The study adopted muscle injection of pVAX1-SP-GHRH(1-44)expression plasmid and electrostimulation to determine its effects on mouse growth and sow production performance.One hundred and fifty four-week-old C57 BL/6 female mice were randomly divided into 6 groups of 5 replicates each.Muscle single-injection followed by electrostimulation was performed.The con-trol group received an empty plasmid injection(80 μg/kg),while the treatment groups received pVAX1-SP-GHRH(1-44)plasmid injections(20,40,80,120,160 μg/kg).Twenty healthy preg-nant sows were randomly divided into 2 groups,each with 10 sows.Electrostimulation treatment was applied to the semimembranosus muscle of the pregnant sows after a single injection.The con-trol group received physiological saline injection,while the plasmid group received a 2 mg pVAX1-SP-GHRH(1-44)expression plasmid injection.Mouse weight,feed intake,and serum GHRH and IGF-1 levels were measured at days 0,7,14,21,and 28 after injection.Pregnant sows were bled via the tail vein at days 0,14,28,and 42 after injection,and their serum was separated to measure serum GHRH and IGF-1 levels.The birth weight,placental weight,number of piglets born,number of healthy piglets,number of weak piglets,number of deformed piglets,number of stillborn piglets,and number of mummified piglets were recorded at day 14.The mouse study re-sults showed that muscle injection of pVAX1-SP-GHRH(1-44)plasmid followed by electrostim-ulation could promote mouse feeding and increase weight gain(P＜0.05),significantly increase mouse serum GHRH and IGF-1 levels(P＜0.05),and maintain its effects until day 21.The results of the pregnant sow study showed that the average birth weight of the piglets in the plasmid group was significantly increased(P＜0.01),and the placenta weight was significantly increased(P＜0.05).The serum GHRH and IGF-1 concentrations in the plasmid group sows were significantly increased(P＜0.01).The study results showed that muscle injection of pVAX1-SP-GHRH(1-44)expression plasmid followed by electrostimulation could promote mouse feeding and increase weight gain,and also significantly improve the average birth weight and placental weight of the piglets in pregnant sows.

Objective To analyze thein vivo three-dimensional dose verification using electronic portal imaging device(EIVD)for intensity-modulated radiotherapy(IMRT)of cervical cancer for investigating the differences between the measured and planned doses,and explore the optimal threshold for gamma passing rate in EIVD quality control based on dosimetric sensitivity.Methods A retrospective analysis was conducted on a cohort of 45 patients with cervical cancer who underwent IMRT at Women's Hospital,School of Medicine,Zhejiang University.During the treatment,all patients underwent EIVD to obtain the measured doses.The passing rate was analyzed using global gamma criteria of 2 mm/2%,2 mm/3%,and 3 mm/3%.Additionally,dose-volume histogram parameters were utilized to evaluate any differences between the measured and planned doses.Pearson correlation analysis was employed to investigate the relationship between the gamma passing rate and dosimetric differences.Furthermore,receiver operating characteristic(ROC)curve was generated to determine the optimal threshold for the gamma passing rate.Results The average gamma passing rates for the criteria of 2 mm/2%,2 mm/3%,and 3 mm/3%were 83.07%±5.25%,91.69%±3.52%,and 95.02%±2.46%,respectively.The Dmean deviation between EIVD measurement and planned dose in the planning target area was 2.43%(P=0.016),while the Dmean deviations in the bladder,rectum,and small intestine were 0.35%,0.46%,and 0.30%,respectively(P>0.05).Pearson analysis revealed a strong correlation between the 3 gamma indexes and dosimetric differences in the PTV(r>0.7),but a weak correlation with organs-at-risk(r<0.7).ROC analysis indicated that the optimal gamma passing rate thresholds for the criteria of 2 mm/2%,2 mm/3%,and 3 mm/3%were 79.06%,90.04%,and 94.19%,respectively.Conclusion The implementation of EIVD can ensure the accuracy of dose delivery within the PTV during IMRT for cervical cancer.Moreover,establishing a gamma passing rate threshold provides a valuable clinical basis for subsequent adaptive IMRT for cervical cancer.

