Objective To investigate the influence of Wnt-3a signaling on aggregative growth of dermal papilla cells.Methods Recombinant adenovirus pAdEasy-1/Wnt-3a was constructed at first,and used to transfect cultured dermal papilla cells(DPCs).Then,the growth pattem of DPCs was observed by inverse microscopy;laser confocal microscopy and Western blot were utilized to detect the expressions of Wnt-3a,β-catenin and versican in low-passage DPCs,high-passage DPCs and transfected high-passage DPCs.Results The over-expression of Wnt-3a protein could effectively induce the aggregative growth of DPCs.Laser confocal microscopy showed that the fluorescence intensity of β-catenin and versican increased from 34.16±9.62 and 18.81±9.59 in high-passage DPCs to 87.35±17.28 and 92.18±18.39 in transfected high-passage DPCs(t=11.98,17.88,both P＜0.0 1),respectively.Also,Western blot proved a significant increase of expression intensity of β-catenin(15.27±2.17 vs 60.09±7.24,t=10.10,P＜0.01)and versican(19.32±2.46 vs 58.46±2.78,t=20.63,P＜0.01)in transfected high-passage DPCs compared with untransfected high-passage DPCS.ConclusionsWnt-3a signaling plays an important role in modulating aggregative growth of DPCs,which is mainly through the classical pathway with β-catenin as the key protein.As an important downstream gene of Wnt signaling,vesican is an indispensable part for the modulation of aggregative growth of DPCs.

Objective To better understand the mechanisms of cone opsin transport , we set to generate a trans-genic zebrafish line with red fluorescence proteins expressing in the cone photoreceptors .Methods We used sws1 promot-er to drive the expression of a chimerical protein , in which the last 44 amino acids of rhodopsin of Xenopus laevis were fused to the C-terminus of tdTomato to restrict its localization to the outer segment of photoreceptors .Results We successfully i-solated such a transgenic zebrafish line and confirmed the localization of tdTomato by immunostaining analysis .Conclu-sions This transgenic zebrafish line will help us to better understand the transport mechanisms of cone opsins , especially the transport in live photoreceptors .

Objective:To study whether autoimmune regulator (Aire) affects macrophage polarization. Methods: The mouse mononuclear macrophage cell line RAW264. 7 cells, stable expressing GFP-Aire protein RAW264. 7 cells ( A33-3 ) and stable expressing GFP protein RAW264. 7 cells (C1-6) were stimulated with LPS,IL-4 and LPS combined with immune complex respectively to make the macrophage polarize to M1 (LPS),M2a (IL-4) and M2b (LPS with immune complex). To investigate the effects of Aire on the various types of macrophages polarization,the M1 related molecules (IL-1α,iNOS and IL-6),M2a related molecules (Arg-1) and M2b related molecule (IL-10) were detected by Real-time PCR. Results:The expression of IL-1α,IL-6 and inducible nitric oxide synthase ( iNOS) ,the products of M1 macrophages, were significantly upregulated in RAW264. 7 cells treated with LPS at 0. 5 μg/ml. The expression of Arg-1 and IL-10 mRNA in RAW264. 7 cells increased in a dose-dependent manner after stimulation with IL-4 or LPS combined with immune complexes, respectively. M1 macrophage-related marker iNOS and IL-1 increased, whereas IL-6 levels decreased in A33-3 cells compared with C1-6 cells after treatment with LPS. The expression of Arg1 and IL-10 were downregulated in A33-3 cells compared with C1-6 cells after IL-4 and LPS combined with immune complexes stimulation,respectively. Conclusion:Aire may promote M1 macrophage polarization while inhibit M2a and M2b macrophage polarization.

Objective:To study the effect of seeding cells of tissue-engineered skins,keratinocytes and fibroblasts,on the phenotypes of langerhans cells (LC).Methods:Peripheral blood mononuclear cells were induced by cytokines,granulocyte macrophage-colony stimulating factor (GM-CSF),interleukin 4 (IL-4),transforming growth factor β1 (TGF-β1) to generate LCs,which were subsequently cocultured with allogenic keratinocytes or fibroblasts.Then the phenotypes of langerhans cells was detected by flow cytometer (FCM),and the degree of proliferation of autogeneic lymphocytes was assessed after stimulation with LCs.Results:Cultured LCs had low level expression of HLA-DR,CD80,CD86 and no expression of CD83.Cocultured with allogenic keratinocytes or fibroblasts,phenotypes of LCs did not change significantly.Simultaneously,LCs could not stimulate autogeneic lymphocytes proliferation.Conclusion:These data indicate that LCs cocultured with seeding cells of tissue-engineered skins remain immature state.It is suggested that seeding cells of tissue-engineered skins have low immunogenicity and are unable to induce significant immunological rejection of host after transplantation.

