Background Congenital aniridia is a rare congenital autosomal dominant disease,which is shown as aniridia of double eyes,and the paired box gene 6 (Pax6) gene mutation is now known to be associated with congenital aniridia.Objective This study was to screen the Pax6 gene mutation in patients with congenital aniridia.Methods Eleven patients with congenital aniridia were enrolled in Tianjin Eye Hospital from August 2012 to October 2015,including 6 patients from 3 congenital aniridia family and 5 sporadic patients.All patients received routine ophthalmic examination.Peripheral venous blood of 3 ml was collected from the patients for DNA extraction according to the standard process of DNA isolation instructions,and all the exons of Pax6 gene,Elp4 gene,exon 5 ' and 3',intron splice sequence and SIMO sequence were amplified by PCR.Pax6 genes of the patients were sequenced using Sanger direct sequencing and multiplex ligation dependent probe amplification (MLPA) and compared with those of 500 ocular trauma patients.This study complied with Helsinki declaration,and written informed consent was obtained from each patient prior to any medical examination.Results Iris absence was found in all the patients,and the visions acuity was hand motion to 0.2.Lens dislocation was seen in 1 patient.Direct sequencing results found that three patients in AN-O1 family were c.688g＞t (p.E230X) mutation of Pax6 gene,and 3 of 5 sporadic patients carried c.468g＞a (p.W156X),c.613c＞t (p.Q205X) and c.141 +2t＞c mutant of Pax6 gene,and the c.688g＞t (pE230X) mutation was a novel-discovered mutation.No any mutation in Pax6,Elp4 gene and SIMO fragment was detected in 1 patient from AN-02 family,2 patients from AN-03 family and 2 sporadic patients by both direct sequencing and MLPA validation.No above-mentioned mutation was found in 500 normal individuals.Conclusions The mutation of Pax6 gene is a pathogenic mutation in congenital aniridia patients,and c.688g＞t (p.E230X) is a novel Pax6 mutant,which expanded the mutation spectrum of Pax6 gene.

Background Oculocutaneous albinism (OCA) is a hereditary disease of pigment absence in eyes,skin and hair due to the lack of congenital melanocyte.OCA is classified into 7 types based on different genetic mutations,and the mutation of tyrosinase (TYR) gene causes OCA type 1 (OCA1).OCA has obvious genetic heterogeneity and phenotypic heterogeneity.The molecular diagnosis of the mutant gene is helpful for the classification and molecular pathogenesis study of OCA.Objective This study was to screen the TYR mutation in OCA patients,and to analyze the association between the gene mutation type and clinical phenotype.Methods Ten patients with OCA were enrolled in Tianjin Ophthalmological Hospital from January 2011 to December 2014.The clinical and ocular manifestations of the patients were examined.Peripheral venous blood 3 ml was collected in the patients and their lineal relatives for the extraction of genomic DNA.Extracted DNA was amplified by PCR and the TYR gene sequence was analyzed,including all 5 exon coding sequence and exon 5 ' and 3' end and the non-coding region sequence of intron splicing in TYR gene.This study complied with Helsinki Declaration and the protocol was approved by Ethic Committee of Tianjin Eye Hospital.Informed consent was obtained from each subject.Results All the patients showed white or reddish hair and snow-white skin,and different degrees of pigment lack was seen in iris.The best corrected visual acuity of the patients was 0.05-0.2,and 3 patients complicated with nystagmus.Fundus findings showed a sunset-like change and dysplasia of macula.The TYR gene sequencing revealed that patient 1 was OCA1A subtype,with the compound heterozygous mutant of c.832C＞T (p.R278X) and c.1217C＞T (p.P406L),and his/her parents occurred the heterozygous mutation of exons P406L and R278X.The phenotype of the patient 1 was white hair and white iris.The patient 3 was OCA1B subtype,with the compound heterozygous mutations of c.1265G＞A (p.R422Q) and c.1217C＞T (p.P406L),showing an appearance of reddish brown hair and sallow iris.TYR gene mutant was not detected in other 8 patients.Conclusions The mutation of TYR gene is the main cause of OCA1 type.The phenotype of OCA1A subtype is no pigment in eyes and hair,and one of OCA1B subtype was obviously lessening of pigment.The difference of mutant genes of OCA is the cause of genetic and phenotypic heterogeneity.

Enterovirus type 71(EV71) causes severe hand-foot-and-mouth disease(HFMD) resulting in hundreds of deaths of children every year; However,currently,there is no effective treatment for EV71.In this study,the EV71 poly-protein(EV71-P1 protein) gene was processed and cloned into the eukaryotic expression vector pPIC9k and then expressed in Pichia pastoris strain GS115.The EV71 P1 pretein with a molecular weight of 100 kD was produced and secreted into the medium.The soluble EV71 P1 protein was purified by column chromatography with a recovery efficiency of 70％.The result of the immunological analysis showed that the EV71 P1 protein had excellent immunogenicity and could stimulate the production of EV71-VP1 IgG antibody in injected rabbits.We suggest that EV71-P1 protein is an ideal candidate for an EV71 vaccine to prevent EV71 infection.

