Objective The stability and other characteristics of the active recombinant human anionic trypsin(hT 2) with site-mutation R 122 L(mhT 2) were investigated. Methods An active human anionic trypsin and its R 122 L mutate were produced with E.coli BL 21(DE 3) and purified with ion-exchange chromatography. The properties of mutant were studied and compared with the wild type. Results The optimal pH for mhT 2 was 7~11. mhT 2 was active over a broad temperature range (4℃~80℃) and owned a little better thermal stability than the wild type. The inhibition of typical metal chelating agent(EDTA), Fe 3+, denaturant, reducer(β-ME) on activity of mhT 2 was the same as the wild type. Michaelis constant Km of mhT 2 was 0.010 mmol/L with BAEE as a substrate, a little lower than wild type. Conclusion Compared with the wild type, the R 122 L site mutate significantly enhanced tolerance to acidic pH、denaturants、reductions and autolysis.

Objective To investigate recurrent spontaneous abortion patients received bends the clinical application effect of nursing intervention during progesterone treatment. Methods Two patients with recurrent spontaneous abortion group were treated with progesterone treatment, the control group in the conventional nursing service only provides flexor progesterone treatment on the basis of comprehensive nursing service study group progesterone treatment on the basis of routine the combination of nursing and nursing intervention in the 2 groups of patients with recurrent spontaneous abortion. The success rate of pregnancy were recorded, the rate of abortion, the data will be given the statistical analysis (using SPSS software) after the conclusion. Results The study group of patients with recurrent spontaneous abortion pregnancy success rate (95.45%) and control group (90.91%) there is no significant difference between the study group of patients with recurrent spontaneous abortion abortion rate (6.82%) was significantly lower than the control group (34.09%), there was significant difference between the groups, P<0.05. Conclusion Recurrence During the treatment of flexor progesterone abortion were given routine nursing and nursing intervention combined with the comprehensive clinical nursing service can obtain ideal success rate of pregnancy, to reduce the rate of abortion is also of positive significance, is conducive to the protection of patients' quality of life, physical and mental health.

Objective To construct a shRNA lentiviral vector targeting the gene encoding tumor necrosis factor alpha-induced protein 8 (TNFAIP8) in RAW264. 7 cells, a mouse macrophage cell line, and to investigate the effects of TNFAIP8 gene silencing on the functions of mouse macrophages. Methods The shRNA sequence targeting TNFAIP8 gene was designed and DNA oligos containing small hairpin frame was synthesized. The double-stranded DNA was cloned into pLKO. 1-TRC vector after annealing. The recombi-nant vector was verified by using double enzyme digestion and gene sequencing. Lentiviruses were prepared by transfecting the constructed vector into 293T cells. Fluorescent quantitative RT-PCR and Western blot as-say were performed to detect the expression of TNFAIP8 at mRNA and protein levels after infecting the RAW264. 7 cells with lentiviruses. Flat dish adhesion experiment and wound-healing assay were used to evaluate the effects of TNFAIP8 gene silencing on the adhesion and migration of RAW264. 7 cells. Results The recombinant lentiviral vector was successfully constructed as indicated by double enzyme di-gestion and gene sequencing analysis. The expression of TNFAIP8 in RAW264. 7 cells at both mRNA and protein levels were significantly down-regulated after lentivirus infection (P<0. 05). Moreover, TNFAIP8 gene silencing significantly impaired the cell adhesion ability of RAW264. 7 cells after 15 min, 30 min or 2 hours of culture. Compared with the cells in control group, the RAW264. 7 cells harboring silenced TN-FAIP8 gene looked round with a smaller number of cellular extensions. The wound-healing assay showed that less TNFAIP8 gene-silenced RAW264. 7 cells migrated into the wounded area as compared with the cells in control group after 24 hours of culture (P<0. 05). The wound-healing rates of the experimental and control groups were 25% and 50%, respectively. Conclusion The recombinant lentiviral vector containing shRNA targeting the TNFAIP8 gene was successfully constructed. Transfecting the RAW264. 7 cells with the con-structed vector significantly silenced the expression of TNFAIP8 gene and inhibited the adhesion and migra-tion of these cells.

