ABSTRACT:Objective To assess the effect of prolonged laparoscopic surgery on peritoneal mesothelial cells and fibrinolysis in humans.Methods We examined prospectively 1 6 consecutive patients who underwent laparoscopic surgery (LAP)and 2 1 patients who underwent conventional open surgery (OP)for high-medium rectal cancer with curative intent.During the procedure,biopsy of the parietal peritoneum was made before operation and at 45 min,90 min,and 120 min after operation.The tissue-type plasminogen activator (tPA)and plasminogen activator inhibitor-1 (PAI-1 )were determined by enzyme-linked immunosorbent assay in peritoneal tissues.The cellular injury was detected by LDH assay.The proliferation was quantified by MTT assay.Results PAI-1 activity in the peritoneal tissue was significantly lower in LAP group than in the OP group.tPA activity decreased after 45min of open surgery,but there was no significant change in the LAP group.With time extension,the LDH activity increased and the proliferation of the mesothelial cells decreased.Conclusion Preservation of a prolonged hypofibrinolytic state by inhibition of PAI-1 up-regulation during LAP may predispose patients to less postoperative peritoneal adhesion. The cellular injury becomes apparent and the proliferation is inhibited during prolonged laparoscopic surgery.

ABSTRACT:Objective To investigate the clinical value and application indications of the second biopsy-cervical conization in the diagnosis and treatment of cervical lesions.Methods We selected 413 patients from Department of Gynecology and Obstetrics,General Hospital of Jinan Military Command,who received cervical multi-point biopsy pathology IA for cervical cancer and cervical conization (LEEP knife)of the second biopsy between January 2012 and October 2015.Their data were retrospectively analyzed.Results Compared with cervical multi-point biopsy, the second biopsy-cervical conization after operation had pathological upgrade in 10.65% (44/413),agreement in 73.37% (303/413),and reverse (pathological levels drop and overcast)in 15.98%(66/413).Cervical multi-point biopsy and the second biopsy-cervical conization significantly differed in the diagnosis of cervical low-level intraepithelial lesion,high-grade intraepithelial lesions,and early invasive cancer (IA)(T=21.740,v=3-1=2,P<0.05).Before conization high-risk type HPV infection positive rate was 71.91%(297/413);after operation it was 86.36% (38/44)in upgrade pathology and 70.20% (259/369)in non-upgrade one.And thin prep cytologic test (TCT)showed that the positive rate (ASC-US and above)was about 87.89% (363/413), of which about 11.85% (43/363)was pathological upgrade after conization.The positive rates of high-risk type HPV infection and TCT result (ASC-US and above)in cervical lesions differed significantly between pathological upgrade and non-upgrade after conization (χ2=5.092,P<0.05,χ2=4.476,P<0.05).Conclusion The second biopsy-cervical conization technique as a means of reevaluation of cervical pathological changes in diagnosis and treatment of cervical biopsy under colposcopy can significantly improve diagnosis rate,reduce misdiagnosis and occult cervical cancer,but its clinical application has some indications.

ABSTRACT:Objective To compare the prognostic value of cystatin-C (Cys-C)and creatinine (Cr)in chronic heart failure (CHF)in old people.Methods We recruited 183 old patients (>75 years)diagnosed with CHF hospitalized at the Department of Cardiology and the Department of Geriatrics of our hospital from 2010 to 2012. Venous blood was withdrawn to detect the expressions of Cys-C and Cr.After three-year follow-up,the patients’ three-year survival was recorded.Logistic regression model was used to examine the relationship of Cys-C,Cr and other risk factors with the CHF patients’mortality.ROC curve was used to compare the prognostic value of Cys-C and Cr in CHF in old people and statistical method was adopted to analyze the combined use of Cys-C and Cr for determining their value in evaluating CHF prognosis.Results At the end of the follow-up,74 (40.44%)patients died.Serum cys-C and Cr levels of the patients in death group increased compared with those in survival group (P<0.05).Multivariate logistic regression showed that age (OR=1.11,95% CI=1.04-1.19),SBP (OR=1.06,95%CI=1.01-1.10),LVEF (OR=0.89,95% CI=0.82-0.95),BNP (OR=4.74,95% CI=1.77-12.69), creatinine (OR=2.04,95% CI=1.03-4.08),and Cys-C (OR=2.97,95% CI=1.44-6.12)were the independent risk factors for the prognosis of CHF.ROC analysis showed that Cys-C was superior to creatinine in AUC value (0.71 vs.0.65;95% CI,0.64-0.78;95% CI,0.58-0.72)(P<0.001).The AUC value of Cys-C and Cr in combination was 0.73 (95% CI,0.66-0.79).Conclusion Cys-C is a stronger predictor of the prognosis of CHF patients than creatinine and the combination of the two can increase the sensitivity and specificity in evaluating the prognosis of patients with CHF.

