The desired DNA product of KGPcd and KGP-hag was obtained from the total DNA of Porphyromonas gingivalis by PCR with two pairs of gene specific primers. The segment of KGPcd and KGP-hag (about 1.5kb and 1.6kb) was inserted into pGEM-T easy Vector. The double-stranded DNA of the postitive clone was analyzed by restriction endonuclease mapping and DNA sequenceing. The sequences of KGPcd and KGP-hag were consistent with those of the references appeared. The proteins of KGPcd and KGP-hag will be obtained for further study.

OBJECTIVETo investigate the changes in proliferation, invasiveness, clone sphere formation and chemosensitivity of human gastric cancer cell lines of KATO-III CD133(+) cells transfected with small interfering RNA (siRNA) against CD133 gene.

METHODSCD133(+) cells of KATO-III cell lines were isolated by magnetic activated cell sorting (MACS). CD133 siRNA was designed and synthesized, and then transfected into KATO-III CD133(+) cells. Cell fluorescence counting under confocal laser scanning microscope was used to determine the transfection efficiency after transfection with the CD133 FITC-siRNA. The knock-down effect of the CD133 gene and expression of epithelial-mesenchymal transition (EMT)-related factors were detected by RT-PCR and Western blotting. Cell counting kit-8 assay (CCK-8), transwell chamber and colony sphere forming assay were performed to measure the variation of cell proliferative, invasive, colony formation viability and chemosensitivity to 5-FU after the above-mentioned treatment.

RESULTSThe transfection efficiency was (87.7±8.1)%. The CD133 mRNA and protein expression levels in the interference group were lower than those in negative control group. Twenty-four, 48 and 72 hours after transfection, cells proliferation activity was significantly inhibited in the interference group compared with negative control group, (all P<0.01). Seventy-two hours after transfection, compared with negative control group, cells proliferation activity was reduced by (52.1±8.0)%. The invasive cell number reduced (41.7±6.0 vs. 130.3±11.0, P<0.05) and clone formation rate decreased significantly [(24.3±4.3)% vs. (45.1±6.4)%, P<0.01] in the interference group. EMT-related gene E-cadherin protein expression increased, while the Snail and N-cadherin protein expression reduced in the interference group (all P<0.01). The cells sensitivity to 5-FU was significantly enhanced in the interference group, and the cell inhibition rate of 5-Fu was (62.4±3.3)%, higher than that in negative control group [(21.5±2.2)%, P<0.01].

CONCLUSIONSThe expression of CD133 gene plays an important role in cell proliferation, invasiveness, colony formation and resistance to chemotherapy of KATO-III CD133(+) gastric cancer cells. It suggests that CD133 can be used as one of surface markers for detection of gastric cancer stem cells. Inhibition of CD133 expression may be a promising way for gastric cancer biotherapy.

AC133 Antigen ; Antigens, CD ; genetics ; metabolism ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Fluorouracil ; pharmacology ; Glycoproteins ; genetics ; metabolism ; Humans ; Peptides ; genetics ; metabolism ; RNA Interference ; RNA, Small Interfering ; genetics ; Stomach Neoplasms ; genetics ; metabolism ; pathology ; Transfection

OBJECTIVETo analyze the early and long-term results after mitral-aortic valve replacement for rheumatic valvular disease and the determinant factors involved and subsequent therapies.

METHODS1 154 patients receiving combined mitral-aortic valve replacement for rheumatic valvular disease from May 1981 to May 2001 were reviewed. The mean age of the patients was 41.48 +/- 10.00 years. Concomitant valve plasty was performed for associated tricuspid organic or significant functional lesions. Lateral tilting disc or bileaflet valve prostheses were used for replacement. New York Heart Association functional status showed Class III or IV in 91.77% of the patients. Moderate to severe pulmonary hypertension occurred in 29.38% of the patients. The duration of follow-up varied from 8 months to 20 years.

RESULTSThe hospital mortality was decreased from 6.50% to 4.45%. The 5-, 10- and l5-year survival rates were 89.46% +/- 1.35%, 86.50% +/- l.91% and 67.86% +/- 6.16%, respectively. The 5-, 10- and l5-year thromboembolic event free rates were 97.80% +/- 0.74%, 88.31% +/- 2.20% and 94.08% +/- 2.29%, respectively. the 5-, 10- and l5-year anticoagulant related bleeding free rates were 94.80% +/- 1.09%, 89.32% +/- 2.10% and 83.12% +/- 3.57% respectively. Cardiac functional status returned to Class II in 98% patients and to Class III in 2% during follow-up.

