Association of Ureaplasma urealyticum colonization with development of bronchopulmonary dysplasia: a systemic review and meta-analysis.

There is controversy regarding the roles of Ureaplasma urealyticum (U. urealyticum) colonization in the development of bronchopulmonary dysplasia (BPD). This study explored the association between U. urealyticum and bronchopulmonary dysplasia at 36 weeks post-menstrual age (BPD36). Studies published before December 31, 2013 were searched from Medline, Embase, Ovid, Web of Science, and Cochrane databases, with the terms "Ureaplasma urealyticum", "chronic lung disease", or "BPD36" used, and English language as a limit. The association between U. urealyticum colonization and BPD36 was analyzed with RevMan 4.2.10 software, using the odds ratio (OR) and relative risk (RR) for dichotomous variables. Out of the enrolled 81 studies, 11 investigated the BPD36 in total 1193 infants. Pooled studies showed no association between U. urealyticum colonization and subsequent development of BPD36, with the OR and RR being 1.03 (95% CI=0.78-1.37; P=0.84) and 1.01 (95% CI= 0.88-1.16, P=0.84), respectively. These findings indicated no association between U. urealyticum colonization and the development of BPD36.

Over-expression of testis-specific expressed gene 1 attenuates the proliferation and induces apoptosis of GC-1spg cells.

The effects of over-expression of testis-specific expressed gene 1 (TSEG-1) on the viability and apoptosis of cultured spermatogonial GC-1spg cells were investigated, and the immortal spermatogonial cell line GC-1spg (CRL-2053™) was obtained as the cell model in order to explore the function of TSEG-1. We transfected the eukaryotic vector of TSEG-1, named as pEGFP-TSEG-1 into cultured spermatogonial GC-1spg cells. Over-expression of TSEG-1 inhibited the proliferation of GC-1spg cells, and arrested cell cycle slightly at G0/G1 phase. Transfection of TSEG-1 attenuated the transcript levels of Ki-67, PCNA and cyclin D1. In addition, over-expression of TSEG-1 induced early and late apoptosis, and reduced the mitochondrial membrane potential of GC-1spg cells. Moreover, transfection of TSEG-1 significantly enhanced the ratio of Bax/Bcl-2 and transcript levels of caspase 9, and decreased the expression of Fas and caspase 8 in GC-1spg cells. These results indicated over-expression of TSEG-1 suppresses the proliferation and induces the apoptosis of GC-1spg cells, which establishes a basis for further study on the function of TSEG-1.

Laparoscopic choledochal cyst excision, hepaticojejunostomy, and extracorporeal Roux-en-Y anastomosis: a technical skill and intermediate-term report in 62 cases.

BACKGROUND: This study aimed to present the authors' technique and the intermediate-term outcome for laparoscopic choledochal cyst excision with Roux-en-Y hepatoenterostomy. METHODS: This retrospective study investigated 62 children (39 girls and 23 boys) who had undergone laparoscopic resection of choledochal cyst. The average age of the children was 2.3 years. The retrospective data and the following investigations about type of choledochal cyst, surgical technique, conversion rate, morbidity, and mortality were analyzed. RESULTS: Of the 62 patients, 43 (69.4%) showed type 1a choledochal cysts, 16 (25.8%) showed type 1c, 2 (3.2%) showed type 4a, and 1 (1.6%) showed type 4b. Total cyst excision could be performed for 51 of the patients (82.3%). The large cysts were opened on the front wall, then divided circumferentially in 29 cases. The small cysts did not need to be opened before excision in 22 cases. For 11 patients (17.7%), Lilly's (Surg Gynecol Obstet 146:254-256, 1978) technique was adopted, and for 5 patients with a huge cyst, the duodenum together with the head of the pancreas had to be mobilized for visualization of the cyst's lower limit. The hepatic duct was excised, and plastic operation of bile duct was performed for two patients. The mean operative time was 226±41.2 min. Eight patients needed blood transfusion, and conversion was required for one patient. The mean hospital stay was 8±1.5 days, and the mean follow-up period was 38 months. The overall morbidity rate was 8.2% (5/61) including bile leakage (n=1), adhesive small bowel obstruction (n=1), intestinal necrosis (n=1), and cholangitis (n=1). Inflammatory edema anastomotic narrowing occurred in one patient. None of the patients needed surgery due to anastomotic stricture. CONCLUSIONS: Laparoscopic choledochal cyst excision, hepaticojejunostomy, and extracorporeal Roux-en-Y anastomosis can be safely and quickly performed for children, with satisfactory intermediate-term results. Extracorporeal Roux-en-Y anastomosis could shorten the operative time.

