Objective To obtain sufficient recombinant multiepitope peptide of ESAT-6.Methods DNA encoding ESAT-6 peptide was amplified by PCR using a pair of primers which were designed in accordance with the reported ESAT-6 gene sequence.The ESAT-6P gene was inserted into the prokaryotic expression vector pET-30a with the N-terminal 6His-tag.The recombinant peptide vector pET30a-ESAT-6P was transformed into E.coli BL21(DE3) via chemical transformation.The positive clone was screened by means of PCR.The target peptide was expressed in E.coli after induction with IPTG and analyzed by SDS-PAGE.The immunogenicity of the peptide was evaluated by dot immunobinding assay.Results The target peptide was successfully expressed and purified.The solubility analysis showed that the recombinant peptide existed as inclusion body in the E.coli.DIBA indicated that the ESAT-6P was specifically reactive to sera from TB patients.Conclusions The recombinant multiepitope peptide of ESAT-6 engineering bacteria has been obtained,which will be quite helpful to develop novel specific diagnostic reagents and effective vaccine.

This paper presents a new method of separating the junction between the epidermis and the dermis with enzyme,and then,observing the small wound of the papillary of the dermis by means of SEM.The result showed that this new method would be advantageous to the identification of small stab wounds and providing some concrete identify targets.

Objective To compare the changes of serum levels of heme oxygenase‐1(HO‐1) ,C‐reactive protein(CRP) and Chitinase‐3‐1ike protein 1(YKL‐40) in patients with rectal cancer after laparoscopic‐assisted radical operation versus open radical surgery .Methods According the integration and elimination standard ,60 cases of patients with rectal cancer were selected and di‐vided into laparoscopic group (30 cases) and open group (30 cases) ,the concentrations of HO‐1 ,YKL‐40 and CRP in peripheral blood of patients were detected on the first and third postoperative day and preoperative day(P<0 .05) ,and were compared between the two groups .Results The serum levels of HO‐1 ,YKL‐40 and CRP in the laparoscopic group were significantly lower than those in the open group on the first and third postoperative day(P<0 .05) .The serum levels of HO‐1 ,YKL‐40 and CRP were significant‐ly increased after surgery ,and serum levels of HO‐1 and CRP on third postoperative day were significantly increased compared with that on the first postoperative day in the two groups after surgery ,while serum level of YKL‐40 on the third postoperative day was significantly lower than that on the first postoperative day in the two groups after surgery .Conclusion The stress reaction is com‐paratively mild in patients with rectal cancer after laparoscopic assisted radical operation ,and the postoperative period may be shor‐ter in patients treated with laparoscopic assisted radical operation than those treated with open radical surgery .

Objective To analyze the pathogen distribution and drug resistance of candida isolated from children with blood infections in our hospital,and to provide reference for clinical effective prevention and treatment.Methods The blood specimens of pediatric patients were collected between January 2009 and December 2015,and were cultured using BacT/ALERT 3D and BD9140 instruments.The candida were separated with Sobaurandps agar culture medium,and identified with chromogenic medium,API 20CAUX test strips or VITEK-2 compact YST card.The minimal inhibitory concentration of 5 drugs were determined by ATB FUNGUS 3 system.Results In 176 cases,92 strains (52.3％) were from neonatal ward,and 46 strains (26.1％) were from PICU.In newborn group,85 strains were isolated from premature,which contained the low and very low birth weight infants (37 strains),pneumonia(20 strains),neonatal respiratory distress syndrome(9 strains).In PICU,the strains were commonly isolated from children with severe infection.Among 176 strains of candida,71 strains (40.3％) were C.albicans,62 strains (35.2％) were C.parapsilosis,16 strains(9.1％) were C.glabrata,9 strains(5.1％) were C.tropicalis,and 18 strains(10.2％) belonged to other candida.Conclusion Candida blood infections can happen at all age of chlidren.The most common strains detected from blood were C.albicans,followed by C.parapsilosis.Most of these strains are susceptible to antifungal drugs,such as fluconazole,except C.glabrata.The sensitive rates to commonly used antifungal drug are more than 93％.The selection of antifungal drugs should be based on the species of strains.

