Objective To explore the cultural method of lingual keratinocytes and make preparations for further investigation in using lingual keratinocytes as a new choice of seed cells for the reconstruction of tissue engineered corneal epithelium.Methods Keratinocytes were enzymatically dissociated from lingual tissue specimens using 0.25%disepase and 0.05%trypsin.The biological characteristics of the cells were observed through phase microscope.Meanwhile.1ingual kemtinoeytes were immtmohistochemically labeled with broad spectrum keratin antibody (AE1/AF3).Results The cells revealed the form of "road metal" and could be passed for 3 or 4 generations.Immunohistochemistry indicated that lingual keratinocytes were positive for AE1/AE3 staining.Condusion Lingual kemtinocytes of rabbits can be cultured with KSFM in vitm and magnitude quantity can be attained,laying a favorable foundation for lingual keratinocytes as a new choice of seed cells for corneal epithelium reconstruction with tissue engineering.

BACKGROUND:Currently, bone graft is mainly used for repair of bone defects, and tricalcium phosphate is the most used artificial bone material. But the effectiveness of the tricalcium phosphate bone graft is stil controversial, and there is also no detailed report about its function during the healing of bone defect. ＜br> OBJECTIVE:To observe the concentration changes of bone morphogenetic protein-2 and vascular endothelial growth factor as wel as bone healing after tricalcium phosphate graft in bone defects. ＜br> METHODS:Forty-eight C57 mice were randomly divided to experimental group and control group. A 2-mm-long diaphyseal segment and periosteum from the middle of the right femur was cut to prepare unilateral bone defect models. Tricalcium phosphate bone graft was used in the experimental group, and no bone graft in the control group. During the fol owing 4 weeks, X-ray examination was done once a week to observe the bone healing, and then the animals were executed for col ecting samples in the graft area. The concentrations of bone morphogenetic protein-2 and vascular endothelial growth factor in samples which were taken from the bone graft area were determined by using ELISA assay. ＜br> RESULTS AND CONCLUSION:X-ray films showed that 2 weeks later, bone fracture healed mostly in the experimental group except a smal part of cortical bone;3 weeks later, bone fracture was basical y healed, and only a smal amount of tricalcium phosphate remained;4 weeks later, bone fracture was completely healed, and the cal us grew obviously, and the tricalcium phosphate was nearly absorbed. In the control group, the fracture line was stil visible at 1-2 weeks, but it became vague at 3 weeks;then, the fracture was healed at 4 weeks except some of the cortical bone. The levels of bone morphogenetic protein-2 and vascular endothelial growth factor were significantly higher in the experimental group than in the control group at different time points (P＜0.05). These results suggest that tricalcium phosphate bone graft can up-regulate the expression of bone morphogenetic protein-2 and vascular endothelial growth factor and accelerate bone healing.

Objective To study the possible influencing factors in the formation of coronary collateral circulation in patients with chronic total occlusion (CTO). Methods Patients were enrolled having at least 1 major coronary artery angiography revealed as CTO of 144 patients. According to the Rentrop classification, patients with grade 0 and grade 1 filling were catogorized as insufficient collateral circulation group (n=72) and patients with grade 2 and grade 3 filling as collateral circulation group (n=72). Serum biomarkers and insulin-resistance by HOMA model were also studied in all patients. Results In the insufficient collateral circulation, BMI,TC,ApoB, lipoprotein a, fasting insulin HOMA-IR,HOMA- beta, CRP was significantly higher than the well collateral circulation group and ApoA-Ⅰ, ISI lower than the well collateral group ( all P ﹤0. 05 ) . Bivariate correlation alaysis showed. Rentrop score, BMI, TC, ApoB, lipoprotein a, fasting insulin, HOMA-IR,HOMA- beta and CRP are positively correlated to the formation of collateral circulation ( P ﹤ 0. 05 ); ApoA-Ⅰ and ISI were negatively correlated ( P ﹤0. 05 ) . Logistic regression analysis after calibration with weight, ApoA-Ⅰ and HOMA-beta factors, lipoprotein a ( OR 7. 575,P=0. 009), TC (OR 2. 154,P =0. 001) were found to be the independent factors of coronary collateral circulation formation. Conclusions TC, lipoprotein a, obesity, CRP, and HOMA-IR are correlated with the formation of coronary collateral circulation and may predict formation of collateral circulation in patients with CTO.

OBJECTIVETo analyze the related factors of the infection after operation of open tibiofibula fractures.

