Many somatic cell typos were obtained by in vitro differentiation or teratoma formation of human embryonic stem ceLls (hESCs). However, it is unclear whether specific cell types can be obtained from hESCs-derived teratoma. It was reported that many kinds of cells, including neural progetfitor/precursor cells (NPCs) and mesenchymal stem cells (MSCs) were isolated efficiently from the teratoma of hESCs through in vitro selection. The teratoma-derived NPCs and MSCs showed specific characteristics of molecular markers similar to the primary NPCs and MSCs. Moreover, these teratoma-induced NPCs and MSCs can be further induced to differentiate into neurons, astrocytes, or adipose and bone cells, reflecting their inherent multi-potencies. Given that teratoma normally contains a mixture of ectoderm, mesodenn, and endoderm lineage cells at different differentiation stage, it was suggested that hESCs-derived teratoma could be an alternative source to generate a variety of uncommitted progenitor cells or terminally differentiated somatic cells, which may be otherwise difficult to obtain through direct in vitro differentiation.

Objective To determine the effect of signal transducers and activators of transcription 3 (STAT3) gene silencing by shRNA mediated by lentiviral vector for the treatment of colorectal cancer. Methods The recombinant lentiviral vector pRNAT-shSTAT3, empty lentiviral vector pRNAT-GFP, and lentiviral packaging plasmids in supernatant were collected to transfect HT-29 cells for harvesting the HT-29-shSTAT3 cells and HT29-GFP cells. Fifteen male rats were divided into three groups (n = 5 ), and then they were inoculated with HT-29cells, HT-29-GFP cells and HT-29-shSTAT3 cells, respectively. Cell growth was assessed by MTT assay and the changes in cell cycle were detected by flow cytometry. The changes in microvessel density (MVD) of tumors were detected by immunohistochemistry. All data were analysed by one-way analysis of variance. Results The growth of HT-29-shSTAT3 cells was significantly suppressed compared with HT-29 and HT-29-GFP cells (F = 632.50,P ＜ 0. 05 ). The proportions of cells at the G0/G1 phase were 68.7% ± 2.9% in HT-29-shSTAT3 cells, 38.5% ±1.6% in HT-29-GFP cells and 38.7% ± 2.3% in HT-29 cells, with a significant difference among the three groups (F = 166.53, P ＜ 0.05 ). The MVDs of HT-29 cells, HT-29-GFP cells and HT-29-shSTAT3 cells were 29 ±5, 28 ±4 and 10 ±3, respectively, with a significant difference among the three groups (F=31.60, P ＜0.05). Conclusion STAT3 gene silencing by shRNA mediated by lentiviral vector can significantly inhibit the growth of colorectal cancer cells.

OBJECTIVETo construct a human source vector containing minidystrophin-EGFP fusion gene and investigate its expression in Cos-7 cells.

METHODSThe recombinant human source vector named pHrnDysG was constructed with PCR-clone methods. Three fragments of dystrophin gene were PCR amplified from normal human dystrophin gene cDNA (GenBank NM04006). These three fragments were ligated to generate a minidystrophin gene. The enhanced green fluorescent protein (EGFP) gene was fused to the C terminal of the minidystrophin gene, and then the pHrnDysG was finally obtained by cloning the fusion gene to pHrneo. Fluorescence microscope and RT-PCR were used to detect the expression of minidystrophin-EGFP fusion gene after the recombinant construct was transfected into Cos-7 cells by lipofectamine.

RESULTSRestrictive enzyme digestion analysis and sequencing confirmed that pHrnDysG vector was constructed successfully. After the recombinant pHrnDysG was transfected to Cos-7 cells, RT-PCR demonstrated that the fusion gene was successfully transcribed, and the green fluorescence was observed at the cell membrane.

CONCLUSIONThe minidystrophin-EGFP fusion gene mediated by pHrneo vector could express in Cos-7 cells and its products' localization in the cell membrane was the same as that of full length dystrophin. These results suggested that the recombinant human source vector pHrnDysG might be potentially used in studies on the gene therapy of Duchenne muscular dystrophy.