Objective To investigate the changes of the electrocardiograms (ECG) and the cardiac markers in patients with acute insular infarction,and analyze the relationship between them and the prognosis.Methods A total of 202 patients with acute middle cerebral artery territory infarction (patients group) and 150 control subjects (control group) was selected in this study.Patients included insular infarction (insular infarction group,136 cases),non-insular infarction (non-insular infarction group,66 cases),left-side insular infarction(71 cases) and right-side insular infarction(65 cases).ECG recordings and plasma cardiac troponin I (cTnI),creatine kinase-MB (CK-MB) were measured and compared.Death in 6 months was followed-up.Results There was significant difference in the incidence of abnormal changes of ECG and plasma cTnI,CK-MB increasing between patients group and control group (P ＜0.01).The incidence of abnormal changes of ECG and fatality rate were higher in insular infarction group than those in non-insular infarction group [80.88％(110/136) vs.46.97％(31/66) and 11.76％ (16/136) vs.3.03％ (2/66),P ＜ 0.05 or ＜ 0.01].The incidences of ectopy and prolonged QT were higher in right-side insular infarction patients than those in left-side insular infarction patients [44.62％(29/65) vs.11.27％ (8/71),P ＜0.01 ; 55.38％ (36/65) vs.35.21％ (25/71),P ＜ 0.05].The incidences of sinus bradycardia and ST segment deviation were higher in left-side insular infarction patients than those in right-side insular infarction patients [22.54％(16/71) vs.7.69％(5/65),P ＜ 0.05 ;47.89％(34/71) vs.13.85％ (9/65),P ＜ 0.05].The increased rates plasma cTnI and CK-MB level were mainly seen in insular infarction [insular infarction group:47.79％ (65/136),34.56％ (47/136); non-insular infarction group:4.55％ (3/66),1.52％ (1/66),P ＜ 0.01].The incidence of plasma cTnI increasing in right-side insular infarction patients was higher than that in left-side insular infarction patients [67.69％(44/65) vs.29.58％(21/71),P＜ 0.05].There was no significant difference in the incidence of plasma CK-MB increasing between left-side insular infarction patients and right-side insular infarction patients(P ＞ 0.05).The fatality rates in plasma cTnI,CK-MB increasing patients were higher than those in normal plasma cTnI,CK-MB patients [16.18％ (11/68) vs.5.22％ (7/134),P ＜0.05;29.17％ (14/48) vs.2.60％ (4/154),P ＜0.01].Conclusions The effects of acute hemispheric cerebral infarction on heart are mainly associated with destruction of insula.Patients with insular infarction have more abnormal changes of cardiac markers and ECG,which is correlated with poor prognosis.

Objective To investigate the role of p38 mitogan-activated protein kinase signal transduction pathway in lipopolysaccha-ricle-indaced acute lung injury (ALI) in rats. Methods 48 wistar rats were randomly divided into 3 groups, saline control group (group A), lipopolysaccharide (LPS) group(group B) and SB203580 group(group C). Models of lipepelysaccharide-imluced ALI were used to oh-serve the expression of p38MAPK in rat lung, protein content, the ratio of neutrophiles in bronchoalveolar lavage fluid (BALF) , pulmonary MDA content and the activities of serum NO. After LPS dripping for 6 hours, arterial blood was drawn for analysis and lung tissue was detec-ted. Results Compared with those in group A, expression of p38MAPK were markedly increased in group B and C (P<0.01). But in group C, expression of P38MAPK was significantly lower than that in group B (P<0.05). The protein content, the ratio of neutrophiles in bronchoalveolar lavage fluid (BALF), content of pulmonary MDA and the activities of serum NO in group B, C were significantly higher than those in group A (P<0.01). There was a significant decrease in the level of arterial bicarbonate and partial pressure of oxygen in group B and C (P<0.01). Compared with those in group B, these indexes of lung injury were significantly lower while the level of arterial bicar-bonate and partial pressure of oxygen was increased in group C(P <0. 05 orP <0. 01). Under light microscope, the pathologic changes in-duced by LPS were significantly attenuated by SB2035g0. Conclusion The activation of P38MAPK play an important role in the mechanism of lipopolysaccharide-induced ALI.

Objective To investigate the effect of radix paeoniae rubra (RPR) on expressions of p38MAPK, NF-κB and iNOS in ltpopolysaccharide (LPS)-induced acute lung injury (ALI) and explore its protective mechanism. Methods Forty Wistar rats with ALI were divided randomly into five groups: saline control group (Group A) , LPS group (Group B), RPR for treatment group (Group C), RPR for prevention group (Group D) and SB203580 group (Group E). The effects of RPR on protein content and the ratio of neutrophiles in bronchoalveolar lavage fluid (BALF), contents of malondidehyde (MDA) and serum NO in lung tissue were observed. Arterial blood was collected for blood gas analysis. Expressions of p38MAPK, NF-κB and iNOS in rat lung tissues were detected. Results Compared with Group A, expressions of p38MAPK, NF-κB and iNOS were significantly increased (P <0.01 orP< 0.05) , the protein content and the ratio of neutrophiles in BALF, contents of MDA and serum NO were significantly higher (P<0.01 or P<0.05) in Groups B, C, D and E. There was a significant decrease in the level of arterial bicarbonate and partial pressure of oxygen in Groups B, C, D and E (P<0.01 or P<0.05). Compared with Group B, the expressions of p38MAPK, NF-κB and iNOS were significantly lower, and the protein content and the ratio of neutrophiles in BALF, contents of MDA and serum NO were significantly decreased, while the levels of arterial bicarbonate and partial pressure of oxygen were significantly higher in Groups C, D and E (P<0.05 or P<0.01). Compared with Group B, the lunginjury of Groups C, D and E was significantly alleviated, while there was no statistical difference among Groups C, D and E in the indices of lung injury. Conclusion Protective effect of RPR on LPS-induced ALI is closely related to the inhibited expressions of p38MAPK, NF-κB and iNOS.