METHODSThe clinical data of 141 patients with open tibiofibula fractures underwent surgical treatment from June 2009 to December 2012 were retrospectively analyzed. All the patients were male and aged from 18 to 61 years old with an average of 39.2 years; all the fractures were unilateral. According to Gustilo typing of fracure, 5 cases were type I, 44 cases were type II, 27 cases were type III a, 56 cases were type III b and 9 cases were type III c. These clinical data included patients' age, gender, body mass index (BMI), underlying diseases, time of operation, fracture site, fracture type, fixation method, postoperative drainage, debridement condition and so on. The postoperative infection conditions were recorded, and the correlation between above factors and infections were analyzed. Finally, the significant related variables were introduced into a Logistic regression model to evaluated their risk.

RESULTSAmong the 141 patients, 22 cases developed with infection (15.6%). The significant relative factors with infection contained fixation method, debridement condition, underlying diseases and postoperative drainage (P < 0.05). The correlativities were stepped up in order, their odds ratios value was 2.451, 3.164, 3.414, 5.117, respectively.

CONCLUSIONActive treatment for underlying diseases before operation, thorough debridement for open wound, suitable fixation method for fracture and thorough draining are effective measures in preventing postoperative infection.

Adolescent ; Adult ; Debridement ; Fibula ; injuries ; Fractures, Open ; surgery ; Humans ; Logistic Models ; Male ; Middle Aged ; Postoperative Complications ; etiology ; Retrospective Studies ; Risk Factors ; Surgical Wound Infection ; etiology ; Tibial Fractures ; surgery

Objectives:The present study was undertaken to investigate the effects of recombinant growth hormone(rhGH) on protein metabolism during intraabdominal infection.　 Methods : The study was made up of two parts: animal experiments and clinical studies. All animals were divided into three groups: normal control, sepsis control and rhGH therapy groups. All patients were divided into two groups: infection control and rhGH therapy groups. Expressions of hepatic albumin mRNA and muscle myosin heavy chain mRNA were detected.　 Results: In septic animals, rhGH accelerated nitrogen retention and recovery of body weight, improved concentrations of plasma albumin and muscle protein. Expressions of hepatic albumin mRNA and myosin heavy chain mRNA decreased significantly after infection. By using rhGH, expression of albumin mRNA and myosin heavy chain mRNA increased obviously. In clinical study, plasma concentrations of albumin, prealbumin and transferrin were higher than infectious controls after rhGH administration.　 Conclusions: rhGH improved nitrogen balance and protein synthesis of liver and muscle during sepsis .More studies were needed to determine weather rhGH was an effective metabolic intervention method during sepsis.

Objective To explore the function of infliximab in inducing remission in Crohn's disease and the effect of the inducing remission were followed up. Methods Ten patients with Crohn's disease received a infliximab, 5-aminosalicylic acid (5-ASA) and Azathioprine (AZA) therapy for inducing and maintenance remission. Crohn' s disease activity index (CDAI), C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), alanine aminotransferase (ALT), apartate aminotransferase, (AST), total bilirubin (TBil), conjugated bilirubin, (CB), creatinine (Scr) were evaluated at week 0, 10, 22 and 50. Simple endoscopic score for Crohn's disease (SES-CD) were evaluated at week 0, 10 and 50. Adverse reactions were also evaluated. Results At week 10, all patients achieved remission. The indicators of CDAI, CRP, ESR and SES-CD were significantly declined than those at week 0 (P＜0.01). The follow-up was terminated in one patient due to the relapse at week 30. At week 50, the indicators of CDAI, CRP, ESR and SES-CD in six patients a little bit increased compared with those at week 10, but no statistic significant (P=0. 2001、0. 0600、0. 1328、0. 4230 respectively), but significantly declined compared with those at week 0 (P =0.0005、0.0087、0.0054、0. 0163 respectively). No severe adverse reaction was observed in all patients.Conclusions Infliximab showed an exact efficacy in inducing remission in Crohn's disease. And 5-ASA and AZA were effective for maintenance remission in part of the patients after infliximab induced remission.