Objective:To explore the pathogenesis of flunarizine-induced parkinsonism from gut-brain axis perspective.Methods:Thirty male C57BL/6 mice were randomly divided into control group and flunarizine group ( n=15). Mice in the control group were given 0.1 mL 50% polyethylene glycol 400+50% saline by gavage once/d for 2 weeks, while mice in the flunarizine group were given 6 mg/mL flunarizine+50% polyethylene glycol 400+50% saline by gavage at a daily dose of 30 mg/kg for 2 weeks. Body mass was recorded 1, 3, 5, 7, 10 and 14 d after drug administration, and motor function was assessed by rotarod test 14 d after drug administration; 16s RNA sequencing was performed in the feces to observe the intestinal flora; intestinal transit function was detected by Evans blue by gavage; and then, the mice were sacrificed and homogenate or frozen sections (brain and intestinal tissues) were prepared; dopamine-ergic neuron expression was detected by Western blotting; RT-qPCR was applied to detect the expressions of inflammatory factors in the substantia nigra, and immunofluorescent staining was used to detect the expressions of ZO-1 and Claudin-5 in the intestinal epithelial tissues. Results:Compared with the control group, the flunarizine group had lower body mass ratio 1, 3, 5, 7, 10 and 14 d after drug administration (ratio to body mass before drug administration). Compared with the control group, the flunarizine group had significantly shortened residence time in rod rotating and lower rotational speed when falling ( P<0.05). Compared with the control group, the flunarizine group had decreased tyrosine hydroxylase protein in the substantia nigra without significant difference ( P>0.05). Compared with the control group, the flunarizine group had significantly increased interleukin-6 and tumor necrosis factor-α in the substantia nigra (1.00±0.00 vs. 2.79±0.83; 1.00±0.00 vs. 3.39±1.37), significantly lower intestinal Evans blue propulsion rate (80.67%±4.51% vs. 50.67%±6.03%), and statistically decreased ZO-1 and Claudin-5 expressions in the colonic epithelial tissues (27.01±1.41 vs. 16.32±2.83; 37.00±2.80 vs. 24.52±2.12, P<0.05). Totally, 576 microorganisms were noted in both control group and flunarizine group, 744 in the control group alone, and 634 in the flunarizine group alone. The intestinal flora β diversity indices in the 2 groups were significantly different based on weighted Unifrac-principle coordinates analysis (PCoA, PCoA1: 39.88%; PCoA2: 30.69%). Compared with the control group, the microbial colony structure of mice in flunarizine group was dominated by phylum thick-walled bacteria and phylum warty microbacteria, and by families Muribaculaceae, Lachnospiraceae and Akkermansiaceae. Compared with the control group, the flunarizine group had significantly decreased relative abundance of Ackermannia spp. and Lactobacillus spp. in the intestinal flora ( P<0.05). Conclusion:Flunarizine may contribute to the pathogenesis of DIP by causing structural disturbances in the intestinal flora and inducing neuroinflammation based on the gut-brain axis.