Helicobacter pylori Infection Facilitates the Expression of Resistin-like Molecule Beta in Gastric Carcinoma and Precursor Lesions.

Helicobacter pylori (H. pylori) was reported to be associated with gastric carcinogenesis. Resistin-like molecule beta (RELMß), a recently described goblet cell-specific protein, was demonstrated to aberrantly express in gastric cancer and correlated with its clinicopathological features. This study aimed to examine the association between H. pylori and RELMß expression in gastric carcinoma and precursor lesions. H. pylori infection and RELMß expression were immunohistochemically evaluated in gastric biopsies from 230 patients. The biopsies consisted of normal gastric mucosa (n=20), mucosa with chronic gastritis (n=41), intestinal metaplasia (n=42), dysplasia (n=31), intestinal-type adenocarcinoma (n=56), and diffuse-type adenocarcinoma (n=40). RELMß expression was measured in gastric biopsies after H. pylori eradication therapy in a subgroup of 32 patients. Cultured gastric cancer cell line SGC-7901 was infected with H. pylori strains, and RELMß expression was detected by reverse transcription PCR, real-time PCR and Western blotting. Higher RELMß immunoreactivity was observed in H. pylori-positive intestinal metaplasia (P=0.003), dysplasia (P=0.032), intestinal-type (P=0.037) and diffuse-type adenocarcinomas (P=0.001) than in H. pylori-negative specimens. Expression rates of RELMß in dysplasia (P=0.005), intestinal-type adenocarcinoma (P<0.001), and diffuse-type adenocarcinoma (P=0.001) were significantly correlated with the grade of H. pylori density. In addition, H. pylori eradication reduced the RELMß intensity in intestinal metaplasia (P=0.001). Infection of gastric cancer SGC-7901 cells with cag pathogenicity island (PAI)-positive H. pylori TN2, but not with its PAI totally deleted mutant (TN2-ΔPAI) for 4-8 h, resulted in enhanced protein and transcript levels of RELMß (P<0.05). In summary, our study suggested that H. pylori infection facilitated the expression of RELMß in gastric garcinoma and precursor lesions.

Enhanced expression of resistin-like molecule beta in human colon cancer and its clinical significance.

Previous studies have indicated that resistin-like molecule beta (RELM beta), an intestinal goblet cell-specific protein, is markedly increased in the intestinal tumors of min mice and over-expressed in a human colon cancer cell line. We hypothesized that RELM beta might be enhanced in human colon cancer. The aim of this study was to examine the clinical importance of RELM beta expression in colon cancer patients and to correlate its expression with various clinicopathological parameters, upstream regulatory molecule expression, tumor proliferative capacity, and patients' survival. Of the 80 colon cancer patients studied, 65 (81.25%) tested positive for RELM beta, mainly in the cytoplasm of colon mucosa. Contrasting sharply with the strongly RELM beta-positive tumors, normal colon mucous membrane was negative or weakly positive. RELM beta positivity in colon cancer was correlated with histological grade of differentiation and lymph node metastasis, but not with age, gender, tumor location and size, tumor infiltration, Dukes' stage, liver metastasis, and venous invasion. RELM beta expression was significantly correlated with the expression of transcription factor CDX-2 (P < 0.01) but not with that of proliferative index Ki-67 (P > 0.05). The mean postoperative survival time (2.76 years) of RELM beta-positive patients was significantly longer than that (1.26 years) of RELM beta-negative patients (P = 0.032). These findings support evidence of the enhanced RELM beta expression in colon cancer patients and suggest that further investigation is warranted to explore the role of RELM beta in colon cancer.

Gastric carcinoma with osteoclast-like giant cells: a case report and review of the literature.

