Cancer-testis antigen lactate dehydrogenase C4 as a novel biomarker of male infertility and cancer.

A unique lactate dehydrogenase (LDH) isoenzyme designated as lactate dehydrogenase C4 (LDH-C4) is found in mammalian mature testis and spermatozoa. Thus far, LDH-C4 has been well studied with regard to its gene and amino acid sequences, structure, biological properties, and peptide synthesis. Accumulating evidence has shown that LDH-C4 is closely related to spermatic energy metabolism and plays a critical role in sperm motility, capacitation, and fertilization. Defects in the catalytic activity of LDH-C4 are key to pathophysiological abnormalities underlying infertility. LDH-C4 was originally thought to be present only in mature testis and spermatozoa; however, recent studies have implicated LDH-C4 as a cancer-testis antigen (CTA), owing to its aberrant transcription in a broad spectrum of human neoplasms. This review highlights the recent findings on LDH-C4 with particular emphasis on its role in male infertility and tumors.

Genetic mutation analysis of 22 patients with congenital absence of vas deferens: a single-center study†.

Congenital absence of the vas deferens (CAVD), a congenital malformation of the male reproductive system, causes obstructive azoospermia and male infertility. Currently, the cystic fibrosis transmembrane conductance regulator (CFTR) has been recognized as the main pathogenic gene in CAVD, with some other genes, such as adhesion G-protein-coupled receptor G2 (ADGRG2), solute carrier family 9 isoform 3 (SLC9A3), sodium channel epithelial 1 subunit beta (SCNN1B), and carbonic anhydrase 12 (CA12), being candidate genes in the pathogenesis of CAVD. However, the frequency and spectrum of these mutations, as well as the pathogenic mechanisms of CAVD, have not been fully investigated. Here, we sequenced all genes with potentially pathogenic mutations using next-generation sequencing and verified all identified variants by Sanger sequencing. Further bioinformatic analysis was performed to predict the pathogenicity of mutations. We described the distribution of the p.V470M, poly-T, and TG-repeat CFTR polymorphisms and identified novel missense mutations in the CFTR and SLC9A3 genes, respectively. Taken together, we identified mutations in the CFTR, ADGRG2, SLC9A3, SCNN1B, and CA12 genes in 22 patients with CAVD, thus broadening the genetic spectrum of Chinese patients with CAVD.

[Prokaryotic expression and purification of mouse recombinant myelin proteolipid protein polypeptide].

Objective To establish and optimize the prokaryotic expression method for the recombinant mouse myelin proteolipid protein (PLP, 139-208 aa) which is a critical immunogenic polypeptide of PLP. Methods The sequence coding for PLP139-208 polypeptide was cloned into pET-32a(+) vector. Afterwards, the expression vector prepared in this research was transformed into E. coli BL21, and the recombinant PLP polypeptide was induced to express by isopropyl-ß-D-thiogalactoside (IPTG). Two key prokaryotic expression conditions, IPTG's induction length and temperature, were analyzed for further optimization. The recombinant PLP polypeptide was induced to express by the expression method under the optimal expression conditions, and then was purified by Ni-NTA agarose and amylose resin. Finally, the gain of PLP139-208 polypeptide was verified by Western blot analysis. Results The results in the combinatorial optimization revealed that the expression of PLP139-208 was obtained at a satisfactory level when it was incubated at 23DegreesCelsius for 20 hours with the IPTG concentration of 0.5 mmol/L. Conclusion The optimized prokaryotic expression method for the recombinant mouse PLP139-208 was successfully established and effectively performed. This will shed light on the further researches on the improved preparation for experimental autoimmune encephalitis (EAE, an animal model of multiple sclerosis) and the underlying mechanism underlying PLP-induced autoimmune demyelination.

3D-cultured adipose tissue-derived stem cells inhibit liver cancer cell migration and invasion through suppressing epithelial-mesenchymal transition.