AIM:To investigate the role of mesenchymal stem cell-induced regulatory dendritic cells ( MSC-DCregs) in mouse acute graft-versus-host disease( aGVHD) model.METHODS: Bone marrow cells from BALB/c ( H-2 d ) mice were isolated and were induced to differentiate into DCs.The DCs were selected by flow cytometry, and after 10 d co-culture with MSCs, they were induced to be MSC-DCregs.Male 8-week-old C57BL/6 (H-2b) mice were used as do-nor mice.The female 8-week-old BALB/c (H-2d) mice, who had received 100 cm source-skin distance, 30 cm ×30 cm radiation field, 700 cGy total body irradiation (TBI) pretreatment were used as recipient mice.The recipients were divided into 5 groups:control group, TBI group ( injected with medium only) , bone marrow transplantation group ( injected with 1 ×107 bone marrow cells), aGVHD group (injected with 1 ×107 bone marrow cells and 1 ×107 spleen cells), and MSC-DCregs group (injected with 1 ×107 bone marrow cells, 1 ×107 spleen cells and 1 ×106 MSC-DCregs).The white blood cell count, recipients’ chimerism, clinical evaluation of aGVHD, survival analysis and pathological changes were deter-mined.RESULTS:Hematopoieic recovery was seen at 10 d after transplantation.The recipients’ chimerism was parallel to the donors’ at 30 d.The median survival time of the mice in aGVHD group and MSC-DCregs group was 27 d and 33 d, and the survival rates at 30 d were 20% and 100% (P<0.01), respectively.The clinical scores of the mice in MSC-DCregs group were lower than those in aGVHD group ( P<0.01) .Moreover, the pathological changes in the skin and liver of the mice in MSC-DCregs group were less serious than those in aGVHD group.CONCLUSION: The MSC-DCregs in-duce an aGVHD tolerance in vivo, and further research of its mechanism is still in great necessary.

Objective:In order to get recombinant protein OCILRP 2-Fc and anti-OCILRP2 antibody for further study of OCILRP2.Methods:Eukaryotic expression vector pIg-CD5-OCILRP2 which fused with extracellular domain of OCILRP 2 and human IgG1 Fc fragment was constructed.G418 was used for stable expression cell strain after pIg-CD5-OCILRP2 transfected into CHO cells.Recombinant protein OCILRP 2-Fc purified from CHO cell supernatant was used to immunize rabbit and anti-OCILRP2 polyclonal antibody was purified from rabbit serum by using protein G column.Results: ELISA data showed that we got a high-titer anti-serum and anti-OCILRP2 antibody purified from the rabbit serum.Western blot indicated this antibody could specifically bind to OCILRP 2-Fc and OCILRP2 in NIH/3T3 lysate.OCILRP2 expression in murine bone marrow derived dendritic cells ( DC) was detected by this polyclonal antibody ,too.Compared to immature DC ,OCILRP2 expression was elevated in LPS induced-mature DC.Conclusion: This study has offered an available tool and provided a clue for further study of the roles of OCILRP 2 in immune response.

Objective To investigate the effect of human telomerase reverse transcriptase C27 polypeptide (hTERTC27) over-expression on orthotopic implanted tumor growth which induced by nasopharyngeal carcinoma cells C666-1 in vivo.Methods Stably transfected C666-1 cell lines were used to establish subcutaneously transplanted tumor mouse model.The tumor growth was observed and the tumor growth curve was made by measuring the volumes of tumors every day.HE staining was used to observe the change of histomorphology.The expressions of cleaved Caspase-3,Caspase-9,PARP,bax and bcl-2 were detected by Western blot analysis.Results The in vivo mouse model showed that over-expression of hTERTC27 significantly reduced tumor volume and tumor weight (P ＜ 0.05),and decreased the local muscle infiltration.Over-expression of hTERTC27 significantly up-regulated the expressions of cleaved Caspase-3,Caspase-9,PARP and bax (P ＜ 0.01),and down-regulated the expression of bcl-2 (P ＜ 0.01).Conclusion The results indicate that over-expression of hTERTC27 can obviously inhibit the growth of transplanted tumor in nude mice,which is possibly related to the regulation of bax and bcl-2 expression.

