Objective To investigate the effectsof quercetin on human umbilical vein endothelial cell (HUVEC)-12 undergoing endothelial-to-mesenchymal transition (EndMT) induced by TGF-β1; To discuss its mechanism of action. Methods Cell activity of intervening by quercetin with different concentrations and TGF-β1 for 72 h was detected by CCK-8 method;RT-PCR and immunofluorescence staining were used to detect the transition of endothelial and stromal markers; Western blot was used to detect the signal transduction pathway; RT-PCR was performed to detect the transcription factors that play crucial roles in the process of transformation. Results The results of RT-PCR and immunofluorescence double staining showed that TGF-β1 (10 ng/mL) stimulated HUVEC-12 cells for 72 h to induce fibroblast phenotype, showing more interstitial markers and less endothelial markers;Western blot and RT-PCR results showed that quercetin inhibited the phosphorylation of smad2/3 in a concentration-dependent manner;After TGF-β1 stimulation, the downstream transcription factors EndMT of snail1, twist1, twist2, ZEB1, and ZEB2 significantly increased, while 100 μmol/L quercetin could down-regulate the five downstream transcription factors. Conclusion Quercetin has anti-fibrosis effects through inhibiting HUVEC-12 cells undergoing EndMT.

Objective To investigate the risk factors for vascular cognitive impairment and the effet of cerebral microbleeds (CMBs) on cognitive function in patients with ischemic stroke.Methods The data of patients with ischemic stroke over the age of 50 were collected.The Montreal cognitive assessment (MoCA) scale and Alzheimer's disease assessment scale-cognitive subscale were used to evaluate cognitive function.Hamilton depression scale (HAMD) was used to evaluate the depression status in order to exclude the patients with depression.The patients with ischemic stroke were divided into either a cognitive impairment group or a non-cognitive impairment group according to the scale evaluation results.The demographic and clinical characteristics in both groups were compared,and the multivariate logistic regression analysis was used to look for the independent risk factors for cognitive impairment in patients with ischemic stroke.The Spearman rank correlation method was used to analyze the degree of CBMs,total score of MoCA,and the correlations of all cognitive domains scores.Results A total of 169 patients with ischemic stroke were enrolled in the study.There were 80 patients in the cognitive impairment group and 89 in the non-cognitive impairment group; 34 patients had CMBs and 135 had no CMBs.The age was older (71.99 ±6.01 years vs.64.47 ±6.15 years; t =8.014,P =0.000),years of education were fewer (4.51 ± 1.534 years vs.6.94 ±2.357 years; t =8.023,P =0.000),systolic blood pressure was higher (156.19± 17.53 mm Hg vs.142.04± 16.03 mmHg,1 mmHg=0.133 kPa; t =5.479,P =0.000),scale of white matter lesion was higher (7.33 ± 2.04 vs.4.39 ± 2.17; t =8.951,P =0.000),cerebral infarction volume was larger (7 123.8 ± 1 587.1 mm3vs.5 628.4 ± 1 017.8 mm3;t =7.201 ; P =0.000),proportion of the patients with history of previous stroke or transient ischemic attack was higher (46.2％ vs.28.1％;x2 =5.982; P=0.014),and number of CBMs was larger (x2 =17.565; P=0.000) in the cognitive impairment group.Multivariate logistic regression analysis showed that the age (odds ratio [OR] 1.115,95％ confidence interval [CI] 1.013-1.227; P =0.026),years of education (OR 0.490,95％ CI0.325-0.793; P=0.001),systolic blood pressure (OR 1.048,95％ CI 1.014-1.083; P =0.005),scale of white matter lesion (OR 2.044,95％ CI 1.466-2.851; P =0.000),and cerebral infarction volume (OR 2.204,95％ CI 1.386-3.503; P =0.001) were all the independent risk factors for cognitive impairment in patients with ischemic stroke.Compared to the non-CBM group,the age was older (72.06 ± 5.59 years vs.67.01 ±7.15 years; t =4.427; P =0.000),years of education were fewer (3.97 ± 1.381 years vs.6.25 ±2.317 years; t =7.367,P =0.000),systolic blood pressure was higher (155.03 ±20.16 mm Hgvs.147.16 ±17.32 mm Hg; t =2.290,P =0.023),scale of white matter lesion was more higher (7.03 ±2.139 vs.5.47 ±2.591; t =3.247,P =0.001),cerebral infarction volume was larger (6 968.5 ± 1 507.4 mm3 vs.6 177.0 ±1 477.1 mm3; t =2.735,P =0.007),and proportions of hypertension (82.4％ vs.41.5％ ;x2 =18.149,P =0.000),hyperlipidemia (88.2％ vs.39.3 ％ ;x2 =26.067,P =0.000),history of previous stroke or transient ischemic attack (70.6％ vs.28.1％ ;x2 =21.061,P =0.000) and coronary heart disease (94.1％ vs.45.2％ ;x2 =26.278,P=0.000) were higher in the CBM group.The MoCA total score (M[Q1 ～ Q3]; 24 [24 ～25]vs.28 [27 ～ 28] ; Z =-7.092,P =0.000) as well as the scores of attention (6 [5 ～ 6] vs.6 [6 ～ 6] ; Z =-2.502,P =0.012),abstraction (2[1 ～2] vs.2[2 ～2] ; Z =-2.382,P =0.017) and visuoexecutive (2[1 ～2] vs.4[4 ～5]; Z=-7.321,P=0.000) in the CMB group were significantly lower than those in the nonCBM group.The Spearman rank correlation analysis showed that the CMB grade was negatively associated with the MoCA total score (rs =-0.879,P =0.000) as well as the scores of visuoexecutive (rs =-0.895,P =0.000),attention (rs =-0.337,P =0.005),and abstraction (rs =-0.333,P=0.006).Conclusions The age,years of education,systolic blood pressure,degree of white matter damage,and cerebral infarction volume are the risk factors for vascular cognitive impairment.The visuospatial executive dysfunction,attention and abstract thinking decline significantly in ischemic stroke patients with CBMs.CMBs and their numbers are closely associated with cognitive impairment.The more the CMB numbers are,the more obvious the cognitive impairment will be.