Objective To investigate the clinical efficacy of Shenmai injection (SMI) combined with heart transplantation of autologous bone marrow stem cells in the treatment of refractory heart failure(RHF). Methods Two hundred patients with RHF were selected from the Emergency Department of the First Affiliated Hospital of Zhengzhou University and the Cardiology Department of the First Affiliated Hospital of Henan College of Traditional Chinese Medicine. They were randomly divided into control group and treatment 1,2 and 3 groups(each 50 cases). In the control group,heart failure standard treatment was given;group 1 received a standard treatment for heart failure combined with autologous bone marrow stem cell heart transplantation;group 2 received a standard treatment for heart failure with SMI;group 3 received a standard treatment for heart failure combined with autologous bone marrow stem cell heart transplantation and SMI. The mean follow-up was 24 months. The prognosis,readmission rate,clinical efficacy,cardiac function and the change in levels of B-type natriuretic peptide(BNP)of patients in each group were observed during the therapeutic course and observation period. Results During the course of treatment,there were 10 cases dead in the control group,4 in each group 1 and 2 respectively,and 3 in group 3. Readmission rates in group 1,2 and 3 were significantly lower than that in control group(38%,36%,24%vs. 48%, P<0.05 or P<0.01). The rates of total efficiency in group 1,2 and 3 were obviously higher than that in the control group(88%,86%,94%vs. 76%,all P<0.05). After treatment,the left ventricular ejection fraction(LVEF),left ventricular fractional shortening(FS)and left ventricular end-systolic diameter(LVESD)in three therapeutic groups were significantly higher than those before treatment,while the left ventricular end-diastolic diameter(LVEDD)and BNP level were significantly lower than those before treatment. All the above indexes in three therapeutic groups after treatment were much more remarkably improved than those in the control group during the same period,and the group 3 being the most significant〔LVEF:0.477±0.099 vs. 0.396±0.098,FS:(30.0±5.1)%vs.(26.8±7.5)%,LVESD (mm):40.6±9.1 vs. 45.8±9.4,LVEDD(mm):44.9±9.8 vs. 52.8±10.1,BNP(ng/L):515±400 vs. 1 875±400, all P<0.05〕. Conclusion SMI combined with heart transplantation of autologous bone marrow stem cells has obvious therapeutic effect for treatment of RHF.

Objective To compare surgical gastrojejunostomy and endoscopic stenting in palliation of malignant gastric outlet obstruction.Methods This retrospective study investigated patients treated for malignant gastric outlet obstruction from January 2007 to January 2014 in the first affiliated hospital of Nanjing Medical University.Endoscopic stenting was placed in 29 patients and surgical gastrojejunostomy was performed in 42 patients.The outcomes assessed included diet scores,time to diet,length of hospital stay,treatments fees and complications.Results Both endoscopic stenting and surgical gastrojejunostomy can relieve patients' syndrome with significant higher GOOSS score compared with that before treatment (P ＜0.05),but score improves faster in stenting group.Clinical success for endoscopic stenting and surgical gastrojejunostomy was 96.6％ and 92.9％ respectively,and technical success was 100％ for both of them.Endoscopic stenting group was found to have lower early complication rate(3.4％ VS 23.8％,P ＜0.05),higher late complication rate(24.1％ VS 6.9％,P ＜0.05),less time to diet,hospital stay and treatment fees(all P value ＜ 0.05)than surgical gastrojejunostomy group.The major complication after endoscopic stenting is re-obstruction while it is infection and leak of anastomotic site for surgical group.There were no significant differences in complication between two groups (27.6％ VS 11.9％,P ＞ 0.05).Conclusion Both endoscopic stenting and surgical gastrojejunostomy can relieve patients' syndrome effectively and safely,but endoscopic stenting improves GOOSS scores more rapid with less time to diet,less early complication rate and easy-dealing late complications,also it needs less hospital stay and fees.It's a better choice for patients with less survival expectation.

Objective To investigate the effect of hepatocyte growth factor (HGF) on migration and apoptosis of,as well as phosphorylated-Akt (p-Akt) expression in cultured human eccrine sweat gland epithehal cells (hESGc).Methods The first generation of hESGc were cultured in keratinocyte serum free medium (KSFM) and treated with various concentrations (2,20,40μg/L) of HGF for different durations.Then,cell scratch test was performed to detect cell migration,a double staining flow cytometry assay using annexin VFITC/propidium iodide to detect cell apoptosis.and Western blot to measure the expression of p-Akt.Results HGF of 2μg/L had no effect on the migration of hESGc,while that of 20 μg/L and 40μg/L could promote the migration of hESGc by 33.2% and 228.2%.respectively.The average number of cells migrating into the scrach zone was significantly lower in untreated cell group than that in 20 and 40μg/L HGF-treated cell group (17.3±5.5 vs 23.0±6.3 and 56.7±7.9,t=2.653, 15.858,P<0.05,0.01, respectively).The apoptosis rate was 14.76% in untreated cells,14.16%,13.5% and 8.87% in cells treated with HGF of 2,20 and 40μ/L, respectively;there was a significant difference between untreated cells and 40μg/L HGF-treated cells (t=7.852,P<0.01).HGF could activate the phosphorylation of Akt protein and increase the expression of p-Akt.Conclusion HGF could promote the migration of,inhibit the apoptosis of,and stimulate the p-Akt expression in.hESGc.

