Integrative molecular characterization of Chinese prostate cancer specimens.

Prostate cancer (PCa) exhibits epidemiological and molecular heterogeneity. Despite extensive studies of its phenotypic and genetic properties in Western populations, its molecular basis is not clear in Chinese patients. To determine critical molecular characteristics and explore correlations between genomic markers and clinical parameters in Chinese populations, we applied an integrative genetic/transcriptomic assay that combines targeted next-generation sequencing and quantitative real-time PCR (qRT-PCR) on samples from 46 Chinese patients with PCa. Lysine (K)-specific methyltransferase 2D (KMT2D), zinc finger homeobox 3 (ZFHX3), A-kinase anchoring protein 9 (AKAP9), and GLI family zinc finger 1 (GLI1) were frequently mutated in our cohort. Moreover, a clinicopathological analysis showed that RB transcriptional corepressor 1 (RB1) deletion was common in patients with a high risk of disease progression. Remarkably, four genomic events, MYC proto-oncogene (MYC) amplification, RB1 deletion, APC regulator of WNT signaling pathway (APC) mutation or deletion, and cyclin-dependent kinase 12 (CDK12) mutation, were correlated with poor disease-free survival. In addition, a close link between KMT2D expression and the androgen receptor (AR) signaling pathway was observed both in our cohort and in The Cancer Genome Atlas Prostate Adenocarcinoma (TCGA-PRAD) data. In summary, our results demonstrate the feasibility and benefits of integrative molecular characterization of PCa samples in disease pathology research and personalized medicine.

Curcumin protects intestinal mucosal barrier function of rat enteritis via activation of MKP-1 and attenuation of p38 and NF-κB activation.

BACKGROUND: Intestinal mucosa barrier (IMB) dysfunction results in many notorious diseases for which there are currently few effective treatments. We studied curcumin's protective effect on IMB and examined its mechanism by using methotrexate (MTX) induced rat enteritis model and lipopolysaccharide (LPS) treated cell death model. METHODOLOGY/PRINCIPAL FINDINGS: Curcumin was intragastrically administrated from the first day, models were made for 7 days. Cells were treated with curcumin for 30 min before exposure to LPS. Rat intestinal mucosa was collected for evaluation of pathological changes. We detected the activities of D-lactate and diamine oxidase (DAO) according to previous research and measured the levels of myeloperoxidase (MPO) and superoxide dismutase (SOD) by colorimetric method. Intercellular adhesion molecule-1 (ICAM-1), tumor necrosis factor α (TNF-α) and interleukin 1ß (IL-1ß) were determined by RT-PCR and IL-10 production was determined by ELISA. We found Curcumin decreased the levels of D-lactate, DAO, MPO, ICAM-1, IL-1ß and TNF-α, but increased the levels of IL-10 and SOD in rat models. We further confirmed mitogen-activated protein kinase phosphatase-1 (MKP-1) was activated but phospho-p38 was inhibited by curcumin by western blot assay. Finally, NF-κB translocation was monitored by immunofluorescent staining. We showed that curcumin repressed I-κB and interfered with the translocation of NF-κB into nucleus. CONCLUSIONS/SIGNIFICANCE: The effect of curcumin is mediated by the MKP-1-dependent inactivation of p38 and inhibition of NF-κB-mediated transcription. Curcumin, with anti-inflammatory and anti-oxidant activities may be used as an effective reagent for protecting intestinal mucosa barrier and other related intestinal diseases.

Soluble intercellular adhesion molecule-1, D-lactate and diamine oxidase in patients with inflammatory bowel disease.

