Objective: The current study was designed to examine whether Sijunzi decoction could induce apoptosis in human gastric cancer xenografts in nude mice. Methods: A human gastric adenocarcinoma cell line SGC-7901 was grafted into 30 nude mice. The animals in two experimental groups received either Sijunzi decoction for 40 days or 5-fluorouracil (5-FU) for 6 days beginning 2 days after grafting. The control animals received saline according to an identical schedule. The animals were sacrified 41 days after grafting. The therapeutic effect was determined by measuring of tumor size and tumor weight. Apoptotic indices were examined with terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate fluorescence nick end labeling (TUNEL) method and flow cytometry analysis. Morphological changes were observed by electron microscopy. Results: Comparing with controls, tumor growth was significantly inhibited by Sijunzi decoction ( 34.33％ ， P<0.05) or 5 FU( 50.00％ ， P< 0.05) . Apoptotic index(AI) was significantly higher(16.24％ ± 3.21％ as determined by TUNEL and 11.38％ ± 6.46％ by FACScan) in Sijunzi decoction treatment group than that in the control group (TUNEL:2.63％ ± 1.03％ ,P< 0.01; FACScan:7.15％ ± 1.32％ ,P< 0.05); while there was no significant increase of AI in 5 FU treatment group. Cell shrinkage, nuclear chromatin condensation, formation of membrane blebs and apoptotic bodies were frequently observed on tumor sections in Sijunzi decoction group, while only few apoptotic cells/bodies and necrotic cells were found in the 5-FU treatment group, and in control group cancer cells were in proliferation. Conclusions: The data demonstrated enhanced apoptosis in human gastric cancer xenografts in nude mice after treatment with Sijunzi decoction, suggesting that Sijunzi decoction may be a potential anticancer agent for gastric cancer treatment.

Objective:To present our initial experience with robot-assisted laparoscopic prostatectomy (RALP) for complicated cases.Methods:Clinical and pathological data from 4 complicated prostate cancer cases,who underwent RALP from October to November in 2015,were analyzed retrospectively.All the cases were conducted transurethral plasmakinetic enucleation of prostate and hormonal therapy before RALP.Results:All surgeries were done successfully.The age,baseline prostatic special antigen,clinical tumor stage,operation time and estimated blood loss were 58-70 years,6.04-70.15 ng/mL,T2bT3b,210-360 min and 50-250 mL,respectively.No blood transfusion was needed.All surgical margin were negative.Conclusion:Although previous transurethral surgeries and hormonal therapies may increase the difficulty for operations,RALP is still appropriate for the complicated cases of prostate cancer.

Objective To construct multiple myeloma mucin MUC1-2VNTR gene eukaryotic expressing vector,which provided the basic material for further study of multiple myeloma DNA vaccine.Methods MUC1-2VNTR coding gene as target gene,and a KOZAK sequence was inserted before it.Hind Ⅲ and Xba Ⅰ restriction enzyme site were inserted on both ends.Then the whole sequence was synthesized and cloned into pcDNA3.1/myc-his B vector,and the recombinant vector was identified by restriction enzyme digestion and DNA sequencing.Results Synthesized MUC1-2VNTR gene was 140 bp.Restriction enzyme digestion and DNA sequencing confirmed pcDNA3.1/MUC1-2VNTR/myc-his B including the whole exact translation frame region and MUC1-2VNTR gene.Condnsion The pcDNA3.1/MUC1-2VNTR/myc-his B has been successfully constructed,which provides the basic material for further studies of MUC1 mucin function and multiple myloma DNA vaccine.

Background Aniridia is a rare congenital hereditary eye disease.Studies determined that PAX6 gene mutation is closely associated with congenital aniridia,but the mutation locus are varied.Objective This study was to identify virulence mutation locus of PAX6 gene of a Chinese family pedigree with autosomal dominant aniridia.Methods A Chinese family affected with autosomal dominant aniridia was collected and examined in Affiliated First Hospital of Zhengzhou University in August 2014.Periphery blood of 10 ml was collected from all the families and 100 unrelated health controls.The genomic DNA was extracted by standardized phenol-chloroform method,and all exons and splicing junctions of PAX6 were amplified by PCR.Real-time fluorescence quantitative PCR was performed to examine the relative expression of PAX6 mRNA in patients and normal phenotype families and heahh controls.This study protocol was approved by Ethic Committee of Affiliated First Hospital of Zhengzhou University and complied with Helsinki Declaration.Written informed consent was obtained from subjects or custodian before any medical examination.Results This Chinese family inclued 3 generations and 9 members,with a classic autosomal dominant inheritance mode.Five patients were found,showing the absence of iris and cataract in 3 adult patients and only absence of the iris in 2 children,and other 4 members showed the normal phenotype.A novel heterozygous PAX6 deletion mutation c.796 del G (p.A266 fs) (GenBank ID:KP255960) in exon 10 was exclusively found in all affected individuals but not in any of the unaffected families or unrelated health controls.PAX6 mRNA level in lymphocytes was about 50％ lower in aniridia patients than in unaffected family members,indicating that this mutation caused nonsense-mediated mRNA decay.Conclusions A novel deletion mutation in PAX6 gene results in an abnormal PAX6 COOH-terminal extension in the Chinese aniridia family.This finding expands the mutation spectrum of PAX6 gene.

