Objective To study the damage of hyperhomocysteinemia (HHcy) and asymmetric dimethylarginine (ADMA) on the structure of rats cerebral vessels,and to explore the treatment effect of probucol,folic acid and vitamin B12.Methods Totally,70 male Sprague-Dawley(SD) rats were randomly divided into seven groups:normal control (A),HHcy model (B),low-dose-probucol (C),middle-dose-probucol (D),high-dose-probucol (E),folic-acid plus vitamin B12 (F),folic-acid plus vitamin B12 plus probucol(G)group.Group A was fed plain feedstuff and water.Other groups were fed homomethionin feedstuff and water,probucol (340,510,680 mg· kg-1 · d-1),folic acid and vitamin B12,folic acid plus vitamin B12 plus probucol respectively.The plasma concentrations of homocysteine(Hcy) and ADMA were assayed by high-performance liquid chromatography method (HPLC) at the beginning,and at the end of 8 and 12 weeks.The analysis of rats cerebral vessels was performed with electron microscope at the end of 12 weeks.Results (1) After adding high concentrations of methionine feedstuff for 8 weeks,the levels of Hcy in group B (60.4±11.9) μmol/L were significantly raised compared with group A(7.1±1.4) μmol/L.Meanwhile,ADMA levels in group B(1.3±0.4)μmol/L were higher than group A(0.4±0.1) μmol/L.At the end of 12 week,the levels of Hcy and ADMA were lower in groups C[(49.9±11.4)μmol/L,(1.0±0.2)μmol/L],D [(46.8± 10.9)μmol/L,(0.9±0.1)μmol/L],E[(44.3±11.4)μmol/L,(0.9±0.1)μmol/L],F [(16.3 ± 4.9)μmol/L,(0.7 ± 0.2)μmol/L] compared with group B [(69.5 ± 11.4)μmol/L,(1.6 ±0.5)μmol/L],but higher than group A[(6.7±1.3)μmol/L,(0.5±0.1)μmol/L].The levels of Hcy and ADMA in group G[(8.9±1.9) μmol/L,(0.6±0.2) μmol/L]were also lower than those in group B (P＜0.05],but they did not show statistically significant differences compared with group A(P＞0.05).(2) According to correlative analysis,Hcy levels were positively correlated with ADMA levels (P＜0.05).(3) On electron microscope,we observed that the cerebral artery and cerebral cortex of group B were seriously damaged.After the intervention with probucol,folic acid and vitamin B12,the structure of rats cerebral vessels were improved in group C,D,E,F and G as compared with group B,and group G were especially improved more obviously.Conclusions The HHcy and ADMA could damage the structure of rats cerebral vessels.Probucol,folic acid and vitamin B12 may alleviate the cerebral vascular damage by decreasing the levels of Hcy and ADMA,and the effect is more significant by the combined drug regimens of Probucol,folic acid and vitamin B12.

Objective To investigate the distribution of hepatitis B virus(HBV)genotypes in Zhejiang.Methods HBV genotyps were detected by fluorimetric real-time PCR in 240 HBVDNA positive patients who were born in Zhejiang(Hangzhou,Huzhou,Jinhua,Shaoxing,Taizhou,Ningbo each 40 positive patients).Results of the 240 HNBDNA positive patients,82(34.17%)were genotype B,and 140(58.33%)were genotype C,15(6.25%)were genotype B and C,3(1.25%)were genotype D.No genotype A、E and F found in the studied subjects.Conclusions HBV genotype B,genotype C,genotype B and C,genotype D existed in Zhejiang and there is no difference in ferms of the distribution of genotypes in six areas metioned above.