ABSTRACT:Objective To investigate the clinical features of seronegative rheumatoid arthritis (RA)in western China and its outcomes after one-year treatment with disease modifying anti-rheumatoid drugs (DMARDs) so as to provide evidence for effective therapy.Methods We made a retrospective analysis of 240 RA patients treated in our department from May 2013 to June 2014.We compared the 47 seropositive and 25 seronegative RA patients in clinical features,laboratory parameters and outcomes after one-year DMARDs medication.Results The percentage of seronegative RA was 10.4% (25/240).The number of swollen small joints was significantly smaller in seronegative RA group (P<0 .0 1 ).Compared with those in seropositive RA,the level of hemoglobin was lower,the level of platelets was higher,and the level of alkaline phosphatase was lower in seronegative RA (P<0 .0 5 ).The remission rate was higher in seronegative RA group than in seropositive RA group after one-year DMARDs administration (P<0.05).Conclusion Seronegative RA is not rare in clinic.Even though seronegative RA patients often present fewer swollen small joints, it is difficult to distinguish between seronegative and seropositve RA just based on the clinical features.Besides,hematological damage is more severe in some patients with seronegative RA.Only after one-year treatment with DMARDs,the remission rate is higher in seronegative RA patients than in seropositve RA ones.

ABSTRACT:Objective To explore the changes of serum cytokine interleukin-17 (IL-17)in patients with chronic hepatitis B of different clinical types and its clinical significance.Methods We selected 30 cases of mild chronic hepatitis B,34 cases of moderate one,29 cases of severe one,38 cases of liver cirrhosis,and 21 cases of acute on chronic liver failure.Another 30 cases over the same period served as the healthy control group.Cytokine IL-1 7 level in peripheral blood was detected in each group,and all the groups except the control group were detected for liver function and HBV-DNA.These related serum markers were detected and the results were statistically analyzed.Results ① IL-17 in the peripheral blood was (8.103±2.061)ng/mL in healthy control group;(25.551 ±7.078)ng/mL,(45.442±18.358)ng/mL and (75.378±19.05)ng/mL in the groups with mild,moderate and severe chronic hepatitis B;and (97.16±17.066)ng/mL in acute on chronic liver failure group.Its levels gradually increased with the severity;and there were significantly different among the five groups and between every two groups (P<0.01).The peripheral blood level of IL-17 was (8.103±2.061)ng/mL in the healthy control group, (34.517±8.905)ng/mL in compensatory cirrhosis group,and (45.615±15.623)ng/mL in the decompensated cirrhosis group.These three groups had pairwise comparison,and the difference between every two groups was significant (P<0 .0 1 ).The peripheral blood level of IL-1 7 in the decompensated cirrhosis group increased compared with that in the compensatory group.③ In the 1 5 2 cases detected,serum IL-1 7 level and serum ALT,AST,TBIL,HBV-DNA levels were positively correlated,and serum PTA had negative correlation with the level of IL-1 7 .④ In the 2 1 cases of acute on chronic liver failure,the peripheral blood level of IL-1 7 did not significantly differ between antigen-e positive and negative groups (P=0.654).⑤ In 21 patients with chronic on acute liver failure,the level of IL-1 7 in peripheral serum and MELD scores showed a positive correlation by Pearson correlation analysis (r=0.533,P=0.013).Conclusion In patients with chronic hepatitis B,the level of IL-17 in peripheral serum increased with disease severity.Moreover,the level of IL-1 7 in peripheral blood may play a role in promoting the progression of cirrhosis and the development of acute on chronic liver failure.