Gastric carcinoma with osteoclast-like giant cells (OGCs) is an extremely rare tumor. So far, only six cases have been reported in the literature. Here we report an additional case of this tumor in a Chinese 78-year-old man presented with abdominal pain, vomiting, and hematemesis. Physical examination and gastroscopy revealed a tumor in the gastric antrum. The biopsy and pathological findings indicated a gastric adenocarcinoma with OGCs, which were present in both the tumor and the metastatic lymph nodes. Further immunohistochemical staining indicated that OGCs were reactive with CD68, CD45, and vimentin protein, but not with pancytokeratin, carcinoembryonic antigen, or epithelial membrane antigen, suggesting the monocytic/histiocytic derivation of these OGCs. In situ hybridization for Epstein-Barr virus showed no nuclear positivity in either adenocarcinoma or OGCs. Postoperative follow-up showed that the patient had survived for at least 6 months without recurrence. Further investigation is warranted to clearly define the prognostic significance of OGCs in gastric carcinoma.

[Cloning and expression of a novel mouse testis gene TSEG-2].

OBJECTIVE: To clone the mouse testis specific gene TSEG-2 via a bioinformatic approach. METHODS: The expressed sequence tags (EST) in the normal mouse testis were obtained from the online EST database ZooDDD. Their highly homologous EST sequences were retrieved through the dbEST database to construct contigs and spliced with the biomedical software Biolign. The corresponding exons and introns within the genome sequences were predicted with the software GeneScan. Primers were designed according to the open reading frame. RT-PCR was applied in cloning the cDNA of the novel gene from the mouse testis tissue and analyzing its expression patterns in the undescended testis and various organ tissues as well as in different developmental stages of the mouse testis. The sequencing results of TSEG-2 underwent bioinformatic analyses. RESULTS: The novel mouse testis gene TSEG-2 was successfully cloned, with full-length sequence of 451 bp. The open reading frame was 267 bp, coding a protein of 88 amino acid residues, and demonstrated to be correct by RT-PCR. The expression of TSEG-2 was high in the mouse testis, regular in the testis cDNA samples of different postnatal days, and down-regulated in the cryptorchidism model. No obvious homology with other mouse cDNA was found for TSEG-2. The GenBank accession number EU079025 was achieved. Function prediction showed that mouse TSEG-2 was probably a soluble non-secretary protein located at chromosome 15qE3, or a nucleoprotein with 2 phosphorylation sites of protein kinase C (PKC) and 1 of casein kinase II (CK2). CONCLUSION: A novel mouse testis specific gene TSEG-2 was successfully cloned, which could be down-regulated by cryptorchidism-inducible 17-beta estradiol. This has prepared the ground for further researches on the biological function and expression regulation of TSEG-2.

Natural jasmonates of different structures suppress the growth of human neuroblastoma cell line SH-SY5Y and its mechanisms.

AIM: Recent evidence has indicated that members of natural jasmonates, a family of plant stress hormones, exhibit anticancer activity. The current study was undertaken to investigate the effects of jasmonates on the in vitro growth of human neuroblastomas, one of the most common solid tumors in children. METHODS: Cellular proliferation was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide colorimetry and colony formation assay. Apoptosis was detected by Hoechst 33258 staining and flow cytometry. Western blotting was applied to assay gene expression. RESULTS: The administration of natural jasmonates, methyl jasmonate, cis-jasmone, and jasmonic acid to cultured neuroblastoma cell line SH-SY5Y, resulted in a decrease of cell proliferation in a doseand time-dependent manner. However, the in vitro growth of cultured human embryonic kidney (HEK) cell line HEK 293 was not affected by jasmonates. The cell cycles of jasmonate-treated SH-SY5Y cells were arrested at the G2/M phase. The incubation of SH-SY5Y cells with jasmonates resulted in characteristic changes of apoptosis. The anticancer activities of natural jasmonates on SH-SY5Y cells are as follows: methyl jasmonate>cis-jasmone>jasmonic acid. In addition, the expressions of proliferating cell nuclear antigen and N-myc were downregulated by methyl jasmonate. Moreover, methyl jasmonate decreased the expression of the Xlinked inhibitor of apoptosis protein and survivin, critical members of inhibitors of the apoptosis protein family, in SH-SY5Y cells. CONCLUSION: Jasmonates suppress the growth of human neuroblastoma cell line SH-SY5Y via inhibiting cell proliferation and inducing apoptosis, which lays the groundwork for further investigation into the anticancer activities and its mechanisms of natural jasmonates on human neuroblastomas.

