ObjectiveTo detect human papilloma virus (HPV)infection with fluorescent quantitative real-time polymerase chain reaction (FQ-PCR)in Minnan population, and explore the correlation between HPV infection and carcinogenesis of esophageal carcinoma (EC)of Minnan patients. MethodsFQ-PCR was performed to examine HPV-6, HPV-11, HPV-16 and HPV-18 in 100 healthy Minnan people (healthy group, 66 males and 34 females with their age of 52.35±6.72 years)and 100 Minnan patients with squamous EC (EC group and tumor-adjacent normal tissue group, 64 males and 36 females with their age of 51.62±6.37 years)between October 2009 and December 2012. ResultsThe incidences of HPV infection in 100 EC tissues, 100 tumor-adjacent normal tissues and 100 esophageal mucosa tissues of healthy people were 22/100, 8/100 and 6/100 respectively, which were statistically different (χ2=10.63, P < 0.01). Positive infection of HPV-6, HPV-11, HPV-16 and HPV-18 was observed in 11 cases, 11 cases, 14 cases and 15 cases in EC group respectively, 5 cases, 6 cases, 7 cases and 8 cases in tumor-adjacent normal tissue group respectively, and 5 cases, 5 cases, 6 cases and 6 cases in the healthy group respectively (P > 0.05). Positive HPV infection was observed in 1 patients with well differentiated squamous EC, 21 patients with moderately differentiated squamous EC and 5 patients with poorly differentiated squamous EC (P > 0.05). ConclusionHPV infection may exist in tumor tissue of Minnan patients with squamous EC, and may be correlated with carcinogenesis and development of squamous EC.
【摘要】 目的 通过比较两种原代人脐静脉内皮细胞的分离培养方法并对细胞特异性抗原进行鉴定,探索提高原代内皮细胞体外培养存活率及纯化率的方法。 方法 采用一次性无菌注射器向人脐静脉灌注消化液,消化液的浓度和消化时间分别025%(质量体积比)胰蛋白酶,10 min和01%(质量体积比)胶原酶Ⅱ,15 min。通过在倒置显微镜下观察细胞的形态特点和用免疫荧光染色的方法对细胞进行鉴定,比较两种消化方法的优劣。 结果 01%胶原酶Ⅱ,15 min的消化方法较025%胰蛋白酶,10 min对原代人脐静脉内皮细胞有更好的分离效果,活细胞数量多且细胞纯度较高。免疫荧光染色结果表明细胞内有Ⅷ因子相关抗原表达。结论 胶原酶Ⅱ可以有效分离脐静脉内皮细胞,最佳消化条件是01%胶原酶Ⅱ,37℃,15 min。【Abstract】 Objective To explore the optimal method for primary culture of human umbilical vein endothelial cells (HUVECs). Methods HUVECs were prepared from human umbilical cords by 01% collagenase Ⅱ digestion for 15 minutes and 025 trypsinase digestion for 10 minutes,respectively. HUVECs were observed under inverted microscope and identified by immunofluorescence.The two methods of digestion were compared. Results More HUVECs were harvested through the method of 01% collagenase Ⅱ for 15 minutes,which expressed Ⅷ related antigen. Conclusion The method of 0.1% collagenase Ⅱ digestion for 15 minutes is a better choice to isolate HUVECs.
Objective To provide references for the rational allocation of health personnel in rural hospitals through understanding the status of health human resources of rural hospitals in remote and poor areas of Sichuan Province. Methodes This study used cluster sampling method, combined with questionnaire survey and qualitative interviews. A total of 711 health workers of 29 rural hospitals in Pengzhou and Baoxing of Sichuan Province were interviewed. SPSS16.0 was used for descriptive analysis.Results The average age of rural hospitals health personnel in remote and poor areas of Sichuan Province was 30 years old. Post-secondary education accounted for 58.12%, and Bachelor degree or above accounted for 7.2%. The number of medium and senior professional titles account for 8.4 %. The ratio of doctors to nurses was 1:0.55. In the survey of health workers, those doctors with practice (assistant) license accounted for 38.5%, and those without any qualification occupied 27.1 %. Conclusions The professional titles of medical personnel of rural hospitals in remote and poor areas in Sichuan province are generally low. The distribution of professional categories is irrational. The staff in charge of prevention and care are inadequate. There exist a large number of unqualified medical workers. Therefore, the government should increase the investment in rural health and take measures to stabilize the team structure, introduce the talented, and strengthen the training for health personnel of rural hospitals to improve their overall quality.
Schwann cells (SC) play an important role in nerve regeneration. The cultures of both human and rabbit SC (gt;99%) were obtained, and were separately derived from the sciatic nerve of the human fetus and the rabbit respectively by "the method of reexplantation". In addition, the cryostore and resuscitation of SC were carried out, and the resuscitated cells could retain their growth properties.
