ObjectiveTo establish an allele-specific polymerase chain reaction （PCR） method for identifying Scolopendra dispensing granules， so as to ensure the quality and therapeutic effects of Scolopendra and its preparations. MethodThe primer interval suitable for the PCR was selected based on the cytochrome c oxidase subunit 3（COX-3） gene sequence of Scolopendra， and the single nucleotide polymorphism （SNP） loci of Scolopendra and its adulterants were mined from the interval for the design of specific primers. The samples of Scolopendra and its adulterants were collected. The PCR system was established and optimized regarding the annealing temperature， cycles， Taq enzymes， DNA template amount， PCR instruments， and primer concentrations， and the specificity and applicability of this method were evaluated. ResultThe PCR system was composed of 12.5 μL 2×M5 PCR Mix， 0.4 μL forward primer （10 μmol·L^-1）， 0.4 μL reverse primer （10 μmol·L^-1）， 2.5 μL DNA template， and 9.2 μL sterile double distilled water. PCR parameters： Pre-denaturation at 94 ℃ for 3 min， 30 cycles （94 ℃ for 20 s， 62 ℃ for 20 s， 72 ℃ for 45 s）， and extension at 72 ℃ for 5 min. After PCR amplification with the system and parameters above， the electrophoresis revealed a bright band at about 135 bp for Scolopendra and no band for the adulterants. ConclusionThe established allele-specific PCR method can accurately identify the medicinal materials， decoction pieces， and standard decoction freeze-dried powder of Scolopendra， as well as the intermediates and final products of Scolopendra dispensing granules， which is of great significance for ensuring the quality and clinical efficacy of Scolopendra and its preparations.

Objective To investigate the causal association between bronchial asthma and bone mineral density at different sites using a two-sample Mendelian randomization (MR) approach. Methods Summary data for exposure factors and outcome were obtained from different genome-wide association studies.Single nucleotide polymorphisms strongly associated with bronchial asthma were selected as instrumental variables,and those in linkage disequilibrium were excluded.The inverse-variance weighted (IVW) method was used as the primary method for MR analysis,complemented by weighted median,simple mode,weighted mode,and MR-Egger regression methods.Sensitivity analyses were conducted to assess the stability of the results. Results The random-effects model of IVW analysis showed that heel bone mineral density (OR=0.986;95% CI,0.974 to 0.998;P=0.023) as the outcome dataset had a reverse causal effect with bronchial asthma,while lumbar spine bone mineral density (OR=1.031;95% CI,0.984 to 1.081;P=0.195),femoral neck bone mineral density (OR=1.014;95% CI,0.973 to 1.057;P=0.505),and forearm bone mineral density (OR=1.011;95% CI,0.935 to 1.094;P=0.775) as outcome datasets showed no causal effect with bronchial asthma.The MR-Egger intercept test results indicated that the P-values for the intercepts of lumbar bone mineral density,femoral neck bone mineral density,forearm bone mineral density,and calcaneal bone mineral density were all over 0.05,suggesting no horizontal pleiotropy and relatively stable results. Conclusion MR analysis reveals a reverse causal effect between bronchial asthma and heel bone mineral density,suggesting that clinicians should strengthen the monitoring of heel bone mineral density in patients with bronchial asthma to timely detect and intervene osteoporosis.

Objective:To investigate the current situation of moisture-associated skin damage (MASD) in patients with permanent colostomy, and analyze its influencing factors.Methods:From January 2019 to January 2020, 115 patients received permanent colostomy in Henan Provincial People's Hospital were selected by convenience sampling method as the research objects. The patients were investigated by the self-designed General Information Questionnaire and Disease Factor Questionnaire.Results:Finally, 107 patients with permanent colostomy were included, and the incidence of MASD was 39.25% (42/107) . One-way analysis of variance showed that age, body mass index, stoma site, stoma height, excrement characteristics, defecation regularity, chassis leakage times were the influencing factors for the incidence of MASD in patients with permanent colostomy ( P<0.05) . Conclusions:The incidence of MASD in patients with permanent colostomy is high. Age, stoma site, stoma height, defecation and so on are the influencing factors of MASD in patients with permanent colostomy. Medical and nursing staff can formulate targeted preventive measures according to the influencing factors to reduce the incidence of MASD.

Objective To study effects of backpack gravity center position on kinetics and kinematics of lower-extremity joints in parachuting landing and evaluate the injuries. Methods Seven participants performed parachuting landing with backpack gravity center on three positions: low-back (position 1), upper-back (position 2) and abdomen (position 3). Results The peak vertical ground reaction force (GRF) with backpack on position 2 was significantly lower than that on position 1. The joint moment on sagittal plane of the hip with backpack on position 2 was significantly higher than that on position 1 and position 3. The joint energy absorption of the hip with backpack on position 2 was significantly higher than that on position 1. The angular displacement of the hip on sagittal plane with backpack on position 2 was significantly higher than that on position 1 and was significantly lower than that on position 3. The angular velocity of the hip on sagittal plane with backpack on position 2 was significantly lower than that on position 3. Conclusions Different positions of backpack gravity center could significantly influence kinetic and kinematic parameters of the hip. Backpack gravity center on upper-back position could decrease the lower-extremity injuries. The results can provide evidences for evaluating backpack gravity center and decreasing injuries in parachuting landing.

