Background The in vitro culture of retinal vascular endothelial cells is the foundation of experimental study of retinal vascular disease. Shortage of human donor eyeballs is a main limiting for the laboratory work. The culture method of rat-derived vascular endothelial cells has been established. However, this method is not enough effective because of severer cellullar injury. Objective Present study was to establish a simple and high effective method for the culture of vascular endothelial cells in vitro. Methods The retinas from 5 SPF SD rats was digested by 0. 1% collagenase and cultured with explant culture method. 20% fetal bovine serum, vascular endothelial growth factor ( VEGF) , insulin-transferrin-selenium( ITS) were composed into the endothelial cell culture medium, and enough blowing was performed to get the cells and fragments from retinal tissue. The cellular suspension was prepared and cultured consequently on human fibronectin-coated culture flasks. Cultured vascular endothelial cells were identified by anti-von Willebrand staining factor. Results The cells emerged from the tissue mass,and cells and some tissue fragments attached to the wall after 24 hours of seeding. The cells grew to show the fusiform in 4 days and merged together in 5 to 6 days,and a cell monolayer was seen in the 14th day after culture. The endothelial cells showed the positive response for von Willebrand factor. After passage, the merging-growth statue of the cells was regained in 2 hours after culture. Conclusion Use of retinal pieces and collagenase-digestion can get the vascular endothelial cells with better activity in vitro. The culture method based on highly selective endothelial cell culture medium associated to FN adhesion-promoting is helpful for gaining the purified of endothelial cells.

Objective To investigate the clinical efficacy of meloxicam combined with glucosamine sulfate in the treatment of patients with osteoarthritis.Methods Ninety cases of knee osteoarthritis were randomly divided into control group and treatment group,each group 45 cases.Control group was given meloxicam tablets 7.5 mg,bid,oral.On the basis of control group,treatment group was treated with glucosamine sulfate capsules 0.5 g,tid,oral.All the patients were treated for 6 weeks.The clinical symptoms,signs,knee joint motor function and pain indexes of all patients were evaluated before and after treatment.The blood samples and joint fluid specimens were collected,to determine the matrix metalloproteinase-3 (MMP-3),matrix metalloproteinase-13 (MMP-13) and prostaglandin E2 (PGE2) levels by enzyme linked immunosorbent assay (ELISA).The clinical efficacy and adverse drug reactions in two groups were compared.Results After treatment,the levels of Lequesne indexes in treatment group and control group were 5.78 ± 3..04 and 8.24 ± 3.62,Lysholm scores were 85.82 ± 10.01 and 72.46 ± 13.58,VAS scores were 2.45 ± 1.08 and 3.86 ± 1.17,serum MMP-3 levels were (114.69 ± 13.54) and (130.52 ± 16.48) ng · L-1,serum MMP-13 levels were (141.06 ± 21.55) and (179.82 ± 24.37) ng · L-1,serum PGE2 levels were(69.82 ± 18.67)and(91.30 ±20.09)ng · L-1,joint fluid MMP-3 levels were(17.89 ±5.12) and (23.10 ±5.46)ng · L-1,joint fluid MMP-13 levels were(27.35 ±6.52)and(38.47 ±6.85)ng · L-1,joint fluid PGE2 levels were (82.16 ± 22.64) and (118.02 ± 23.41) ng · L-1,all with significant difference (all P ＜ 0.05).The improvement rates of control group and treatment group were 73.33％ (33 cases/45 cases),91.11％ (41 cases/45 case),with significant difference (P ＜ 0.05).No adverse drug reactions occurred during the treatment.Conclusion The clinical effect of meloxicam combined with glucosamine sulfate in the treatment of knee osteoarthritis is exact,the treatment is conducive to the improvement of clinical symptoms,can reduce serum MMP-3,MMP-13,PGE2 levels,with high safety.

AIMTo explore the molecular biological mechanism of hemoglobin with high oxygen affinity in Tibetans by determining the sequence of globin cDNA in Tibetans living at high altitude.

METHODSTotal RNA was isolated from human bone marrow samples of three Tibetans who live in Qinghai-Tibet plateau. cDNA fragments coding for alpha and beta genes of human hemoglobin were obtained through RT-PCR and were ligated to plasmid pGEM-T easy vectors, and then the ligation liquid were transformed to Escherichia coli and cloned and sequenced. Nucleotide sequences were compared with GenBank data by BLAST method.

RESULTSsequence of a globin cDNA in Tibetans were the same with the registering globin genes in the GenBank, and Hb Abruzzo (beta143 (H21), His- > Arg) gene mutation, a high oxygen affinity beta globin mutation, was found in one Tibetan' beta goblin coding gene (CAC- > CGC).

CONCLUSIONThis hemoglobin gene mutation may be associated with high altitude adaptation of Tibetans living at high altitude.