[Methyl jasmonate induces apoptosis of human neuroblastoma cell line BE(2) -C and its mechanism].

This study is to explore the inhibitory effect of methyl jasmonate on cell proliferation and expression of XIAP and survivin of human neuroblastoma cell line BE(2)-C. After cultivation of 1 - 2 mmol x L(-1) jasmonates with BE (2) -C cells for 6 - 24 h, the growth inhibiting rates of BE (2) -C cells were studied by MTT colorimetry. Cell proliferation was detected by colony formation assay. Cell cycle phases were assayed by propidium iodide staining flow cytometery. Cell apoptosis was inspected by acridine orange-ethidium bromide fluorescent staining, Hoechst 33258 fluorescent staining, and Annexin V-FITC and propidium iodide staining flow cytometry. Expressions of cyclin D1, XIAP and survivin were determined by RT-PCR and real-time RT-PCR. Methyl jasmonate inhibited the growth of BE(2)-C cells in a dose- and time-dependent manner. After addition of 1, 1.5 and 2 mmol x L(-1) of methyl jasmonate for 24 h, the inhibiting rates of cell growth reached 20.6% - 85.5% (P < 0.01), and the IC50 was 1.35 mmol x L(-1). The cell cycles were arrested at S phase. A part of cells presented the characteristic morphological changes of apoptosis. The early apoptotic rates were 13.51%, 17.32%, 24.59% (P < 0.01) and the cell death rates were 29.36% , 54.73% , 75.52% (P < 0.01), respectively. The expression of XIAP and survivin mRNA were downregulated by 18.5% - 68.9% , 22.4% - 48.7% (P < 0.05), respectively, without change in that of cyclin D1. The results indicated that methyl jasmonate could significantly inhibit the growth of BE(2) -C cells through inducing cell cycle arrest and apoptosis, downregulating the expression of XIAP and survivin might be one of its molecular mechanisms of action.

[Strategies for cloning of novel genes based on EST].

Expressed sequence tags (EST) are short, randomly selected single-pass nucleotide sequence reads derived from cDNA libraries and represent a small part of a gene. Along with the development of bioinformatics and genetic localization, EST has already become a powerful tool for mapping, cloning and expression profiling of genes. Recently, because of the fast distension of EST databases, application of EST in gene mapping and cloning leads to revolutionary change in the strategies for cloning of novel genes. Despite of some insufficiencies, it has been proved that EST could promote the discovery and research of novel genes. In this article, an introduction about EST, especially EST-based strategies for cloning of novel genes will be given in details.

[Cloning and sequence analysis of TSEG-1, a novel gene specifically expressed in mouse testis].

The expressed sequence tags (ESTs) of normal mouse testis were obtained from online EST database ZooDDD. Their highly homologous EST sequences were found through the dbEST database to construct contigs, and spliced by the biomedical software Biolign. The corresponding exons and introns within genome sequences were predicted by software GeneScan. According to the open reading frame, the primers were designed. RT-PCR was applied in the cloning of novel gene from mouse testis and analyzing its expression pattern in various mouse tissues. The bioinformatics analysis on the sequencing results of TSEG-1 was conducted. Results indicated that a novel gene TSEG-1 was cloned from 1 668-2 011 kb of mouse X chromosome, with full-length sequence of 510 bp. The open reading frame (ORF) is 336 bp in length and en-codes a deduced amino acid sequence of 111 residues. The molecular weight of TSEG-1 protein is 12.84258 kDa, and its pI is 11.4000. RT-PCR demonstrated the correctness of its ORF. TSEG-1 was distinctively expressed in testis, but not in other tissues of mouse. No obvious homology with other mouse cDNA was found for TSEG-1. The GenBank accession number EU079024 was achieved. It was predicted that TSEG-1 is a kind of transmembrane protein, and the transmembrane domain is from 41 amino acid residue to 61 amino acid residue. Blastn analysis revealed its high homology to human testis-specific gene H2AX. Computational prediction of the 5'-untranslated region of TSEG-1 gene revealed a 680 bp-length promoter region. There are four antigen binding sites and two phosphorylation sites of specific protein kinase in TSEG-1 protein, with subcellular localization in mitochondria. The cloning of mouse testis specific gene TSEG-1 laid a foundation for subsequent research of its biological function and expression regulation.