Objective To investigate the clinical outcomes of surgical excision combined with recom binant interferon aipha-2b in the treatment of acral malignant melanoma (MM). Methods Fifteen patients with acral MM admitted to the department since 2004 were recruited into this study. The tumors varied from 1.8 mm to 3.9 mm in invasion depth. Thin tumors with an invasion depth of 1.8 - 2.0 mm were excised with a margin of lcm beyond the tumors, and those with an invasion depth of 2.0 - 3.9 mm were excised with a margin of 2 cm beyond the tumors. After excision, 4 cases of minor excision were sutured directly, 10 cases of large excision were repaired with adjacent skin grafts and flaps, 1 patient with the involvement of the first metatarsophalangeal joint underwent toe amputation followed by the repair of planta wound surface with the remaining skin on the dorsa of toes. Patients received intramuscular recombinant interferon alpha-2b for 3 months (3 million units daily for the first 3 days and 6 million units for the remaining days) following operation. Results There were 6 cases of MM in situ and 9 cases of invasive MM in this study. All the skin grafts and flaps survived. Within the 3-year follow up, relapse was observed only in 1 patient with invasive MM. Recovery was achieved in the functions of feet in all patients. Conclusion The excision of tumors with a margin determined by tumor thickness plus intramuscular interferon alpha-2h may improve the survival of patients with cutaneous MM in planta pedis with avoidance of amputation.

Objective To introduce an operative method of rhytidectomy with W-incision and fewer complications for facial rejuvenation.Methods A total of 84 patients were treated with rhytidectomy by multi-methods and followed up from January 2007 to December 2012.The rhytidectomy was done along the temporal hairline edge through the combined procedures,including sharp and blunt separation,liposuction and filling with autologous fat,tighting and fixing the fascial flap of superficial musculoaponeurotic system,as well as the suspension of the zygomatic cheek fiber fat pad.Results Follow-up for 3 moths to 2 years after surgery in 84 patients showed that the wound was healing with good concealment and without complications such as facial nerve injury or local uneven.All the cases achieved good results in facelift with high satisfaction rate after the combined operations.Conclusions This combined rhytidectomy is safe,effective,simple and easy.And it is a good and ideal approach of facial rejuvenation worthy of clinical application.

Objective To assess the effect of continuous early enteral nutrition(EEN)combined with intestinal mucosal protective agents on gut barrier function in patients with severe acute pancreatitis.Methods A total of 79 patients with severe acute pancreatitis selected from four centers between May 2004 to June 2006 were enrolled and divided into EEN combined with intestinal mucosal protective agents group(combined group,n=39)and total parenteral nutrition(TPN)group(n=40).The patients were received either EEN or TPN when homeostasis were achieved within 72 hours after onset.The patients in combined group were administered pepti-2000 variant combined with glutamine,arginine and intestinal mucosal protective agents.The patients in TPN group were administered through a central vein.APACHE-Ⅱ score was recorded every week;The concentration of serum amylase,plasmic diamine oxi dase(DAO)and endotoxin were mesured on day 1,7,14 and 21 as well as urinary excretion of lactulose (L)and mannitol(M).Complications,lenth and charges of hospital stay were recorded.Results There was no death in both groups.The APACHE-Ⅱ score decreased on day 7,but lower in combined group (6.00±1.60)than that in TPN group(7.08±2.34)(P＜0.05).On day 7,14 and 21,the concentrations of endotoxin in combined group was(39.30±15.82),(22.64±14.31),(14.81±10.93)Eu/L,respectively,urinary L/M ratio was 0.28±0.25,0.21±0.18 and 0.08±0.04,respectively,IFABP-c was 15.62±5.26),(5.46±1.18)and(3.26±0.94)pg/ml,respectively.All of these parameters were significantly lower than those in TPN group(P＜0.05).The infectious rates including pancreatic,peritoneal and respiratory infection in TPN group were much higher than that in combined group(26.47% vs 3.44%,P＜0.01).The composition of flora fecal remained unchange in combined group rather than TPN group.The mean hospital stay was shorter in combined group[(20.0±5.7)days]compared to TPN groups[(34.5±12.9)days].The charges were also significantly lower in combined group,with average cost of RMB 25,900±14,200,while it was 46,800±4,030 in TPN group.Conclusions EEN combined with intestinal mucosal protective agents can improve gut barrier function via reducing the gut permeability,improving the hypoperfusion,maintaining the integrity and gut fecal flora.It might reduce the course and charges of hospital stay.