Brain-computer interface (BCI) can establish a direct communications pathway between the human brain and the external devices, which is independent of peripheral nerves and muscles. Compared with invasive BCI, non-invasive BCI has the advantages of low cost, low risk, and ease of operation. In recent years, using non-invasive BCI technology to control devices has gradually evolved into a new type of human-computer interaction manner. Moreover, the control strategy for BCI is an essential component of this manner. First, this study introduced how the brain control techniques were developed and classified. Second, the basic characteristics of direct and shared control strategies were thoroughly explained. And then the benefits and drawbacks of these two strategies were compared and further analyzed. Finally, the development direction and application prospects for non-invasive brain control strategies were suggested.
ObjectiveTo detect the expression of human transforming growth factor β1 (hTGF-β1) gene mediated by adenovirus (Ad) in hamstring tendon after anterior cruciate ligament (ACL) reconstruction in rabbits. MethodsAd-hTGF-β1 and Ad-green fluorescent protein (GFP) were diluted to 5×108 PFU/mL with DMEM. Forty-eight New Zealand white rabbits were divided into 3 groups randomly (n=16) for ACL reconstruction with hamstring tendon autograft. Hamstring tendon was cultured and transfected with Ad-hTGF-β1 (group A) and Ad-GFP (group B) for 12 hours before ACL reconstruction, and was cultured with DMEM in group C. After 12 hours of transfection, green fluorescence was observed in groups A and B under fluorescence microscopy. At 2, 4, 6, and 8 weeks after operation, the hamstring tendon was harvested to detect the mRNA and protein expressions of hTGF-β1 by real time fluorescence quantitative PCR and Western blot. ResultsGreen fluorescence was observed after 12 hours of transfection in groups A and B. TGF-β1 protein level reached (221.0±12.2) ng/mL at 12 hours in group A. The hTGF-β1 mRNA expression could be detected in group A, but it could not be detected in group B and group C. The mRNA expression levels of hTGF-β1 were 1.004±0.072 at 2 weeks, 0.785±0.038 at 4 weeks, 0.469±0.053 at 6 weeks, and 0.172±0.021 at 8 weeks in group A, showing significant difference (P<0.05). Western blot results showed weakly positive band in groups B and C; the protein expression of TGF-β1 in group A was significantly higher than that in groups B and C (P<0.05), but no significant difference was found between groups B and C P>0.05). The protein expression of TGF-β1 gradually reduced with time, showing significant difference between different time points (P<0.05). ConclusionAd-hTGF-β1 can transfect the hamstring tendon successfully, and it can effectively express for a long time after ACL reconstruction.
Objective To investigate the effects of matrine on cell proliferation and expression of connective tissue growth factor( CTGF) and hypoxia inducible factor-1α( HIF-1α) of human lung fibroblast ( WRC-5) in normoxia ( 21% O2, 74% N2 , 5% CO2 ) and hypoxia ( 1% O2, 94% N2 , 5% CO2 )conditions. Methods MRC-5 cells were cultured and divided into differrent groups interfered with different dose of Matrine ( final concentration of 0 ～3. 2 mmol / L) in normoxia or hypoxia for 24 h. Cells were dividedinto 8 groups according to culture conditions, ie. normoxiagroup( N0 group) , normoxia + matrine 0. 2 mmol / L group( N0. 2 group) , normoxia + matrine 0. 4 mmol / L group( N0. 4 group) , normoxia + matrine 0. 8 mmol / L group( N0. 8 group) , hypoxia group( H0 group) , hypoxia + matrine 0. 2 mmol /L group( H0. 2 group) , hypoxia +matrine 0. 4 mmol /L group( H0. 4 group) , and hypoxia + matrine 0. 8 mmol / L group( H0. 8 group) . The MTT assay was used to measure the cell proliferation activity. Western-blot assay was used to examine the expression of CTGF and HIF-1α. Results Hypoxia promoted the cell proliferation in all groups( P lt;0. 05) .Matrine inhibited the proliferation of WRC-5 cells in a concentration-dependent manner in hypoxia or normoxia conditions( P lt;0. 05) . The expression of CTGF andHIF-1αwas lower in normoxia and higher in hypoxia( P lt;0. 01) . Matrine inhibited the expression of CTGF and HIF-1αin a concentration-dependent manner in hypoxiaand normoxia( P lt;0. 05) . Conclusion Matrine can inhibit the cell proliferation and the expression of CTGF and HIF-1αof WRC-5 cells in normoxia and hypoxia in a concentration-dependent manner.