OBJECTIVETo explore a safe and effective scheme of analgesic for labor.

METHODSEighty-four primiparas without contraindication of vaginal delivery were divided into an observation group and a control group, 42 cases in each one. In the observation group, the acupoint injection was given at Zusanli (ST 36) combined with electroacupuncture (EA) at Hegu (LI 4) and Sanyinjiao (SP 6) till the cervical opening at the end of first stage labor. In the control group, the routine respiratory instruction was applied. In 5 min, 10 min and 60 min of acupuncture (the same time points in the control group) as well as at the end of the first and second stage labor, the analgesic effect was assessed for the primiparas of the two groups. The labor stages, adverse reactions, postpartum hemorrhage, postpartum urine retention, newborn asphyxia rate and usage rate of oxytocin were compared between the two groups.

RESULTSIn the observation group, in 5 min, 10 min and 60 min of acupuncture as well as at the end of the first and second stages, the visual analogue scale (VAS) was lower apparently as compared with the control group at the corresponding time points (all P < 0.05). The differences in the time limit in the active period, the second and third stages were not significant between the two groups (all P > 0.05). The incidence of adverse reactions and the usage rate of oxytocin were lower than those in the control group [2.4% (1/42) vs 31.0% (13/42); 2.4% (1/42) vs 23.8% (10/42), both P < 0.05]. The differences in postpartum hemorrhage, postpartum urine retention and newborn asphyxia rate were not significant between the two groups (all P > 0.05).

CONCLUSIONThe combination of acupoint injection and EA is the effective analgesic scheme for labor. This scheme effectively alleviates labor pain and has no maternal and child complications.