Growth inhibition and apoptosis inducing mechanisms of curcumin on human ovarian cancer cell line A2780.

OBJECTIVE: To explore the growth inhibition effects and apoptosis inducing mechanisms of curcumin on human ovarian cancer cell line A2780. METHODS: After treatment with 10 - 50 micromol/L curcumin for 6 - 24 h, the growth activity of A2780 cancer cells were studied by [4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide (MTT) colorimetry. Cellular apoptosis was inspected by flow cytometery and acridine orange-ethidium bromide fluorescent staining methods. The fragmentation of cellular chromosome DNA was detected by DNA ladder, the ultrastructural change was observed under a transmission electron microscope, and the protein levels of nuclear factor-kappa B (NF-kappaB, P65) and cysteinyl aspartate specific protease-3 (Caspase-3) in ovarian cancer cells were measured by immunohistochemistry. RESULTS: After treatment with various concentrations of curcumin, the growth inhibition rates of cancer cells reached 62.05% - 89.24%, with sub-G(1) peaks appearing on histogram. Part of the cancer cells showed characteristic morphological changes of apoptosis under fluorescence and electron microscopes, and the rate of apoptosis was 21.5% - 33.5%. The protein expression of NF-kappaB was decreased, while that of Caspase-3 was increased in a time-dependent manner. CONCLUSION: Curcumin could significantly inhibit the growth of human ovarian cancer cells; inducing apoptosis through up-regulating Caspase-3 and down-regulating gene expression of NF-kappaB is probably one of its molecular mechanisms.

Downregulation of XIAP expression induces apoptosis and enhances chemotherapeutic sensitivity in human gastric cancer cells.

X-linked inhibitor of apoptosis (XIAP) is the most potent member of the inhibitor of apoptosis protein (IAP) gene family in terms of its ability to inhibit caspases and suppress apoptosis. Recent evidence has suggested that XIAP is a key determinant in chemoresistance of cancer cells. To explore a novel approach for ameliorating chemotherapy of gastric cancer, the antisense expression vector for the XIAP gene was constructed and transferred into gastric cancer cell lines, MKN-45 (wild-type p53) and MKN-28 (mutant-type p53). This transfer resulted in significant downregulation of XIAP expression, decreased in vitro cell viabilities, and induced apoptosis. In transferred cells, inactive caspase-3 precursors were cleaved into the active subunits (p20 and p17) during apoptosis induced by downregulation of XIAP. The inhibitory effects of cisplatin and mitomycin C on the growth of XIAP downregulated cancer cells were significantly enhanced. In addition, this process occurred only in wild-type p53 (MKN-45), but not in mutant-type p53 (MKN-28) gastric cancer cells. The data presented suggest that downregulation of XIAP via antisense RNA can lead to apoptosis of gastric cancer cells in vitro, correlating with cellular p53 status and activation of caspase-3. This finding could lead to a potential strategy for improving the efficiency of therapies for gastric cancer.

Stable transfection of extrinsic Smac gene enhances apoptosis-inducing effects of chemotherapeutic drugs on gastric cancer cells.