Animals ; COS Cells ; Cercopithecus aethiops ; Dystrophin ; genetics ; metabolism ; Genetic Vectors ; genetics ; Green Fluorescent Proteins ; genetics ; metabolism ; Humans ; Microscopy, Fluorescence ; Models, Genetic ; Recombinant Fusion Proteins ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection

OBJECTIVE: To investigate the biological characteristics of endothelial progenitor cells (EPCs) from the umbilical cord blood (UCB), and to evaluate their oncogenicity after long-term culture in vitro. METHODS: The mononuclear cells (MNCs) were isolated from the UCB and cultured in MCDB131 medium supplemented with 20% FBS, VEGF and other growth factors. Morphology of the EPCs was observed, and the growth curve of the EPCs was investigated. Surface antigens of the EPCs were analyzed by the flow-cytometer. The capability of intaking the acetylated low-density lipoprotein (acLDL) of the EPCs was detected using fluoresencent chemical method. The vasoformative capability and genetic stability of EPCs were cultured in matrigel, and examined by karyotype analysis. The oncogenicity of EPCs was verified by the tumorigenesis test in athymic mouse and soft agar. RESULTS: EPCs were successfully derived from the UCB, and could be passaged to at least 42(nd) generation and had strong abilities of proliferation, acLDL intake and vasoformation, but there was not oncogenicity. They expressed endothelial cell-surface antigens and maintained normal karyotype. CONCLUSION The EPCs with proliferative potential can be isolated from the UCB. They can be passaged in long-term cultures without oncogenicity, and can maintain normal karyotype. The EPCs can be served as a new type of cells in cell and gene therapy.
Animals ; Antigens, Surface ; analysis ; Cell Line ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; metabolism ; Fetal Blood ; cytology ; Flow Cytometry ; HeLa Cells ; Humans ; Infant, Newborn ; Intercellular Signaling Peptides and Proteins ; pharmacology ; Karyotyping ; Mice ; Mice, Nude ; Neoplasms, Experimental ; pathology ; Stem Cells ; cytology ; metabolism ; Vascular Endothelial Growth Factor A ; pharmacology

OBJECTIVETo determine the prevalence of herpes simplex virus (HSV) and its correlates among HIV/AIDS patients in a county of Shanxi.

METHODSAll HIV-infected patients in a county in Shanxi province who were receiving antiretroviral treatment (ART) were included in this study. Participants were interviewed using standard questionnaires. Serum samples were tested to determine HSV-1 and HSV-2 infections.

RESULTSA total of 195 AIDS patients were recruited and 195 blood samples were collected. Among 195 AIDS patients, 189 (96.9%) were farmers; 116 (59.5%) were men while 79 were women; 146 (74.9%) were between 20 - 50 years old; 180 (92.3%) were married. The major routes of HIV transmission were blood/plasma donation or transfusion (176 patients, 90.3%). CD(4)(+) T cell counts were between (1 - 1531) × 10(6) cells/L ((323.6 ± 14.8) × 10(6) cells/L), with 44 (26.5%) patients' CD(4)(+) T cell counts less than 200 × 10(6) cells/L. Of which, 154 patients (79.0%) had sexual partners. 86.8% (118 patients) consistently used condoms during the past 6 months, while for the last sexual act, 91.8% (123 patients) used condoms. For anti-HSV-1 status, there were about 164 patients (84.1%) were positive, and 26 (13.3%) were positive for anti-HSV-2. While, 14 (7.2%) were positive for both anti-HSV-1 and anti-HSV-2. Logistic regression analysis indicated that marital status were correlated with HSV-2 infection (OR = 7.41; 95%CI: 2.42 - 22.73; P < 0.01). No socio-demographic and sexual characteristics were identified to be correlated with HSV-1 infection.

CONCLUSIONA substantial proportion of AIDS patients in a rural county of Shanxi province of China were co-infected with HSV-1 and/or HSV-2. Marital status was correlated with HSV-2 infection.