Acupuncture Analgesia ; Acupuncture Points ; Adult ; Analgesics ; administration & dosage ; Combined Modality Therapy ; Electroacupuncture ; Female ; Humans ; Labor Pain ; drug therapy ; therapy ; Labor, Obstetric ; Pregnancy ; Young Adult

Objective:To explore the effect of autoimmune regulator to TLRs expressions on peripheral antigen presenting cells(APC).Methods:①pEGFPC3-AIRE plasmid was transfected with liposome.②Confocal microscopy was used to observe the effect of transfection.③RT-PCR assay was used to detect the expressions of AIRE and TLR1-9 in RAW264.7cells at 36,48,72,96 h after transfection.Results:①The plasmid was transfected into RAW264.7 cells successfully,and the efficiency of transfection was 60~70%.②AIRE transfected RAW264.7 cells were achieved,and the best time was 72 h.③At 72 h after transfection,the expressions of TLR1,4,5,9 increased,and TLR3,7,8 reduced.The expression of TLR2,6 increased at 96 h.Conclusion:AIRE may regulate the immune response by control TLR expression in APC.It maintain the effective response to pathogen and tolerance state to self tissues through the effects to different TLRs.

Objective To evaluate four tumor markers of insulin-like growth factor 1((IGF-1), CEA, cytokeratin fragment antigen 21-1 (CYFRA21-1), neuron-specific enolase (NSE) for the diagnosis and prediction of treatment response in human lung cancer. Methods Serum samples were taken from three groups: 91 patients with lung cancer, 30 healthy adults and 15 patients with benign pulmonary diseases. Serum IGF-1 was assayed by radioimmunoassay and CEA, CYFRA21-1, and NSE by electrochemiluminescence immunoassay. The differences among the three groups were determined by Kruskal-Wallis one-way analysis of variance (ANOVA) and with Mann-Whitney rank-sum test. Diagnostic efficacy was evaluated by ROC curves. Results The four serum tumor marker levels were significantly higher in lung cancer group, as compared with the benign and the healthy (IGF-1:χ2=26.95,P<0.001, CEA:χ2=49.11,P<0.001; CYFRA21-1:χ2=40.63,P<0.001; NSE:χ2=14.76;P<0.001). The diagnostic sensitivities of IGF-1, CEA, CYFRA21-1 and NSE was 75.6% (34/45), 53.3% (24/45), 66.7% (30/45) and 42.2% (19/45) respectively for lung cancer. The diagnostic sensitivity of IGF-1 combined with CYFRA21-1 was 95.5 %( 43/45) and that of IGF-1 combined with CEA and CYFRA21-1was 97.8%(44/45). Only IGF-1 and CYFRA21-1 showed significant changes before and after treatment (IGF-1: χ2=5.99,P=0.014; CYFRA21-1:χ2=4.99, P=0.025) in cancer group. Conclusions Serum IGF-1, CEA, CYFRA21-1 and NSE are all valuable for lung cancer diagnosis and the combination of those parameters can enhance the diagnostic efficiency. Serum IGF-1 and CYFRA21-1 may also be useful for evaluating the treatment response in lung cancer.

Objective To assess the value of combined detection of enterovirus nucleic acid and antibody in early etiological diagnosis for hand-foot-and-mouth disease ( HFMD).Methods A case-control study was conducted.A total of 1 066 cases of children clinically diagnosed with HFMD from Hangzhou Children′s Hospital were involved into the research group from January to June 2014, consisting of 401 common cases and 665 severe cases; Throat swabs and serum samples from these children underwent combined detection for EV71/CA16/EV of enterovirus nucleic acid by fluorescence quantitative RT-PCR and for EV71/CA16-IgM by ELISA.All data were analyzed with SPSS 16.0.Results The total positive rate of enterovirus nucleic acid EV71/CA16/EV by fluorescence quantitative RT-PCR in the 1 066 cases of children clinically diagnosed with HFMD was 75.52%( 805/1 066 ) ( 95%CI: 72.80%-78.05%).But the total positive rate of combined detection was 91.46%( 975/1 066 ) ( 95%CI:89%.58-93.04%).The total positive rate of combined detection is higher than that of RT-PCR test(χ2 =98.338,P=0.000).The positive rate of EV71 type of combined detection was 64.63%(689/1 066)(95%CI:61.67%-67.49%),which is 15.38%higher than that of RT-PCR test 49.25%(525/1 066)(95%CI:46.21%-52.29%)(χ2 =51.453, P=0.000).In 665 severe cases of HFMD, the total positive rate of combined detection was 96.69%(643/665)(95%CI:94.95%-97.87%), which is higher than that of RT-PCR test 79.25%(527/665)(95%CI:75.92%-82.22%)(χ2 =95.607, P =0.000).In the severe cases, the positive rate of EV71 type of combined detection was 87.52%( 582/665 ) ( 95%CI:84.71%-89.89%) , which is 18.95% higher than that of RT-PCR test 68.57%(456/665) (95%CI:64.87%-72.06%) (χ2 =69.665, P=0.000).In the fatal cases, the positive rate of EV71 type of combined detection was 95.92%(94/98) (95%CI:89.28%-98.68%).Conclusions The combined detection of enterovirus nucleic acid and specific IgM antibody can significantly increase the positive rate of HFMD, especially for severe cases.The combine detection increases both the total positive rate and EV71 positive rate.Thus it has a high potential for becoming a new guidelines for laboratory diagnosis of HFMD.

