With the rapid development of Next-generation Sequencing(NGS)technologies, and its high throughput and low cost is applied widely in the field of life science, the increase in the depth of sequencing together decrease in the consumption of time and cost, makes a wide application of NGS in the research of microbiological research, ancient DNA study, clinical diagnosis, forensic science research, etc. The article discusses the second generation sequencing technology platform and its genetic markers in the forensic application. Included STR typing, SNP typing, HLA genotype prediction and the application in the degradation of the material.

Objective To test the hypothesis of autophagy that silencing PHD2 gene could increase hypoxia inducible factor (HIF)-1α levels in the renal medulla and attenuate hypoxia injury in cultured human renal proximal tubular epithelial cell (HK-2) under cobalt dichloride (CoCl2) exposure.Methods HK-2 cells were harvested at hour 0,6,12,24,36 and 48 after exposure to CoC12 (200 μmol/L).The role of HIF/PHD pathway in CoCl2-induced cell apoptosis/autophagy was studied by employing small-interfering RNA (siRNA).Dynamic profiles of apoptosis markers (Bax,Bcl-xl) and autophagy marker (LC3) of HK-2 cells within 48 h after exposing to CoCl2 were recorded.Alamar Blue assay was used for quantitative analysis of cellular growth and viability.Electron microscopy analysis was employed to evaluate the changes in autophagic structures.Results The protein expressions of PHD2 were gradually increased after exposing to CoCl2 (200 μmol/L),with statistics significance at 24 h and reached the peak at 48 h (both P ＜ 0.01).PHD2 siRNA reduced PHD2 levels by ＞ 60％ and significantly increased HIF-1α protein levels (P ＜ 0.01),but had little effect on HIF-2α.The protein expression of Bcl-xl was significantly up-regulated,while the level of Bax and LC3-Ⅱ/LC3-Ⅰ were down-regulated in PHD2 siRNA group (all P ＜ 0.01),compared with the negative control group.Meanwhile,either 3-Methyladenine (an autophagy inhibitor) treatment or PHD2 knockdown rescued cell death and increased cell viability through autophagy inactivation.The ratio of LC3-Ⅱ/LC3-Ⅰ and the quantity of autophagosomes were decreased,and the cell ultrastructure was also relatively intacter than the negative control group.Of interest,co-administration of HIF-1α siRNA with PHD2 siRNA abrogated renoprotective effect conveyed by PHD2 siRNA alone,suggesting that activation of endogenous HIF-1α-dependent pathways mediated the autophagy inactivation effects of PHD2 silencing.Conclusions Direct inhibition of PHD2 promotes renal epithelia cell survival against CoCl2-induced cell apoptosis/autophagy.Activation of the HIF-1α signaling pathway is required to reduce apoptosis and autophagy via up-regulating the expression of Bcl-xl protein.

Objective To investigate the molecular mechanism of protection of ischemia preconditioning on renal ischemia reperfusion injury. Methods Male C57/BL6N mice were randomly divided into two groups: in IR group, 35 min ischemia was induced by occlusion of both renal pedicles followed by 24 h perfusion (I/R). 15 min ischemia was induced 4 days before I/R in IPC group. Blood sample and kidney were collected in IR and IPC group after 24 h perfusion. Serum creatinine (Scr) and histological changes were used to evaluate the renal injury. PHD2 and HIF-1αwere evaluated by Western blotting, miR-21 expression was confirmed by real-time PCR. In vitro, hypoxic model was established by 1% O2 in HK-2 cells. Knockdown of miR-21 in hypoxic model was perfermed by locked nucleic acid modified-anti-miR-21 transfection. The levels of miR-21, HIF-1α and PHD2 mRNA were confirmed by real-time PCR. The levels of HIF-1α and PHD2 proteins were tested by Western blotting. Results In vivo, Compared with IR group, the renal function and histological changes were improved in IPC group (P<0.01). Compared with IR group, the expression of miR-21(P<0.01) and HIF-1α(P<0.05) were increased in IPC group, while PHD2 was reduced (P<0.01). In vitro, hypoxia reduced miR-21. The inhibition of miR-21 could increased the expression of PHD2 (P<0.05). Conclusions Ischemia preconditioning may exert protection against renal ischemia reperfusion injury by inhibiting PHD2.