Acquired Immunodeficiency Syndrome ; complications ; epidemiology ; virology ; Adolescent ; Adult ; Child ; China ; epidemiology ; Female ; Herpes Simplex ; complications ; epidemiology ; Herpesvirus 1, Human ; Herpesvirus 2, Human ; Humans ; Logistic Models ; Male ; Middle Aged ; Prevalence ; Risk Factors ; Rural Population ; Young Adult

OBJECTIVETo evaluate the genetic constitution and mutation status of the immunoglobulin variable heavy chain region (IGVH) gene expression in Chinese patients with chronic lymphocytic leukemia (CLL).

METHODSThe IGVH mutation was detected by multiplex PCR and direct sequencing of the purified PCR products from 64 CLL patients. The segments of VH, DH and JH family and mutations were analyzed by IMGT/V-QUEST and IGBlast.

RESULTSIn the 64 patients, the most common usage was VH3 (31/64, 48%), followed by VH4 (26/64, 41%), VH1 (4/64, 6%), VH2 (2/64, 3%) and VH7 (1/64, 2%). The results also showed that 44 patients (69%) had mutated VH, 6 cases (9%) had identical germline sequences. Among the 64 sequences of DH segments, DH3 gene family was used most frequently (25/64, 39%), among which 11 cases had unmutated VH. The most frequent usage of the JH segments was JH6.

CONCLUSIONThere is significant difference in the frequency of the IGVH gene family in Chinese CLL patients compared to Western patients, suggesting the involvement of antigen selection in different ethnic and/or environmental factors in CLL disease initiation, and its prognostic significance needs further investigation.

Adult ; Aged ; Aged, 80 and over ; Asian Continental Ancestry Group ; genetics ; Female ; Gene Expression ; Humans ; Immunoglobulin Class Switching ; genetics ; Immunoglobulin Heavy Chains ; genetics ; Immunoglobulin Variable Region ; genetics ; Leukemia, Lymphocytic, Chronic, B-Cell ; genetics ; immunology ; Male ; Middle Aged ; Mutation

OBJECTIVETo investigate ESI-MS/MS fragmentation pattern of bile acids contained in Qingkailing injection, and the rapid identification method.

METHODHPLC-ESI-MS/MS technology was adopted to investigate ESI-MS/MS fragmentation pattern of five bile acids contained in Qingkailing injection, and rapidly identify bile acids contained in Qingkailing injection from eight different manufacturers.

RESULT5 bile acids showed similar ESI-MS/MS fragmentation patterns, based on which 14 bile acids were rapidly separated from Qingkailing injection. Among them, 11 were found in injections of all of the manufacturers, and the rest three were found in individual manufacturers.

CONCLUSIONHPLC-ESI-MS/MS is used to rapidly identify bile acids from complex material systems, which provides an effective method for interpreting the complex material base of compound traditional Chinese medicine preparations and enhance the quality standards.

Bile Acids and Salts ; chemistry ; Chromatography, High Pressure Liquid ; methods ; Drugs, Chinese Herbal ; chemistry ; Spectrometry, Mass, Electrospray Ionization ; methods ; Tandem Mass Spectrometry ; methods

Objective To investigate the prevalence and influencing factors related to abuse and negligence against the elderly in the rural areas. Methods 975 elderly over 60 years from 41counties in Anhui province were included. All participants completed an anonymous questionnaire including items as: educational background, marital condition, income, child-discipline, rude action to parents, daily activities, physical functions, having chronic illness, abuse and negligence against the elderly, etc. Results In the last year, rates of common physical abuse, serious physical abuse,emotional abuse, financial exploitation, negligence, overall abuse and negligence against the elderly were 4.5%, 1.5%, 26.9%, 4.9%, 7.2%, 29.9% respectively. Among the 281 victims, 80.4% reported that they were suffered more than 3 times of abuse and neglect episodes, and 34.9% reported that they were suffered more than 2 forms of abuse and negligence. The primary sadism was carried out by the daughter-in-law or son-in-law (43.2%) of the elderly. Low activity on daily life and having chronic illness were the risk factors causing common physical abuse while better education was the protective factor to it, Low ability in managing daily activity of living was the risk factor causing serions physical abuse. Less active on daily life and having rude action to parents were the risk factors to emotional abuse, but being strict with their children was the protective factor to emotional abuse. Less active on daily life, often beating their children and having rude action to parents were the risk factors related to financial exploitation. Less active on daily life, having rude action to parents and having bad physical functions were the risk factors causing negligence. Less active on daily life and having rude manner to parents were the risk factors of overall elderly abuse and negligence, but being strict with their children was protective factor to the abuse and negligence against the elderly. Conclusion High prevalence on abuse and negligence against the elderly was seen in the rural areas of China. Different forms on elderly abuse and negligence were affected by different factors that called for more attention to be paid to those elderly with lower ability in managing their daily life.