ObjectiveTo evaluate prognostic factors affecting local tumor progression after radiofrequency ablation (RFA) of hepatocellular carcinoma (HCC).MethodsA total of 246 HCC patients (343 lesions) underwent RFA treatment in our department and were enrolled into this study.The average tumor size was 3.7 cm ( range 0.9 ～ 3.7 cm).Regular follow-up with enhanced CT was performed to evalutate the treatment results.Kaplan-Meier model and log-rank test were used in univariate analysis and COX regression model was used in multivariate analysis to identify risk factors for local tumor progression.ResultsThe local tumor progression rate was 11.4％ (39/343 lesions),and the average time from initial RFA to local tumor progression was 12.0 months.Univariate analysis indicated tumor size ( P ＜0.001 ),close to intrahepatic vessels ( P ＜0.001),tumor boundary ( P =0.020),pathological grade( P =0.010) and CEUS before RFA ( P =0.001) as risk factors for local progression.The following factors were identified as independent prognostic factors for local tumor progression by multivariate model:tumor size (P ＜ 0.001),isolated or close to intrahepatic vessels( P ＜0.001) and CEUS before RFA(P =0.018).ConclusionsTumor size,CEUS before RFA and close to intrahepatic vessels are the most important factors for local progression after RFA.Being awaring of possible risk factors for local tumor progression may increase the treatment efficacy and help to promote the use of RFA technique.

Objective To examine the effects of IL-10 on cardiac fibroblasts ( CFBs) proliferation and phenotype transformation to myofibroblasts (MyoFbs) induced by transforming growth factor-β1 (TGF-β1);and to investigate the regulating pathways .Methods Cardiac fibroblasts were isolated from cardiac ventricles of neonatal SD rats . The passage 2~4 were used and divided into the following groups for treatment:1) control group, 2) IL-10 reac-tion group, 3) TGF-β1 reaction group, and 4) IL-10 plus TGF-β1 reaction group (TGF-β1 treatment followed with IL-10 pretreatment ) .Cells proliferation was assessed by MTT assay and immunocytochemistry staining for prolifera-ting cell nuclear antigen (PCNA);the phenotype transformation into MyoFbs was assessed by immunocytochemistry of α-smooth muscle actin (α-SMA);extracellular signal related kinase ( ERK1/2) and P38 kinase pathways were assessed by western-blot.Results TGF-β1 (10 μg/L) treatment boosted the proliferation and the expression ofα-SMA significantly (P<0.01), while IL-10 (10, 50 or 100 μg/L) plus TGF-β1 co-treatment induced lower cell proliferation and expression of α-SMA than treating with TGF-β1 alone ( P<0.05 ) , with the inhibitory effect of IL-10 being concentration dependent .TGF-β1 could significantly stimulate the ERK 1/2 and P38 kinase phospho-rylation ( P<0.01 ) , however IL-10 (100 μg/L) plus TGF-β1 co-treatment failed to down-regulated the phospho-rylation of ERK1/2 and P38 kinase compared with TGF-β1 alone ( ERK1/2:P<0.05;P38:P<0.01 ) .Conclu-sions IL-10 can attenuate TGF-β1-induced CFBs proliferation and phenotype transformation to MyoFbs .The in-hibitory effects may explained by a mechanism of inhibiting the activation of ERK 1/2 and P38 kinase .

Objective To investigate the expressions of heparanase(Hpa) and basic fibroblast growth factor(bFGF) as well as their relationship with angiogenesis in human ovarian carcinoma.MethodsForty-one ovarian carcinoma specimens resected from the patients diagnosed with ovarian carcinoma from January 2003 to November 2007 were examined by immunohistochemistry for the expressions of Hpa and bFGF and microvessel density(MVD).ResultsHpa highly expressed in ovarian carcinoma,and its expression was positively correlated with MVD and bFGF expression(r=0.351,P

Objective To explore the effect of application of hospital-school combined three-dimensional teaching mode in the operation nursing practice teaching.Methods 129 students were collected by cluster sampling,then based on class unit,they were randomized into the experimental group (69 students) and the control group (60 students).The experimental group used hospital-school combined threedimensional teaching mode and the control group used traditional teaching mode.Then,both groups had finished an operation nursing comprehensive theoretical examination and a self-report questionnaire for mastery of basic knowledge of operation nursing.Besides,the experimental group had a questionnaire survey of cognitive evaluation for the new teaching mode.Results The average score of operation nursing comprehensive theoretical examination of the experimental group was (77.81 ±9.87),higher than the control group (70.35±12.37); The self-report score for mastery of basic knowledge of operation nursing of the experimental group was (3.94±0.45),higher than the control group (3.68±0.46).The students of the experimental group also considered that the new teaching mode improved their comprehensive knowledge and innovative abilities,critical thinking ability,humanistic care spirit,communication ability,team cooperation ability and innovation ability,etc.Conclusions The effect of application of hospital-school combined three-dimensional teaching mode in the operation nursing practice teaching was good,but it still had some limitations which needed us to pay attention to and to perfect.