Adipose tissue-derived stem cells (ADSCs) are considered promising candidates for stem cell therapy; however, the tumorigenicity of ADSCs remains controversial. The present study aimed to investigate the association between ADSCs and liver cancer cells, and to determine whether culture methods could influence the effects of ADSCs on liver cancer cell growth in vitro. Liver cancer cells were treated with ADSCs-conditioned medium (CM) that was collected using the two-dimensional (2D) culture method, sphere culture method, or three-dimensional (3D) culture method. After that, cell viability and apoptosis were measured using CCK-8 and Annexin V-FITC assay, respectively; the cell motility and adhesive capacity were analyzed by scratch wound healing and cell adhesion assay, respectively; the cell migration and invasion were examined by Transwell units; and the molecular mechanisms of ADSCs on effecting epithelial mesenchymal transition signaling pathway were further analyzed. The results demonstrated that ADSCs­CM was able to inhibit the growth of liver cancer cells by inhibiting cell proliferation and promoting cell apoptosis, as well as by suppressing cell motility, adhesive capacity, migration and invasion. In addition, ADSCs­CM was able to suppress cell growth via the downregulation of epithelial­mesenchymal transition signaling. Notably, the enhanced inhibitory effects of ADSCs on liver cancer cell growth could be achieved after cultu-ring using a 3D approach. These findings suggested that ADSCs may provide a novel promising therapeutic approach for the treatment of patients with liver cancer, and the 3D culture method may provide a novel approach to explore the association between ADSCs and cancer.

A new bioinformatic insight into the associated proteins in psychiatric disorders.

BACKGROUND: Psychiatric diseases severely affect the quality of patients' lives and bring huge economic pressure to their families. Also, the great phenotypic variability among these patients makes it difficult to investigate the pathogenesis. Nowadays, bioinformatics is hopeful to be used as an effective tool for the diagnosis of psychiatric disorders, which can identify sensitive biomarkers and explore associated signaling pathways. METHODS: In this study, we performed an integrated bioinformatic analysis on 1945 mental-associated proteins including 91 secreted proteins and 593 membrane proteins, which were screened from the Universal Protein Resource (Uniport) database. Then the function and pathway enrichment analyses, ontological classification, and constructed PPI network were executed. RESULTS: Our present study revealed that the majority of mental proteins were closely related to metabolic processes and cellular processes. We also identified some significant molecular biomarkers in the progression of mental disorders, such as HRAS, ALS2, SLC6A1, SLC39A12, SIL1, IDUA, NEPH2 and XPO1. Furthermore, it was found that hub proteins, such as COMT, POMC, NPS and BDNF, might be the potential targets for mental disorders therapy. Finally, we demonstrated that psychiatric disorders may share the same signaling pathways with cancers, involving ESR1, BCL2 and MAPK3. CONCLUSION: Our data are expected to contribute to explaining the possible mechanisms of psychiatric diseases and providing a useful reference for the diagnosis and therapy of them.

Bioinformatics analysis of gene expression profiles of dermatomyositis.

Dermatomyositis (DM) is a type of autoimmune inflammatory myopathy, which primarily affects the skin and muscle. The underlying mechanisms of DM remain poorly understood. The present study aimed to explore gene expression profile alterations, investigate the underlying mechanisms, and identify novel targets for DM. The GSE48280 dataset, which includes data from five DM and five normal muscle tissue samples, was obtained from the Gene Expression Omnibus. Firstly, differentially expressed genes (DEGs) were screened by limma package in R. Subsequently, functional and pathway enrichment analyses were performed using ClueGO from Cytoscape. Finally, protein­protein interaction (PPI) networks were constructed using STRING and Cytoscape, in order to identify hub genes. As a result, 180 upregulated and 21 downregulated genes were identified in the DM samples. The Gene Ontology enrichment analysis revealed that the type I interferon (IFN) signaling pathway was the most significantly enriched term within the DEGs. The Kyoto Encyclopedia of Genes and Genomes pathway analysis identified 27 significant pathways, the majority of which can be divided into the infectious diseases and immune system categories. Following construction of PPI networks, 24 hub genes were selected, all of which were associated with the type I IFN signaling pathway in DM. The findings of the present study indicated that type I IFNs may have a central role in the induction of DM. In addition, other DEGs, including chemokine (C­C motif) ligand 5, C­X­C motif chemokine 10, Toll­like receptor 3, DEXD/H­Box helicase 58, interferon induced with helicase C domain 1, interferon­stimulated gene 15 and MX dynamin­like GTPase 1, may be potential targets for DM diagnosis and treatment.