ABSTRACT:Objective To investigate the role of PAR2 in the visceral sensitivity of IBS patients by observing the expression of rectal PAR2 in patients with irritable bowel syndrome and the effect of exogenous PAR2 on visceral sensitivity.Methods Based on Rome III criteria,16 patients with constipation predominant IBS (IBS-C),18 patients with diarrhea predominant IBS (IBS-D ),and 1 8 controls were selected from our hospital inpatient and outpatient departments. All the patients received colonoscopic examination and rectal mucosa biopsies. The expression of rectal mucosa PAR2 was observed by immunohistochemistry and Real-time fluorescence quantitative PCR.We studied the abdominal reactions by administering the PAR2 agonist in the rectal mucosa of rats to explore whether PAR2 is involved in visceral sensitivity.Results The results of PAR2 immunohistochemistry showed that PAR2 was mainly expressed in intestinal epithelial cells,especially in the villi;in addition,endothelial cells were also found positive.While the integral optical density (IA)of PAR2 expression did not significantly differ between IBS-D or IBS-C patients and controls according to the IPP image analysis.The PAR2 mRNA level in IBS-D or IBS-C patients was not significantly different from that of the control group by Real-time PCR analysis.There was no significant difference between the IBS-D and IBS-C groups.The EMG activity significantly increased in a volume-dependent manner during the rectal balloon expansion in the PAR2 agonist group.However,there was little EMG activity when the balloon was not dilated.The area under the curve in the PAR2 agonist group with 0 mL,0.4 mL and 0.6 mL of distension volume did not differ compared with that of the vehicle group.When the balloon volume increased to 0.8 mL,1.0 mL and 1.2 mL,the EMG activity was statistically significant (P<0.01)in the PAR2 agonist group compared with the control group.Conclusion PAR2 is highly expressed in the rectal mucosa of IBS patients.Administration of exogenous PAR2 agonist increases visceral sensitivity,suggesting that PAR2 is involved in visceral sensitivity.

ABSTRACT:Objective To analyze the effects of antisurvivin oligonucleotide (Survivin ASODN)on IL-6/STAT3 signaling pathway and down-stream cancer genes of ovarian cancer SKOV3 cell line and to detect the changes of invasive ability of cell line in order to explore the role of Survivin ASODN in reducing metastasis and recurrence of ovarian cancer and its potential clinical value.Methods ASODN was transfected into SKOV3 cells by lipofectamineTM2000 in ASODN group,and lipofectamineTM 2000 was introduced in control group.The invasive ability in ASODN and control groups was detected by Transwell chamber.The expressions of interieukin-6 (IL-6 ), signal transducer and activator of transcription 3 (STAT3 ), Survivin, and vascular endothelial growth factor (VEGF)in ovarian cancer cell line at mRNA level were detected by Real-time PCR.The expressions of IL-6 , STAT3 ,signal transducer and activator of transcription 3 phosphorylation (p-STAT3 ),Survivin,and VEGF-A in ovarian cancer cell line at protein level were detected by Western blot.Results The expressions of IL-6,STAT3, Survivin,and VEGF in ovarian cancer cell line at mRNA level were significantly down-regulated in ASODN group (P<0 .0 5 ).IL-6 ,STAT 3 ,p-STAT 3 ,Survivin and VEGF-A protein levels in ovarian cancer cell line were significantly down-regulated in ASODN group (P<0.05).The invasive ability was significantly reduced by ASODN (P<0.01).Conclusion ASODN targeting Survivin could reduce the invasive ability of ovarian cancer cell line. The mechanism may be blocking IL-6/STAT3 signaling pathway and its down-stream cancer genes of ovarian cancer cell line.ASODN targeting Survivin may have clinical value in preventing and treating the metastasis and recurrence of ovarian cancer.