AIM: To explore the feasibility of enhancing apoptosis-inducing effects of chemotherapeutic drugs on human gastric cancer cells by stable transfection of extrinsic Smac gene. METHODS: After Smac gene was transferred into gastric cancer cell line MKN-45, subclone cells were obtained by persistent G418 selection. Cellular Smac gene expression was determined by RT-PCR and Western blotting. After treatment with mitomycin (MMC) as an apoptotic inducer, in vitro cell growth activities were investigated by trypan blue-staining method and MTT colorimetry. Cell apoptosis and its rates were determined by electronic microscopy, annexin V-FITC and propidium iodide staining flow cytometry. Cellular caspase-3 protein expression and its activities were assayed by Western blotting and colorimetry. RESULTS: When compared with MKN-45 cells, the selected subclone cell line MKN-45/Smac had significantly higher Smac mRNA (3.12+/-0.21 vs 0.82+/-0.14, t = 7.52, P<0.01) and protein levels (4.02+/-0.24 vs 0.98+/-0.11, t = 8.32, P<0.01). After treatment with 10 microg/mL MMC for 6-24 h, growth inhibition rate of MKN-45/Smac (15.8+/-1.2-54.8+/-2.9%) was significantly higher than that of MKN-45 (5.8+/-0.4- 24.0+/-1.5%, t = 6.42, P<0.01). Partial MKN-45/Smac cancer cells presented characteristic morphological changes of apoptosis under the electronic microscope with an apoptosis rate of 36.4+/-2.1%, which was significantly higher than that of MKN-45 (15.2+/-0.8%, t = 9.25, P<0.01). Compared with MKN-45, caspase-3 expression levels in MKN-45/Smac were improved significantly (3.39+/-0.42 vs 0.96+/-0.14, t = 8.63, P<0.01), while its activities were 3.25 times as many as those of MKN-45 (0.364+/-0.010 vs 0.112+/-0.007, t = 6.34, P<0.01). CONCLUSION: Stable transfection of extrinsic Smac gene and its over-expression in gastric cancer cell line can significantly enhance cellular caspase-3 expression and activities, ameliorate apoptosis-inducing effects of mitomycin C on cancer cells, which is a novel strategy to improve chemotherapeutic effects on gastric cancer.

Selection of optimal antisense accessible sites of survivin and its application in treatment of gastric cancer.

AIM: To select the optimal antisense accessible sites of survivin, a highly expressed gene in tumor tissues, in order to explore a novel approach to improve biological therapy of gastric cancer. METHODS: The 20 mer random oligonucleotide library was synthesized, hybridized with in vitro transcribed total survivin cRNA, then digested by RNase H. After primer extension and autoradiography, the antisense accessible sites (AAS) of survivin were selected. Then RNADraw software was used to analyze and choose the AAS with obvious stem-loop structures, according to which the complementary antisense oligonucleotides (AS-ODNs) were synthesized and transferred into survivin highly- expressing gastric cancer cell line MKN-45. Survivin expression was detected by RT-PCR and Western Blotting. Cellular growth activities were assayed by tetrazolium bromide (MTT) colorimetry. Cellular ultrastructure was observed by electronic microscopy, while apoptosis was detected by annexin V-FITC and propidium iodide staining flow cytometry. RESULTS: Thirteen AAS of survivin were selected in vitro. Four AAS with stem-loop structures were chosen, locating at 207-226 bp, 187-206 bp, 126-145 bp and 44-63 bp of survivin cDNA respectively. When compared with non-tranfection controls, their corresponding AS-ODNs (AS-ODN(1), AS-ODN(2), AS-ODN(3) and AS-ODN(4)) could reduce Survivin mRNA levels in MKN-45 cells by 54.3+/-1.1% (t = 6.12, P<0.01), 86.1+/-1.0% (t = 5.27, P<0.01), 32.2+/-1.3% (t = 7.34, P<0.01) and 56.2+/-0.9% (t = 6.45, P<0.01) respectively, while survivin protein levels were decreased by 42.2+/-2.5% (t = 6.26, P<0.01), 75.4+/-3.1% (t = 7.11, P<0.01), 28.3+/-2.0% (t = 6.04, P<0.01) and 45.8+/-1.2% (t = 6.38, P<0.01) respectively. After transfection with 600 nmol/L AS-ODN1-AS-ODN(4) for 24 h, cell growth was inhibited by 28.12+/-1.54% (t = 7.62, P<0.01), 38.42+/-3.12% (t = 7.75, P<0.01), 21.46+/-2.63% (t = 5.94, P<0.01) and 32.12+/-1.77% (t = 6.17, P<0.01) respectively. Partial cancer cells presented the characteristic morphological changes of apoptosis, with apoptotic rates being 19.31+/-1.16% (t = 7.16, P<0.01), 29.24+/-1.94% (t = 8.15, P<0.01), 11.87+/-0.68% (t = 6.68, P<0.01) and 21.68+/-2.14% (t = 7.53, P<0.01) respectively. CONCLUSION: The AAS of survivin could be effectively selected in vitro by random oligonucleotide library/RNase H cleavage method combined with computer software analysis, this has important reference values for further studying survivin-targeted therapy strategies for gastric cancer.