BACKGROUND:The chemical compositions and structure of calcium phosphate bone cement are similar to those of human bone, which can fil the bone colapse caused by fracture and induce osteogenesis, but its degradation rate is slow. OBJECTIVE:To evaluate the biocompatibility of the calcium phosphate cement/fibrin glue and the feasibility of repairing radius defects. METHODS: In vitro cytotoxicity experiment: Mouse fibroblasts were cultured in the calcium phosphate bone cement/fibrin glue extracts, phenol solution, and RPMI 1640 culture medium containing 10% fetal bovine serum, respectively, to detect the cytotoxicity grade. Hemocompatibility experiment: Calcium phosphate bone cement/fibrin glue extracts, normal saline and distiled water were respectively added into the rabbit anticoagulation, to detect the hemolytic rate. Forty-five New Zealand white rabbits were enroled and modeled into bilateral radius defects, folowed by randomly alotted into three groups: blank control group without any intervention, experimental and control groups were given the implantation with calcium phosphate bone cement/fibrin glue and autologous radius, respectively. X-ray, histology, bone mineral density and biomechanical test were performed at postoperative 4, 8 and 16 weeks. RESULTS AND CONCLUSION:The toxicity grade of the calcium phosphate cement/fibrin glue was 0 to 1. The hemolytic rate of the calcium phosphate cement/fibrin glue was 3.15%. At 16 weeks postoperatively, X-ray showed that in the experimental and control groups, the fracture line disappeared completely, pulp cavity was recanalized, and in plastic completely. Histology showed that the reconstructed bone trabecular was obvious, plate layer of bone was mature, and medulary cavity recanalization appeared in the control group; there were a large number of new grid-shaped woven bone tissues growing into the material in the experimental group, with overt degradation, and degradation rate was in paralel to bone ingrowth. The bone density, the maximum load, maximum stress and failure energy in the experimental and control groups were significantly higher than those in the blank control group (P < 0.05), and al above indicators showed no significant differences between the experimental and control groups. These results manifest that the calcium phosphate bone cement/fiber protein glue composite material holding a good biocompatibility can promote bone tissue regeneration for bone defect repair, achieving similar curative effect with autologous bone transplantation.

Objective To explore the differentiation of allogeneic naive T cells to regulatory T cells (Tregs) and T helper (TH) 1/2/17 cells by coculture with bone marrow-derivedimmature dendritic cells (irnDC).Method Bone marrow-derived imDC were cultivated from Balb/c mice.Lipopolysaccharide-stimulated DC were harvested as mature dendritic cells (mDC) and unstimulated cells were collected as imDC.Then irnDC or mDC were cocultured with allogeniec naive T cells,respectively.TH1 cytokines [interferon-γ (IFN-γ) and interleukin (IL)-2],TH2 cytokines (IL-4 and IL-10),and TH17 cytokine (IL-17) of co-cultured cells were detected by enzyme linked immunospot assay.CD4+ Forkhead box p3 (FoxP3) + Treg proportion in CD4+ cells in the co-cultured system with IL-2 and transforming growth factor-β1 (TGF-β1) was analyzed by flow cytometry.Result As compared with mDC,na(i)ve T cells cocultured with imDC secreted much less IFN-γ (11.67 ± 2.18 vs.182.00±23.71,P＜0.01),IL-2 (26.67±2.96 vs.318.30± 18.62,P＜0.01),IL-4 (17.00±3.78 vs.45.33±3.48,P＜0.01),IL-10(7.00±1.00vs.158.70±10.90,P＜0.001) and IL-17 (0.66 ± 0.33 vs.238.30 ± 24.39,P＜0.001).Furthermore,imDC induced more CD4+ FoxP3+ Tregs than mDC after adding IL-2 and TGF-β1 in the coculture system for 7 days (22.70 ± 1.53 ％ vs.5.42 ± 1.27％,P＜0.01).Conclusion imDC are more effective to induce na ve T cells to Tregs,but not differentiate to TH 1/TH 2/TH 17 cells.These findings provide in vitro experimental evidence for induction of transplant tolerance by adoptive transfer of imDC.

