Objective To establish and identify the induced pluripotent stem cell(iPSC) line reprogrammed from human umbilical cord mesenchymal cells(HuMSCs).Methods HuMSCs were cultured by adhesion method,and OCT4,SOX2,KLF4,c-Myc,NANOG,LIN-28 were transfected into HuMSCs with lentiviral victor to reprogramme HuMSCs into iPSC.Morphological observation,pluripotency genes (SOX2,TDGF1,THY-1,OCT4,REX1 and TERF1) expression,alkaline phosphatase detection,karyotype analysis,embryonic stem cells (ESC) specific proteins (NANOG,OCT4,SSEA-4,TRA-1-81) immunofluorescence staining,differentiated into teratomas in vivo(inject the iPSC into SCID mice) and embryniod bodies in vitro were performed to exam the pluripotency of the iPSC.Results Four days after being infected by lentivirus,the HuMSCs became round-shape; 10 days after infection,some embryonic stem(ESC)-like colonies appeared.Fourteen days after infection,picked up the regularly shaped colonies and cultured several passages.About 1.25％ HuMSCs were reprogrammed into iPSC.The iPSC presented clone-like growth like ESC.All the cells were positive to alkaline phosphatase staining and expressed the pluripotency genes.The iPSC also expressed the ESC specific proteins,and karyotype analysis showed normal chromosome caryotype (46,XY).Furthermore,the iPSC could form embryoid bodies in vitro,expressed alpha fetoprotein(AFP),smooth muscle actin(SMA) and β-tubulin.The iPSC could alsoform teratomas in vivo.Conclusion OCT4,SOX2,KLF4,c-Myc,NANOG,LIN-28 can reprogram HuMSCs into iPSC efficiently.

BACKGROUND:Traditional Chinese medicine compound prescription has a long history in the treatment of primary osteoporosis,and the curative effect is definite,but the medication rule and mechanism are not clear. OBJECTIVE:Using the methodology of data mining and network pharmacology,to explore and verify the law of drug use and molecular mechanism of modern traditional Chinese medicine in the treatment of primary osteoporosis. METHODS:The relevant documents included in CNKI,WanFang,VIP and PubMed were used as data sources,and the relevant data were statistically counted and extracted by Microsoft EXCEL2019,IBMSPSS25.0 and other software.The high-frequency drugs obtained from the data statistics were analyzed by association rules analysis and cluster analysis,and the core drug combination of traditional Chinese medicine compound prescription in the treatment of primary osteoporosis was obtained by combining the two results.The therapeutic mechanism of this combination was explained by network pharmacology and verified by molecular docking. RESULTS AND CONCLUSION:Finally,151 articles were included and 207 prescriptions were selected,involving 285 flavors of Chinese herbs.(1)Ten groups of important drug combinations were obtained through the above two analyses,among which the core drug combination with the highest confidence and improvement was"Drynaria-Eucommia-Angelica."The key components of the combination in the treatment of primary osteoporosis were quercetin,kaempferol,naringenin and so on.The core targets were SRC proto-oncogene,phosphoinositide-3-Kinase regulatory subunit 1 and RELA proto-oncogene.The main pathways were cancer signaling pathway,JAK-STAT signaling pathway,VEGF signaling pathway,and NF-κB signaling pathway.(2)The key active components were docked with the core targets,and the two showed a good combination.To conclude,Chinese herbal compound therapy in the treatment of primary osteoporosis can use a variety of active components to exert its efficacy through multiple signal pathways and acting on multiple targets,which can provide a theoretical basis for the research and development of new drugs for the follow-up treatment of primary osteoporosis.

Objective:To establish virus culture method and full genome sequencing method for severe acute respiratory syndrome coronavirus (2019-nCoV), and to illuminate the variation of 2019-nCoV.Methods:The pharyngeal swab specimens were inoculated into Vero-E6 cells for isolation, and the cytopathic effect were observed day by day, and the result of virus isolation were confirmed by Real-Time RT-PCR. Some isolated viruses were diluted 10 times to detect the virus titer. Full-genome of 2019-nCoV was sequenced with the whole genome capture technology and next generation sequencing technology. Phylogenetic analysis and molecular characterization for obtained 2019-nCoV sequences were undertaken.Results:Twenty-two 2019-nCoV strains were isolated with Vero-E6 cell line in the Biosafety Level-3 Laboratory (BSL-3). Eighteen full-genome sequences obtained (almost 29 000 nucleotide) were analyzed with 99.94%~100% identity and 28 nucleotides and 20 amino acids variations. Phylogenetic analysis showed that the 2019-nCoV sequences belonged to different clades, including clade S and clade L with Europe lineage L. I and Europe lineage L. II.1. Furthermore, 5 of 18 2019-nCoV harbor the D614G mutation.Conclusions:2019-nCoV were successfully isolated with Vero-E6 cells, and partially illuminated the phylogenetic characteristics, which provided a basis for subsequent drug screening and biological characteristics research. Continuous monitoring and analysis of the sequences of new cases would be vital to understand the genetic evolution and rates of substitution of the 2019-nCoV.

Objective:The investigation and research on the application status of Hepatic Venous Pressure Gradient (HVPG) is very important to understand the real situation and future development of this technology in China.Methods:This study comprehensively investigated the basic situation of HVPG technology in China, including hospital distribution, hospital level, annual number of cases, catheters used, average cost, indications and existing problems.Results:According to the survey, there were 70 hospitals in China carrying out HVPG technology in 2021, distributed in 28 provinces (autonomous regions and municipalities directly under the central Government). A total of 4 398 cases of HVPG were performed in all the surveyed hospitals in 2021, of which 2 291 cases (52.1%) were tested by HVPG alone. The average cost of HVPG detection was (5 617.2±2 079.4) yuan. 96.3% of the teams completed HVPG detection with balloon method, and most of the teams used thrombectomy balloon catheter (80.3%).Conclusion:Through this investigation, the status of domestic clinical application of HVPG has been clarified, and it has been confirmed that many domestic medical institutions have mastered this technology, but it still needs to continue to promote and popularize HVPG technology in the future.