Objective To explore the feasibility of 256-slice whole-brain CT perfusion (CTP) in evaluate graft reperfusion after surgical revascularization and hemodynamic alterations before and after surgery in Moyamoya disease. Methods Twenty-five cases with Moyamoya disease were scanned on a 256-slice CT.CTP was performed pre- and post- surgical revascularization. The wolumetric CT angiographic ( CTA ) images were generated from volumetric data acquired at the arterial phase of CTP. CBF, CBV, TTP and MTT were measured in functional maps at the operated side within middle cerebral artery perfusion areas and contralateral mirroring areas. Relative CBF( rCBF), relative CBV (rCBV), relative TTP (rTTP), relative MTT (rMTT) were also obtained. Differences in perfusion CT values pre- and post operation were assessed with the paired t test or matched-pairs signed-ranks test. Data with normal distribution was present as : (x-)± s,while those with the non-normal distribution were present as M ( P25-P75 ). Results All the direct graft patencies were displayed on volumetric CTA. No significant differences were found between volumetric CTA and conventional CTA. Postoperative CBF, rCBF and rCBV values of the operated side [ 72. 86 (55.54-112. 19) ml · 100 g-1 · min-1 , 1. 31 ( 1.05-1.73), 1.45 ±0. 62] were significantly higher than those before operation [46.72(28.57-57.67) ml · 100 g-1 · min-1, 0.53(0.33-0.82), 1.01 ±0.36](Z=- 2.72, - 2. 98, t = - 2. 85, P ＜ 0. 05 ). Postoperative MTT, TTP and rTTP values of the operated side [ (3.98 ± 2. 36 ) s, ( 17.56 ± 4. 38 ) s, 1.01 ± 0. 09 ] were significantly lower than those before operation [(5.43±2.07) s,(19.40±3.87) s,1.14±0.28] (t=2.41,2.17,2.17, respectively, P＜0.05).However, no significant differences were detected for changes of CBV and rMTT after revascularization ( P ＞0. 05). Conclusion 256-slice CT has the potential value for the non-invasive assessment of both the graft patency and cerebral hemodynamics changes in moyamoya disease after surgery with administration of one contrast medium bolus in a single examination.

Objective To identity whether there is muscle atrophy phenomenon in end-stage kidney disease patients and to detect the level of transcription factor Foxo1 and the activity of ubiquitin-proteasome system.Methods Twenty-two patients in chronic kidney disease (CKD) stage 5 were selected and their mean muscle cross sectional area was measured.mRNA and protein levels of Foxo1,Atrogin-1,MuRF1 in rectus abdominis biopsies obtained from consecutive patients were detected.Control biopsies were obtained from 8 healthy subjects during elective surgery for abdominal wall hernias and 6 subjects during elective surgery for adenomyosis.Results Compared with the control group,cross sectional area of muscle fibers decreased and the transcription and protein levels of Foxo1,Atrogin-1,MuRF1 were upregulated in CKD group(P＜0.05).Protein level of p-Foxo1 decreased in CKD group(P＜0.05).Conclusion There exist muscle atrophy phenomenon in CKD patients,which may associate with the upregulation of Foxo1 and activation of ubiquitin-proteasome system.

Objective To study skeletal muscle atrophy and the change of autophagy in skeletal muscle of patients with chronic kidney disease.Methods Mean muscle cross sectional area,mRNA and protein expression of autophagy markers Bcl-2-adenovirus E1B interacting protein 3 (LC3B),Bcl-2-adenovirus E1B interacting protein 3 (Bnip3),Beclin-1 were measured in rectus abdominis biopsies obtained from 22 consecutive patients with stage 5 CKD scheduled for peritoneal dialysis from 4 hospitals in Shanghai.Control biopsies were obtained from another 8 healthy subjects during elective surgery for adenomyosis and 6 subjects during elective surgery for abdominal wall hernias.Rectus abdominis muscles were obtained at the beginning of surgery.HE staining was performed and mean cross sectional area (CSA) was calculated.Electron microscopy was used to confirm the changes of autophagy.mRNA levels of LC3B,Beclin-1,Bnip3 were evaluated by RT-PCR and protein levels of those parameters were evaluated by Western blotting.Results Compared with control group,mean CSA of muscle fibers was decreased and the transcript levels of LC3B,Beclin-1,Bnip3 were up-regulated in CKD group.Similarly,protein levels of LC3BⅠ,LC3B Ⅱ,Beclin-1 and Bnip3 were increased in CKD group.Additionally,activation of autophagy was confirmed by the appearance of autophagosomes by electron microscopy.Conclusion Chronic kidney disease may cause skeletal muscle atrophy and lead to activation of autophagy,which may contribute to muscle atrophy.

Under the guidance of strengthening medical undergraduates' practical ability and their competence in future jobs, Dalian Medical University has established a clinical medical talents practical ability training mode. This model has implementedThree Turnsclinical skills training pattern to strengthen the three stage (before, during and after practice) clinical skills training and concentrated on humanistic quality education through overall evaluations and set up clinical skill examination system to evaluate teach-ing effectiveness comprehensively and truly , which has effectively improved the quality of education by perfecting safeguard mechanisms and guaranteeing teaching quality.