Aim To screen peptides binding specifically to anti-human PTA1mAbs from a random twelve-peptide phage-disp-layed library. Methods Series of PTA1mAbs(LeoA1、 1B11、 C9、 2D1、 2E9、 2G8、 2H2 and E8)were purified using protein-A affinity column. PTA1mAbs which could bind PTA1-Fc fusion protein binding to its ligand were confirmed by flow cytometry, and then used as target to screen phage library. After three rounds of affinity screening, the peptide sequences of positive phage clones were determined and analyzed. Results LeoA1 could block PTA1-Fc fusion protein binding to its ligand. 13 phages which could bind specifically to LeoA1 were isolated from phage library and further confirmed by ELISA. Conserved motifs were found among the sequences of the peptides. Conclusion It was shown that the conserved motifs were candidafe regions binding to PTA1 ligand,which is important to identify functional epitopes for seeking ligand of PTA1 and further investigation of biological function of PTA1.

OBJECTIVEThe effect of advanced glycation end products (AGEs) on the osteogenic differentiation of humanperiodontal ligament stem cells(hPDLSCs) was discussed. Changes in the Wnt signaling pathway during glycation were also determined.

METHODSIn vitro tissue explanting method was primarily applied. Limiting diluted clone was cultured to obtain hPDLSCs in vitro. The subjects were divided into two groups: the healthy group (N-hPDLSCs) and the AGEs-stimulating group (A-hPDLSCs). Osteoblast mineralization was induced in the experimental groups. The following processes were performed: alizarin red staining; alkaline phosphatase (ALP) staining; real time polymerase chain reaction (real time PCR) for detecting osteogenic genes and Wnt classical pathway-related factors, DKK-1 and β-catenin; Western blot analysis. Bone protein and β-catenin were correlated in the nuclear expression.

RESULTSThe cells were osteogenically induced. ALP staining showed that the N-hPDLSCs displayed the deepest color. Alizarin red staining indicated that the A-hPDLSCs group had less calcified nodules than the N-hPDLSCs group. The real time PCR results suggested that the expression of relative osteogenic genes in A-hPDLSCs was quite low. Statistically significant differences in differentiation were found between groups (P < 0.05). The Western blot result was similar to that of real time PCR. Classical Wnt signaling pathway-related factor β-catenin was higher in A-hPDLSCs than in N-hPDLSCs. By contrast, DKK-1, which is an inhibitor in the Wnt pathway, had a significantly lower expression rate in A-hPDLSCs than in N-hPDLSCs. The Western blot result also showed that β-catenin expression in the nucleoprotein in A-hPDLSCs was notably higher than in N-hPDLSCs.

CONCLUSIONAGEs can inhibit hPDLSCs osteogenic differentiation. AGEs induce changes in the normal periodontal ligament stem cells classical Wnt pathway. Canonical Wnt pathway is reactivated because of AGEs stimulation.

Cell Differentiation ; Glycation End Products, Advanced ; Humans ; In Vitro Techniques ; Osteoblasts ; Osteogenesis ; Periodontal Ligament ; Stem Cells ; Wnt Proteins ; Wnt Signaling Pathway ; beta Catenin

Objective To explore surgical opportunity and management of high-energy Pilon fractures.Methods Twenty-five cases of the high-energy Pilon fractures were treated with anatomic type plate one-staged or ses of type III fractures.8 cases were open fractures and 17 cases were close fractures.8 cases open fractures and 6 cases close fractures were treated with anatomic type plate one-staged.11 cases close fractures with serious soft tissue swell were treated two-staged delayed.The combined fibular fractures were fixed accordingly.All cases were followed up for 32 months after operation in average( range 17 to 76 months).Results According to Mazur's criteria,the surgical result was evaluated as excellent in 10 patients,goed in 11 ,fair in 3 and poor in 1.Postoperative complication ineluded infection in 2 cases,necrosis of skin in 1 case and traumatic arthritis of the ankle joint in 2 cases.Conclusion In order to get a satisfactory result,it is important to choose the right timing of surgery and correct procedure in high-energy Pilon fracture.