Single Nucleotide Polymorphism of the Enhancer of Zeste Homolog 2 Gene rs2072408 is Associated with Lymph Node Metastasis and Depth of Primary Tumor Invasion in Gastric Cancer.

BACKGROUND: The single nucleotide polymorphism (SNP) rs2072408 is located in an intron of the enhancer of zeste homolog 2 (EZH2) gene. Its role in gastric cancer (GC) has not been determined. METHODS: In the present study, the genotype of (EZH2) rs2072408 and the relationship of genotype with lymph node metastasis (LNM) and the invasive depth of gastric wall (T stage) was determined in 330 patients with GC, and the association between the genotype and recurrence and survival was determined in 253 patients with GC. RESULTS: The TT genotype was identified to be significantly associated with LNM [P = 0.013; odds ratio (OR), 6.49 (95% confidence interval (CI), 1.47 - 28.59)] and T stage [T1 + T2 vs. T3 + T4, p = 0.017, OR, 3.02 (95% CI, 1.22 - 7.44)]. The overall rates of LNM and T3 + T4 stage in the present study were 70.6% and 60.6%, respectively, the rates increased 23.8% and 20.0% in patients with the TT genotype. CONCLUSIONS: These results suggest that the TT genotype of EZH2 (rs2072408) was associated with LNM and the depth of gastric wall invasion in patients with GC.

[Advances in the studies of spermatogonial stem cells in primate mammals].

Spermatogonial stem cells(SSCs) are a type of spermatogonial cells that play an important role in the spermatogenesis of males. SSCs not only possess the properties of stem cells but also differentiate into sperm. They are the unique adult stem cells that transmit genetic information to subsequent generations. Therefore,SSCs are regarded as an ideal alternative source of pluripotent stem cells according with moral and ethical issues,legality,and safety. Long-term in vitro culture systems and identification of SSCs have paved the ground for the studies of their transplantation and pluripotency. Early relevant studies mainly focused on non-primate mammals. Recently,researches on SSCs have made great progress in primate mammals,especially in humans. This review focuses on the characterization,isolation,purification,cultivation,identification,and biological markers of SSCs,with a discussion on their unlimited pluripotency and application,and meanwhile provides an insight into the application potential of SSCs in the treatment of male infertility and human regenerative medicine.

Expression and clinical significance of long non-coding RNA HNF1A-AS1 in human gastric cancer.

BACKGROUND: Increasing evidence has demonstrated that long non-coding RNAs (lncRNAs) play essential roles in the occurrence and development of human cancers, including gastric cancer (GC). However, the functional and clinical significance of lncRNAs are still poorly understood. METHODS: In this study, the expression of LncRNA HNF1A antisense RNA 1 (HNF1A-AS1) was first examined by lncRNAs microarray analysis in 6 GC tissues, and was then further verified by real-time quantitative reverse transcription PCR (qRT-PCR) both in 3 GC cell lines and 161 cases of GC tissues. We also evaluated the association between HNF1A-AS1 expression and clinicopathological features of patients with GC. RESULTS: LncRNAs microarray analysis results exhibited that HNF1A-AS1 was downregulated in GCs tissues (mean fold change 2.06, p < 0.05), which was further confirmed by qRT-PCR. The results from qRT-PCR showed that the expression of HNF1A-AS1 was not only downregulated in three GC cell lines (AGS, BGC-823, and MKN-45) relative to that in a normal gastric mucosal epithelial cell line (GES-1), but also decreased in GC tissues relative to that in paired adjacent non-neoplastic tissues (low expression, 94 of 161; low expression rate, 58.38%). Furthermore, low HNF1A-AS1 expression was associated with tumor size/diameter (p = 0.005, multivariate analysis), levels of serum carcinoembryonic antigen (CEA), and carbohydrate antigen 19-9 (CA19-9), and RRM1 expression in tissue samples (p = 0.028, p = 0.009, and p = 0.006, respectively). CONCLUSIONS: Taken together, our data indicate that lncRNA HNF1A-AS1 may be a regulator of GC, and thus, it may have potential as a novel biomarker and treatment target for this type of cancer.

MicroRNAs as Biomarkers for the Diagnostics of Bladder Cancer: a Meta-Analysis.