Adult ; Altitude ; Asian Continental Ancestry Group ; genetics ; Cloning, Molecular ; DNA, Complementary ; genetics ; Hemoglobins, Abnormal ; genetics ; Humans ; Male ; Middle Aged ; Molecular Sequence Data ; RNA ; Sequence Analysis, DNA ; Tibet ; alpha-Globins ; genetics ; beta-Globins ; genetics

OBJECTIVETo study the role of serum from asphyxiated neonates in the inducement of human renal proximal tubular epithelial cells (HK-2) adhesion to neutrophils and possible mechanisms.

METHODSHK-2 cells were cultured randomly with 20% serum from neonates (1, 3, and 7 days after asphyxia), pyrrolidine dithiocarbamate (PDTC) or placebo. The activity of myeloperoxidase (MPO), an indicator of adhesion ability of HK-2 cells to neutrophils in suspensions, was detected by the biochemistry assay. Intercellular adhesion molecule-1 (ICAM-1) and nuclear factor-kappaB (NF-kappaB) of HK-2 cells were examined with the immunohistochemical staining.

RESULTSThe expression of MPO in the post-asphyxial 1-day serum treatment group were significantly higher than that in the PDTC treatment and the control groups as well as the post-asphyxial 3 and 7-day serum treatment groups (P<0.01). The expression of ICAM-1 and NF-kappaB in the post-asphyxial 1-day serum treatment group was also significantly higher than that in the other groups (P<0.01).

CONCLUSIONSSerum from asphyxiated neonates can induce HK-2 cell adhesion to neutrophils, possibly through activating NF-kappaB and increasing the synthesis and expression of ICAM-1 on the surface of renal tubular epithelial cells.

Asphyxia Neonatorum ; blood ; complications ; Cell Adhesion ; Cells, Cultured ; Humans ; Infant, Newborn ; Intercellular Adhesion Molecule-1 ; analysis ; biosynthesis ; Kidney Tubules ; pathology ; NF-kappa B ; analysis ; metabolism ; Neutrophils ; physiology

With the development of molecular pathology,many types of epidermal growth factor receptor (EGFR) mutations have been identified.The efficacy of EGFR tyrosine kinase inhibitors (EGFR-TKIs) in non-small cell lung cancer (NSCLC) patients with different types of EGFR mutations,especially in patients with single rare mutations or complex mutations (co-occurrence of two or more different mutations),has not been fully understood.This study aimed to examine the efficacy of EGFR-TKIs in NSCLC patients with different types of EGFR mutations.Clinical data of 809 NSCLC patients who harbored different types of EGFR mutations and treated from January 2012 to October 2016 at Renmin Hospital and Zhongnan Hospital,Wuhan,were retrospectively reviewed.The clinical characteristics of these patients and the efficacy of EGFR-TKIs were analyzed.Among these patients,377 patients had only the EGFR del-19 mutation,362 patients the EGFR L858R mutation in exon 21,33 patients single rare mutations and 37 patients complex mutations.Among these 809 patients,239 patients were treated with EGFR-TKIs.In all the 239 patients,the disease control rate (DCR) was 93.7％ with two patients (0.2％) achieving complete response (CR),the median progression free survival (PFS) was 13.0 months (95％ confidence interval [CI],11.6-14.4 months),and the median overall survival (OS) was 55.0 months (95％ CI,26.3-83.7 months).Subgroup analysis revealed that the DCR in patients harboring single rare or complex mutations of EGFR was significantly lower than in those with del-19 or L858R mutation (P＜0.001).Patients with classic mutations (del-19 and/or L858R mutations) demonstrated longer PFS (P＜0.001) and OS (P=0.017) than those with uncommon mutations (single rare and/or complex mutations).Furthermore,the patients with single rare mutations had shorter median OS than in those with other mutations.Multivariate Cox regression analysis identified that the type of EGFR mutations was an independent risk factor for PFS (hazard ratio [HR]=0.308,95％ CI,0.191-0.494,P＜0.001) and OS (HR=0.221,95％ CI,0.101-0.480,P＜0.001).The results suggest that the single rare or complex EGFR mutations confer inferior efficacy of EGFR-TKIs treatment to the classic mutations.The prognosis of the single rare EGFR mutations is depressing.EGFR-TKIs may be not a good choice for NSCLC patients with single rare mutations of EGFR.Further studies in these patients with uncommon mutations (especially for the patients with single rare mutations) are needed to determine a better precision treatment.

Objective To evaluate the situation of HIV prevalence,incidence and drug resistance(DR)among recent HIV infected men who have sex with men(MSM)during the year of 2006.Methods A transect investigation was conducted through snowballing to recruite volunteers.HIV infected status was confirmed by ELISA and Western Blot(WB),the recent infected within 155 days were confirmed by BED-Capture ELISA Assay(BED-CEIA),which based on the principle that characteristics of the initial HIV antibody response in recent infections differs from those of established or long-term infections either by antibody titer,proportion,specificity,isotype or avidity.Bayer Trugene was applied to PCR the target sequence of HIV pol gene,and the sequences were analyzed to detect the prevalence of HIV gene mutation.Furthemore.HIV antiretroviral drug resistance among MSM was evaluated.The results were analyzed through three methods:Trugene DR 3.7 IVD(2006/07/27)Guidelines 1 1.0(2005/12/15)provided byBayer company,Stanford HIVdb and Gen02pheno.Results The prevalence and incidence of HIV among MSM were 10.4%and 7.98% PY,by three methods.There was only 1 sample found to have occurred resistant Drimary mutation which could be inferred that the average DR ratio was 4.55%(1/22).Conclusion The incidence and prevalence of MSM in Chongqing were notable,and the mutation rate stands at a low level when comparing to researches conducted in developed countries.Secondary mutations appeared frequently,suggesting that more research need to be conducted to understand how HIV was transmitting among the carriers.