CONCLUSIONSBoth left and right ventricular functions may be impaired as a result of rheumatic valvular disease. Tricuspid valve should be explored during surgery and any significant tricuspid annular enlargement and regurgitation showed be corrected in concomitance. Long-acting penicillin regimen is needed for 3 - 5 years for the prevention of rheumatic fever relapse. A low intensity anticoagulant regimen after valve replacement with prothrombin time targeting at 1.5 - 2.0 times is advisable in lessening anticoagulant related bleeding yet optimizing sufficient prevention against thromboembolic complications.

Adolescent ; Adult ; Aged ; Aortic Valve ; surgery ; Female ; Follow-Up Studies ; Heart Valve Diseases ; etiology ; surgery ; Heart Valve Prosthesis Implantation ; methods ; mortality ; Humans ; Male ; Middle Aged ; Mitral Valve ; surgery ; Postoperative Complications ; prevention & control ; Recurrence ; Retrospective Studies ; Rheumatic Heart Disease ; complications ; prevention & control ; Survival Analysis ; Survival Rate ; Treatment Outcome ; Tricuspid Valve ; surgery ; Young Adult

OBJECTIVETo study the changes of genomic density polymorphism, quantitative expression and the adhesion activity of complement receptor type 1 (ECR1) on erythrocytes in patients with chronic hepatitis.

METHODSPolymerase chain reaction (PCR) and Hind restriction enzyme digestion, the quantitative assay of ECR1 and the activity of erythrocytes immune adhesion test were applied.

RESULTSThe spot mutation rate (25.0%-30.3%) of ECR1 density gene in patients with chronic hepatitis was not significantly different from that of healthy individuals (28.0%). The amount of ECR1 in patients with chronic hepatitis, except for the diseases with normal liver function, was significantly lower than that of healthy individuals (t=9.87,P<0.000 1). The quantitative expression of ECR1 in decompensated cirrhosis was obviously lower than that of compensated cirrhosis (t=2.21,P<0.05).

CONCLUSIONSDefective expression of ECR1 in chronic hepatitis B may be acquired through central and/or peripheral mechanisms. It is very important to study the quantitative expression in the patients with chronic hepatitis.

Erythrocytes ; immunology ; metabolism ; Hepatitis B, Chronic ; genetics ; Humans ; Liver Cirrhosis ; genetics ; Polymerase Chain Reaction ; Polymorphism, Genetic ; Receptors, Complement ; analysis ; genetics ; metabolism ; Tissue Adhesions

OBJECTIVETo examine the association between CD133 expression and invasion of gastric cancer, and to elucidate whether CD133 can promote the invasion and metastasis of gastric cancer via epithelial-mesenchymal transition (EMT).

METHODSThe CD133(+) and CD133(-) KATO-III( cells were sorted by magnetic activated cell sorting (MACS). The invasion ability was detected by Transwell method. RT-PCR and Western blot were used to detect the expression of EMT-related factors in KATO-III( cells before and after CD133 was knocked out by siRNA method. The expressions of CD133 and EMT-related proteins of cancer and adjacent normal tissues in 50 patients with gastric cancer were detected by Western blot, and correlations among protein expressions were also analyzed.

RESULTSAs compared to CD133(-) cells, the number of broken-membrane cells was significantly higher (67.7±10.5 vs. 13.3±6.8, P=0.001) and the invasion ability was stronger (P<0.05) in CD133(+) cells, while the mRNA expression levels of Snail and N-cadherin were significantly higher in CD133(+) cells (0.311±0.015 vs. 0.223±0.016, P=0.040; 0.581±0.020 vs. 0.270±0.018,P=0.004), and the protein expression levels of Snail and N-cadherin were significantly higher in CD133(+) cells as well (0.513±0.015 vs. 0.179±0.023, P=0.030; 0.538±0.028 vs. 0.202±0.032, P=0.020), but E-cadherin mRNA and protein levels were significantly lower in CD133(+) cells (0.231±0.009 vs. 0.460±0.015, P=0.040; 0.426±0.030 vs. 0.748±0.027, P=0.040). After CD133 knock-out, the expressions of Snail and N-cadherin were down-regulated (P<0.05) and the expression of E-cadherin was up-regulated (P<0.05). As compared to normal mucosal tissues, the protein expression levels of Snail, N-cadherin and CD133 in gastric cancer tissues were significantly higher(0.635±0.119 vs. 0.485±0.116, P=0.029; 0.599±0.114 vs. 0.259±0.108, P=0.020; 0.754±0.154 vs. 0.329±0.134, P=0.001), while the protein expression of E-cadherin in gastric cancer tissues was lower (0.378±0.123 vs. 0.752±0.156, P=0.003). The protein expressions of Snail and N-cadherin were positively correlated with CD133 expression (r=0.278, P=0.048; r=0.406, P=0.003) and the protein expression of E-cadherin was negatively correlated with CD133 expression (r=-0.504, P=0.000).