BACKGROUND: Bladder cancer (BCa) is the fifth most common cancer with significant morbidity and mortality. Recently, numerous studies demonstrated that microRNAs (miRNAs) are emerging as diagnostic biomarkers for BCa. However, the findings in these studies are inconsistent. To systematically assess the potential diagnostic value of miRNAs for BCa, a meta-analysis was performed in this study. METHODS: Relevant literature was researched in PubMed, Embase, Cochrane Library, Chinese National Knowledge Infrastructure (CNKI), and WanFang databases up to September 1, 2014. The pooled sensitivity, specificity, positive likelihood ratio (PLR), negative LR (NLR), diagnostic odds ratio (DOR), and area under the SROC curve (AUC) value were analyzed by the random-effects model, whose parameters reflected the overall diagnostic performance of miRNAs. RESULTS: Thirty studies from 10 individual publications, including 1019 BCa patients and 690 controls, were included in this meta-analysis. The pooled sensitivity, specificity, PLR, NLR, DOR, and AUC were 0.80 (95% CI: 0.78 - 0.81), 0.74 (95% CI: 0.72 - 0.76), 3.22 (95% CI: 2.68 - 3.87), 0.26 (95% CI: 0.21 - 0.32), 15.20 (95% CI: 10.25 - 22.53) and 0.85, respectively, indicating a moderate diagnostic accuracy for BCa. Moreover, our subgroup analyses showed that analysis of multiple miRNAs (AUC, sensitivity, and specificity of 0.913, 0.86, and 0.80, respectively) yielded a higher diagnostic accuracy than of single miRNAs (AUC, sensitivity, and specificity of 0.84, 0.78, and 0.73, respectively) in BCa diagnosis. In addition, as biomarkers, miRNAs are more suitable for the diagnosis of non-muscle-invasive BCa (NMIBCa) with AUC of 0.84, sensitivity of 0.74, and specificity of 0.77 than muscle-invasive BCa (MIBCa) with AUC of 0.79, sensitivity of 0.73, and specificity of 0.73. CONCLUSIONS: The present meta-analysis suggests that miRNAs are potential novel biomarkers for detection of BCa. However, further validation studies are still needed to confirm our findings.

Prognostic value of differential CCND1 expression in patients with resected gastric adenocarcinoma.

Cyclin D1 (CCND1) plays essential roles in cancer progression. In this study, CCND1 expression patterns in 211 cases of resected gastric adenocarcinoma (RGA) tissue were determined by immunohistochemistry, and the association between CCND1 expression levels and RGA prognosis was analyzed. RGA tissues displayed differential CCND1 expression (high expression, 52.1 %; n = 110, and low expression, 47.9 %; n = 101). CCND1 expression levels were related with median overall survival time (MST). MST in patients with high CCND1 expression was 43 months, whereas with low CCND1 expression it was 62 months (P = 0.013). When data were stratified by postoperative treatments and CCND1 expression levels, the MST for patients treated with fluoropyrimidine plus platinum (n = 140) was significantly longer than for those treated with fluoropyrimidine only (n = 71) in both high and low CCND1 expression groups (65.0 vs. 29.0 months, P = 0.041; and 74.5 vs. 33.0 months, P = 0.003, respectively). Cox multivariate analyses further confirmed that high CCND1 expression was related with poor prognosis in both treatment groups [hazard ratio (HR) 1.91, 95 % confidence interval (CI) 1.12-3.23; P = 0.017, and HR 2.14, 95 % CI 1.08-4.25; P = 0.029] and that fluoropyrimidine plus platinum was more effective than fluoropyrimidine only in high CCND1 (HR 0.47, 95 % CI 0.28-0.78; P = 0.004) and low CCND1 (HR 0.44; 95 % CI 0.23-0.82; P = 0.01) expression patients. Therefore, CCND1 may be used as a prognostic biomarker for patients with RGA.

[Analysis of APC gene mutation in a familial adenomatous polyposis pedigree].

