Evidence for GALNT12 as a moderate penetrance gene for colorectal cancer

Characterizing moderate penetrance susceptibility genes is an emerging frontier in colorectal cancer (CRC) research. GALNT12 is a strong candidate CRC‐susceptibility gene given previous linkage and association studies, and inactivating somatic and germline alleles in CRC patients. Previously, we found rare segregating germline GALNT12 variants in a clinic‐based cohort (N = 118) with predisposition for CRC. Here, we screened a new population‐based cohort of incident CRC cases (N = 479) for rare (MAF ≤1%) deleterious germline GALNT12 variants. GALNT12 screening revealed eight rare variants. Two variants were previously described (p.Asp303Asn, p.Arg297Trp), and additionally, we found six other rare variants: five missense (p.His101Gln, p.Ile142Thr, p.Glu239Gln, p.Thr286Met, p.Val290Phe) and one putative splice‐altering variant (c.732‐8 G>T). Sequencing of population‐matched controls (N = 400) revealed higher burden of these variants in CRC cases compared with healthy controls (P = 0.0381). We then functionally characterized the impact of substitutions on GALNT12 enzyme activity using in vitro‐derived peptide substrates. Three of the newly identified GALNT12 missense variants (p.His101Gln, p.Ile142Thr, p.Val290Phe) demonstrated a marked loss (>2‐fold reduction) of enzymatic activity compared with wild‐type (P ≤ 0.05), whereas p.Glu239Gln exhibited a ～2‐fold reduction in activity (P = 0.077). These findings provide strong, independent evidence for the association of GALNT12 defects with CRC‐susceptibility; underscoring implications for glycosylation pathway defects in CRC.     

bcl-2 protein expression is associated with better prognosis in colorectal cancer

AIMS: To investigate the expression of bcl-2 in colorectal carcinoma and to examine its association with mediators of apoptosis (p53 and mdm-2), clinicopathological features and long-term outcome. METHODS AND RESULTS: We determined by immunohistochemistry the expression of bcl-2 in 102 colorectal carcinomas with 10-year follow-up. In 66 of these cases in which we had previously assessed p53 status, no correlation was seen between bcl-2 and p53. The mdm-2 protein was not detected in any of the 66 cases. Cytoplasmic staining of the bcl-2 gene product was seen in the tumour cells of 22 cases (22%). Using a polymerase chain reaction technique we showed that overexpression of bcl-2 was not due to rearrangement of the bcl-2 gene. Expression of bcl-2 protein was related to tumour grade but was unrelated to patient age, sex, tumour site, tumour size or Dukes' stage. There was a trend towards increased survival in those whose tumours expressed bcl-2 protein (P = 0.055). When entered into a multivariate analysis, this survival difference was independent of tumour stage (P = 0.05). CONCLUSIONS: These results suggest that bcl-2 expression in colorectal carcinoma is associated with a better long-term prognosis.     

GCS通过MEK/ERK通路调控白血病多药耐药细胞凋亡相关基因bcl-2的表达

目的:探讨葡萄糖神经酰胺合成酶(GCS)是否通过MEK/ERK信号通路调控凋亡相关基因bcl-2的表达,从而诱导人白血病K562/A02细胞多药耐药。方法:用小干扰RNA(siRNA)靶向干扰K562/A02细胞中GCS的表达,real-time PCR、Western blotting检测Bcl-2、磷酸化及总ERK水平;用MEK特异性化学抑制剂U0126抑制MEK/ERK信号通路的活化,real-time PCR与Western blotting技术分别检测Bcl-2 mRNA与蛋白水平;CCK-8试剂盒检测细胞存活情况。结果:与阴性对照组比较,GCS siRNA明显抑制K562/A02细胞GCS和Bcl-2的表达,并抑制MEK/ERK信号通路的活化;U0126使Bcl-2 mRNA及蛋白水平呈浓度依赖性下降,并使K562/A02细胞ADM敏感性增加。结论:GCS通过MEK/ERK信号通路调控K562/A02细胞株中凋亡相关基因bcl-2的表达,从而诱导白血病细胞多药耐药。     

Retention of imprinting of the human apoptosis-related gene TSSC3 in human brain tumors

Genomic imprinting is the result of a gamete-specific modification leading to parental origin-specific gene expression in somatic cells of the offspring. Several embryonal tumors show loss of imprinting of genes clustered in human chromosome 11p15.5, an important tumor suppressor gene region, harboring several normally imprinted genes. TSSC3, a gene homologous to mouse TDAG51, implicated in Fas-mediated apoptosis, is also located in this region between hNAP2 and p57 (KIP2). TSSC3 is the first apoptosis-related gene found to be imprinted in placenta, liver and fetal tissues where it is expressed from the maternal allele in normal human development. This study investigated the imprinting status of TSSC3 in human normal, adult brain and in human neuroblastomas, medulloblastomas and glioblastomas. A polymorphism in exon 1 at position 54 was used to analyze the allelic expression of the TSSC3 gene by a primer oligo base extension (PROBE) assay using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). We found that the TSSC3 gene is not imprinted in human normal, adult brain and blood. In contrast, strong allelic bias resembling imprinting could be detected in most examined tumor specimens. The results demonstrate for the first time that the tumors under investigation are associated with a retention of imprinting of a potential growth inhibitory gene.     