AIM: To study the levels of serum soluble intercellular adhesion molecule-1 (sICAM-1), plasma D-lactate and diamine oxidase (DAO) in patients with inflammatory bowel disease (IBD), and the potential clinical significance. METHODS: Sixty-nine patients with IBD and 30 healthy controls were included in this study. The concentration of sICAM-1 was detected with enzyme-linked immunosorbent assay, the level of D-lactate and DAO was measured by spectroscopic analysis, and the number of white blood cells (WBC) was determined by routine procedure. RESULTS: The levels of sICAM-l, DAO, and WBC in IBD patients were significantly higher than those in the control group (P < 0.01). sICAM-l in IBD patients was found to be closely related to the levels of DAO and D-lactate (212.94 +/- 69.89 vs 6.35 +/- 2.35, P = 0.000), DAO 212.94 +/- 69.89 vs 8.65 +/- 3.54, P = 0.000) and WBC (212.94 +/- 69.89 vs 7.40 +/- 2.61, P = 0.000), but no significant difference was observed between patients with ulcerative colitis and patients with Crohn's disease. The post-treatment levels of sICAM-l, D-lactate and WBC were significantly lower than before treatment (sICAM-l 206.57 +/- 79.21 vs 146.21 +/- 64.43, P = 0.000), (D-lactate 1.46 +/- 0.94 vs 0.52 +/- 0.32, P = 0.000) and (WBC 7.24 +/- 0.2.33 vs 5.21 +/- 3.21, P = 0.000). CONCLUSION: sICAM-1, D-lactate and DAO are closely related to the specific conditions of IBD, and thus could be used as a major diagnostic index.

[Changyanqing decoction produces anti-inflammatory effect by inhibiting the activation of nuclear factor-kappaB].

OBJECTIVE: To detect the changes in intestinal mucosal permeation in rats with methotrexate-induced small intestinal damage and investigate the protective effects of Changyanqing decoction. METHODS: Rat enteritis model was established by methotrexate (MTX) and sodium chloride. The rats were randomly divided into normal control group, model group, N-acetylcysteine (NAC) group and Changyanqing decoction group, and Changyanqing decoction (100 mg/kg) or saline was administered daily in the corresponding groups by gastric irrigation for 6 days. The disease activity index (DAI), colonic mucosal damage index (CMDI) and histological score (HS) of the rats were observed and evaluated. The levels of mRNA expressions of TNF-alpha and IL-1beta were detected by semi-quantitative RT-PCR. The expression of IL-10 was detected by enzyme linked immunosorbent assay, and IkappaB expression was determined with Western blotting. RESULTS: Compared with the normal control group, the model group showed significantly increased DAI, CMDI and HS. The DAI, CMDI, and HS in rats treated with Changyanqing decoction were significantly decreased in comparison with those in the model group (P<0.01). The expressions of TNF-alpha and IL-1beta were significantly higher in MTX-treated group than in the control group. The expression of TNF-alpha and IL-1beta mRNA in the Changyanqing group and NAC group were significantly lower, but IL-10 significantly higher than those of the MTX group. In MTX group, obvious NF-kappaB activation was observed, whose expression was significantly stronger in the cell nuclei, and the IkappaB in the cytoplasm was markedly degraded. CONCLUSION: Changyanqing decoction offers protection on intestinal mucosa by inhibiting NF-kappaB activation to reduce TNF-alpha and IL-1beta mRNA expressions and increase IL-10 expression.

[Value of prostate specific antigen in early diagnosis of prostatic cancer].

OBJECTIVE: To evaluate the value of serum prostate specific antigen (PSA), free PSA (FPSA) and PSA density (PSAD) in early diagnosis of prostatic cancer. METHODS: Sixty-eight patients with benign prostate hyperplasia (BPH), 28 with prostatic intraepithelial neoplasia (PIN) without canceration, and 32 with prostatic cancer, all diagnosed by prostatic biopsy, were enrolled in this study. Serum PSA and FPSA were measured and FPSA/PSA ratio and PSAD calculated for each patient, and the data analyzed to explore the association of these indices with prostatic cancer. RESULTS: Serum PSA level and PSAD were markedly different between the cancer patients and non-cancer patients (P<0.001 and P<0.01, respectively). FPSA/PSA ratio also differed between them (P<0.05). The same results were also found between BPH and cancer patients. Significant difference was noted in serum PSA and PSAD between PIN and cancer patients (P<0.01), but not in FPSA/PSA ratio (P>0.05). No marked difference was observed in serum PSA, FPSA/PSA ratio and PSAD between BPH and PIN patients. CONCLUSION: Serum PSA provides a very important clue for early diagnosis of prostatic cancer, and more accurate diagnosis can be obtained by considering also FPSA/PSA ratio. PSAD may also assist in the early diagnosis of prostatic cancer.