OBJECTIVE: To analyze mutation of adenomatous polyposis coli (APC) gene in a family affected with familial adenomatous polyposis. METHODS: The diagnosis was made based on clinical manifestations, family history, presence of numerous polyps in the colon as well as pathological examination. Peripheral blood samples were collected, and genomic DNA was extracted. Potential mutation of the APC gene was detected by polymerase chain reaction (PCR) and DNA sequencing. After finding the mutation in the proband, the same mutation was screened among other family members. The mutation was also confirmed with PCR-restriction fragment length polymorphism (RFLP), with which 100 unrelated healthy controls were examined. RESULTS: A novel heterozygous nonsense mutation c.2891T>G (L964X) of the APC gene was identified in this pedigree. The mutation has led to premature termination of translation. The same mutation was not detected among the 100 healthy controls. CONCLUSION: The c.2891T>G (L964X) of the APC gene probably underlies the familial adenomatous polyposis in this pedigree. The combined DNA sequencing and PCR-RFLP method is efficient and accurate for the diagnosis.

[A novel mutation of NTRK1 gene in a family with congenital insensitivity to pain with anhidrosis].

OBJECTIVE: To screen for mutations in the neurotrophic tyrosine kinase receptor type 1 (NTRK1) gene in a Chinese family affected with congenital insensitivity to pain with anhidrosis (CIPA). METHODS: With informed consent obtained, peripheral blood samples were obtained from the patient and his family members. Seventeen coding exons and intron-exon boundaries of the NTRK1 gene were amplified with PCR and analyzed by direct sequencing. RESULTS: A novel mutation c.2086_2087insC (p.Arg696 fsx) was identified in exon 16 of the NTRK1 gene in the proband. This insertion has caused open reading frame shifting and a premature termination has occurred just one codon downstream. Truncation of 72 amino acids at the C terminus has wiped out part of the tyrosine kinase domain (TKD) of the protein. Both of the proband's parents and two grandmothers have carried the c.2086_2087insC (p.Arg696 fsx) mutation. No mutation was found in the NTRK1 gene of other siblings. CONCLUSION: Mutation analysis of the NTRK1 gene has been carried out in a Chinese family affected with CIPA, and a novel NTRK1 gene mutation was identified.

BAX and CDKN1A polymorphisms correlated with clinical outcomes of gastric cancer patients treated with postoperative chemotherapy.

This study aims to examine the correlation between two single nucleotide polymorphisms (SNPs) of apoptosis-related genes and clinical outcomes in gastric cancer. A total of 221 patients with stage T2 and T3 gastric cancer treated with postoperative chemotherapy between 2003 and 2008 were retrospectively collected in this study to explore the association of rs4645878 located in BAX gene and rs1801270 located in CDKN1A gene with survival, recurrence, and toxicity to chemotherapy. Additionally, immunohistochemistry was used to detect the BAX expression in gastric cancer tissues. Patients carrying at least one variant genotype in BAX SNP (rs4645878) showed a significantly increased recurrence risk [hazard ratio (HR) 2.63; 95 % confidence internal (95 % CI) 1.71-4.03] and poor survival (HR 2.89; 95 % CI 1.88-4.44). Moreover, the recurrence and survival rate in patients with GA genotype was 72.7 and 24.7 %, respectively, compared with total recurrence rate of 54.8 %, P = 0.006, and compared with total survival rate of 46.6 %, P = 0.001. In addition, the GA genotype was related to lower BAX expression in gastric cancer tissues. The CDKN1A (rs1801270) mutant genotype was associated with a significantly decreased risk of hematologic toxicity [odds ratio (OR) 0.28; 95 % CI 0.12-0.63]. SNPs located in BAX and CDKN1A genes are closely associated with clinical outcomes in patients with gastric cancer.

[Genetic diagnosis and analysis of related genes for a pedigree with 2p25 and 12p13 cryptic rearrangements].