Objective To investigate if the administration of CORM-2 can provide protection against renal ischemia-reperfusion injury (IRI).Method Murine renal ischemia was induced by clamping left renal pedicles for 40 min with vascular micro damps at 32 C,then the contralateral kidney was removed.CORM-2 or vehicle was administered via intravenous infusion 1 h before the onset of ischemia.The blood plasma and renal samples were obtained at 24 h after reperfusion to assess renal function and cellular injury.Plasma Cr and BUN levels,HE and TUNEL were performed to estimate the magnitude of renal damage.Kidneys were retrieved from indicated animals at various time points after renal IRI,and the sections were prepared for histological evaluation.MPO staining procedures were performed to assess the neutrophils infiltration in the renal IRI.Besides,Immunofluorescent stain of TNF-α was performed on the kidneys which were retrieved from indicated animals to determine the production of inflammatory mediators in renal I/R.Results The plasma Cr and BUN were significantly increased at 24 h after reperfusion in IRI control mice,and CORM-2 treatment could markedly diminish the increase of plasma Cr and BUN in mice subjected to I/R.In parallel,histological analysis demonstrated that CORM2 treatment markedly reduced apoptosis of the renal tubular epithelium cells and hemorrhage.IRI caused marked infiltration and accumulation of the MPO-positive neutrophils in renal interstitium.Administration of CORM-2 before ischemia dramatically inhibited neutrophils infiltration as compared with IRI or iCORM-2 group.Furthermore,we confirmed that CORM-2 markedly decreased production of TNF-α.Conclusion Carbon monoxidereleasing molecule CORM-2 could ameliorate inflammation to protect against the renal IRI in mice.

Objective To investigate whether rat adipose-derived stem cells (rASCs) could resist human xenoantibody-dependent complement-mediated lysis and to explore its possible mechanisms.Method SD rat ASCs were isolated,rASCs at passage 2 to 8 were used for the following studies and rat lymphocytes were harvested as control cells.α-Gal expression was detected by flow cytometry.After incubation of rASCs with 20％ normal human serum (NHS) or heat inactivated normal human serum (HINHS),flow cytometry was used to detect cytotoxicity,IgG or IgM binding,and C3c,C4c and C5b-9 deposition.Result We successfully established the method to isolate and culture rASCs.The morphology of rASCs remained unchanged after passages.rASCs were positive for tell surface markers of CD44 and CD90,while negative for CD45 and MHC-Ⅱ.As compared with rLCs,rASCs significantly resisted human natural antibody and complement-mediated lysis when incubated with 20％ NHS in vitro (20.42％ ± 2.80％ vs 51.84％ ± 6.70％,P ＜ 0.01).Mechanistically,rASCs expressed lower level of α-Gal (13.97 ± 0.33 vs.24.47 ± 3.03,P＜0.05),which was correlated with decreased binding of human xenoreactive IgG and IgM (IgM:9.4 ± 2.0 vs.107.2± 4.8,P＜0.01; IgG:5.73 ± 1.0 vs.27.49 ± 3.9,P＜0.01) and reduced deposition of complements C3c,C4c and C5b-9 (C3c:294.6 ± 38.02 vs.1924 ± 509.4,P＜0.05; C4c:35.23 ± 3.1vs.177.3 ± 37.17,P＜0.05; C5b-9:5.63 ± 1.74 vs.37.05 ± 7.4,P＜0.01).Conclusion These data demonstrated that the resistance of rASCs to human xenoantibody and complement-mediated lysis is associated with low expression of xenoantigen a-Gal and inhibition of MAC (membrane attack complex) formation.

