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Objective. To reveal the effect of enzyme digestion into cell clumps on protein levels of octamer-binding protein 4 (OCT4) and sex determining region Y-box 2 (SOX2) in the human embryonic stem cells (hESCs) and its possible molecular mechanism. Methods. 1 Western blotting was conducted to detect the protein levels of OCT4 and SOX2 in the hESCs treated by enzyme digestion into cell clumps [Dispase or collagenase type IV (COL4)], enzyme digestion into single cells (Accutase or trypsin), or mechanical scraping dissociation. 2 Ethylene glycol tetraacetic acid (EGTA) was used to destruct hESC-hESC interaction, and protein levels of OCT4 and SOX2 in the hESCs treated with EGTA alone or combined with COL4 were detected by Western blotting. 3 Quantitative proteomics analysis based on mass spectrometry was carried out to identify differentially expressed total proteins and phosphorylated proteins in the hESCs treated with COL4 alone or in combination with EGTA. 4 hESCs were pre-treated with proteasome inhibitor (bortezomib) and lysosome inhibitor (chloroquine), respectively, followed by Western blotting and immunofluorescence staining to measure their effects on COL4-induced down regulation of OCT4 and SOX2. 5 Western blotting analysis was conducted to test the effect of COL4 treatment on cofilin as well as Rac1 inhibitors (EHT1864 and EHop-016) treatment on the protein level of SOX2 in the hESCs. Results. 6 COL4 or Dispase treatment led to protein levels decrease of OCT4 and SOX2, while Accutase, trypsin and mechanical dissociation did not elicit such effect. 7 EGTA rescued OCT4 and SOX2 protein levels decrease indcued by COL4 treatment. 8 Quantitative mass spectrometry and functional enrichment analysis suggested that microtubule, actin filament (F-actin) and lysosome-associated proteins might be related to OCT4 and SOX2 protein levels decline in the suspended clumps of hESCs upon COL4 treatment. 9 Chloroquine, instead of bortezomib, could block COL4-induced protein levels decrease of OCT4 and SOX2. 10 The treatment of COL4 or Rac1 inhibitors could downregulate protein levels of cofilin and SOX2. Conclusion. Upon hESC detachment in cell clumps induced by enzyme treatment, the protein levels of SOX2 and OCT4 decline, which may be related to Rac1/cofilin/F-actin signaling pathway and lysosome. [ABSTRACT FROM AUTHOR]

Objective To study the muscular pathological characteristics in riboflavin-responsive lipid storage myopathy before and after treatment. Methods A 10-year follow-up visit was made on a patient with riboflavin - responsive lipid storage myopathy, and the changes of serum enzymes, and both histological and ultrastructural data acquired by general muscular pathology, immunohistochemistry and electron microscope were observed before and after treatment by using levocarnitine. ETFDH gene were detected in the patient and his family. Results The patient presented limb weakness, difficulty in raising head and dysphagia, which were typical clinical features of lipid storage myopathy (LSM). The serum creatine kinase (CK) level and lactic dehydrogenase (LDH) level elevated evidently. EMG showed myogenic abnormality, and muscular pathology revealed numerous lipid droplets deposited in the fibers. ATPase staining showed predominant atrophy of type- fibers and relative increasing of the portion of type- fibers. Modified Gomori trichrome (MGT) staining did not observe ragged red fibers. Immunohistochemical staining showed positive expression of dystrophin. Sultan - staining revealed multiple vacuolated myofibers. ETFDH gene test showed two heterozygous mutations in the patient. After treating with levocarnitine for several years, the patient could live a normal life. The muscular pathological result returned to normal. Conclusions After appropriate therapy, patient with lipid storage myopathy can not only gain complete remission clinically, but also the reversion of lesion pathologically. [ABSTRACT FROM AUTHOR]

Aiming at the synergistic effects between different kinds of cellulose ethers, hydroxyethyl cellulose (HEC) and carboxymethyl cellulose (CMC) mixed systems in aqueous medium were studied. Viscosity and oscillatory measurements were employed to monitor the parameters of apparent viscosity, theology and viscoelasticity of HEC/CMC mixed systems with various mass mixing ratios, so as to analyze the mechanism of interactions between ionic and nonionic cellulose ethers. The synergistic viscosity effects were observed in HEC/CMC mixed systems with thixotropy decreased, and the microscopic structures of mixed systems were further stabilized. The synergistic effects were more obviously revealed as m (HEC) : m (CMC) approaching to 1 : 1. When m(HEC) : m(CMC) became 2 : 1, the sharing electron field effect was formed. A relatively fragile network structure is presented as m(HEC) : m(CMC) is 1 : 2, due to the weaker interactions between HEC and CMC molecules. [ABSTRACT FROM AUTHOR]

