 15 fetuses with cardiac rhabdomyoma, 8 cases (53.3%) were single and 7 cases (46.7%) were multiple, with left ventricle as most common favorite site, and prenatal ultrasound showed uniform hyperechoic mass. The other structural malformation were not detected in 13 fetuses (86.7%), 1 case accompanied with ventricular septal defect and 1 case with multiple nodules in the brain parenchyma. Pregnancy outcome: 9 fetuses (60.0%) were terminated, 5 fetuses (33.3%) delivered at term, and 1 fetus lost follow-up. In the 5 full-term delivered fetuses, cardiac rhabdomyoma still presented in 3 cases, and tuberous sclerosis complex (TSC) were not found. Cardiac rhabdomyoma disappeared in 1 case and no other abnormalities were found, and TSC in many organs were found in 1 case. Conclusion: Prenatal ultrasound of fetal cardiac rhabdomyoma has typical features: single or multiple round-like, uniform echogenic masses in the heart cavity, well-demarcated, and attached to the ventricular wall or septum. The condition without TSC has good prognosis, and its prognosis can be guided by strengthening doctor-patient communication in clinical work, combined with magnetic resonance imaging (MRI) and genetic test.

Abstract:Objective:　To investigate serum proprotein convertase subtilisin/kexin type 9 (PCSK9) concentrations in childbearing-aged female patients with systemic lupus erythematosus (SLE) combined with acute myocardial infarction (AMI), and to analyze the correlation between PCSK9 levels and clinical indexes. Methods:　Thirty childbearing-aged female patients with SLE combined with AMI (SLE-AMI group), thirty age-matched female SLE patients without AMI (SLE-non-AMI group), and thirty female healthy individuals (control group) were included in our research. Cardiovascular disease (CVD) risk factors and serum PCSK9 levels were compared among three groups. Changes of PCSK9 concentrations, lipids profile and inflammatory indexes before and after therapies were compared in SLE-AMI group. Univariate correlational analysis of serum PCSK9 levels before treatment and disease parameters in SLE-AMI patients was conducted. Results:　No difference of lipids profile was found among the three groups. Patients in SLE-AMI group had significantly higher PCSK9 levels, proportion of antiphospholipid antibody positivity and inflammatory markers than patients in SLE-non-AMI group. And serum PCSK9 levels declined vitally after therapies. In addition, higher PCSK9 levels were observed in SLE-AMI patients who had positive result of antiphospholipid antibody or antiphospholipid syndrome (APS) than those patients with negative results or without APS, respectively. Univariate and multivariate regression analysis suggested that serum levels of PCSK9 in SLE-AMI patients were correlated with C-reactive protein level and coexisted APS, without significant correlation with disease activity and lipids profile. Conclusion: Significant elevation of PCSK9 concentrations can be observed in childbearing-aged female patients with SLE combined with AMI. Moreover, PCSK9 levels are associated with clinical outcomes, inflammatory markers, and combination of APS, but not lipid profile.

Abstract:Objective:　To explore the clinical treatment effects of two surgical methods of typeⅢcesarean scar pregnancy (CSP) vaginal and laparoscopic focal excision repair (FER) with fertility requirements. Methods:　Retrospective analysis was made on 55 cases of typeⅢCSP with fertility requirements taking non-invasive assisted vaginal FER and 36 cases of laparoscopic FER admitted to Chongqing Health Center for Women and Children from January 2017 to December 2019. Results:　The operation went smoothly among 88 cases. The bleeding during vaginal FER operation was (116.6±33.3) mL, and the operation time was (68.0±13.3) min; the bleeding during laparoscopic FER operation was (150.6±31.6) mL, and the operation time was (107.0±13.3) min. There were significant differences in the bleeding volume and operation time among the two kinds of operations (P<0.05). The hospitalization cost of vaginal FER was significantly lower than that of laparoscopic FER (P<0.05). The scar thickness of female FER incision [(4.66±0.30) mm]was significantly greater than that of laparoscopic FER [(4.06±0.37) mm]in one-year of ultrasound follow-up after operation (P<0.05), and there was no significant difference in pregnancy after 18 months (P>0.05). Conclusion: There are advantages and disadvantages of the two surgical methods for typeⅢCSP, and uterus artery embolization (UAE) can be avoided in non-emergency situations.