Objective:To investigate the effects of isoflurane anesthesia on the transcription levels of clock genes RORα and Rev-erbα and their circadian rhythm changes in the hippocampus and cerebral cortex of mice. Methods:Thirty-six 7-week-old male C57BL/6J mice were divided into control group and isoflurane anesthesia group ( n=18) according to random number table method. They were subjected to 12 h light/12 h dark environment for 1 week. Mice in the isoflurane anesthesia group were anesthetised with 1.2% isoflurane at 22:00 pm on the night before the experiment for 6 h; 4 h after anesthesia, the light time of 8:00 am was taken as the zeitgeber time (ZT0), and then, every 8 h was taken as the recorded time (ZT8, ZT16, ZT24, ZT32 and ZT40). The RORα and Rev-erbα mRNA expressions were measured by real-time fluorescent quantitative PCR (RT-qPCR) at each time point in the cerebral cortex and hippocampus of the two groups. The cosine curves of RORα and Rev-erbα mRNA expressions were analyzed by Chronos-Fit software. Results:As compared with the control group, the isoflurane anesthesia group had significantly down-regulated RORα mRNA expression in the hippocampus, significantly up-regulated RORα mRNA expression in the cerebral cortex, and significantly down-regulated Rev-erbα mRNA expression in the cerebral cortex ( P<0.05). The RORα and Rev-erbα mRNA expressions in the hippocampus and cerebral cortex of the control group showed a rhythmic cycle of about 24 h; while the circadian rhythm of RORα mRNA shifted to the right by 5.41 h, and the circadian rhythm of Rev-erbα mRNA shifted to the left by 0.89 h in the hippocampus of the isoflurane anesthesia group. The peak circadian rhythm of RORα mRNA shifted to the right by 0.35 h, and the peak circadian rhythm of Rev-erbα mRNA shifted to the left by 0.56 h in the cerebral cortex of the isoflurane anesthesia group. Conclusion:Isoflurane anesthesia can induce the changes of RORα and Rev-erbα transcription levels and circadian rhythm changes in the hippocampus and cerebral cortex of mice.

Objective:To investigate the expressions and correlation of bcl-2, programmed death-1 (PD-1) and programmed death ligand-1(PD-L1) in diffuse large B-cell lymphoma (DLBCL) tissue specimens, and the relationship between chemotherapy efficacy and prognosis of DLBCL patients.Methods:The expressions of bcl-2, PD-1 and PD-L1 in 82 patients with DLBCL who were admitted to Chenzhou First People's Hospital of Hunan Province from May 2011 to April 2014 were detected by using immunohistochemistry, and the correlation of the expressions of bcl-2, PD-1 and PD-L1 with clinicopathological characteristics and prognosis was analyzed.Results:The positive rate of bcl-2, PD-L1 and PD-1 in cancer tissues of DLBCL patients was 53.7% (44/82), 56.1% (46/82) and 32.9% (27/82), respectively. There was a correlation between bcl-2 and PD-L1 expression ( r = 0.306, P = 0.005). Bcl-2 was highly expressed in patients with international prognosis index (IPI) score 3-4 points, non-germinal center B-cell-like (non-GCB) subtype and B symptoms (all P < 0.05); PD-1 was highly expressed in patients with IPI score 3-4 points ( P < 0.05); PD-L1 was highly expressed in patients with IPI score 3-4 points, tumor stage Ⅲ-Ⅳ, B symptoms, ≥60 years old, and non-GCB (all P < 0.05). The overall survival (OS) and progression-free survival (PFS) in bcl-2-negative group were better than those in the bcl-2-positive group, and the differences were statistically significant (both P < 0.05); OS and PFS in PD-L1-negative group were better than those in PD-L1-positive group, and the differences were statistically significant(both P < 0.01). There were no statistical differences in OS and PFS between PD-1-positive group and PD-1-negative group (both P > 0.05). OS and PFS in bcl-2 and PD-L1 co-expression group were worse than those in both negative or any negative group (all P < 0.01), and PFS in bcl-2 and PD-1 co-expression group was worse than those in both negative or any negative group ( P = 0.044). Cox multivariate analysis showed IPI score 3-4 points and B symptoms were the independent influencing factors of OS in DLBCL patients (both P < 0.01); IPI score 3-4 points, B symptoms and PD-L1-positive were the independent influencing factors of poor PFS in DLBCL patients (all P < 0.05). Conclusion:The positive expressions of bcl-2 and PD-L1 are the independent factors for the poor prognosis of DLBCL patients, which may become new targets for the treatment of DLBCL.