Synthesized a novel kind of enteric-coating material-HPMCT. Its structure and application properties such as dissolving ability in different organic solvent, pH sensibility, coating technical properties and dissolution rate in buffer solution(pH=6.86)were analyzed by 1R and UV. The results indicated that ethanol/methylene chloride, acetone/ethanol/water, acetone/ ethanol and ethanol /water were all good solvent systems for the HPMCT; the pH sensibility value range of the HPMCT is 3.8 ∼ 4.0, the HPMCT can dissolve rapidly in buffer solution (pH=6.86), achieving tight, smooth and having metallic brightness coating without plasticizer. Dissolution rate of the coated tablet in buffer solution(pH=6.86) in 20 minutes were 92.60%, which meet the necessity of enteric coating material. [ABSTRACT FROM AUTHOR]

To predict the required minimum collapse distance along impact direction, the scoring principle for the head injury criteria (HIC) is analyzed, and an ideal square wave deceleration curve is used to replace the actual impact deceleration curve. By taking the limit values of head injury criteria specified in GB/T24550-2009 The protection of motor vehicle for pedestrians in the event of a collision as objective control parameters, the minimum collapse distance for engine hood is analyzed based on the square wave deceleration curve. Finally the validity of the method is verified by tests. [ABSTRACT FROM AUTHOR]

A clinical case of ectopic thyroid carcinoma in front of hyoid bone was reported in this paper. The patient, a 17-year-old female, presented with an enlarging neck mass of 1-week history. Physical examination revealed a 3 cm×2 cm neck mass in front of the hyoid bone. Ultrasonographic depicted as a cystic solid mixed echogenic mass with punctate strong echogenicity. CT scan showed a cystic-solid mass in front of the hyoid bone with punctate calcifications. The patient was misdiagnosed as a thyroglossal duct cyst and underwent surgery. The final pathological diagnosis was papillary thyroid carcinoma with cyst formation., Competing Interests: The authors of this article and the planning committee members and staff have no relevant financial relationships with commercial interests to disclose., (Copyright© by the Editorial Department of Journal of Clinical Otorhinolaryngology Head and Neck Surgery.)

Objective: To evaluate the colorectal cancer (CRC) prevention effect of metformin in comparison with that of other T2DM medications from a Markov model perspective., Methods: Literature concerning CRC morbidity of T2DM patients with metformin or other diabetes medications treatment was reviewed in PubMed and Cochrane Library database from September 2010 to December 2016., Inclusion Criteria: (1)enrolled population was adult patients with T2DM but without CRC; (2) any of the parameters applied in our model was reported; (3) randomized clinical trials (RCTs), quasi-randomized trials, prospective or retrospective cohort studies. With CRC morbidity as endpoint, parameters were extracted to construct Markov model to assess CRC morbidity and cumulative tumor-free survival in each group over 11 years' follow-up period. Finally, Monte Carlo analysis was performed to evaluate the influence of parameter instability on the model., Results: Seven literatures were recruited and 10 000 patients were virtually allocated for each arm. In contrast with non-metformin group, T2DM patients treated with metformin had a lower rate of CRC(1.670% vs. 2.146%, P=0.016). Moreover, cumulative tumor-free survival of metformin group was, slightly but significantly, better than that of non-metformin group (10.908 years vs. 10.882 years, P=0.000)., Conclusions: T2DM patients treated with metformin have a lower morbidity of CRC and a better cumulative tumor-free survival than those of non-metformin group. Large scale RCTs are needed to illustrate the role of metformin in the prevention of CRC.

Objective: To investigate the effect of aquaporin 5(AQP5) on proliferation and migration of ectopic endometrial epithelial cells., Methods: AQP5 shRNA interference fragments were designed and transfected into ectopic endometrial epithelial cells stably by lentivirus technology. Fluorescence quantitative RT-PCR and Western blotting were used to detect the AQP5 mRNA and protein expression, respectively. The cell proliferation and migration were determined by using MTT method and Transwell system, respectively. Levels of phosphorylated AKT(p-AKT) and total AKT were examined by Western blotting. The nude mice model of endometriosis was constructed and the endometrial cell nodule formation was observed., Results: AQP5 shRNA transfection inhibited cell proliferation and migration compared with control group (both P<0.05). The activation of AKT in AQP5 shRNA transfected cells was lower than that in control cells (P<0.01). Compared to control group, the endometrial cells nodule formation was suppressed in mice inoculated with AQP5 shRNA-silencing ectopic endometrial epithelial cells., Conclusion: Down-regulation of AQP5 expression can suppress the proliferation and migration of ectopic endometrial epithelial cells and endometrial cell nodule formation in nude mice, in which AKT pathway may be involved.

