Copy number variations (CNVs) refer as a DNA segment that is 1 kb or larger and is presented at a variable copy number in comparison with a reference genome. Classes of CNVs include insertions, deletions, duplications and their complex combinations. Because they widely distributed in the genome with some important characteristics, such as heritable, relative stable and heterogeneity, CNVs are considered as novel genomic polymorphism markers. And the alteration of gene dosage which resulted from CNVs could change phenotype, so a novel CNV genome-wide association analysis (CNV-GWAS) strategy appeared recently and began to used for identifying susceptible genes of complex diseases. It was approved that it could complement the tranditional genome-wide association studies based on single nucleotide polymorphisms. Therefore, genomic structure variances are favorable for revealing the molecular mechanisms and genetic foundation of complex diseases.

Objective To investigate the relationship between osteoprotegerin ( OPG ) gene polymorphisms and ischemic stroke etiological subtypes, as well as the extent and distribution of cerebral atherosclerosis ( AS) lesions.Methods Patients with ischemic stroke included 285 cases of large-artery atherosclerosis (LAA), 91 cases of small-artery occlusion (SAO) and 42 cases of purely AS, and 165 healthy controls were enrolled in this study.The LAA group was respectively divided into 3 subgroups according to the number and the distribution of stenostic vessels.Genotyping of three single nucleotide polymorphisms (SNPs;rs2073617, rs3134069, and rs3102735) in the promoter region of the OPG gene was performed by polymerase chain reaction-restriction fragment length polymorphism.Results Regarding the three SNPs of OPG gene, the frequence of genotype CC/CT and the prevalence of allele C of rs3102735 were higher in the LAA group contrasting with the control group ( 24.04% vs 14.85%, 44.21% vs 27.88%,χ2 =10.758, 11.804, P =0.001,0.024).However, comparisons of other frequences of genotypes or alleles did not reveal any significant differences among the LAA group, the SAO group, the AS group and the control group, as well as among different subgroups of LAA group.Haplotype analysis revealed that the frequencies of haplotype C-C-T in LAA group and SAO group were significantly lower ( 0.023, 0.017 vs 0.068,χ2 =10.399, 5.841,P=0.001, 0.016), while that of haplotype T-A-C was significantly higher in SAO group(0.043 vs 0.016,χ2 =4.708, P=0.030) compared with controls.Conclusions Our findings indicate that OPG gene polymorphisms might be associated with increased susceptibility to LAA ischemic stroke.But we fail to show association of OPG gene with the extent and distribution of AS.

0.05). Conclusion Enough high-purified LECs can be isolated by collagenase digestion procedure followed by immunomagnetic beads sorting. Post-thawed endothelial cells are proved to have high vitality and growth potential in vitro without significant morphological changes. Cryopreserved LECs may serve as a cell choice for research of lymphangiogenesis and lymphatic patterning.

Background Mesenchymal stem cells (MSCs) have been applied in basic and clinical researches of organ transplantation.Our previous study showed that intravenous injection of MSCs prolonged corneal allograft survival in rat.However,the effect of local administration of MSCs on corneal allograft rejection remains unclear.Objective The aim of this study was to investigate the effect of subconjunctival injection of MSCs on corneal allograft rejection in rat model of keratoplasty.Methods MSCs were isolated and cultured from femur and tibia bone marrow of clean Wistar rats,and then the cells were identified by induced differentiation of osteoblast and adipocyte.The third generation of MSCs was used in subsequent experiment.Allogenic penetrating keratoplasty was performed with the bilateral corneas of 20 Wistar rats as donor grafts and the right eyes of 40 Lewis rats as recipients.PBS 0.1 ml containing 2 × 106 MSCs and 0.1 ml PBS only was subconjunctivally injected immediately and postoperative 3 days respectively in randomized two groups,and another 6 normal Lewis rats served as the normal control group.Corneal rejection response was evaluated under the slit lamp after surgery based corneal opacity,edema and neovascularization,and the grafts were scored according to the criteria of Larkin.The corneal samples were extracted from 12 rats of the PBS control group and the MSCs group separately 10 days after surgery.The relative expressions of Th1 cytokines (interferon-γ [IFN-γ] mRNA and interleukin-2 [IL-2] mRNA) and Th2 cytokines (IL-4 mRNA and IL-10 mRNA) were detected by real-time quantitative PCR.Protein levels of IL-4 and IL-10 proteins in the corneas were assayed by ELISA.All experimental protocols involving rats were approved by the laboratory animal care and use committee of the Tianjin Medical University and treated with the ARVO statement for the use of animals in ophthalmic and vision research.Results The cells grew well with the orange stain for alizarin red in differentiated the osteoblasts and red stain for Oil red O in differentiated adipocytes.The survival time of corneal graft in the MSCs group was (11.8±1.6) days,it was significantly longer than (9.6±1.4) days in the PBS control group (P=0.004).The levels of IL-4 mRNA and IL-10 mRNA in the MSCs group were significantly higher than those in the PBS control group (both at P =0.00);while the levels of IFN-γ mRNA and IL-2 mRNA were not significantly different between the groups (both at P＞0.05).The IL-10 protein contents were (22.74 ±7.06),(68.40±12.83) and (215.41 ±44.66)pg/ml in the normal control group,PBS control group and MSCs group,showing significant difference among the three groups (F =55.06,P =0.00) and a significant increase in the MSCs group compared with the PBS control group and the normal control group (both at P ＜ 0.05).Conclusions Subconjunctival injection of MSCs prolongs the survival time of cornea allograft in penetrating keratoplasty probably by modulating the balance between Th1 and Th2 cytokines,especially by up-regulating Th2 cytokines.