Objective To examine HIV prevalence and related risk factors among men who have sex with men(MSM) in Chongqing, and to explore the feasibility of using respondent-driven sampling (RDS) in the survey. Methods Based on results from formative research, a RDS survey was designed and conducted to collect demographic, behavioral and serologic data. RDSAT was used to calculate point estimation and confidence intervals. SPSS was used for bi-variate analysis using RDSAT exported weighed data. NETDRAW was used to draw network diagram. Results Among 617 subjects recruited, the adjusted HIV and syphilis prevalence rates were 16.8% and 10.9%, respectively. 73.0% of the subjects were 20 to 29 years old and 72.9% were officially registered residents of Chongqing. 83.4% were single, with the proportion of students the highest, accounting for 24.6%. During the last six months, 83.4% of them reported ever having anal sex, and 54.0% reported having unprotected anal sex. Conclusion This survey confirmed that Chongqing had a higher reported HIV prevalence among MSM than from other Chinese cities. Comprehensive intervention services were required to address this alarmingly high prevalence, with focus on intervention through internet and those having syphilis infection. RDS seemed one of the effective ways of recruiting hidden MSM populations in Chongqing which had a large population of active MSM who did not frequently visit MSM venues as compared with snowball method.

OBJECTIVETo study the betulonic acid on the cell cycle and related protein expressions on mice of bearing H22 tumor cells.

METHODFlow cytometray was used to observe the changes of betulonic acid on the cell cycle and P53 of H22 tumor cells. Immunohistochemistry was determined the espressions of PI3K and AKT.

RESULTIncreasing the doses of betulonic acid, the number of H22 cells in S phase and G2 phase was increasing gradually, it can speculate that when the betulonic acid act on cells, the cells were blocked in S and G2 phase and inhibited the protein expressions of PI3K and AKT.

CONCLUSIONBetulonic acid may be up-regulate the activity of P53 and inhibite the expressions of PI3K and AKT, so that it inhibited the survival pathway of tumor cells.

Animals ; Betula ; chemistry ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Female ; Flow Cytometry ; G2 Phase ; drug effects ; Gene Expression Regulation ; drug effects ; Immunohistochemistry ; Male ; Mice ; Neoplasm Transplantation ; Phosphatidylinositol 3-Kinases ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Tumor Suppressor Protein p53 ; metabolism

Objective To examine the effect of aqueous and ethanol extracts of Coix lacryma-jobi stem on the growth of mouse S180 sarcoma cells in mice.Methods Tumor growth inhibition assays were performed using both aqueous and ethanol extracts of Coix lacryma-jobi stem.Specific-pathogen-free (SPF) Kunming mice,6-8 weeks old,were inoculated with 1 × 107 cells · mL-1 S180 sarcoma cells in a volume of 0.4 mL The next day,the mice were randomly divided into blank group,control group,experimental-H,experimental-M,experimental-L doses groups,and administered with 0.9％ NaC1,cyclophosphamide (20 μg · g-1 · d-1) or Coix lacryma-jobi stem extract (1 000,500,250 mg · kg-1 · d-1) respectively by gavage or tail vein injection for 10 d.The tumor and spleen were dissected and weighed.The tumor inhibition rate was calculated.Results Tumor growth inhibition ratios in the experimental-H,experimental-M,experimental-L groups by the aqueous extract of Coix lacryma-jobi stem were 32.11％,75.63％ and 11.41％,respectively.Tumor growth inhibition ratios for experimental-H,experimental-M,experimental-L groups by the ethanol extract of Coix lacryma-jobi stem groups were 77.09％,80.03％ and 73.52％,respectively.Conclusion Each dose of water or ethanol extract of Coix iacryma-jobi stem was able to inhibit the growth of S180 sarcoma to a degree,and exhibited few adverse reactions.The optimal dose of the aqueous extract was 500 mg · kg-1 · d-1.

Two female patients with rectal tumor undergoing proctectomy via vagina, namely natural orifice transluminal endoscopic surgery (NOTES), are reported. The operations were performed on June 8 and August 10, 2010, respectively. No Trocar was used in the abdomen except for the transumbilical incision. There were no visible scars in the abdomen. Tubulovillous adenoma and moderately differentiated adenocarcinoma were diagnosed respectively through postoperative pathological examination. Both patients resumed normal work and life at the most recent follow up. Sexual life was satisfactory.
Adult ; Female ; Humans ; Natural Orifice Endoscopic Surgery ; methods ; Rectal Neoplasms ; surgery ; Vagina ; surgery