Objective: To evaluate the efficacy and safety of low dose sublingual nifedipine dripping pills (5 mg) in treating moderate and severe hypertension in comparison with normal dose (10 mg) of sublingual nifedipine dripping pills. Methods: This study was designed as a randomized, double-blind, positive drug parallel controlled, multi-center, non-inferiority clinical trial. Patients with moderate and severe hypertension were enrolled by 14 clinical trial centers, randomly divided into the trial group (sublingual 5 mg nifedipine dripping pills) and the control group (sublingual 10 mg nifedipine dripping pills). The changes in blood pressure were monitored continuously within 2 hours after the initial administration, repeated the dose in 20 minutes interval after the initial administration for up to additional 3 doses (maximum 4 doses) if the antihypertensive efficacy was not satisfactory. The efficacy of antihypertensive therapy between the two groups was evaluated by repeated administration rates and blood pressure changes at 60 minutes post the initial administration, and the safety of treatment was evaluated by recording adverse event rate of the two groups. Results: The anti-hypertensive effective rates at 60 minutes after sublingual administration were 83.5% (202/242) and 86.7% (208/240) respectively between the trial group and control group (χ²=1.307, P=0.253) . On the aspect of antihypertensive effectiveness at 60 minutes after single dose of sublingual administration, the anti-hypertension effective rates of the trial group and the control group were 85.6% (154/180) and 87.2% (164/188) respectively (χ²=0.221, P=0.639). Prevalence of the repeated administration was also similar between the two groups (25.6%(62/242) in the trial group and 21.7% (52/240) in the control group, χ²=1.043, P=0.307). On the safety aspect, there was no adverse events/reactions in the trial group, but there were 15 cases of adverse events/reactions occurred in control group (6.25%, χ²=15.611, P<0.001). Conclusions In the treatment of moderate to severe hypertension, the antihypertensive efficacy of low dose nifedipine dripping pills is similar to that of conventional dosage, and the safety profile of low dose nifedipine dripping pills is better than that of the conventional dose.

Objective To study the changes of immune function of endotoxin tolerant dendritic cell (ETDC) and to observe its effect on sepsis in mouse model.Methods ETDC were prepared by pretreated bone marrow dendritic cells derived from BALB/c mice with lipopolysaccharide stimulation.The cells were collected and the expressions of surface markers including major histocompatibility complex ( MHC)Ⅱ, CD86 and CD11c were detected by flow cytometry.The proliferation rate of T lymphocytes was evaluated by cell counting kit-8 and the concentrations of cytokines in the supernatant were detected by enzyme linked immuno sorbent assay.Afterwards, 36 mice were randomly assigned into 4 groups.The blank control group did not receive any treatment, the sham-operated group underwent simple incision suture, the sepsis group and ETDC reinfusion group underwent cecal ligation and puncture to establish sepsis.Before sepsis model establishment, 0.9% sodium chloride solution or suspension of ETDC and 0.9%sodium chloride were reinfused by tail vein.The serum levels of alanine aminotransferase (ALT) and aspartate transaminase (AST), tumor necrosis factor (TNF)-α, interleukin(IL)-6 and IL-10, and the proportion of help T cell ( Th) 17/regulatory T cell ( Treg) in spleen of each group were detected.The pathological manifestations of liver, kidney and ileum in each group were observed.T test and χ2 test were used for comparisons between groups.Results The results of flow cytometry showed that MHCⅡ, CD86 and CD11c of ETDC were 70.4%, 43.1%, and 73.1%, respectively, which were significantly lower than those of mature dendritic cell (mDC) (96.1%, 89.5%, and 84.6%, respectively) (χ2 ＝56.47, 83.78, and 23.29, respectively, all P＜0.01).The concentrations of IL-10, TNF-αand IL-6 in the supernatant of ETDC were (978.04 ±56.70), (980.34 ±111.96) and (12 743.03 ±865.81) ng/L, respectively, and those of mDC were (741.35 ±99.23), (1 703.11 ±117.00) and (19 052.28 ±1 145.84) ng/L, respectively.The differences were all statistically significant (t＝5.073, 10.93, and 10.76, respectively, all P＜0.01).The proliferation rates of T lymphocytes co-cultured with ETDC in 1∶5 and 1∶10 ratio group were (676.95 ±85.99)%and (514.00 ±106.39)%, respectively, which were lower than those of the mDC group (956.25 ±127.12)%and (772.07 ±214.08)%, respectively.The pathological injuries of liver, kidney and ileum in ETDC treatment group were significantly lighter than those in sepsis group.Serum ALT and AST levels in the ETDC reinfusion group were (299.71 ±36.91) and (690.39 ±154.92) U/L, respectively, and TNF-α, IL-6 and IL-10 were (0.067 ±0.005), (0.428 ±0.051) and (0.058 ±0.005) ng/L, respectively.Serum ALT and AST levels in the sepsis group were (620.67 ±69.27) and (1 430.17 ±134.05) U/L, respectively, and TNF-α, IL-6 and IL-10 were (0.085 ±0.007), (0.774 ±0.088) and (0.036 ±0.005) ng/L, respectively.The differences were all statistically significant (t＝11.60, 10.96, 5.991, 8.657, and 8.04, respectively, all P ＜0.01).The proportion of Treg/Th17 in the ETDC reinfusion group was 23.4%, and that in the sepsis group was 60.8%(χ2 ＝28.69, P＜0.01).Conclusion The reinfusion of ETDC has a protective effect on sepsis in mouse model, which may play a negative immune regulatory role by regulating the differentiation of T cells.