Objective To examine the effect of rapid infusion test guided by extravascular lung water index (EVLWI) on hemodynamics in critically ill patients at different states in order to guide volume resuscitation.Methods A prospective observation was conducted.Forty critically ill patients admitted to Department of Critical Care Medicine of Tianjin Third Central Hospital from June 2012 to April 2014 were enrolled.Based on the levels of EVLWI and pulmonary vascular permeability index (PVPI) and the cardiac function,the patients were divided into four groups:septic patients with normal EVLWI and PVPI (n =17),septic patients with increased EVLWI and PVPI (n =3),septic patients with increased EVLWI and normal PVPI (n =4),and coronary heart disease and heart failure patients with normal EVLWI and PVPI (n =16).The rapid infusion test was conducted in all patients using lactated Ringer solution 250 mL,followed by infusion of crystalloid with rate of 150 mL/h.The conditions of mechanical ventilation and vasoactive drugs were not changed during study.The changes in EVLWI,intrathoracic blood volume index (ITBVI),and cardiac index (CI) before capacity load,at immediate capacity load,and 15,45,105 minutes after load were determined by pulse indicator continuous cardiac output (PiCCO).On the base of volume status before and after the liquid infusion,the standard for the changes were:stroke volume (SV) increased by 12％-15％,central venous pressure (CVP) greater ≥ 2 mmHg (1 mmHg =0.133 kPa),CI ＞ 15％,and ITBVI change greater than 10％.Results There were no statistically significant differences in the observed indicators at the each time point before and after rapid infusion test among the four groups (all P ＞ 0.05).In septic patients with normal EVLWI and PVPI group,ITBVI was slightly increased by 5.4％-9.7％ from 15 minutes to 45 minutes after rapid infusion test.In coronary heart disease and heart failure patients with normal EVLWI and PVPI group,the EVLWI was increased by 11.9％,5.9％,and 14.7％ respectirely at 15,45,and 105 minutes,ITBVI was slightly increased by 6.4％ at 45 minutes,CI was increased by 29.5％ immediately after rapid infusion.In septic patients with increased EVLWI and PVPI group,CVP was increased by 8 mmHg immediately,EVLWI was increased significantly by 15.8％ at 45 minutes,ITBVI was slightly decreased by 10.0％ at 45 minutes,CI was increased by 24.7％ immediately,and increased by 17.0％ at 105 minutes,and PVPI was increased by 15.6％-28.1％ at 15-105 minutes after rapid infusion.In septic patients with increased EVLWI and normal PVPI group,CVP was increased by 1.5 mmHg at 15 minutes,EVLWI was increased immediately,which was increased by 17.4％,24.0％,and 31.4％ respectively at 15,45,and 105 minutes,ITBVI was increased by 13.9％ at 15 minutes,CI was increased by 16.1％ at 15 minutes after rapid fluid infusion.Conclusions Rapid fluid replacement in critically ill patients with crystalloid,regardless of whether the EVLWI was normal or increased,the short-term response was affected by the volume and cardiac function of patients.Different status of patients showed different volume effect curve:no significant changes in hemodynamic parameters were found in patients with normal EVLWI and volume parameters.In patients with potential cardiac dysfunction,CI and EVLWI increased significantly; regardless of PVPI increased or normal,EVLWI and CI were increased in patients with elevated EVLWI; two different changes could be found in the two types of pulmonary edema while ITBVI was increased.

Objective To compare selective coverage and chimney technology of the left subclavian artery (LSA) in thoracic endovascular aortic repair (TEVAR).Methods Eighty-one cases were retrospectively reviewed from January 2005 to January 2013 with two different management of the LSA during TEVAR:selective coverage (SC) vs chimney (C).Technical success rates,operating time,endoleaks,and perioperative cerebrovascular accidents,spinal cord ischemia (SCI),left arm ischemic symptoms and mortality were analyzed.The survival rate was also evaluated.Statistical analysis was performed using the x2 test,t-test and Kaplan-Meier survival curve.Results There were 37 patients in group SC and 34 in group C.Operating time was (61 ± 22) min for SC,and (101 ± 20) min for C (P =0.000) ; left arm ischemic symptoms occurred in 23.4％ in group SC,and 2.9％ in group C (P =0.011).Technical success rates were 100％ and SCI were 0 for both SC and C.Endoleaks,perioperative cerebrovascular accidents,and mortality were similar between the two groups.Conclusions During TEVAR,selective coverage of the left subclavian artery is safe,effective; meanwhile,chimney technology is safe,effective for patients whose left subclavian arteries need revascularization.