CONCLUSIONCD133(+) cells in primary lesion of gastric cancer have relatively higher invasion ability, which may promote the metastasis of gastric cancer via up-regulation of EMT-related factors.

AC133 Antigen ; Adult ; Aged ; Aged, 80 and over ; Antigens, CD ; metabolism ; Cell Line, Tumor ; Epithelial-Mesenchymal Transition ; Female ; Glycoproteins ; metabolism ; Humans ; Male ; Middle Aged ; Neoplasm Invasiveness ; Neoplasm Metastasis ; Peptides ; metabolism ; Stomach Neoplasms ; metabolism ; pathology

OBJECTIVETo explore the feasibility and efficacy of the vacuum sealing drainage (VSD) technique combined with skin flap for the treatment of chronic ulcerative wounds.

METHODSFrom June 2009 to Aug. 2011, the VSD technique combined with skin flap has been applied in the treatment of 15 patients with chronic ulcerative wounds caused by various reasons. The VSD was applied to the wound for 1-6 times. When infection was controlled and fresh granulation grew, skin flap was used to cover the wound.

RESULTSFlap necrosis happened in a small area at the distal end in one case, which healed after skin graft. All the other flaps survived with primary healing. The patients were followed up for 6-24 months postoperatively with no recurrence of infection.

CONCLUSIONSVSD combined with skin flap is an ideal choice for reconstruction of chronic ulcerative wounds. It has the advantages of low complications, reliable flap survival rate, and low infection recurrence.

Adult ; Aged ; Female ; Humans ; Male ; Middle Aged ; Negative-Pressure Wound Therapy ; Skin Transplantation ; methods ; Surgical Flaps ; Treatment Outcome ; Ulcer ; surgery

Objective To analyze the dynamic changes and the influencing factors of T lymphocyte subsets in recipients with stable graft status within 1 year after lung transplantation. Methods Clinical data of 41 recipients with stable graft status after allogeneic lung transplantation were analyzed. The absolute value and ratio of T lymphocyte subsets in peripheral blood from recipients were measured by flow cytometry before operation, 2 weeks and each month (within 1 year) after operation, respectively. The effects of age, gender, body mass index (BMI), surgical method, incidence of primary graft dysfunction (PGD) after operation, and primary disease upon the absolute values of T lymphocytes were evaluated. Results Within 1 year after lung transplantation, the absolute values of CD3⁺, CD3⁺CD4⁺, CD3⁺CD8⁺T lymphocytes and CD4⁺/CD8⁺ ratio were changed over time (all P < 0.001). Compared with preoperative values, there was no statistical significance in the absolute values of CD3⁺ and CD3⁺CD4⁺T lymphocytes at 12 months after operation (P=0.659, 0.109), whereas the absolute value of CD3⁺CD8⁺T lymphocytes was increased (P=0.02) and the CD4⁺/CD8⁺ ratio was decreased (P < 0.001). Age, gender, BMI, surgical method and incidence of PGD after operation exerted no significant effect on the dynamic changes of absolute values of CD3⁺CD4⁺ and CD3⁺CD8⁺T lymphocytes (all P > 0.05). Primary disease before lung transplantation exerted no effect on the changes of CD3⁺CD4⁺T lymphocytes, whereas the postoperative absolute value of CD3⁺CD8⁺T lymphocytes was higher in recipients with infectious lung diseases (P < 0.05). Conclusions The absolute values of CD3⁺, CD3⁺CD4⁺, CD3⁺CD8⁺T lymphocytes in recipients with stable graft status after lung transplantation are relatively low in the early stage after lung transplantation, then gradually restore, and stabilize at 6 months after operation. Dynamic changes are not associated with age, gender, BMI, surgical method and incidence of PGD after operation of recipients.

