Objective To enrich pancreatic cancer stem cells through culturing mammospheres, and to detect the expressions of epidermal growth factor receptor(EGFR) and a disintegrin and metalloprotease 9 (ADAM9) and investigate their significance. Methods PANC1 cells were cultured in serum-free conditioned medium to continuously generate mammospheres, and parts of mammospheres were cultured on a collagen substratum to induce differentiation. Mammospheres cells and differentiated cells were collected, flow cytometry was used to detect the proportion of side population (SP) cells, and the expressions of EGFR, ADAM9 mRNA and protein were detected by real-time PCR and Western blotting. Results PANC1 cells mammospheres were successfully generated and could be passed continuously. After differentiation, mammospheres cells could regain the ability of adherent growth. The proportion SP cells in mammospheres cells and differentiated cells were ( 5.40 ± 0.38 ) % and (2.80 ± 0.42 ) %, and the difference was statistically significant ( P ＜ 0. 05 ).Compared with differentiated cells, the expression of EGFR and ADAM9 mRNA of mammospheres cells upregulated 2.5 and 3.0 folds ( P ＜ 0. 05 ). The expressions of EGFR and ADAM9 protein of mammospheres cells were 0.90 ± 0. 09 and 0.64 ± 0.07, which were significantly higher than those in differentiated cells (0.62 ±0.11 and 0.48 ±0.09, P ＜0.05). Conclusions Mammosphere cells contained higher proportion of pancreatic cancer stem cells. ADAM 9 may play an important role in the occurrence and development of pancreatic cancer through the EGFR signaling pathway.

ObjectiveToevaluatethefeasibilityandresultsof transorallaparoscopic thyroidectomy. MethodThyroidectomy was attempted in 5 cases,including 4 females and 1 male with the average age of 42 years( range 35 -60 years).All patients was diagnosed as single nodule of the thyroid gland confirmed by B-mode ultrasound examination before the operation.The average diameter of nodule was 2.5 cm (range 2 - 3.4 cm). ResultTransoral laparoscopic thyroidectomy was perfoormed successfully in all 5 patients.The operation time was 120 - 210 min,averaging 170 min,blood loss during the operation was 15-60 ml,the postoperative hospitalization was 4 days. There was no conversion to open surgery,no recurrent laryngeal nerve injury,nor parathyroid glands dysfunction. ConclusionsTransoral laparoscopic thyroidectomy is feasible and safe,giving excellent cosmetic results.

Previous studies have shown that miRNAs participate in a wide range of biological functions and play important roles in various human diseases including cancer. We found miR-146b-5p significantly dysregulated in human pancreatic cancer cells by qRT-PCR. To demonstrate its function and regulation mechanism, we overexpressed miR-146-5p by transfecting the mimics. Our data showed that miR-146b-5p overexpression significantly reduced the abilities of migration and invasion of MIA PaCa-2 pancreatic cancer cells. Furthermore, we found that matrix metalloproteinase 16 (MMP16) was a downstream target of miR-146b-5p by dual-luciferase reporter assay. Altogether, our findings suggest that miR-146b-5p may be involved in pancreatic cancer cell migration and invasion by targeting MMP16, and miR-146b-5p may be a potential therapeutic target for the pancreatic cancer.

Objective To prepare polyclonal antibody of transactivated protein 5 of hepatitis C virus nonstructural 5A (NA5ATP5) and to explore its expression in the liver tissues. Methods In Escherichia coil BL21, the prokaryotic expression vector pET32a(+)-NS5ATP5 was induced by isopropyl-β-D-thiogalactoside (IPTG), and it was analyzed with sodium dodecyl sulfate-polyaerylamide gel electrophoresis (SDS-PAGE) and Western blotting. And the purified protein was used to immunize the rabbit to prepare polyelonai antibody, with which we studied the function of NSSATP5 by determining the different liver tissues with the streptavidin-perosidase (SP) immunohistochemistry method. Results Recombinant NS5ATP5 (molecular weight: 65 kD) and polyclonal antibody were successfully prepared. NS5ATP5 expression in the liver of patients with chronic HCV infection was much higher than that of a normal person, and it was detected mainly in the cytoplasm. Conclusion The findings of the expression difference between HCV patients and normal people led to a novel diagnostic marker to detect HCV infection.