Objecitv e To investigate the effects of a recombinant protein osteoclast inhibitory lectin related protein 2( OCILRP2)-Fc on LPS-induced endotoxemia by blocking OCILRP 2 signaling pathway and to in-vestigate the roles of OCILRP2 during inflammation.Methods Real-time PCR was used to detect OCILRP2 ex-pression at mRNA level in RAW264.7cells before and after in vitro stimulation with LPS.A mouse model of en-dotoxemia was established by intraperitoneal injection of BALB /c mice with a median lethal dose of LPS .Two hours prior to LPS treatment, mice were intraperitoneally injected with OCILRP2-Fc, human IgG or PBS, re-spectively .Several parameters including the survival rate of BALB/c mice with and without LPS treatment , spleen weight for arterial hyperemia analyzing , histopathological changes of lung and liver by HE staining , serum levels of inflammatory cytokines (IL-6, IL-12, TNF-αand IFN-γ)by ELISA , NF-κB activity by Western blot, were analyzed .Results Real-time PCR showed that LPS elevated in vitro OCILRP2 expression at mRNA level in macrophages (P<0.05).Upon the treatment of OCILRP2-Fc, BALB/c mice suffered from endotoxemia showed obviously increased survival rate , decreased spleen hyperemia , attenuated pathological injury of lung and liver, reduced levels of IL-6, IL-12, TNF-αand IFN-γin serum samples (P <0.05) as compared with mice treated with human IgG and PBS .LPS induced NF-κB p65 phosphorylation and IκB degradation were inhibited by OCILRP2-Fc treatment.Conclusion OCILRP2-Fc protects mice from endotoxemia by blocking OCILRP 2 signaling, which suggests that OCILRP2 plays an important role in LPS induced inflammation.

Objective To explore the feasibility of using contralateral breast as the donor for immediately breast reconstruction or chest wall defect repair after mastectomy in breast cancer patients. Methods From Jul. 2013 to Mar. 2016, contralateral breast fat flap was used as the donor for 8 breast cancer patients with immediate autologous non-microsurgical breast reconstruction or chest wall defect repair after mastectomy. All participants in this study received preoperative oncological screening with ultrasound, mammography, and magnetic resonance imaging which revealed the absence of pathological abnormalities in the donor breast. Results Among the 8 pa-tients, 4 patients underwent immediate breast reconstruction and 4 received chest wall defect repair. Only 1 pa-tient undergoing breast reconstruction had minor complications with little or no effect on the final outcome. No patient undergoing chest wall defect repair had postoperative complications. The functional and aesthetic out-comes were very satisfactory. Regular follow-ups were from 3 to 34 months with no recurrence found up to the present. Conclusions This article presents the first case for immediately breast reconstruction or chest wall de-fect repair using contralateral breast as the donor. The surgical method has some complications but with good aesthetic outcomes, which can be an option for breast cancer patients with hypertrophic and ptotic breast.

Objective: To observe the efficacy and safety of micro-plasma technology for the treatment of stable scars. Methods: 63 cases were divided into 4 groups, including burn scar group (28 cases), traumatic scar group(15 cases), acne scar group(10 cases), and hemangioma scar group (10 cases). Micro-plasma was performed every 2 months, 4 times as one session. Vancouver scar scale(VSS) was used to record the scar score and evaluate its clinical efficacy. Results: All of the scar points in four groups dropped significantly after treatment, showing significant difference between groups(P<0.05). The point decrease rate in acne scar group and burn scar group was significantly higher than that in traumatic scar group and hemangioma scar group (P<0.05). Efficacy evaluation also showed obvious better effect in acne and burn scar than in traumatic scar (P<0.05). Conclusions The micro-plasma has a good effect on burn, trauma, acne and hemangioma scar, especially for burn and acne scar.

Adenocarcinoma ; metabolism ; pathology ; Adenocarcinoma, Clear Cell ; metabolism ; pathology ; Adult ; Carcinoma, Endometrioid ; metabolism ; pathology ; Cervix Uteri ; metabolism ; pathology ; surgery ; Cyclin-Dependent Kinase Inhibitor p16 ; metabolism ; Diagnosis, Differential ; Electrosurgery ; Endometrial Neoplasms ; metabolism ; pathology ; Female ; Humans ; Hyperplasia ; Keratin-7 ; metabolism ; Mesonephros ; metabolism ; pathology ; surgery ; Neprilysin ; metabolism ; Uterine Cervical Neoplasms ; metabolism ; pathology