Objective To establish information management systems of drinking water defluoridation project in water-related endemic fluorosis areas and investigate the status of drinking water defluoridation project in Liaoning, provide the basis for the development of control measures. Methods Global positioning systems (GPS)and geographic information systems(GIS) were used in the study in August 2006 - July 2008. Water defluoridation projects of 1234 in 48 counties(cities, districts) in drinking water type of fluorosis areas were positioned. Latitude and longitude, water samples, water fluoride content were collected or tested. GIS was used to establish information management system of water defluoridation projects. Results We have established information management system for the facilities of decreasing water fluorine in drinking water type of endemic fluorosis regions in Liaoning. One thousand two hundred and thirty four defluoridation facilities distributed in east longitude between 39.39° - 43.37°,north latitude between 119.25° - 125.50°, and altitude between - 6.60 and 801.14 meter in 48 endemic fluorosis counties in 13 cities. Nine hundred and twenty seven facilities for decreasing fluorine were able to supply water regularly, accounting for 75.1% of investigated projects;29 facilities was .not yet completely rebuilt, accounting for 2.4%;278 facilities(supply water for 344 villages) were out of order or discarded for 22.5% of investigated projects.Water fluorine contents of 63 facilities were greater than 1.2 mg/L, accounting for 6.8% of investigation project.Facilities working regularly and water fluorine was in accord with hygienic standard for drinking water facilities were 70.0%. Conclusions The establishment of Liaoning province defluoridation project information management system in the whole province of drinking water type of fluorosis areas provides scientific basis for accurate decision-making on prevention and control of the disease.

OBJECTIVE: To detect two exons of Duchenne muscular dystrophy (DMD) gene and a gender discrimination locus amelogenin gene by single cell triplex PCR, and to evaluate the possibility of this technique for preimplantation genetic diagnosis (PGD) in DMD family with DMD deletion mutation. METHODS: Single lymphocytes from a normal male, a normal female, two DMD patients (exon 8 and 47 deleted, respectively) and single blastomeres from the couples treated by the in vitro fertilization pre-embryo transfer (IVF-ET) and without family history of DMD were obtained. Exons 8 and 47 of DMD gene were amplified by a triplex PCR assay, the amelogenin gene on X and Y chromosomes were co-amplified to analyze the correlation between embryo gender and deletion status. RESULTS: In the normal single lymphocytes, the amplification rate of exons 8 and 47 of DMD and amelogenin gene were 93.8%, 93.8%, and 95.3% respectively. The false positive rate was 3.3%. In the exon 8 deleted DMD patient, the amplification rate of exon 47 of DMD and amelogenin gene was 95.8%, and the false positive rate was 3.3%. In the exon 47 deleted DMD patient, the amplification rate of exon 8 of DMD and amelogenin gene was 95.8%, and the false positive rate was 0. In the single blastomeres, the amplification rate of exons 8 and 47 of DMD and amelogenin gene was 82.5%, 80.0% and 77.5%, respectively, and the false positive rate was 0. CONCLUSION The single cell triplex PCR protocol for the detection of DMD and amelogenin gene is highly sensitive, specific and reliable, and can be used for PGD in those DMD families with DMD deletion mutation.
Amelogenin ; genetics ; Blastomeres ; cytology ; metabolism ; Chromosomes, Human, X ; genetics ; Chromosomes, Human, Y ; genetics ; Cytogenetic Analysis ; methods ; Exons ; genetics ; Female ; Gene Deletion ; Humans ; Lymphocytes ; cytology ; metabolism ; Male ; Muscular Dystrophy, Duchenne ; blood ; diagnosis ; genetics ; Polymerase Chain Reaction ; methods ; Pregnancy ; Preimplantation Diagnosis ; methods