Objective To investigate whether human amniotic mesenchymal stem cells (hAMSCs) have the characteristics of mesenchymal stem cells (MSCs) and the differentiation capacity into ligament fibroblastsin vitro. Methods The hAMSCs were separated through trypsin and collagenase digestion from placenta, the phenotypic characteristics of hAMSCs were detected by flow cytometry, the cytokeratin-19 (CK-19) and vimentin expression of hAMSCs were tested through immunofluorescence staining. The hAMSCs at the 3rd passage were cultured with L-DMEM/F12 medium containing transforming growth factor β1 (TGF-β1) and vascular endothelial growth factor (VEGF) as the experimental group and with single L-DMEM/F12 medium as the control group. The morphology of hAMSCs was observed by inverted phase contrast microscope; the cellular activities and ability of proliferation were examined by cell counting kit-8 (CCK-8) method; the ligament fibroblasts related protein expressions including collagen type I, collagen type III, Fibronectin, and Tenascin-C were detected by immunofluorescence staining; specific mRNA expressions of ligament fibroblasts and angiogenesis including collagen type I, collagen type III, Fibronectin, α-smooth muscle actin (α-SMA), and VEGF were measured by real-time fluorescence quantitative PCR. Results The hAMSCs presented monolayer and adherent growth under inverted phase contrast microscope; the flow cytometry results demonstrated that hAMSCs expressed the MSCs phenotypes; the immunofluorescence staining results indicated the hAMSCs had high expression of the vimentin and low expression of CK-19; the hAMSCs possessed the differentiation ability into the osteoblasts, chondroblasts, and lipoblasts. The CCK-8 results displayed that cells reached the peak of growth curve at 7 days in each group, and the proliferation ability in the experimental group was significantly higher than that in the control group at 7 days (P<0.05). The immunofluorescence staining results showed that the expressions of collagen type I, collagen type III, Fibronectin, and Tenascin-C in the experimental group were significantly higher than those in the control group at 5, 10, and15 days after culture (P<0.05). The real-time fluorescence quantitative PCR results revealed that the mRNA relative expressions had an increasing tendency at varying degrees with time in the experimental group (P<0.05). The relative mRNA expressions of collagen type I, collagen type III, Fibronectin, α-SMA, and VEGF in the experimental group were significantly higher than those in the control group at the other time points (P<0.05), but no significant difference was found in the relative mRNA expressions of collagen type I, collagen type III, and VEGF between 2 groups at 5 days (P>0.05). Conclusion The hAMSCs possesses the characteristics of MSCs and good proliferation ability which could be chosen as seed cell source in tissue engineering. The expressions of ligament fibroblasts and angiogenesis related genes could be up-regulated, after inductionin vitro, and the synthesis of ligament fibroblasts related proteins could be strengthened. In addition, the application of TGF-β1 and VEGF could be used as growth factors sources in constructing tissue engineered ligament.
ObjectiveTo test the expressions of human mammary gland globin (hMAM) mRNA in the peripheral blood of breast cancer and breast benign lesions patients, try to provide the theory basis for the choice of breast cancer molecular marker. MethodsPolymerase chain reaction (PCR) technology was used to detect the expressions of hMAM mRNA in peripheral blood of 78 cases of breast cancer patients, 15 cases of hyperplasia of mammary gland, and 15 cases of breast fibroadenoma. The relationship between the expressions of hMAM-mRNA in peripheral blood of breast cancer patients with patient's age, tumor size, pathological type, tumor stage, axillary lymph node metastasis, and the ER, PR and HER-2 status were analyzed. ResultsThe expressions of hMAM-mRNA in peripheral blood were not detected in breast hyperplasia and breast fibroadenoma patients, but the peripheral blood hMAM-mRNA expression rate in breast cancer patients was 48.72% (38/78), the difference was statistically significant (χ2=12.357, P=0.000). The expression of peripheral blood hMAM mRNA was not related to the patient's age, tumor size, pathological type, and ER, PR and HER-2 status (P > 0.05), but the expression of peripheral blood hMAM mRNA was related to the clinical staging of tumor (Z=-2.214, P=0.027) and lymph node metastasis status (Z=-2.754, P=0.006). ConclusionPeripheral blood hMAM-mRNA detected is a sign of breast cancer, further research is needed to confirm whether hMAM mRNA detection in peripheral blood correlates with poor prognosis of breast cancer patients.
【Abstract】ObjectiveTo develop a new singletube polymerase chain reaction amplification (ST Amp) protocol for the efficient sequencing-based typing (SBT) of human leukocyte antigen DRB1（HLA-DRB1）.MethodsA set of 7 group-specific exonic 5′ amplification primers and a single generic 3′ primer were included together in a single PCR mix to facilitate a single PCR amplification per sample for HLA-DRB1 typing.ResultsAll samples were successfully typed, the typing result was accurate and repeatable.ConclusionST Amp technique has resulted in the ability to perform high-resolution, high-specificity and high-throughput HLA-DRB1 typing by DNA sequencing.