Objective:To explore the initial motivation of nursing students to engage in voluntary service for the aged, and to provide scientific basis for the formulation of strategies and measures of voluntary service for the aged.Methods:Totally 25 nursing students volunteers who regularly participated in the volunteer service for the aged in Changsha First Welfare Home were selected by objective sampling method for semi-structured in-depth interviews, and the data were analyzed by Colaizzi phenomenological 7-step analysis method.Results:Four themes of nursing students' initial motivation to participate in voluntary service for the aged were extracted: self-interest motivation, altruistic motivation, affinity motivation and achievement motivation. Among them, self-interest motivation included two sub-themes: enriching college life and life experience, improving one's own ability and gaining professional experience; altruistic motivation included two sub-themes: accompanying and helping the elderly and eliminating loneliness of the elderly; affinity motivation included two sub-themes: establishing emotional sustenance and making up for the lack of emotion; achievement motivation included two sub-themes: facing challenges bravely, acquiring a sense of achievement, serving society and others, and realizing one's own value.Conclusion:Nursing students have a clear initial motivation to participate in the voluntary service for the aged of the aged care facilities, and schools can strengthen the relevant education for students to participate in voluntary service in the training of nursing talents. The voluntary service for the aged can formulate corresponding development strategies and incentives according to the initial motivation of nursing students, so that promote the expansion and stability of the voluntary service for the aged team, and promote the development and improvement of voluntary service for the aged.

Objective To explore a simplified method of coagulation test for the individuals with erythrocytosis. Methods The antico-agulants, blood volume and fixed blood collecting volume were adjusted by the formula: anticoagulants (mL)＝(100－HCT×100)× blood (mL)×0.001 85. A total of 124 blood samples for coagulation testing in which the calcium ( Ca2+) interval was designated and hematocrit (HCT) was more than 55% were tested with calibrated anticoagulants, adjusted blood volume and fixed blood collection vol-ume [anticoagulant(mL)/0.055 5]. The results of plasma prothrombin time (PT), international standardization ratio (INR) and acti-vated partial thromboplastin time (APTT) before and after adjustment were compared. The results of the samples from 3 groups after adjustment were also compared. The relationship of HCT with unadjusted PT and APTT were simultaneously observed. Results The unadjusted results of PT, INR and APTT were significantly higher than those after anticoagulants adjustment (27.52±16.37 vs 12.49± 1.35, 2.31±1.47 vs 0.99±0.11 and 50.09±13.32 vs 33.37±5.05) with statistically significant difference in paired comparison (P＜0.05). No statistical difference was found in the comparison of the results for PT, INR and APTT after adjustment within the 3 groups ( PT: 12.49±1.35 vs 12.84±1.54 vs 12.82±1.76, INR:0.99±0.11 vs 1.02±0.13 vs 1.02±0.15, APTT: 33.37±5.05 vs 33.49±5.09 vs 32.83±5.06) (P＞0.05). HCT values of the patients were positively correlated with unadjusted PT (r＝0.461, P＜0.05) and APTT (r＝0.571, P＜0.05). Conclusion The coagulation test of the individuals with erythrocytosis may use to adjust the blood volume and the fixed blood collection volume provided calcium concentration in reference interval.