Objective To establish a technical route for the efficient expression and purification of PprI protein from Deinococcus radiodurans R1 by using eukaryotic Pichia pastoris.Methods The encoding sequence of the Deinococcus radiodurans pprI gene was modified according to the preference of Pichia pastoris' codon.Modified pprI gene was fully synthesized with PCR and a 6 × His tag was added at its Nterminal.The PCR products were purified and then cloned into Pichia pastoris expression vector pHBM-905A.After utilizing Cop I and Not I double enzyme digestion and retrievering linear objective fragment,new pprI gene was transformed to the GS115 strain of Pichia pastoris.The obtained Pichia pastoris transformants were induced to express.Culture supernatants were detected by SDS-PAGE,Western blot,and mass spectrometry.A Ni-NTA column was uesd to purify the target protein and the BCA method was used to determine protein concentration.Results The coding sequence of new synthetic Deinococcus radiodurans pprI gene was correct.The purpose protein band of a molecular weight of 43 000 was detected in the culture supernatant of transformed Pichia pastoris strains by SDS-PAGE and Western blot.The mass spectrometry confirmed that it was the Deinococcus radiodurans PprI protein.When the concentration of imidazole was 250 mmol/L,the elution rate of PprI protein was the highest.The purified protein concentration was 0.35 mg/ml measured by BCA method.Conclusions This study has successfully constructed a new pprI gene and the recombinant strain of Pichia pastoris secreting PprI protein,and established a technical route for the efficient expression and purification of PprI protein.

Objective To investigate the long?term effect of nerve?spring radical hysterectomy( NSRH) on anorectal function after radical hysterectomy. Methods Fifty?six cases of uterine cervical carcinoma patients who met the criteria were selected for the study and were randomly divided into RH group and NSRH group. Defecation functional and anorectal manometry were compared at 1 year after surgery. Results There were 2 patients were excluded both in the two groups, and 26 cases were included in the follow up of each group. Compared with RH group, NSRH group had a lower constipation and chronic diarrhea incidence ( 2 (7. 7%),8(30. 8%);1(3. 8%),6(23. 1%);χ2=4. 457,4. 127P<0. 05),a better self?evaluation bowel func?tion(no significant change:10(38. 5%),5(19. 2%);poor:7(26. 9%),3(11. 5%);very poor:9(34. 6%),18 (69. 2%);χ2=6. 267,P=0. 044;P<0. 05),a higher level of maximal anal squeeze pressure((132. 7±43. 6) mmHg,(119. 5±45. 3) mmHg;t=2. 116,P<0. 05),a lower level of threshold perception of distension((38. 6 ±10. 5) mmHg,(45. 8±12. 4) mmHg;t=2. 326,P<0. 05) and threshold perception of evacuative stimulus ((78. 3±33. 2) mmHg,(90. 6±40. 9) mmHg;t=2. 208,P<0. 05). Conclusion RH may cause more serious long?term anorectal dysfunction,while NSRH help to protect defecation function.