[Frequencies of CGT52TGT, GGC54GAC and GGA57GAA point mutations in MBL gene in Chinese Uyghur population].

OBJECTIVE: To investigate the frequencies of three point mutations, CGT52TGT, GGC54GAC and GGA57GAA, in exon 1 of mannan-binding lectin (MBL) structural gene in Chinese Uyghur population. METHODS: Blood samples were collected from a Uyghur population in Xinjiang Uyghur Autonomous Region, and the genomic DNA was extracted from the leucocytes and the target gene fragment amplified by PCR. The three point mutations in exon 1 of MBL gene were detected by fluorogenic probe hybridization technique with visual monitoring. RESULTS: In 95 Uyghur individuals, 2 were identified as homozygous for codon 54 mutations, 28 were heterozygous for codon 54 mutation, and no CGT52TGT and GGA57GAA point mutations were found. CONCLUSION: The frequencies of CGT52TGT, GGC54GAC and GGA57GAA mutant alleles in exon 1 of MBL structural gene are 0, 0.168 and 0 respectively in the Chinese Uyghur population.

[Construction of standard recombinant plasmids for 7 common mannan-binding lectin gene haplotypes].

OBJECTIVE: To construct the standard recombinant plasmids for 7 common haplotypes of mannan-binding lectin (MBL) gene. METHODS: The DNA samples with known haplotypes and genotypes of MBL gene were used as the templates for amplifying the fragments of MBL gene haplotypes including the promoter region and exon 1 with sequence-specific primer-polymerase chain reaction (SSP-PCR) method. The amplified fragments were cloned into T vector and the bases located at codon 52 and codon 57 of exon 1 in MBL gene were mutated respectively by site-directed mutagenesis. All the 7 recombinant plasmids were identified by PCR and direct sequence analysis. RESULTS: From the DNA samples with known haplotypes and genotypes of MBL gene, the standard plasmids of haplotypes HYPA, LXPA, LYQA, LYPA and LYPB of MBL gene were constructed by SSP-PCR and molecular cloning technique. From the recombinant plasmids of HYPA and LYQA, the standard plasmids of haplotypes HYPD and LYQC of MBL gene were constructed by site-directed mutagenesis, respectively. CONCLUSION: The constructed standard plasmids of haplotypes HYPA, LXPA, LYQA, LYPA, LYPB, HYPD and LYQC of MBL gene provide standard controls for detecting the SNPs, haplotypes and genotypes of MBL gene with such genotyping methods us SSP-PCR and real-time PCR.

Molecular cloning and characterization of mannan binding lectin C gene from Balb/c mice.

OBJECTIVE: To obtain full-length cDNA encoding mouse mannan-binding lectin C(MBL-C) polypeptide. METHODS: The cDNA encoding mouse MBL-C was isolated from Balb/c mouse liver cells by reverse transcriptase (RT)-PCR and inserted into pUC-T vector, and the encoding region structure of the DNA was analyzed to understand its evolutionary relationship with single human MBL homologues. RESULTS: The amplified mouse MBL-C cDNA, which was 735 bp in length, encoded 245 amino acid residues, and its structural analysis showed 100% homology with published mouse MBL-C gene sequence and 71.4% homology with MBL gene of Chinese human. CONCLUSION: The gene encoding Balb/c mouse MBL-C has been successfully isolated, which may facilitate further study of the innate immune functions of MBL molecule in vivo.