Objective:To analyze the clinical efficacy of prednisone, hydroxychloroquine (HCQ) combined with plasmapheresis (PE) or not for the treatment of systemic lupus erythematosus (SLE) during pregnancy.Methods:Fourteen patients with SLE during pregnancy were analyzed. Totally 7 patients in the non-PE group were given prednisone and HCQ only while 7 patients in PE group were given prednisone and HCQ combined with PE. The fetus outcomes and clinical data, such as erythrocyte sedimentation tate (ESR), urine protein level, blood cell count and systemic lupus erythematosus disease activity index (SLEDAI) score before and after treatment at 3, 6, 12 months were used to evaluate the efficacy between the two groups. The comparison between groups was performed by repeated measures analysis of varianc (ANOVA).Results:Totally 11 patients delivered successfully in both groups while three of the 7 patients in the non-PE group had stillbirth. The 11 fetuses developed well and were born with an Apgar score of 8 or more at birth in both groups. There was a significant difference in ESR and platelet counts between the two groups ( F=7.838, P<0.05 ; F=32.269 , P<0.05). The ESR of the PE group was lower than that in the non-PE group at 3, 6 and 12 months after delivery, while the platelet count was higher than that in the non-PE group. Although there was no significant difference in the SLEDAI scores between the two groups ( F=2.816, P=0.119), the average of SLEDAI scores in the PE group was lower than that in the non-PE group at 3, 6 and 12 months after delivery. In addition, the urine protein of 7 patients in the PE group turned negative at 6, 12 months after delivery. In the non-PE group, urinary protein-positive patients were present in 3, 6, 12 months after delivery. Conclusion:PE in combination with oral prednisone and HCQ is a more effective than oral prednisone and HCQ alone for patients with active SLE during pregnancy, which reduces pregnancy loss and promote the patient's outcome.

Objective:To establish and apply the electronic further modified early warning score system (e-fMEWS), and explore its role in the condition evaluation and early warning of inpatients in non-critical units, so as to provide clinical nurses with an early and dynamic method to identify the potential deterioration risk of patients' condition.Methods:A retrospective analysis of 262 805 inpatients in multiple non-critical units of the Second Affiliated Hospital of Zhejiang University School of Medicine from January to December 2018 and January to December 2020 was performed. The patients who were hospitalized from January to December 2018 were used as the control group, and the responsible nurse used the traditional single evaluation index to start the emergency response system; the patients from January to December 2020 were used as the research group, and the emergency response system was started using e-fMEWS. The inclusion criteria were as follows: (1) hospitalization time ≥24 h; (2) patient ≥14 years old. Exclusion criteria were as follows: (1) patients had cardiopulmonary resuscitation before admission; (2) patients discontinued treatment or were transferred to another hospital during treatment; (3) patients received palliative care; (4) patients were admitted to non-critical wards in grade I of emergency pre-examination and triage. The activation of the rapid response team (RRT), the activation of the cardiorespiratory arrest team, the incidence of cardiac and respiratory arrest, the number of cases of invasive mechanical ventilation, the number of cases admitted to the intensive care unit, the length of hospital stay and the prognosis were compared. Statistical software SPSS 22.0 was used for data analysis.Results:Under the e-fMEWS assessment, compared with the control group, the rate of initiation of the research group decreased by 0.03%. For patients who initiated RRT, the average length of hospital stay was shortened, and the number of in-hospital respiratory cardiac arrest decreased (12.2% vs. 13.2%) and the number of cases transferred to the intensive care unit was less (42.8% vs. 50.6%), the rate of improvement and recovary increased (58.4% vs. 56.1%).Conclusions:The application of e-fMEWS can help clinical nurses to quickly and accurately identify the potential risk of deterioration of the patient's condition. Through early identification of potentially critically ill patients in non-critical units, early intervention and timely treatment can avoid adverse events and improve the patient prognosis.