BACKGROUND: Reperfusion injury and inadequate perfusion exist,causing the injury of intestinal mucosa during shock / resuscitation.OBJECTIVE: To investigate protective effect of shenfu injection (SFI) in equal dosage for human effect on intestinal mucosal pH (PHi) of sigmoid colon, the contents of nitric oxide (NO), malonialdehyde(MDA) and Ca2+ of intestinal mucosa, activity of serum diamine oxidase(DAO of rabbits during shock/resuscitation.DESIGN:A randomized controlled trial with experimental animals as the subjects.SETTING: DepartmentofAnesthesiology, Renmin Hospital of Wuhan University MATERIALS: The experiment was performed in the Research Institute of Anesthesiology, Renmin Hospital of Wuhan University from August to October in 2003. Twenty-four adult healthy white rabbits were randomly divided into 3 groups with 8 animals for each: Shenfu injection group (SFI group), shock/resuscitation group and control group .INTERVENTIONS: Hemorrhagic shock was induced by blood withdrawal from femoral artery at a rate of 2 mL/kg per minute until MAP dropped to 40 mmHg. MAP was maintained at this level for 60 minutes,then the collected auto-blood and the same volume of balanced salt solution was transfused to the animal for preparing shock/resuscitation model. In SFI group: the animals were given 2.1 mL/kg SFI together with the auto-blood transfusion and followed by a continuous infusion of 5mL/kg SFI per hour.MAIN OUTCOME MEASURES: Intestinal mucosal pH (PHi) of sigmoid colon, NO, MDA and Ca2+ contents of intestinal mucosa, activity of serum DAO were detected before shock (So), 1 hour after shock (S1), 1 hour (R1) and 3 hour (R3) following resuscitation.the R1 reperfusion (7.171±0.102) , and R3 reperfusion (7.194±0.106) of the SFI group were higher than those of the shock/ resuscitation group (6.920±0.155,6.971±0.165,P ＜ 0.05,P ＜ 0.01) and those of the contents of the reperfusion R1 (35.090±1.184) μkat/L and R3 (32.440±2.884) μkat/L of the SFI groups were significantly higher than those of the shock/resuscitation group [(50.994±2.684),(52.377±1.217) μkat/L,P＜ 0.01] and of the control group [(15.970±1.734), (16.620±0.767)μkat/L,Pgroup were significantly lower than those in the shock/resuscitation group [ (61.8±5.3,72.2±5.8 ) μmol/g , (68.2±4.9,96.9±8.5) μmol/L ,P＜ 0.05]. Ca2+ content of intestinal mucosa at reperfusion 3 hours in the SFI group [(2.43±0.27)μmol/L] was lower than that in the shock/resuscitation group [(2.93±0.34)μmol/L,P ＜ 0.05] and higher than that in the control group [(2.26±0.31 )μmol/L, (P ＜ 0.05)].CONCLUSION: SFI in equal dosage for human effect could protect intestinal mucosa from shock/resuscitation injury through improving perfusion and oxygenation, inhibit the activity of NO, reduce oxygen free radical and calcium overload, with a good protective effect on intestinal mucosa during shock/resuscitation.

Objective To evaluate the effects of ischemic postconditioning on brain injury induced by myocardial ischemia-reperfusion (I/R) in diabetic rats.Methods Diabetes mellitus was induced by intraperitoneal streptozotocin 60 mg/kg and confirmed by blood glucose level ＞ 16.7 mmol/L.Thirty male Sprague-Dawley rats,weighing 220-280 g,in which diabetes mellitus was successfully induced,were randomly allocated into 3 groups (n =10 each) using a random number table:group sham operation (group S),group I/R and group ischemic postconditioning (group P).Myocardial I/R was induced by occlusion of the anterior descending branch of the left coronary artery in I/R and P groups.Group P received 3 cycles of 10 s reperfusion followed by 10 s ischemia at the end of myocardial ischemia.The rats were sacrificed at 120 min of reperfusion and the brains were removed for microscopic examination and for determination of cell apoptosis (by TUNEL) and expression of interleukin-6 (IL-6),IL-8,IL-10,glycogen synthase kinase-3 beta (GSK-3β) and phosphorylated GSK-3β (pGSK-3β) (by immuno-histochemistry).Apoptotic index was calculated.Results Compared with group S,apoptotic index was significantly increased,IL-6 and IL-8 expression was up-regulated,and IL-10 and pGSK-3β expression was downregulated in I/R and P groups (P ＜ 0.01).Compared with group I/R,apoptotic index was significantly decreased,IL-6 and IL-8 expression was down-regulated,and IL-10 and pGSK-3β expression was up-regulated in group P (P＜0.01).There was no significant difference in GSK-3β expression among the 3 groups (P ＞ 0.05).The pathologic changes were significantly attenuated in group P as compared with group I/R.Conclusion Ischemic postconditioning can attenuate brain injury induced by myocardial I/R in diabetic rats,and inhition of activity of GSK-3β may be involved in the mechanism.