Preparation of arsenic trioxide-loaded albuminutes immuno-nanospheres and its specific killing effect on bladder cancer cell in vitro.

BACKGROUND: Recently, arsenic trioxide (As2O3) was considered as a novel anti-tumor agent. However, it showed severe toxicity effect on normal tissue at the same time. To improve its therapeutic efficacy and decrease its toxicity,we prepared arsenic trioxide-loaded albuminutes immuno-nanospheres [As2O3-(HAS-NS)-BDI-1] targeted with monoclonal antibody (McAb) BDI-1 and tested its specific killing effect against bladder cancer cell. METHODS: As2O3-HAS-NS was prepared by chemical cross-linking method. Monoclonal antibody BDI-1 was purified with ammonium sulphate saltingout and chromatography. Albuminutes microspheres were conjugated with McAb by SPDP cross-linking method. Concentration of As in As2O3-(HAS-NS)-BDI-1 and As2O3-HAS-NS was measured by atomic fluometry method. As2O3-(HAS-NS)-BDI-1 and its activity were detected by SDS-PAGE reduction electrophoresis, indirect immunofluorescence test, light microscope and scanning electron microscope observation. Acridine orange staining and tritiated thymidine (3H-TdR) incorporation tests were used to indicate specific killing activity of As2O3-(HAS-NS)-BDI-1 in vitro. RESULTS: In As2O3-(HAS-NS)-BDI-1 groups, we saw two protein bands in SDS-PAGE reduction electrophoresis. Albuminutes immuno-nanospheres were rounded with clear green fluorescence by immunofluorescence test. Under microscope, we observed that BIU-87 cells were covered with the As2O3-(HAS-NS)-BDI-1 and that As2O3-(HAS-NS)-BDI-1 moved with the BIU-87 cells. The albuminutes immuno-nanospheres were tightly junctioned with the BIU-87 cells. Specific killing activity of As2O3-(HAS-NS)-BDI-1 on bladder tumor cells was observed by acridine orange staining and 3H-TdR incorporation assays. CONCLUSIONS: As2O3-(HAS-NS)-BDI-1 might bind specifically against BIU-87 cells, thus leading to high activity of killing bladder tumor cells.

[Regulatory effects on apoptotic activities of gastric cancer cell line by over-expression of Smac gene].

OBJECTIVE: To investigate the effects of overexpression of second mitochondria-derived activator of caspases (Smac) gene on apoptosis of gastric cancer cells. METHODS: Under the induction of liposome, MKN-45 cells were transfected by Smac gene and incubated with G418 for subclone selection. Reverse transcriptase-polymerase chain reaction and Western blot were used to determine cellular Smac gene expression. After induction of apoptosis by mitomycin (MMC), cell viabilities were analyzed using trypan blue stain. Apoptosis was measured by electronic microscopy, acridine orange-ethidium bromide fluorescent staining and in situ terminally labelled transferase technique (TUNEL). Western blot and colorimetry were used to assess cellular caspase-3 expression and its activity. RESULTS: The Smac mRNA and protein levels in MKN-45/Smac subclone cells (subclone consistently expressing Smac gene) were significantly higher than those in MKN-45 (P < 0.01). When compared with those in MKN-45, cell viabilities of MKN-45/Smac were reduced by 10.0% to 30.8% (P < 0.01), after treatment with 10 microg/ml MMC for 6 to 24 hours. Some of the MKN-45/Smac cells showed characteristic morphologic changes of apoptosis, their apoptotic rate being increased by 21.2% (P < 0.01). After treatment with MMC, caspase-3 expression and its activity in MKN-45/Smac cells were significantly higher than those in MKN-45 (P < 0.01). CONCLUSIONS: Overexpression of Smac in gastric cancer cell line significantly improves expression and activity levels of caspase-3 after induction by MMC. Such apoptosis-inducing effect establishes a novel strategy for regulating the apoptosis activity of gastric cancer.

A Modified Approach for Inguinal Hernias in Children: Hybrid Single-Incision Laparoscopic Intraperitoneal Ligation.