OBJECTIVE: To analyze chromosome aberration in a child with mental retardation and abnormalities and its parents. METHODS: Chromosome G banding, multiplex ligation-dependent probe amplification, fluorescence in situ hybridization and single nucleotide polymorphisms array were employed for analysis. RESULTS: Karyotype analysis revealed that the child was 46,XX and the father was 46,XY, while the mother was 46,XX, add (12)(p13). Subtelomeric region analysis with MLPA displayed that the child has reduced ACP1 gene copy number in 2p25 region and increased SLC6A12,KDM5A gene copy numbers in 12p11 region. SNP-array has fine mapped the duplication to 12p13.33-p12.3, a 15.142 Mb region, and a deletion to 2p25.3 for 3.194 Mb, which resulted in duplication of 9 genes including SLC6A12 as well as deletion of 11 genes including SNTG2, respectively. FISH analysis revealed that the child was 46,XX,ish,der(2),t(2;12)(p25;p13)mat, or partial monosomy 2p25 and partial trisomy 12p13. In addition,the mother was a carrier with cryptic balanced translocation chromosome, 46,XX,isht(2;12) (p25;p13). Mental abnormalities and retardation of the child may be attributed to heterozygous deletion of SNTG2, MYT1L genes and duplication of SLC6A12 gene. CONCLUSION: Combined use of MLPA, FISH and SNP-array can facilitate accurate diagnosis of cryptic rearrangement at genomic level.

Additive effects of EGF and IL-1ß regulate tumor cell migration and invasion in gastric adenocarcinoma via activation of ERK1/2.

Growth and inflammatory factors are associated with poor prognosis in gastric adenocarcinoma (GA); however, the additive effects of growth and inflammatory factors in GA remain unclear. In this study, we investigated the ability of epidermal growth factor (EGF) and interleukin (IL-1ß) to activate extracellular signal-regulated kinase (ERK)1/2 in GA cells, and correlated the relationships between their roles with the metastatic potential both in GA cells and GA tissues. The effects of EGF, IL-1ß and EGF plus IL-1ß in AGS and MKN-45 GA cells were examined using western blotting, Transwell migration and invasion assays, immunocytochemical staining and an activator protein (AP)-1 luciferase reporter gene assay, and was further characterized in GA tissues by immunohistochemistry. The results exhibited that EGF and IL-1ß additively activated ERK1/2, increased migration and invasion than either EGF or IL-1ß alone in AGS and MKN-45 cells. The mechanisms were involved in upregulating MMP-9 expression through increasing AP-1 transcriptional activity via ERK1/2 pathway; these effects were dose-dependently inhibited by silencing ERK1/2 or using U0126. In vivo data also confirmed that the overexpression of p-ERK1/2 in GA tissues correlated well with the EGF, IL-1ß, EGF plus IL-1ß, and was associated with metastasis, which was well correlation with the expression of MMP-9 and c-fos (AP-1). The results demonstrate that growth and inflammatory factors play an important role in metastasis of GA by additively activating ERK-1/2 and AP-1, and upregulating MMP-9. As both cytokines contribute to the migration and invasion of GA cells, EGF/IL-1ß/ERK1/2 pathways may be key pathways closely associated with GA progression.

A GG allele of 3'-side AKT1 SNP is associated with decreased AKT1 activation and better prognosis of gastric cancer.

PURPOSE: v-akt Murine thymoma viral oncogene homolog (AKT) pathway is critically involved in cancer cell growth, invasion, and survival. We examined the correlation between the genetic variations in molecules of AKT pathway and clinical outcomes of gastric cancer. PATIENTS AND METHODS: Six single nucleotide polymorphisms (SNPs) located in the four core genes of AKT pathway, namely the PIK3CA, PTEN, AKT1, and mTOR, were determined in 221 patients with stage T2 and T3 gastric cancer. Additionally, the activation of AKT1 in gastric cancer tissues was examined by immunostaining. The correlation between SNPs, AKT activation, and the progress of gastric cancer was analyzed after an average of 51-month follow-up. RESULTS: The overall recurrence and survival rate in this study group were 54.8 and 46.6 %, respectively. The recurrence rate was reduced 30.4 %, and the survival rate was increased 33.7 % in patients with GG allele of a 3'-side AKT1 SNP (rs2498804). Significantly, GG allele was associated with lower AKT1 activation in gastric cancer tissues. On the contrary, CC allele of PTEN (rs701848) was associated with the increased risk of recurrence (hazard ratio [HR] 2.06, 95 % CI 1.19-3.58) and patient death (HR 2.01, 95 % CI 1.15-3.53). CONCLUSIONS: The genetic variants in the PI3K/PTEN/AKT especially the GG allele in 3' side of AKT1 are closely related to clinical outcomes of gastric cancer.