OBJECTIVE To investigate the clinical efficacy of Gushen antai pills combined with heparin sodium in the treatment of antiphospholipin antibody （ACA）-positive recurrent abortion. METHODS A total of 203 ACA-positive recurrent abortion patients were randomly divided into control group （n＝101） and combination treatment group （n＝102）. Control group was additionally treated with heparin sodium on basis of basic treatment， and combination treatment group was additionally treated with Gushen antai pills on the basis of control group. Serum samples were collected from patients， the levels of interleukin 6 （IL- 6）， tumor necrosis factor α （TNF-α）， cyclooxygenase 2 （COX-2）， β-human chorionic gonadotropin （β-HCG） and ACA were detected by enzyme-linked immunosorbent assay. The antibody-negative conversion rate was calculated， the clinical efficacy of the patient was observed， and the occurrence of adverse drug reactions was recorded. RESULTS After treatment， the pregnancy success rate and antibody-negative conversion rate of combination treatment group were significantly higher than those in the control group （P＜0.05）， and the abortion rate was significantly lower than that in the control group （P＜0.05）. Compared with before treatment， serum levels of IL-6， TNF-α and ACA in 2 groups were decreased significantly （P＜0.05）， while the levels of COX-2 and β-HCG were increased significantly （P＜0.05）； the changes of above indexes in combination treatment group were more significant（P＜0.05）. There was no statistical significance in the incidence of adverse drug reactions between 2 groups（P＞0.05）. CONCLUSIONS Gushen antai pills can maintain high expression of β-HCG， reduce the level of ACA and improve immune microenvironment in patients with ACA-positive recurrent abortion by increasing the serum level of COX-2 and decreasing the serum level of IL-6， so as to increase the success rate of pregnancy.

Objective: To investigate the role of gut microbiota in the amelioration of liver fibrosis induced by CCl4 in mice by 3-methyladenine (3-MA). Methods: Fifteen mice were randomly divided into normal control group, liver fibrosis group and 3-MA treatment group. The liver fibrosis model was established by injecting CCl4, and the mice in the 3-MA treatment group were injected 3-MA additionally from the third week onwards. After 8 weeks, all of the mice were sacrificed and their blood, liver tissue and fecal samples were collected to analyze serum ALT, AST, GGT levels, liver histopathology and gut microbiota. Results: Compared with the liver fibrosis group, serum ALT and AST levels in 3-MA treatment group decreased obviously ([68.6±4.2] U/L vs [111.0±7.8] U/L, [179.0±12.9] U/L vs [253.2±26.7] U/L, P＜0.01), and the degree of hepatic histopathological changes was reduced. The intestinal flora in three groups were distinguished by principal coordinate analysis (PCoA) and non-metric multi-dimensional scaling (NMDS) analysis. Compared with the normal control group, there were decreased Alpha diversity of intestinal community, reduced significantly abundance of Lachnospiraceae, and increased obviously abundance of Actinobacteria and Desulfovibrionacea in the liver fibrosis group (P＜0.05). Compared with the liver fibrosis group, there were increased Alpha diversity of intestinal community, increased significantly abundance of Lachnospiraceae and Blautia, and reduced abundance of Bifidobacteriaceae in the 3-MA treatment group (P＜0.05). In addition, the abundance of Lactobacillaceae in the 3-MA treatment group was significantly higher than that in the normal control group (P＜0.05). Conclusion 3-MA improves the liver fibrosis of mice induced by CCl4, and gut microbiota may play an active role in this process.

To analyze the interest distribution relationship and symbiotic mode in the construction of urban medi-cal groups,based on the symbiotic theory,a framework for analyzing the interest distribution relationship of urban medical groups is constructed.Based on actual cases,urban medical groups are summarized and divided into four types:one-way supply type,partial benefit symbiosis type,asymmetric reciprocity symbiosis type,and symmetric reciprocity symbiosis type.The focus of a one-way supply oriented urban medical group is on reshaping the profit distribution mechanism,a partial benefit symbiotic urban medical group is on establishing a profit compensation mechanism for benefit medical institutions,and an asymmetric and mutually beneficial symbiotic urban medical group is on building a long-term benefit balance mechanism.