Objective To analyze the risk factors and clinical prognosis of acute kidney injury (AKI) early after lung transplantation. Methods Clinical data of 155 recipients undergoing lung transplantation or combined heart-lung transplantation were retrospectively analyzed, and they were divided into the AKI group (n=104) and non-AKI group (n=51) according to the 2012 Kidney Disease: Improving Global Outcomes Clinical Practice Guideline. The incidence of AKI early after lung transplantation was summarized. The main indexes of recipients were collected. The risk factors of the occurrence of AKI early after lung transplantation were subjected to univariate and multivariate analysis. The clinical prognosis of lung transplant recipients was evaluated and the survival curve was delineated. Results The incidence of AKI early after lung transplantation was 67.1%(104/155), including 47 recipients with stage 1 AKI, 34 recipients with stage2 AKI and 23 recipients with stage 3 AKI, respectively. Sixteen recipients required continuous renal replacement therapy (CRRT) early after lung transplantation. Preoperative complication with diabetes mellitus, preoperative complication with pulmonary hypertension, intraoperative mean arterial pressure (MAP) < 60 mmHg, intraoperative massive blood transfusion, and treatment with excessive therapeutic concentration of tacrolimus (Tac) within postoperative 1 week were the independent risk factors for the occurrence of AKI early after lung transplantation. Up to the end of follow-up, 66 recipients (42.6%) died, including 50 recipients in the AKI group and 16 recipients in the non-AKI group. The cumulative survival rate in the AKI group was significantly lower than that in the non-AKI group (40% vs. 66%, P < 0.05). With the increase of AKI severity, the cumulative survival rate of lung transplant recipients was decreased. Conclusions AKI develops early after lung transplantation with high incidence and poor clinical prognosis. Preoperative complication with diabetes mellitus and pulmonary hypertension, intraoperative MAP < 60 mmHg and massive blood transfusion, and treatment with excessive therapeutic concentration of Tac within postoperative 1 week are the independent risk factors for the occurrence of AKI early after lung transplantation.

Acute cellular rejection (ACR) is a common complication after lung transplantation, which is mainly caused by the immune response of T lymphocytes recognizing the major histocompatibility complex on the cellular surface of grafts. It is currently considered as the main pattern of acute rejection. ACR is not only a direct cause of death of recipients, but also a high-risk factor for chronic rejection after lung transplantation. Nevertheless, it is a challenging task to deliver the diagnosis and treatment of ACR following lung transplantation. In this article, new progresses on the risk factors, pathogenesis, diagnosis and treatment of ACR in lung transplant recipients were summarized, aiming to improve the diagnostic and treatment efficiency of ACR and prolong the survival of recipients.

Objective To explore the association between metabolic syndromes (MS) and carotid atherosclerosis and to estimate the predictive effects of MS under 3 different definitions.Methods A cross-sectional study was conducted in 2 community-based populations in Beijing,in 2008.1266 subjects (598 men,668 women),aged 45-69,were included in the analyses.MS was defined by the criteria of International Diabetes Federation (IDF),the revised NCEPATPm (ATP Ⅲ-R) and "The Guidelines of Dyslipidemia Control for Chinese Adult" ( "Guidelines" ) in 2007.Results The prevalence rates of MS by the 3 criteria were 39.0%,43.3% and 30.9% respectively.The Kappa value for the measure of the agreement between each pair of the 3 definitions were 0.911,0.719 and 0.730 respectively.The intima-media thickness in common carotid artery (CCA-IMT) was significantly higher (P＜0.001) in all MS groups than in non-MS groups,diagnosed with the 3 criteria independent of age,gender,LDL-C,and current smoking status.After adjustment of age,gender,LDL-C,and current smoking status,the classification of MS significantly increased the risk of prevalence of carotid atberosclerotic plaques,compared to the non-MS group.OR value were 1.499 (95% CI:1.157-1.942) for IDF,1.696 (95% CI:1.314-2.189) for NCEP-R,1.763 (95% CI:1.344-2.312) for "Guideline" respectively.Conclusion Our research findings indicated that,when MS were defined with the 3 definitions,prediction on the risk of sub-clinical atberosclerosis would work beyond some of the conventional cardiovascular risk factors such as smoking,LDL-C.There might exist some differences in gender issue on the strength of association between MS when diagnosed by different criteria and carotid plaque.