BACKGROUND: Utilizing tissue engineering technique, various gel systems are served as scaffolds to repair bone defect. The scaffolds should have features of nontoxic and no teratological effects to the body. OBJECTIVE: To observe the effect of sodium alginate-gelatin/osteoblast gel on chromosomal pattern aberration in rabbits. DESIGN, TIME AND SETTING: The in vivo material animal experiments were conducted at the Beijing Shijitan Hospital and Department of Histology and Embryology, Peking University Health Science Center from October 2007 to March 2008. MATERIALS: A total of 12 New Zealand rabbits, aged 2 months, with clean grade, were randomly divided into 2 groups. The experimental group contains 4 female and 4 male rabbits, and the remaining 4 females were served as the control group. Sodium alginate dried powder were purchased from Sigma, USA, and the gelatin dried powder were supplied by Liidao Company, Hebei, China. METHODS: Following numbering, bone marrow was collected from 12 rabbits. Bone marrow stromal stem cells (BMSCs) were isolated by the density gradient centrifugation, and then in vitro cultured with osteoblast inductor. Osteoblasts following passage were an order of magnitude of 10~7. Bright pink gelatiniform liquid with mass ratio of sodium alginate and gelatin at ratio of 2:3 was prepared. Rabbit osteoblasts with final concentration of 5×10~9/L were mixed with CaCb solution to form fruit jelly-shaped sodium alginate-gelatin/osteoblast gel. Critical-sized calvarial defects were created in diameter of 1.5 cm in 12 rabbits. After 1 week, cell/scaffold complex (0.5 mL) was implanted to repair the bone defect in the experimental group. There was no treatment in the control group. MAIN OUTCOME MEASURES: The change of chromosomal pattern was observed at 3 months following reparation. RESULTS: No Chromosome somatotype aberration was found in 100 metaphases in the experimental group. From 400 metaphases of the control group, 4 abnormal cells were found, with 1% chromatid-type aberration ratio. Meantime, 12 abnormal cells in 800 metaphases of the control group were found, with 1.5% chromatid-type aberration ratio. The numerical value was within the normal range. Chromosome karyotype analysis: the chromosome number of each experimental rabbit was 2n=44, karyotype of the control rabbit was 44, XX, which was normal female; or 44, XY, normal male, no abnormal was found. The female rabbit in the experiment group was 44, XX, no abnormal was seen. CONCLUSION: From the cytogenetoxicity point of view, sodium alginate-gelatin/osteoblast gel is safe in repairing bone defects.

Objective:To retrospectively analyze the routine clinical laboratory parameters for hepatocellular carcinoma,in an attempt to search for parameters for diagnosis of hepatocellular carcinoma(HCC).Methods:The pre-operation clinical labo- ratory data,such as tumor makers,and serological biochemical indices,hepatitis B virus(HBV)infection markers,and HBV DNA titers,were collected from 828 patients who were pathologically diagnosed as having HCC;then the correlation between these data with tumor size and the pathological grades of HCC was analyzed.Results:It was found that 97.9% of the 828 pa- tients were infected with HBV and 70.9% of them were accompanied by liver fibrosis.We also found that the tumor size was correlated with albumin(ALB),globulin(GLB),A/G,aspartate aminotransferase(AST),ratio of aspartate to alanine amin- otransferase(AST/ALT),gamma-glutamyl transferase(GGT),alkaline phosphatase(ALP),alpha-L-fucosidase(AFU),al- pha-fetoprotein(AFP)and tumor grades;meanwhile,the pathological grades of tumor was correlated to prealbumin(PALB), GGT and tumor size(all P