OBJECTIVETo assess the association of CYP2C19 gene polymorphisms with the incidence of ischemic stroke among patients receiving clopidogrel therapy following coronary stenting for coronary artery disease.

METHODSClinical data of patients receiving clopidogrel therapy after coronary stenting were retrospectively studied. For a case-control study, 137 patients with acute cerebral infarction and 122 non-stroke patients were selected. Based on the variants of the CYP2C19 gene detected by a DNA microarray assay, the patients were further divided into the wild-type group(CYP2C19*1/*1) and mutant group(defined by the presence of at least one loss-of-function allele, including CYP2C19*1/*2, CYP2C19*1/*3, CYP2C19*2/*2, CYP2C19*2/*3 and CYP2C19*3/*3). The incidences of ischemic stroke in the two groups were compared through a chi-square analysis. The influence of CYP2C19 gene polymorphisms and clopidogrel therapy on the incidence of ischemic stroke was analyzed through multivariable logistic regression.

RESULTSA total of 259 patients were enrolled. The case and control groups showed no difference in terms of gender and age. There were 123 cases (47.5%) in the CYP2C19 wild-type group and 136 cases (52.5%) in the mutant group. The incidence of ischemic stroke of mutant group was significantly higher than that of wild-type group (59.9% vs. 44.3%, X2=6.398, P=0.042). Multivariate analysis revealed that loss-of-function polymorphisms of the CYP2C19 gene carried a 1.13 times greater risk for ischemic stroke compared to wild-type genotype (OR=2.13, 95%CI: 1.23-3.71).

CONCLUSIONThe efficacy of clopidogrel for the prevention of ischemic stroke in post-coronary stent patients may be reduced by the insufficiency of the CYP2C19 gene. The dosage of clopidogrel therapy should be adjusted based on its polymorphisms.

Brain Ischemia ; prevention & control ; Cytochrome P-450 CYP2C19 ; genetics ; Genotype ; Humans ; Percutaneous Coronary Intervention ; adverse effects ; Platelet Aggregation Inhibitors ; therapeutic use ; Polymorphism, Genetic ; Stents ; adverse effects ; Stroke ; prevention & control ; Ticlopidine ; analogs & derivatives ; therapeutic use