Objective:To compare the difference between the combined diagnostic effect of plasma Septin9(SEPT9) and polyligand Syndecan-2(SDC2) methylation with four serum tumor markers in the diagnosis of colorectal cancer.Methods:In this study, 128 patients who were treated in the affiliated Hospital of Xuzhou Medical University from March to December in 2019 were selected for a case-control study.All the subjects were examined by gastroenteroscopy.According to the pathological results, they were divided into three groups: colorectal cancer group( n=74) and colorectal adenoma group( n=7). The patients with no abnormal or inflammatory polyps or proliferative polyps examined by gastroenteroscopy were taken as the control group( n=47). The methylation levels of SEPT9 gene and SDC2 gene were detected by Roche Lightcycler 480 II real-time fluorescence quantitative polymerase chain reaction, and the concentrations of alpha-fetoprotein, carcinoembryonic antigen, carbohydrate antigen 125 and carbohydrate antigen 199 were detected by Roche Cobas 8000 electrochemiluminescence instrument.Chi-square test was used to compare the positive rate of each marker in the three groups.Medcalc was used to draw the receiver operating characteristic curve (ROC) curve of the subjects′ working characteristic curve, and the value of each index in the diagnosis of colorectal cancer was analyzed. Results:The positive rates of SEPT9 gene and SDC2 gene methylation were 81.1%(60/74) and 67.6%(50/74) respectively in colorectal cancer group, and increased to 85.1%(63/74) after combined detection.The positive detection rates of alpha-fetoprotein, carcinoembryonic antigen, carbohydrate antigen 125 and carbohydrate antigen 199 in colorectal cancer group were 1.4%(1/74), 33.8%(25/74), 6.8%(5/74) and 13.5%(10/74), respectively.When the four tumor markers were detected together, the positive detection rates were only increased to 43.2%(32/74), except for AFP and carbohydrate antigen 125(χ 2=3.847, 2.430, all P>0.05). The differences were statistically significant (χ 2=48.230, 30.487, 43.285, 3.847, 8.788, 6.988, 8.722, all P<0.05). The area under the curve (AUC) of SEPT9 methylation, SDC2 methylation, alpha fetoprotein, carcinoembryonic antigen, carbohydrate antigen 125 and carbohydrate antigen 199 were 0.854 (0.781, 0.910), 0.795 (0.715, 0.861), 0.575 (0.485, 0.662), 0.685 (0.597, 0.764), 0.603 (0.513, 0.689) and 0.631 (0.541, 0.715), respectively.The AUC of combined detection of two DNA methylation markers was better than that of alpha fetoprotein, carcinoembryonic antigen, carbohydrate antigen 125 and carbohydrate antigen 199, and the differences were statistically significant (alpha fetoprotein: Z=4.990, P<0.001; carcinoembryonic antigen: Z=3.743, P<0.001; carbohydrate antigen 125: Z=4.951, P<0.001; carbohydrate antigen 199: Z=3.983, P<0.001). The combined detection of two kinds of gene methylation was better than the combined detection of four kinds of serum markers in the diagnosis of colorectal cancer, and the difference was statistically significant ( Z=3.334, P<0.001). Conclusion:The combined detection of SEPT9 gene and SDC2 gene methylation in plasma is more suitable for non-invasive diagnosis of colorectal cancer than the combined detection of 4 serum tumor markers.