Objectives To isolate cancer stem cells (CSCs) in pancreatic cancer cell lines PANC1 and ASPC-1 with serum-free medium( SFM ), and to detect the expression of miR-590-3p in CSCs. Methods PANC1 and ASPC-1 cells was cultured in serum-free medium. The monoclonal formation, differentiation and cell cycle, half inhibitory concentration ( IC50 ), and the expression of the surface markers CD24 + , CD44 + were detected. qRT-PCR was used to detect the expression of miR-590-3p. Results After SFM culture, (0.94 ±0.53 ) ％ of ASPC-1 and (0.57 + 0. 12 ) ％ PANC1 survived, and they formed spheres, and could continuously passage in vitro. Cell spheres differentiation recurred when serum was supplemented in SFM. The G0/G1 stage proportion, CD24+ , CD44 + , CD24+ CD44+ cells proportion, IC50 in ASPC-1 cell were (75.3 ± 5.4)％,0.96％ ～ 2.01％, 27.52％ ～ 34.47％, 0.35％ ～ 0.44％ and (224.37 ± 5.71 ) μg/ml, which were significantly higher than that those in parent cell [ (43.7 ± 3.8 ) ％, 0. 38％ ～ 0.42％, 17.65％ ～ 18.25％,0.05％ ～0.08％, (11.43 ±2.10)μg/ml, P＜0.05]. The G0/G1 stage proportion, CD24+ ,CD44+ ,CD24 +CD44 + cells proportion, IC50 in PANC 1 cell were ( 80. 1 ± 4.7) ％, 5.31％ ～ 9.84％, 72.05％ ～ 93.06％,4.91％ ～5.21％, (296.58±4.27) μg/ml, which were significantly higher than that those in parent cell [ (46.1 ±5.3)％, 4.09％ ～4.97％, 47.71％ ～55.66％, 1.48％ ～2.63％, (26.17 ±3.81)μg/ml, P＜0.05]. The expression of miR-590-3p in ASPC-1, PANC1 spheres was 4.67 and 4.52 times higher than the expression in parent cell lines. Conclusions Pancreatic cancer cell spheres can be isolated from ASPC-1, PANC1 by culture with SFM. miR-590-3p is up-regulated and may play an important role in regulating biological characteristics of pancreatic cancer stem cells.

OBJECTIVE: To investigate the therapeutic effects of oral modified device combined with nasopharyngeal enlargement surgery and evaluate the oral modified device' s adjuvant therapy meaning in severe OSAHS patients after surgery treatment. METHOD: 46 cases with severe OSAHS were diagnosed by PSG according to AHI and the lowest arterial oxygen saturation (LSaO2). We performed the nasal or pharyngeal cavity expansion surgery for them according to the pathological change part correspondingly. Then all subjects were divided into combined group (n=26) and surgery alone group (n=20) according to their personal willingness. We monitored the PSG for all subjects aftter 2 weeks and 3 months respectively, then we calculate the diversity between the two group or intragroup change on the basis of the AHI, LAT, LSaO2, mean arterial oxygen saturation (MSaO2) and sleep structures recorded by PSG. At the same time, we collected the subjective sensations by questionnaire. RESULT: The AHI and LAT in combined group were significantly lower and LSaO2 was significantly higher than these in surgery alone group(P<0. 05), and it's no difference in MSO2 between the two groups (P>0. 05). The N 1% was more shorter and the N2% and N3% were more longer after nasal or pharyngeal operation compared with pre-operative states in both groups(P<. 05), but we didn't find difference in REM%(P>. 05). The data of PSG also showed that the shallow sleep proportion was more shorter and the slow wave sleep proportion was more longer in combined group compared with surgery alone group. The subjective sensations results also showed significantly alleviated in combined group, such as mental state, daytime sleepiness and physical strength. The efficiency ratio of treatment was 85. 0% and 92. 3% in surgery alone group and combined group respectively. CONCLUSION Nasal and pharyngeal cavity enlargement surgery combined with oral modified device is a more effective treatment in patients with severe OSAHS, and it is meaningful for the long-term curative effect of surgery to prevent relapse and improve.
Humans ; Nasopharynx ; surgery ; Nose ; surgery ; Otorhinolaryngologic Surgical Procedures ; Oximetry ; Sensation ; Sleep ; Sleep Apnea, Obstructive ; surgery ; Sleep Stages ; Treatment Outcome

Objective To evaluate the feasibility and results of laparoscopic thyroidectomy by oral and breast approach for the treatment of papillary throid carcinoma.Methods Thyoidectomy was performed in 26 cases,including 24 females and 2 males with the average age of 34 years (range 20-53 years).All patients were diagnosed throid carcinoma confirmed by FNA or B-mode ultrasound examination,a thyroid lobe or total thyroidectomy and central compartment dissection was performed by breast approach,then additional dissection of central compartment was completed through oral approach.Results Laparoscopic thyroidectomy via oral in combination with brest approach was performed successfully in all 26 cases.The mean operative time was (164 ± 13) min,including average time of oral approach of (40 ± 7) min.The mean number of lymph node dissection in central compartment was 7.42 ± 4.88,oral approach achieved additional 1.23 ± 2.21,with metastatic lymph nodes diseccted by oral approach in 3 cases.Conclusions Laparoscopic thyroidectomy via oral in combination with breast approach for the treament of papillary throid carcinoma is better than breast approach alone in central compartment dissection.