ABSTRACT:Objective To investigate the effect and mechanism of Hedgehog signaling pathway on the invasion of breast cancer cells in vitro.Methods The SHH,SMO and Gli-1 expression levels of breast cancer cell line MDA-231 and normal mammary epithelial cell line MCF-10A were detected by Western blot at protein level and by Real-time RT-PCR at mRNA level.Next,shRNA vector was transfected into the MDA-2 3 1 cells with highly expressed SMO,and the stable transfected cells were selected by G4 1 8 .Western blot and Real-time RT-PCR were performed to observe the inhibitory effect of RNAi on SMO expression.MTT assay was used to assess the influence of SMO siRNA on cell proliferation.Transwell assay was applied to observe cell invasion ability.The expressions of Gli-1,Snail,MMP-9,E-cadherin and Vimentin protein were determined by Western blot.Results Compared with those of normal mammary epithelial cell line MCF-10A,the expressions of SHH,SMO and Gli-1 were significantly increased.The invasion of MDA-2 3 1 cells was inhibited significantly after SMO silencing.Additionally, the protein expressions of Gli-1 , Snail, MMP-9 and Vimentin were obviously inhibited, and E-cadherin was significantly increased.Conclusion Mutative activation of Hedgehog signaling pathway in breast cancer cells promotes cell invasion probably through induction of epithelial-mesenchymal transition of the tumor cells.

ABSTRACT:Objective To observe whether N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP)can inhibit rat silicotic fibrosis by regulating stimulatory G proteinα(Gαs)/inhibitory G proteinα(Gαi)signal.Methods Male Wistar rats were randomly divided into three groups (n=1 0 ):control 1 6-w group,silicosis 1 6-w group,and Ac-SDKP pre-treatment group.The pathological changes of the lung tissue was observed by HE staining;the expressions ofα-smooth muscle actin (α-SMA),Collagen Ⅰ,fibronectin (Fn),Gαs,Gαi2 ,Gαi3 and cAMP were detected by Western blot.Immunofluorescence was performed on lung tissue sections to detect the coexpression ofα-SMA/Gαi3 . Results Within silicosis 16-w group,HE staining showed that the silicotic nodule volume increased,nodule fusion and the formation of interstitial fibrosis could be seen,and cell fibrous nodules were visible.Immunofluorescence staining showed the enhanced coexpression ofα-SMA/Gαi3 in fibrosis area.Compared with those in control group, the expressions ofα-SMA,Collagen Ⅰ,Fn,Gαi2 and Gαi3 significantly increased in silicosis 1 6-w group,but the expressions of Gαs and cAMP decreased.Compared with silicosis 1 6-w group,Ac-SDKP pre-treatment group had alleviated lung injury and decreased coexpression ofα-SMA/Gαi3 .The expressions ofα-SMA,Collagen Ⅰ,Fn,Gαi2 and Gαi3 protein significantly decreased in Ac-SDKP pre-treatment group,while the expressions of Gαs and cAMP increased obviously.Conclusion Ac-SDKP can regulate the expressions of Gαs and Gαi and promote the formation of cAMP,thus playing an effective role against silicotic fibrosis.

ABSTRACT:Objective To construct V-set and immunoglobulin domain containing 4 (Vsig4)nanobodies (Nbs) as specific macrophage probes so as to use them as molecular probes of macrophagocytes.Methods A nanobody phage library was generated by using peripheral blood lymphocytes isolated from an alpaca immunized with recombinant Vsig4 protein.After three rounds of selection against recombinant Vsig4.The Nbs were subjected to sequencing and genome alignment to obtain VHH sequence.Nbs were isolated and tested for Vsig4 specificity in an ELISA using recombinant Vsig4.The affinity capacity of Nbs was verified by the cell line stably expressing Vsig4. Results A nanobody phage library with an estimated 7.27 × 107 clones with 70% insertion was successfully constructed.Totally 1 3 6 Vsig4-positive clones were sequenced and aligned according to different CDR3 sequences. In summary,1 5 Vsig4 nanobodies were obtained and grouped into 3 different CDR3 epitopes.The affinity of representing nanobody and Vsig4 was analyzed via ELISA;Nb1 1 9 showed the highest affinity against both recombinant and native Vsig4.Conclusion We successfully constructed and screened Vsig4 specific nanobody number 1 1 9 with high affinity and specificity.It can help with macrophage detection and in vivo monitoring.