IL-1ß-induced activation of p38 promotes metastasis in gastric adenocarcinoma via upregulation of AP-1/c-fos, MMP2 and MMP9.

BACKGROUND: Interleukin-1ß (IL-1ß) has been implicated in the progression of gastric adenocarcinoma (GA); however, the molecular mechanisms of action of IL-1ß in GA are poorly characterized. P38 and JNK are the major MAPK family members that regulate IL-1ß signaling pathways. Here, we investigated the role of both p38 and JNK in IL-1ß-induced GA cell migration, invasion and metastatic potential. METHODS: The effects of IL-1ß-induced p38 and JNK activation in GA cells were determined using in vitro Transwell migration and invasion assays of MKN-45 and AGS cells, or an in vivo metastasis assay in nude mice. The IL-1ß-induced p38 signaling pathway was further characterized in GA cells. Activation of the IL-1ß/p38 signaling pathway was also assessed in human primary GA tissues by immunohistochemistry. RESULTS: IL-1ß-induced activation of p38 increased GA cell migration and invasion in vitro and promoted the metastatic potential of GA cells in vivo; these effects were attenuated by p38 siRNA or the p38 inhibitor SB202190. MMP2 or MMP9 siRNAs and the MMP2/9 inhibitor BiPS also inhibited IL-1ß-induced GA cell migration and invasion in vitro. IL-1ß-induced p38 activation significantly increased MMP2 and MMP9 mRNA and protein expression and activity. Luciferase reporter assays demonstrated that the activator protein-1 (AP-1) and the AP-1 binding sites of the MMP9 promoter (-670/MMP9) were activated by IL-1ß-induced p38 activation. Phospho-p38 was significantly upregulated in human GA tissues (compared to matched non-neoplastic tissues), and significantly associated with lymph node metastasis, and invasion beyond the serosa. Expression of phospho-p38 significantly correlated with IL-1ß, MMP2, MMP9, and c-fos expression in both human GA tissues and GA cell metastases in the lungs of nude mice. IL-1ß was also capable of activating JNK in GA cells, but activation of JNK was not associated with GA cell migration and invasion. Therefore, IL-1ß-induced the migration and invasion in GA cells were regulated by p38, but not by JNK. CONCLUSIONS: IL-1ß-induced p38 activation and the IL-1ß/p38/AP-1(c-fos)/MMP2 & MMP9 pathway play an important role in metastasis in GA; this pathway may provide a novel therapeutic target for GA.

[Molecular diagnosis and prenatal diagnosis in a hereditary multiple osteochondromas family].

OBJECTIVE: To identify the mutation in the disease gene and provide prenatal diagnosis for a hereditary multiple osteochondromas (HMO) family. METHODS: The exons of EXT1 gene in the proband with HMO and his family members were amplified by PCR. The products were analyzed by direct sequencing. Prenatal genetic diagnosis was performed by amniocentesis sampling after genotyping the proband. RESULTS: In the family, the affected proband was heterozygous of the mutation of 1476_1477delTC in the EXT1 gene, and the proband's father carried the same mutation in part of his somatic cells. No mutation was found in the EXT1 gene of the proband's mother and other 11 siblings of his father. CONCLUSION: METHODS for molecular diagnosis and prenatal diagnosis of HMO were established and applied to a family of HMO.

p73 participates in WWOX-mediated apoptosis in leukemia cells.

The WWOX gene is considered to be a tumor-suppressor gene which encodes a protein (Wwox) implicated in various types of solid human cancers. It has been shown that overexpression of WWOX in human tumors promotes apoptosis in vitro and suppresses tumor growth in vivo. Recently, we investigated the effects of WWOX overexpression in vitro and observed marked growth arrest in human leukemia cells; however, the underlying mechanism(s) for this effect is unknown. The present study aimed to elucidate the primary mechanism(s) underlying WWOX-mediated apoptosis in human leukemia. We traced the interactions between WWOX and its associated factors p73 and p53 after WWOX overexpression was induced in Jurkat and K562 cells. Our data revealed that p73 participates in WWOX-mediated apoptosis in Jurkat and K562 cells through binding with Wwox in the cytoplasm without a nuclear-cytoplasmic translocation.