Objective: To explore the diagnosis, treatment and long-term outcome of late acute rejection (LAR) following adult orthotropic liver transplantation (OLT)., Methods: A total of 398 consecutive adult patients who underwent OLT in Organ Transplant Center, the First Affiliated Hospital, Sun Yat-sen University between January 2007 and December 2012 were reviewed retrospectively. There were 48 patients (12. 1%) developed to LAR, including 43 male patients and 5 female patients, with an average age of (52 ± 13) years(18 - 70 years). The mean body mass index was (22.1 ± 4. 5) kg/m2 (15. 4 - 30. 4 kg/m2). The indications of the liver transplantation recipients included 16 cases of end-staged liver cirrhosis after hepatitis B or C(33. 3%), 14 cases with severe hepatitis (29. 2%), 9 cases of primary liver cancer(18. 5%), 5 cases of alcoholic liver cirrhosis (10. 4%), 1 case with autoimmune liver disease (2. 1%) , the other 3 cases (6. 3%). They were followed up by outpatient service, telephone and other means. Survival curves were generated with the Kaplan-Meier method and Cox proportional hazards modeling was used for predictors of mortality. Statistically significant variables found by single factor regression analysis were put into the Cox proportional hazards regression model of multivariate analysis., Results: The time-to-event was 23. 6 months after OLT which were more common in the first year to the third year post-transplant (26/48,52. 4%). Thirty-five cases were assessed as mild, 11 cases were assessed as moderate, and 2 cases were assessed as severe ,based on the Banff schema. After adjustment to the immunosuppressive regimen, the overall recovery rate reached to 81. 3%. The rate of steroid-resistant acute rejection was 11. 8% (4/34). Inadequate immunosuppression and steroid pulsation were two independent risk factors affecting the prognosis of LAR (P = 0. 008, P = 0. 003, respectively)., Conclusions: LAR is an uncommon complication after OLT. Inadequate immunosuppression and steroid pulsation are the major risk factors for prognosis of LAR. Improving patient compliance and strengthening blood concentration surveillance can increase the patient survival.

Objective: To investigate the clinicopathological features of gastric cancer in Southern China, and provide a base of research and therapy for gastric cancer., Methods: A total of 1 879 cases of gastric cancer with radical gastrectomy from Southern China were collected from August 1994 to July 2012. Analyze and summarize the characters of gender, age, tumor location, WHO histopathologic type and grade, pTNM stage and family history, retrospectively., Results: Among all cases, male to female ratio was 2.08: 1, while female was more than male before 40 years (χ(2) = 77.831, P = 0.000). Cases aged over 60 years had a highest incidence of gastric cancer (46.0%), with predilection of sinus (45.7%), body (26.3%) and cardia (20.1%). The common WHO histopathologic types were tubular or papillary adenocarcinoma (81.5%) and signet ring cell carcinoma (11.0%). Most patients were at III or IV stage on pTNM staging (40.5% and 26.5%)., Conclusions: Gastric cancer in Southern China has a predilection on male, while female is more than male before 40 years. Gastric sinus and adenocarcinoma is most common. Most patients are diagnosed at advanced stages.

Objective: Through construction of a lentiviral expression vector of chemokine receptor 3 (CCR3)RNA interference (RNAi) of mouse, to further study the function of CCR3 gene on eosinophils., Methods: Focused on the CCR3 gene sequences, RNAi target sequences were designed, then the target sequences of Oligo DNA were synthesized and annealed to double stranded DNA, which was subsequently connected to pLVX-shRNA2-m vector digested by MluI, SacI, EcoRI, HindIII, BamHI and Xho I, short hairpin RNA lentiviral vectors were constructed. Short hairpin RNA lentiviral vectors were constructed. 293T cells and eosinophils were transfected by shRNA lentiviral vector, and virus titer was determined. The expression of the CCR3 gene in eosinophils was identified by quantitative-PCR., Results: The lentiviral vector of shRNA-mCCR3-oligonucleotide chain was inserted correctly. Infection efficiency of 293T cells observed under fluorescence microscope was more than 90%, the virus titer was 4×10(8) TU/ml. CCR3 interference rate was 86.7%., Conclusion: A lentiviral vector of CCR3-gene RNAi was constructed successfully by the genetic engineering technology, and it provides a condition for further research in vitro and vivo.

Objective: To establish a method for analyzing the PTEN-induced kinase 1 gene (PINK1) exon copy number and apply it to the analysis of PINK1 gene exon copy number variation (CNV) in patients with autosomal recessive early-onset Parkinsonism (AREP)., Methods: Real-time PCR was used to analyze the exon copy number in 22 probands with AREP from unrelated Chinese Han families and 30 healthy controls., Results: Copy numbers of exons 1-8 of the PINK1 gene were analyzed, and satisfactory reaction conditions and primers for exons of the PINK1 gene were obtained. No exon CNV in the PINK1 gene was detected in this group., Conclusion: An analytical method for PINK1 gene exon copy number was established. The exon CNV in the PINK1 gene was rare in Chinese patients with AREP.