Objective To investigate relationship of platelet (PLT) count with clinicopathological features and prognosis of patients with breast cancer,and explore the susceptibility index to evaluate prognosis of patients with breast cancer.Methods A retrospective analysis of clinical data of 498 patients with breast cancer in January 1995 to December 2005 was carried out.PLT count was tested.Those patients were divided into group A (PLT ＜ 150 × 109/L),group B[(150-250) × 109/L],and group C (PLT ＞ 250 × 109/L) according to PLT count level.The relationship of platelet count with clinicopathological features was analyzed.Kaplan-Meier and Cox proportional hazards model were used for univariate analysis and multivariate analysis of PLT impact on survival time.Results There was positively correlated between PLT count and clinicopathological features (Pearson coefficient ＞ 0,P ＜ 0.05).There was negative correlated between PLT count and survival time (Pearson coefficient =-O.583,P ＜ 0.05).The survival time of groups A,B and C were significantly different (P =0.018).Cox proportional hazards model multi-factor analysis showed that PLT count was an independent factors affecting survival time (OR =2.256,P ＜ 0.05).Conclusions Patients with breast cancer associated with increased emphasis and PLT count.PLT count had negative correlation with survival time.PLT count could be a susceptible index to predict the prognosis of patients with breast cancer.

Objective To assess the prevalence of Personality Diagnostic Questionnaire (PDQ)personality deviations in patients referred for functional dyspepsia (FD) with reliable and universal psychological measures, and to explore the relationship between co-occurring PDQ-personality deviations and functional dyspepsia. Methods The sample comprised 246 patients referred for functional dyspepsia. Four groups were divided according to their patterns of gastrointestinal symptoms: the FD group, FD with refluxlike symptom group(FD + RS group), FD with irritable bowel syndrome group( FD + IBS group), and FD with reflux-like symptom and irritable bowel syndrome group ( FD + RS + IBS group). Participants were assessed with the Personality Diagnostic Questionnaire for DSM-Ⅳ ( PDQ-4 ) to evaluate the presence of personality deviations. Results Overall 65% patients scored positive for any personality deviation, male and female alike. Cluster C (anxious/fearful) personality was most commonly found in FD patients (142 patients, 57.7% ). The FD + IBS group and the FD + RS +IBS group had significantly higher total PDQ scores than the FD group (23.39 ± 8. 77 and 24. 22 ± 10. 97 vs 18.98 ± 11.88, P ＜ 0. 05, respectively),indicating that FD patients with greater level of personality deviations tend to report other symptoms involving the esophagus and lower gastrointestinal tract. Reflux-like symptom without actual pathological acid regurgitation indicated cluster A (odd/eccentric) personality deviations. Conclusions The current study shows personality deviations are common in patients referred for functional dyspepsia. Negative emotions,maladaptive coping, and lack of social support, may strongly influence their healthcare-seeking behavior.There is no single personality type specific for some kind of gastrointestinal symptom. But FD patients with personality deviations tend to report other symptoms involving the esophagus and lower gastrointestinal tract.

BACKGROUND:Rapid loss of liver-specific functions of the cultured hepatocytes limits the development of hepatocyte-based therapies. OBJECTIVE:To establish a three-dimensional culture system based on col agen hydrogel that enables to enhance liver-specific functions for a long period during culture of hepatocytes. METHODS:Hepatocytes were isolated from Sprague-Dawley rats and then encapsulated into liquid type Ⅰcol agen solution that was premixed with hepatocyte growth factor and Dulbecco's modified Eagle’s medium to create hepatocyte/col agen hydrogel compounds. The compounds were inoculated into a 96-wel plate. After gelation, culture medium was added. Light microscope, hematoxylin-eosin staining and transmission electron microscopy were used to examine the morphological characteristics and ultrastructure of the cultured hepatocytes. The cellsupernatant was col ected and tested for albumin secretion and urea synthesis. Periodic acid-schiff staining, immunofluorescence staining and quantitative real-time PCR were also used to further clarify liver-specific phenotype or function of the hepatocytes.RESULTS AND CONCLUSION:(1) Light microscope revealed that hepatocytes were round shape and distributed uniformly in col agen hydrogel. The three-dimensional hepatocyte culture system exhibited similarities to liver-like structure and tight junction were formed between hepatocytes after 14 days of culture. (2) Within the three-dimensional culture system, hepatocytes remained positive for periodic acid-schiff staining, albumin and hepatocyte nuclear factor-4αpositive after 14-day culture, which provided the convincing evidence of highly differentiated primary hepatocytes with functions of glycogen and albumin synthesis. (3) The albumin and urea productions in the three-dimensional culture system had a significantly higher level than in the two-dimensional culture, and could remain at a high level at least for 15 days. (4) The expression levels of hepatocyte-specific genes including Albumin, HNF-4α, Claudin-3, CYP1A1, CYP3A1 and G6P were significantly improved in the three-dimensional culture as compared with the two-dimensional culture. The col agen hydrogel based three-dimensional culture system provides a valuable model to enhance the hepatocyte functional maintenance and lay the foundation for the development of hepatocyte-based therapy for liver disease.