Objective To establish the methods for quantitating the circulating tumor DNA with PicoGreen fluorescent nucleic acid stain and investigate the role of the quantitative analysis in diagnosis of breast cancer. Methods Circulating tumor DNA was isolated from serum with QIAmp blood kit and was quantitated by spectrofluorometry with PicoGreen fluorescent stain. Receiver operator characteristic ( ROC) curve and area under the curve were used to estimate the role of DNA quantification in diagnosis of breast cancer. Results The efficiency of QIAamp blood kit isolating DNA from serum was 37. 8%-46. 2%, average 43. 4%. Circulating tumor DNA concentration as low as 1 ng could be detected by PicoGreen spectrofluorometry. and the detected range was 1-500 ng/0. 2 ml. The median concentration of serum DNA in breast cancer group was (169. 70+ 124. 10) μg/L, and that of healthy control and breast benign group were (54. 30±36. 84) μg/L and (51. 70±29. 04) μg/L, respectively (P<0. 01). The area under the ROC curve was 0. 899 (95% CI: 0.848-0.951), and the sensitivity was 78. 2%, the specificity was 90% by using the cutoff value of 96. 0 ng/ml. Conclusions The concentration of circulating tumor DNA can be efficiently quantitated by PicoGreen spectrofluorometry, which indicates the potential of clinical applicability in breast cancer diagnosis.

Objective To explore the relationship between cognitive impairment and arterial stiffness in elderly patients. Methods A total of 142 elderly patients were enrolled. Cognitive function was assessed by mini-mental state examination (MMSE) and arterial stiffness was assessed by pulse wave velocity (PWV). A full score on the MMSE was 30, and cognitive impairment was defined as a score less than 24. All subjects underwent the measurement of PWV and MMSE . The subjects were divided into 2 groups: 93 were assigned to the normal cognitive function group (MMSE score 24), and the remainders (n=49) were assigned to the cognitive impairment group (MMSE score 24). Results The PWV was significantly increased in the cognitive impairment group than in the normal cognitive function group [(13.3±2.4) m/s vs. (11.8±2.2) m/s, t=3. 775, P=0. 000]. Logistic regression analysis showed that the PWV was also independently and significantly associated with the MMSE score. Conclusions The increase of arterial stiffness is an important risk factor for impaired cognitive function in elderly patients.

OBJECTIVETo analyze the fungal composition in Massa Medicata Fermentata based on culture dependent method and independent PCR-SSCP technique.

METHODFungi were directly isolated from Massa Medicata Fermentata samples. The obtained strains were identified according to morphology and DNA sequence. Meanwhile the total fungal DNA was extracted from Massa Medicata Fermentata samples, the cultural independent PCR-SSCP technique based on β-tubulin gene were used to identify the mycobiota.

RESULTAccording to cultural method, Aspergillus flavus and Rhizopus oryzae were present in Massa Medicata Fermentata samples, while A. flavus and A. niger were present in fried Massa Medicata Fermentata samples. In contrast, 5 species were obtained by PCR-SSCP technique, A. flavus was overlapped with fungal taxa derived from culture dependent method; A. ambiguu and A. s ivoriensis were dominant with relative abundance of 57% and 35% respectively, while the relative abundance of A. flavus was as low as 4%. None species was obtained from fried Massa Medicata Fermentata samples.

CONCLUSIONPCR-SSCP based on β-tubulin gene could distinguish fungi into species, culture dependent method combined with culture independent method could better understand the fungal composition associated with Massa Medicata Fermentata fermentation.

Objective To detect the differential expression genes(DEGs)between Barrettg esophagus(BE)and normal esophagus with oligomicroarray,and to explore the target genes related to the development of BE.Methods The total RNAs of matched BE and normal esophagus mucosa from saIne patient were isolated with one step Trizol method.Matched RNAs were qualified with 10g/L agarose gel electrophoresis.After tRNA purification,cRNAs were synthesized and labeled with fluorescence.which were tIlen hybridized with Agilent oligomicroarray containing 30,968 probes.The fluorescence intensity features were detected by Agilent scanner and quantified by software Feature Extraction.Results On average,2 biopsies by disposable jumbo biopsy forceps provided approximately 5μg RNA required for microarray.The total RNA,reverse transcription product and fluorescence labeled cRNA were all of high quality.Among 2-fold DEGs,there were 142 up-regulated genes and 284 down-regulated genes including 15 bel-2 related genes such as bel-2,MCL1,BAX,BIK and BCLAF1 Conclusion Microarray-based studies are feasible in endoscopically obtained tissues.The development of BE is a complicated process involving multi-genes,in which abnormal expression of bel-2 family related genes might be involved,but the exact mechanism needs further research.