PURPOSE: To report a laparoscopic approach for pediatric inguinal hernia repair using a hybrid single-incision laparoscopic (H-SIL) technique and its clinical outcomes. MATERIALS AND METHODS: A retrospective study was carried out in inguinal hernia cases treated with the new H-SIL approach using intracorporeal jumping purse-string sutures. The operative time, length of postoperative hospital stay, efficiency of the operation, and complications were analyzed. RESULTS: In total, 157 inguinal high ligations were performed in 106 children (89 boys, 17 girls). The median age was 1.5 years (range, 25 days-11.6 years). The mean operative time was 15.8±3.4 minutes for the single-side procedure and 20.3±2.5 minutes for bilateral procedures. The mean postoperative hospital stay was 0.99±0.52 (range, 0.25-3 days). No postoperative bleeding, abdominal wall emphysema, abdominal viscera injury, or scrotal edema was found, and there were no known cases of postoperative testicular atrophy or hypotrophy. Ninety-three percent of the patients became fully mobile on the first postsurgical day. The median follow-up period was 17 months (range, 9-21 months), with no recurrence, no visible scars on the abdominal wall, and no foreign body felt in the inguinal region. CONCLUSIONS: This H-SIL approach is a safe and efficient method for pediatric inguinal hernia repair. The maneuverability is the same as that in the triport laparoscopic technique, and the cosmetic results are similar to those of single-port laparoscopic surgery.

BAK overexpression mediates p53-independent apoptosis inducing effects on human gastric cancer cells.

BACKGROUND: BAK (Bcl-2 homologous antagonist/killer) is a novel pro-apoptotic gene of the Bcl-2 family. It has been reported that gastric tumors have reduced BAK levels when compared with the normal mucosa. Moreover, mutations of the BAK gene have been identified in human gastrointestinal cancers, suggesting that a perturbation of BAK-mediated apoptosis may contribute to the pathogenesis of gastric cancer. In this study, we explored the therapeutic effects of gene transfer mediated elevations in BAK expression on human gastric cancer cells in vitro. METHODS: Eukaryotic expression vector for the BAK gene was constructed and transferred into gastric cancer cell lines, MKN-45 (wild-type p53) and MKN-28 (mutant-type p53). RT-PCR and Western Blotting detected cellular BAK gene expression. Cell growth activities were detected by MTT colorimetry and flow cytometry, while apoptosis was assayed by electronic microscopy and TUNEL. Western Blotting and colorimetry investigated cellular caspase-3 activities. RESULTS: BAK gene transfer could result in significant BAK overexpression, decreased in vitro growth, cell cycle G0/G1 arrest, and induced apoptosis in gastric cancer cells. In transferred cells, inactive caspase-3 precursor was cleaved into the active subunits p20 and p17, during BAK overexpression-induced apoptosis. In addition, this process occurred equally well in p53 wild-type (MKN-45), or in p53 mutant-type (MKN-28) gastric cancer cells. CONCLUSIONS: The data presented suggests that overexpression of the BAK gene can lead to apoptosis of gastric cancer cells in vitro, which does not appear to be dependent on p53 status. The action mechanism of BAK mediated apoptosis correlates with activation of caspase-3. This could be served as a potential strategy for further development of gastric cancer therapies.

The roles of microRNAs in neuroblastoma.

BACKGROUND: Neuroblastoma (NB) is the most common extracranial solid tumor in childhood and displays remarkable heterogeneity in clinical behaviors, ranging from spontaneous regression to rapid progression or resistance to multimodal treatment. Recent evidence has shown that microRNAs (miRNAs), a class of small non-coding RNAs, are involved in tumor development and progression. This article aimed to review recent advances in investigating the roles of miRNAs in NB. METHODS: We searched the PubMed/MEDLINE database for articles about the expression profile, functions and target genes of miRNAs in NB. RESULTS: We reviewed the most recent evidence regarding the functional roles of oncogenic and tumor suppressive miRNAs in NB and application of novel miRNA-based methods for diagnostic, prognostic and therapeutic purposes. CONCLUSIONS: Deregulation of miRNAs is associated with the development and progression of NB, suggesting that miRNAs may serve as novel targets for the treatment of high-risk NB patients. However, their precise functions and underlying mechanisms still warrant further studies.