AIM: To investigate whether rs7903146 polymorphisms in transcription factor 7-like 2 (TCF7L2) gene are associated with susceptibility to type 2 diabetes mellitus (T2DM) in Chinese Uygur population.METHODS: In this case-control study, 935 cases of T2DM patients were recruited in T2DM group, and 971 healthy examination individuals were selected as normal control.The TCF7L2 gene polymorphism was detected by matrix-assisted laser desorption/ionization time-of-flight mass spectrum.RESULTS: Significant differences in the frequencies of CC, CT and TT genotypes and the frequencies of C and T alleles on TCF7L2 rs7903146 were found between T2DM group and control group(P<0.05).As compared with C allele, the patients with T allele had a significantly higher risk of T2DM with OR of 1.190 (95% CI: 1.034~1.371).As compared with CC genotype, the patients with CT genotype had a significantly higher risk of T2DM with OR of 1.374 (95% CI: 1.122~1.683), and the patients with CT+TT genotype had a significantly higher risk of T2DM with OR of 1.307 (95% CI: 1.090~1.567).The levels of fasting plasma glucose, serum creatinine and blood urea nitrogen were higher in all participants with CT+TT genotype of rs7903146 than those with CC genotype, which showed a significant difference (P<0.05).CONCLUSION: The polymorphisms of rs7903146 in TCF7L2 gene may be associated with T2DM in Uygur population from Xinjiang region.The T allele and CT genotype of rs7903146 are the risk factors for T2DM.

Objective To investigate the role of miR-592 in the Glioma.Methods We first analyzed the expression of miR-592 in Glioma tissues from patients by quantitative PCR.We transfected U251 cells with miR-592 mimics and then detected the growth of cells by MTT assay.We performed dual-luciferase reporter assay and western blot assay to examine whether Runx2 was the direct target of miR-592 in U251 cells.In order to test whether Runx2 was the functional target of miR-592,we determined the cell growth curve by down-regulating the level of Runx2.Moreover,we also detected the apoptosis of U251 after Runx2 knockdown.Results The expression of miR-592 was significantly reduced in glioma tissues (t=2.752,P--0.013).Over-expression miR-592 remarkably increased the apoptotic rate of U251 cells compared with the control group (t=2.127,P=0.031;t=2.284,P=0.026).Flow cytometry analysis showed that MiR-592 significantly promoted apoptotic cell death of U251 cells Apoptosis rate was 7.2％±0.68％ in miR-592 group and 17.47％±1.45％ in control group (t=3.294,P=0.007).The results of double luciferase assay and Western blot assay showed that miR-592 directly targeted the 3'Runx2 of-UTR to inhibit the level of Runx2 protein.The effect of down-regulation of Runx2 on the growth of U251 cells was detected,the results showed that growth was significantly slower in the cells transfected with Runx2 siRNA than in those without Runx2 siRNA (t=3.124,P=0.O11).Detection of cycle by flow cytometry showed that runx2 down-regulated the apoptosis rate of U251 cells.Tumor growth curve showed that overexpression of miR-592 significantly inhibited tumor growth and the down regulation of Runx2 expresssion also significantly inhibited tumor growth.Conclusion miR-592 suppresses the growth and promotes the apoptotic rate of U251 cells by targeting Runx2.

Objective To assess the clinical value of lung ultrasound (LUS) B-lines in diagnosis of rheumatoid arthritis (RA) associated interstitial lung diseases (RA-ILD).Methods Forty-five consecutive patients with RA who underwent a high resolution computed tomography (HRCT) scan of the chest,were also examined by LUS for detection of B-lines(within 1 month independently in all patients).The B-lines score was obtained by summing the number of total 50 inter-costal spaces (ICSs) of chest wall.Pulmonary fibrosis was quantified by HRCT as previously described by the 30-point Warrick score.Results B-lines score significantly correlated with the Warrick score [(r=0.778,95％CI(0.627,0.872),P＜0.05].Receiver operating characteristic (ROC) curve confirmed that B-lines cut-off point 77[sensitivity of 100％,specificity of 64.3％ respectively,area under curve [AUC] =0.86,95％CI(0.724,0.945)] and 108[sensitivity of 90％,specificity of 88.6％ respectively,AUC=0.879,95％CI(0.747,0.957)] had an optimal power to discriminate mild (Warrick score＜8) and severe fibrosis (Warrick score＞15):Conclusion The data confirm that LUS is a useful technique to identify ILD in RA.In RA-ILD,B-lines correlate significantly with HRCT and are able to identify mild and severe degree of fibrosis.LUS is a promising non-invasive and non-ionizing strategy for screening RA-ILD.