Objective To investigate pathological characteristics and differential diagnosis of desmoplastic melanocytic nevus.Methods Four cases of desmoplastic melanocytic nevus were analyzed based on the clinical manifestations and histological,immunohistochemical and fluorescence in situ hybridization (FISH) features.Results Of the 4 cases,2 were male and 2 were female.Their age ranged from 19 to 30 years with the average age being 26.5 years.The skin lesions were located on the extremities in 3 cases,on the vulva in 1 case.Histologically,the lesions were bilaterally symmetrical intradermal nevus.Nevus cells appeared epithelioid and/or fusiform,some were clustered or scattered in the proliferative fibrous tissue.None of lymphocyte aggregation,necrosis or ulceration was observed.Immunohistochemical examination showed positive staining for S100 and Melan A in 3 cases,positive staining for P16 in 2 cases,Ki-67-1abeling index less than 5％ in all the 4 cases,and negative staining for factor XⅢ (FXⅢ) and CD34 in 2 cases.FISH assay showed no copy-number variations in gene loci 6p25 (RREB1),6q23 (MYB),6p11.1-q11.1 (Cep6) and 11q13 (CCND1) in desmoplastic nelanocytic nevus.Conclusion Desmoplastic melanocytic nevus is a kind of histologically unique,benign melanocytic nevus,and immunohistochemical staining for Ki-67,S-100,Melan A and FXmand FISH assay on melanoma can be helpful for the differential diagnosis between cutaneous fibrous histiocytoma and melanoma.

Effect of p-nitrophenol shock on microbial populations and sludge activity in UASB reactor was studied by DGGE-PCR of 16S rDNA fragments and detection of COD removing and biogas yield.The results showed that p-nitrophenol seriously inhibited the sludge activity,resulting in the drop of biogas and COD removing rate.The 40mg/L p-nitrophenol had more inhibition than 20mg/L p-nitrophenol.It would take 27 and 16 days respectively for reactor to recover after 40mg/L and 20mg/L p-nitrophenol shock.The diversity of eubacteria and methanogens were also effected by the p-nitrophenol shock.The variation of eubacteria was more than that of methanogens after p-nitrophenol shock.The drop of biogas was mainly related to the variation of Methanosaeta sp.and Methanomicrobia sp.after p-nitrophenol shock.Among the eubacteria the population of Chloroflexi sp.、Bacteroide sp.and Anaerovibrio sp.decreased greatly after p-nitrophenol shock.And more,the Rheinheimera sp disappeared after 40mg/L p-NP treatment.But the Flavobacteria sp.appeared after p-nitrophenol shock,which was probably related to the degradation of p-NP.

AIM: To assess the differentiation of rat mesenchymal stem cell (MSC) in the microenvironment of retinitis pigmentosa(RP) induced by the administration of sodium iodate. METHODS: In vitro cultured Lewis rat MSC were injected into the subretinal space of NaIO3 induced RP rat eyes (30g/L NaIO3 100mg/kg). To observe the trace and differentiation of MSC by immuno-fluorescent method successively in 5 weeks after the surgery.RESULTS: The majority of the transplanted cells stay in retinal pigment epithelium(RPE) layer and cones and rods layer. From the 2nd week after transplantation, the engrafted MSC expressed PCK and rhodopsin under fluorescent microscope.CONCLUSION: MSC can survive mainly in the outer layer of retina in the microenvironment of RP and differentiate forward the RPE cell and photoreceptor.

AIM: To analyze the genotype of the allele distribution of a polymorphic G-954C within the 5 upstream promoter region of the nitric oxide synthetase 2A gene (NOS2A) in samples of diabetic retinopathy in patients with cystoid macular edema in the mainland of China.METHODS: Eighty-nine patients with diabetic retinopathy and cystoid macular edema and 90 healthy controls were enrolled in this study. Nest polymerase chain reaction (PCR)was performed, and restriction endonudease digestion and gene fragments sequence were examined to detect the genotype of NOS24 G-954C.RESULTS: The genotypes of the sample population of 89 cases and 90 healthy controls were all detected as GG.CONCLUSION: The distribution of G-954C of NOS2A polymorphism are at a lower frequency in China, with little relevancy to the frequency of diabetic retinopathy combined with cystoid macular edema.