Objective To investigate the correlation of T-lymphocyte expressing HLA-DR with serum HBV DNA and HBeAg contents in chronic hepatitis B. Methods Totally 134 chronic hepatitis B patients and 36 healthy blood donors were enrolled in the study. The T-lymphocytes (CD3 + HLA-DR + ,CD4 + HLA-DR+ and CD8 + HLA-DR+ T) expressing HLA-DR were detected by flow cytometry, the serum HBV viral loads were detected by the real-time quantitative PCR and HBeAg was detected by chemiluminescence method. According to serum HBV DNA viral loads patients were defined as HBV DNA negative (≤ 103 copies/mL), low (＞ 103 - 105 copies/mL), medium (＞ 105 - 107 copies/mL) and high groups (＞ 107 - 109 copies/mL) ; according to serum HBeAg levels, patients were defined as HBeAg negative (≤1 PEIU/mL), low (＞ 1 - 100 PEIU/mL), medium (＞ 100-1 000 PEIU/mL) and high groups (＞ 1 000-10 000 PEIU/mL). T test and one-way ANOVA were performed. Results With HBV DNA loads, HBeAg levels increased, the percentage of CD3 + HLA-DR + , CD4 + HLA-DR + and CD8 + HLA-DR +decreased, especially CD8 + HLA-DR +. Compared with HBV DNA negative group, the percentages of CD3 +HLA-DR + , CD4 + HLA-DR + and CD8 + HLA-DR + were significantly reduced in high group (t = 3. 686,4. 592 and 3. 216, P ＜ 0. 0l); the percentages of CD4 + HLA-DR + and CD8 + HLA-DR + were also reduced in medium group (t = 3. 761 and 2.862, P ＜ 0.01); while in low group, only the percentage of CD8 + HLA-DR + was reduced (t = 2.215, P ＜ 0.05). Compared with HBeAg negative group, the percentages of CD3 +HLA-DR+, CD4 + HLA-DR+ and CD8 + HLA-DR+ were significantly reduced in medium and high groups (thigher =3. 144, 2.222 and 4.035; tmiddle =3.311, 2.362 and 3.374, P ＜0.05), while in the low group,only the percentage of CD8+HLA-DR+ was reduced (t=2.029, P＜0. 05). Conclusion The combined measurement of HBV DNA, HBeAg and T-lymphocytes expressing HLA-DR in chronic hepatitis B patients may not only help to evaluate the immune status of patients, but also can predict the disease progression and clinical outcomes.

Objective: To investigate the clinical value of GeneXpert in the rapid diagnosis of rifampicin resistance in tuberculosis and extrapulmonary tuberculosis. Methods: From June 2018 to March 2019, a total of 122 tuberculosis patients admitted to the People's Hospital of Yueqing were selected.Among them, 109 patients with pulmonary tuberculosis and 13 patients with extrapulmonary tuberculosis.GeneXpert MTB/RIF, smear, LJ solid medium, BACTEC MGIT 960 liquid medium four detection methods were used to detect tuberculosis secretion.GeneXpert MTB/RIF, LJ solid medium, BACTEC MGIT 960 liquid medium three detection methods were used to detect extrapulmonary tuberculosis secretion.The sensitivity and specificity of GeneXpert MTB/RIF, smear, LJ solid medium, BACTEC MGIT 960 liquid medium for tuberculosis specimens were detected and compared, and the monitoring results of rifampicin resistance in extracorporeal tuberculosis specimens in culture medium detected by GeneXpert MTB/RIF, LJ solid medium, BACTEC MGIT 960 liquid were observed. Results: The GeneXpert MTB/RIF, smear, LJ solid medium, BACTEC MGIT 960 liquid medium were used to detect 109 cases of tuberculosis, and the detection rates were 94.50%(103/109), 55.04%(60/109), 57.79% (63/109), 61.47%(67/109), respectively.The detection rates of extrapulmonary tuberculosis were 7.69%(1/13), 0.00%(0/13) and 15.38%(2/13), respectively.The sensitivity and specificity of the four methods for the diagnosis of tuberculosis had statistically significant differences(χ²=121.540, 127.610, all P<0.05). The diagnostic efficiency of GeneXpert MTB/RIF was significantly better than smear, LJ solid medium, BACTEC MGIT 960 liquid medium.The results of stratification analysis of rifampicin resistance using GeneXpert MTB/RIF, LJ solid medium and BACTEC MGIT 960 liquid medium showed that there were no statistically significant differences among the three methods and gold standard(χ²=0.750, 0.942, 0.947, all P>0.05). Conclusion GeneXpert technology has the advantages of simple operation, rapid detection and high detection rate.It has high clinical value for the rapid diagnosis of tuberculosis and extrapulmonary specimens and the detection of rifampicin resistance.