AIM:To investigate the inhibitory effect of small interfering RNA ( siRNA) on the expression of protein kinase Cε( PKCε) in human hepatoma SK-Hep-1 cells, and the biological behaviors of the transduced cells , inclu-ding proliferation and invasion , were investigated.METHODS:The cultured SK-Hep-1 cells were divided into 3 groups, including PKCε-siRNA group , negative control ( NC)-siRNA group and control group .MTT assay was used to analyze the proliferation of the SK-Hep-1 cells in the respective groups , while invasion potency was determined by Transwell assay .The protein levels of functional biomarkers such as Ki 67 and matrix metalloproteinase 9 ( MMP-9 ) were measured by Western blotting .The Luciferase reporter gene assay was used to explore the activity of the NF-κB pathway .RESULTS:PKCεex-pression in SK-Hep-1 cells transfected with PKCε-siRNA was significantly down-regulated at both mRNA and protein levels compared with that in the normal SK-Hep-1 cells (P<0.01), with the decreases in the protein levels of Ki67 and MMP-9. The invasion and proliferation of SK-Hep-1 cells were obviously inhibited in PKCε-siRNA group compared with control group (P<0.01).Furthermore, the transcriptional activity of NF-κB was down-regulated when PKCε was effectively in-hibited by PKCε-siRNA (P<0.01).CONCLUSION:Down-regulation of PKCεinhibits the proliferation and invasion of hepatic carcinoma cells , which might be mediated via the NF-κB signaling pathway .

Alpha-glycerophosphate oxidase (alpha-GPO) from Enterococcus casseli flavus was successfully isolated and purified by using polyethylene glycol (PEG)/(NH4)2SO4 aqueous two-phase system (ATPS). The results showed that the chosen PEG/(NH4)2SO4 ATPS could be affected by PEG molecular weight, pH, concentration of PEG and (NH4)2SO4, and inorganic salt as well as additional amount of crude enzyme. After evaluating these influencing factors, the final optimum purification strategy was formed by 16.5% (m/m) PEG2000, 13.2% (m/m) (NH4)2SO4, pH 7.5 and 30% (m/m) additive crude enzyme, respectively. The NaCl was a negative influencing factor which would lead to lower purification fold and activity recovery. These conditions eventually resulted in the activity recovery of 89% (m/m), distribution coefficient of 1.2 and purification fold of 7.0.
Ammonium Sulfate ; chemistry ; Glycerolphosphate Dehydrogenase ; chemistry ; isolation & purification ; Molecular Weight ; Polyethylene Glycols ; chemistry ; Water

OBJECTIVEThis study aims to detect microRNA-17(mir-17) expression on the osteogenic differentiation of advanced glycation end products (AGEs)-stimulated hunman periodontal ligament stem cells (HPDLSCs) and to analyze the influence of these cells on this process.

METHODSHPDLSCs were isolated using limited dilution technique. After osteogenic differentiation occurred, different time points of mir-17 expression in the experimental groups were detected by real time polymerase chain reaction (PCR). The mir-17 overexpression and inhibition were evaluated using cell transfection technique. Differences in gene expressions were detected by real time PCR; differences in protein expressions were analyzed by Western blot.

RESULTSThe mir-17 expression was reduced after osteogenic differentiation occurred at 3, 7, and 14 d compared with that in the control group (P < 0.05). The expression levels of bone sialoprotein (BSP), Runt-related transcription factor-2 (Runx-2)and alkaline phosphatase (ALP) in the experimental groups were lower than those in the mimic control group when mir-17 expression increased. In addition, the protein expression levels of Runx-2 in the experimental groups were lower than those in the control group. The expression levels of BSP, Runx-2 and ALP in the experimental groups were higher than those in the inhibitor control group when mir-17 expression decreased. Likewise, the protein expression levels of Runx-2 in the experimental groups were higher than those in the control group.

CONCLUSIONAGEs inhibit the osteogenic differentiation of HPDLSCs by affecting mir-17 expression.

Alkaline Phosphatase ; Cell Differentiation ; Glycation End Products, Advanced ; Humans ; MicroRNAs ; Osteogenesis ; Periodontal Ligament ; Stem Cells