Objective:To investigate the effects of subanesthetic concentration of sevoflurane on the plasticity of dendritic spines in the prefrontal cortex neurons of juvenile rats.Methods:Thirty-six clean-grade male Sprague-Dawley rats, aged 24 days, weighing 50-60 g, were divided into control group (group C) and sevoflurane anesthesia group (group S) using a random number table method, with 18 rats in each group.Group S inhaled 1.2% sevoflurane and 50% oxygen (flow rate 1 L/min) for 3 h, while group C inhaled 50% oxygen (flow rate 1 L/min) for 3 h. Open-field test and Morris water maze test were performed at 3 days after anesthesia.Animals were sacrificed, and brain samples were then taken for determination of the number of apoptotic neurons in layer Ⅱ-Ⅲ of the prefrontal cortex, density of dendritic spines, and expression of postsynaptic density protein 95 and gephyrin by TUNEL staining, Golgi staining or Western blot.Results:Compared with group C, no significant change was found in total distance or time of staying at the central region in the open-field test or the average swimming velocity, escape latency or the number of apoptotic neurons in the Morris water maze test ( P>0.05), and the density of dendritic spines was significantly increased, and the expression of postsynaptic density protein 95 and gephyrin was up-regulated in group S ( P<0.05). Conclusion:Subanesthetic concentration of sevoflurane can enhance the plasticity of dendritic spines in the prefrontal cortex neurons of juvenile rats.

Teaching rounds is an important part of clinical teaching practice, therefore, we established a demonstration team for teaching ward rounds. By formulating standard operation procedure for teaching rounds and encouraging innovation on teaching models, the team played a demonstration role in the clinical teaching rounds, which not only made up the shortcomings in teaching, but also improved the teaching level of clinical teachers and the quality of clinical training.

Objective To evaluate the anti-HBV effect of hypericin from the cellular level and to preliminarily explore its po-tential drug target point.Methods Liver cell line HepG2.2.15 cells secreting HBV particles were selected as the experimental ob-jects.Hypericin served as the HY group,lamivudine was taken as 3TC group and deionized water as the blank control group.The cells were grouped and administrated.The HBV-DNA copy level was measured at72 h after medication by Southern blot and fluo-rescent quantitative PCR;the inhibition rate of HBsAg and HBeAg was detected by using ELISA assay;the pgRNA expression level was tested by using Northern blot and fluorescent quantitative PCR;Western blot and fluorescent quantitative PCR were adopted to detect the expression of regulatory factors including HNF3β,HNF4α,PPARαand RXRα.Results Compared to the blank control group,both hypericin and lamivudine had significant inhibiting effect on HBV DNA and expression level of HBsAg and HBeAg in HepG2.2.15 cells (P <0.05).Hypericin could significantly decrease the pgRNA expression compared with the blank control group (P <0.05),while lamivudine had no obvious change (P <0.05).Moreover,hypericin exhibited significant effects on the expression of HNF3βand regulatory factor HNF4αcompared with the blank control group and 3TC group(P <0.05).Conclusion Hypericin represents a strong anti-HBV effect,moreover could increase the negative regulatory factor HNF3βn expression and decreases the positive factor HNF4αexpression,prompting that its drug target point could be pgRNA.

Objective To analyze the effect of GC-rich DNA fragments on the level of transgenic expression in Chinese hamster ovary (CHO) celts and its position effect.Methods The synthetic DNA fragment with GC-rich was cloned into the 5'or 3'or both 5'and 3'ends of expression cassette of expression vector.Three new expression vectors (pIRES-G1,pIRES-G2 and pIRES-G3) which was inserted with the GC-rich DNA fragments in different position were transfected CHO ceils,respectively,and then was observed under fluorescence microscope;the control vector was pIRES-EGFP.Stable transfected cell lines were screened under G418,and enhanced green fluorescent protein(EGFP) expression was analyzed by flow cytometry and the transgenic copy number was detected by quantitative real-time quantitative polymerase chain reaction (qRT-PCR).Results Three expression vectors with a GC-rich DNA fragments in different position were constructed successfully.The insertion of GC-rich DNA fragments at 3'end and both 5',3'ends of the box of expression vector could obviously improve the expression level of vector in CHO cells;and the expression level of the stably transfected CHO cells increased 1.39 fold and 1.32 fold compared to the control vector,respectively;the transgene copy number increased 1.32 fold and 1.24 fold compared with the control vector.While the insertion of GC-rich DNA fragments at 5'end of expression cassette had no obvious effect on the level of gene expression.Conclusion The role of DNA fragment with GC-rich in improving the transgenic expression of CHO cells is related to its position in the vector.The insertion of GC-rich DNA fragments at 3'end and both 5',3'ends of the box of expression vector can improve transgenic expression.