Objective: To investigate the features and diagnosis way of small round cell tumor of nasal sinus and bases of skull., Method: A retrospective analysis of 123 case with small round cell tumor of nasal sinus and bases of skull were carried out in our hospital in past ten year. Clinical, histological, radiological and immunohistochemical characters of these cases were studied., Result: All cases usually complained of nasal obstruction, headache, diplopia, nasal mucus with bleeding, vision or weight loss. Expansible or infiltrative lumps were found in nasal sinus or bases of skull in radiological examination. A lot of small round cells were found in these tumors in histological pathology. At least 5-6 cell, tissue or tumor markers were examined immunohistochemically in most of cases before the final diagnosis were made. In some cases over 20 markers were examined. Five cases were carried out transmission electron microscope examination, special features such as desmosome and myofilament were found., Conclusion: Clinical symptom, physical signs and radiological finding can supply malignant evidences of these tumors. Histological examination can make certain that they are small round cell tumors, but final diagnosis is still hard to make only by these. Immunohistochemical examination of various markers can tell the original characters of the specimen tissues, it is the key for final diagnosis. Transmission electron microscope examination is another helpful way for diagnosis.

In order to investigate Sry regulation network in the development of male embryo, we inhibited the Sry gene expression by RNAi and then examined expression of other sex-related genes. Six genes (Sox9, Wt1, Sf1, Dax1, Gata4 and Amh), which are suggested to be closely related to Sry regulation were studied. Two siRNA expression vectors pSilencer4.1/Sry217 and pSilencer4.1/Sry565 were constructed and injected into gestated mouse through tail vein at 9.5 day of conception (dpc). The inhibition efficiency of Sry and the expression of other six genes were examined in male embryos at 11.5 dpc by RT-PCR and Western-blot. Expressions of the other six genes were analyzed by fluorescence quantity PCR. The results indicated both the pSilencer4.1/Sry217 and pSilencer4.1/Sry565 could inhibit significantly increased after Sry silencing. In contrast, no significant changes were observed in the expression of Sf1, Amh, Gata4, Dax1 and Sox9 when silencing Sry by siRNA. Our results suggested that the Wt1 transcription was regulated by Sry, whereas the Sox9 expression is not directly regulated by Sry in the development of genital ridge.

Background & Objective: The role of oval cells in hepatocarcinogenesis is unclear yet. This study was to explore the correlation of oval cells to hepatocarcinoma through dynamic observation on evolutive characters of oval cells in experimental hepatocarcinogenesis., Methods: Male SD rats were fed with 3o-me-DAB to establish an animal model of experimental hepatocarcinoma. Evolutive characters of oval cells in liver tissue during experimental hepatocarcinogenesis was dynamically observed with routine HE staining, Alcian blue staining, and immunohistochemistry., Results: Oval cells (OV-6-positive) appeared sparsely around the portal tract in the 4th week of tumor-induction. In the 8th and the 14th weeks, OV-6-positive cells were increased gradually and expanded into hepatic lobules; the hepatic tissue was divided as pseudo-lobule-like. Till the 17th and the 24th weeks, carcinoma foci were formed, meanwhile, the total amount of oval cells were decreased, and OV-6-positive cells were observed in carcinoma foci. On Alcian blue-stained preparations, two distinct histologocal types of carcinoma foci could be seen: cholangioepithelial carcinoma foci were positive and hepatocellular carcinoma foci were negative., Conclusion: Oval cells, as intrahepatic stem cells, might play an important role in hepatocarcinogenesis.

Objective: To transfect a short hairpin RNA (shRNA) against survivin gene into human T lymphoblastic leukemia cell line Jurkat, and to explore the effects on apoptosis and proliferation of transfected cells., Methods: The survivin-shRNA expression vector were constructed and transfected into Jurkat cells. Expression of survivin mRNA and protein were assessed by RT-PCR and Western blot analysis respectively. Apoptosis index of transfected Jurkat cells was quantified by flow cytometry. The potential of cell proliferation was described by cell growth curves., Results: In survivin-shRNA transfected Jurkat cells, survivin mRNA levels were significantly reduced by 66.67% ( transient transfection) and 60.69% ( stable transfection) respectively, compared with that in control-shRNA treated group and PBS treated group (P < 0.05); and the levels of survivin protein were significantly reduced by 63.41% (transient transfection) and 60.18% (stable transfection), compared with that in the two control groups (P < 0.05). Apoptosis index was significantly increased during both transient and stable transfection, respectively [(22. 41 +/- 2.83)% and (20.73 +/- 2.56)% (P < 0.05)]. Survivin-shRNA also inhibited the proliferation of Jurkat cells., Conclusions: Vector-based survivin-shRNA can effectively reduce the expression of survivin gene, induce apoptosis