Red blood cell distribution width is an index reflecting the size heterogeneity of circulating red blood cells, which is usually used for diagnosis and treatment of blood system diseases.In recent years, more and more evidences show that the increase of red blood cell distribution width level is closely related to the occurrence, development and prognosis of cardiovascular diseases.We focused on the relationship between red blood cell distribution width and atrial fibrillation (AF) and its possible mechanism, in order to provide some reference value for the clinical treatment of AF.

Objective To collect the basic information on publications in Chinese Journal of Perinatal Medicine,and understand the current situation and set the goals for the future. Methods Information on all publications in this journal from 1998 to 2007 was collected.The distribution of all publications,Price Index and relative data were analyzed. Results There were 1368 publications altogether during the past 10 years.The first six kinds of papers were original articles(508,31.7%),short articles(258,18.9%),case reports(194,14.2%),reviews(122,8.9%),experimental studies(90,6.6%)and brief communications(83,14.2%).47.9% of all publications were from Beijing,Guangdong and Shanghai.The first three areas who had the most submissions were Beijing,Shandong and Guangdong in 2007.The citation frequency analysis showed that 708 publications were cited at 1east once and only two were cited over 80 times.53.4% of all cited publications were original articles and experimental studies. Conclusions Original articles and short articles are the two main types of publications in this journal,however original articles and experimental studies contributed to half of the publications being cited.Publications in this journal are focused in few areas and this might be related to the number of submissions.

Zinc-finger proteins have been widely studied due to their highly conserved structures and DNA-binding specificity of zinc-finger domains. However, researches on the zinc-finger proteins from microorganisms, especially those from prokaryotes, are still very limited. This review focuses on the latest progress on microbial zinc-finger proteins, especially those from prokaryotes and the application of artificial zinc-finger proteins in the breeding of robust strains. Artificial zinc-finger proteins with transcriptional activation or repression domain can regulate the global gene transcription of microbial cells to acquire improved phenotypes, such as stress tolerance to heat, ethanol, butanol, and osmotic pressure. Using the zinc-finger domain as DNA scaffold in the construction of enzymatic system can enhance the catalytic efficiency and subsequently the production of specific metabolites. Currently, zinc-finger domains used in the construction of artificial transcription factor are usually isolated from mammalian cells. In the near future, novel transcription factors can be designed for strain development based on the natural zinc-finger domains from different microbes, which may be used to regulate the global gene expression of microbial cells more efficiently.
Bacteria ; metabolism ; DNA ; chemistry ; Protein Engineering ; Transcription Factors ; chemistry ; Transcriptional Activation ; Zinc Fingers

Objective To construct a eukaryotic expression vector in Pichia pastoris containing human tissue factor( hTF) gene,in order to achieve high level secretory expression in extracellular.Methods Expression plasmid, pGAPZaA-hTF, was constructed by inserting the synthesized sequence encoding human extracellular tissue factor into yeast expression vector pGAPZaA and transformed into Pichia pastoris SMD1168H with electroporation.Having been selected by Zeocin, transformants containing hTF cDNA were expressed in YPD, extracellular proteins were detected by SDS-PAGE and confirmed by Western bolt.Results Successfully constructed the recombinant pGAPZaA-hTF expression system in Pichia pastoris.SDS-PAGE showed that the molecular weight of the expression product was about 37 -40 kDa.Western-blot indicated that it was human extracellular tissue factor.The crude yield of total protein in medium was up to 1 g/L, more than 80% of which was hTF.Conclusion Truncated rTF gene is expressed in Pichia pastoris and the active products are secreted into the medium which have the same activation as the native TF.

Objective:To observe the clinical efficacy of warm needling moxibustion plus tuina in treating knee osteoarthritis (KOA) due to cold-dampness blocking collaterals.
Methods:Forty-eight KOA patients were randomized into 2 groups by their visiting sequence, 24 cases in each group. The treatment group was intervened by warm needling moxibustion plus tuina, while the control group was treated with acupuncture plus tuina. The two groups were both treated once a day, 30 min for each session, 10 d as a treatment course, totally for 3 treatment courses. The visual analogue scale (VAS) and Lysholm knee scoring scale (LKSS) were observed before and after the treatment to evaluate the clinical efficacy.
Results:After 3 treatment courses, the VAS and LKSS scores were significantly changed in the two groups (P<0.01), and the between-group differences were also statistically significant (P<0.01). The markedly-effective rate was 83.3% in the treatment group versus 66.7% in the control group, and the difference was statistically significant (P<0.05).
Conclusion:Warm needling moxibustion plus tuina can produce a more significant efficacy than acupuncture plus tuina in treating KOA due to cold-dampness blocking collaterals.