Traditional Chinese medicine (TCM) preparations are widely used for healthcare and clinical practice. So far, the methods commonly used for quality evaluation of TCM preparations mainly focused on chemical ingredients. The biological ingredient analysis of TCM preparations is also important because TCM preparations usually contain both plant and animal ingredients, which often include some mis-identified herbal materials, adulterants or even some biological con-taminants. For biological ingredient analysis, the efficiency of DNA extraction is an important fac-tor which might affect the accuracy and reliability of identification. The component complexity in TCM preparations is high, and DNA might be destroyed or degraded in different degrees after a series of processing procedures. Therefore, it is necessary to establish an effective protocol for DNA extraction from TCM preparations. In this study, we chose a classical TCM preparation, Liuwei Dihuang Wan (LDW), as an example to develop a TCM-specific DNA extraction method. An optimized cetyl trimethyl ammonium bromide (CTAB) method (TCM-CTAB) and three com-monly-used extraction kits were tested for extraction of DNA from LDW samples. Experimental results indicated that DNA with the highest purity and concentration was obtained by using TCM-CTAB. To further evaluate the different extraction methods, amplification of the second internal transcribed spacer (ITS2) and the chloroplast genome trnL intron was carried out.The results have shown that PCR amplification was successful only with template of DNA extracted by using TCM-CTAB. Moreover, we performed high-throughput 454 sequencing using DNA extracted by TCM-CTAB. Data analysis showed that 3-4 out of 6 prescribed species were detected from LDW samples, while up to 5 contaminating species were detected, suggesting TCM-CTAB method could facilitate follow-up DNA-based examination of TCM preparations.

OBJECTIVETo study the baseline distribution of polymorphisms in the promoter of peroxisome proliferators activated receptor co-activator 1 (PPARGC1A) gene in ethnic Hans from Beijing, and to assess their association with type 2 diabetes (T2DM).

METHODSA 2-stage study was designed. Firstly, the promoter region of PPAGC1A gene was screened with PCRRFLP in a small population (n=216, T2DM/control: 104/112), which was followed by a replication study of a larger group (n=1546, T2DM/control: 732/814). Fasting plasma glucose, insulin, blood lipid, height, weight, waist circumference, and blood pressure were measured in all subjects. Potential association was assessed by logistic regression. Linkage disequilibrium and haplotype analysis were conducted with Haploview software.

RESULTSFive polymorphisms were identified with Sanger sequencing, among which T-2120C (rs3755857), -1999C/G (rs2946386) and -1437T/C (rs2970870) were included for genotypic analysis based on their moderate levels of heterozygosity. No significant difference was found between the two groups. When adjusted for age and gender confounding, we have combined the OR values from population 1 and population 2 based on Mantel-Haenszel fixed model, and recognized a mild contribution of C allele of -1999C/G (rs2946386) to the 1.18-fold risk of T2DM (P=0.03, OR=118). No haplotype was associated with T2DM after permutation correction.

CONCLUSIONThe C allele of -1999C/G ( rs2946386) in the promoter region of the PPARGC1A gene is mildly associated with T2DM. Variations in the promoter region of the PPARGC1A gene seem not to confer the risk of T2DM in our population.

Adult ; Aged ; Asian Continental Ancestry Group ; ethnology ; genetics ; Blood Glucose ; metabolism ; Case-Control Studies ; China ; ethnology ; Diabetes Mellitus, Type 2 ; blood ; ethnology ; genetics ; Ethnic Groups ; genetics ; Female ; Genetic Variation ; Humans ; Lipids ; blood ; Male ; Middle Aged ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha ; Polymorphism, Single Nucleotide ; Promoter Regions, Genetic ; Transcription Factors ; genetics

Previous studies have shown that miRNAs participate in a wide range of biological functions and play important roles in various human diseases including cancer.We found miR-146b-5p significantly dysregulated in human pancreatic cancer cells by qRT-PCR.To demonstrate its function and regulation mechanism,we overexpressed miR-146-5p by transfecting the mimics.Our data showed that miR-146b-5p overexpression significantly reduced the abilities of migration and invasion of MIA PaCa-2 pancreatic cancer cells.Furthermore,we found that matrix metalloproteinase 16 (MMP16) was a downstream target of miR-146b-5p by dual-luciferase reporter assay.Altogether,our findings suggest that miR-146b-5p may be involved in pancreatic cancer cell migration and invasion by targeting MMP16,and miR-146b-5p may be a potential therapeutic target for the pancreatic cancer.