NKX3.1下调前列腺癌PC-3细胞中抗凋亡基因 bcl-2 的表达

目的: 研究前列腺特异转录因子NKX3.1对 bcl-2 基因表达以及前列腺癌细胞凋亡的影响。方法: 转染pcDNA3.1- NKX3.1 于前列腺癌PC-3细胞中,得到瞬时转染NKX3.1的PC-3细胞,通过RT-PCR和Western blotting方法研究NKX3.1对 bcl-2 基因的表达的影响,并通过EMSA技术,研究NKX3.1诱导 bcl-2 基因下调的分子机制;进一步通过流式细胞术、凋亡小体染色等方法研究NKX3.1对细胞凋亡的影响。结果: NKX3.1可明显下调PC-3细胞中 bcl-2 mRNA及Bcl-2蛋白的表达;EMSA结果证明NKX3.1可与 bcl-2 基因上游调控区中的NKX3.1结合元件相互作用;流式细胞术结果显示,PC-3细胞转染NKX3.1后,细胞凋亡数增加了1.41倍;用Hoechst 33258细胞染色发现NKX3.1可使PC-3细胞凋亡小体数量明显增加。结论: NKX3.1可下调抗凋亡基因 bcl-2 的表达,促进前列腺癌细胞凋亡。     

胶质瘤bcd—2基因表达水平与其细胞增殖和凋亡关系的研究

目的 探讨胶质瘤细胞ｂｃｌ－２基因表达水平与肿瘤恶性程度、细胞增殖活性及凋亡程度的关系。方法 以６９例不同级别的人胶质瘤组织为研究对象,用原位杂交及免疫组化染色ＡＢＣ法分别检测ｂｃｌ－２ｍＲＮＡ、ｂｃｌ－２蛋白和增殖细胞核抗原（细胞增殖活性标记物）的表达,并用３’末标记法做原位细胞凋亡检测。结果 ６４例（９２．８％）表达ｂｃｌ－２ｍＲＮＡ,６０例（８７．０％）表达ｂｃｌ－２蛋白,两者的表达水平呈正     

Immunolocalization of bcl-2 in the human corpus luteum

The mechanisms of luteal regression and rescue in women areunknown but forms of programmed cell death may be involved.The proto-oncogene bcl-2 is an important inhibitor of apoptosisbut has not previously been described in the human corpus luteum.Immunohistochemical localization of bcl-2 protein was investigatedin human corpora lutea obtained from women undergoing surgeryduring endocrine monitored menstrual cycles as well as fromwomen who had been treated with human chorionic gonadotrophin(HCG) to prolong the luteal phase. Bcl-2 was found to be localizedin granulosa-lutein, theca-lutein (as identified by co-localizationof P450_{17-hydroxylase}) and the endothelial cells around someblood vessels. Immunoblotting demonstrated the presence of asingle band of MW 26 kDa. There was no apparent change in eitherthe intensity of immunostaining or the histological localizationduring the normal luteal phase or following treatment with humanchorionic gonadotrophin. The product of the proto-oncogene bcl-2is present in the human corpus luteum. It is unlikely that bcl-2expression alone is responsible for prolongation of the lifespanof the corpus luteum in early pregnancy although it is possiblethat the action of the bcl-2 gene present is modified by changesin other members of the bcl-2 family. apoptosis/bcl-2/corpus luteum/granulosa/17-hydroxylase     

Expression of bcl-2 gene product in neuroblastoma

The expression of bcl-2 is associated with inhibition of apoptosis and prolonged cell survival. The purpose of this study was to examine the immunoreactivity of neuroblastoma and ganglioneuroblastoma tissue samples to the bcl-2 gene product in order to see if it was related to prognosis. BCL-2 protein was detected in all the 46 formalin-fixed, paraffin-embedded samples from 34 patients representing all clinical stages and sites of involvement. Immuno-positivity was observed in tumours from the primary and metastatic sites. Moreover, it was demonstrated in the pre-chemotherapy and the post-chemotherapy samples from six cases with stage 4 disease. It was observed in neuroblasts in various stages of differentiation. A small proportion of undifferentiated neuroblasts were negative. As BCL-2 oncoprotein was present in all the cases irrespective of the clinical outcome, it does not appear to be one of the factors influencing prognosis.     

大肠肿瘤中p53和bcl-2蛋白的表达

目的：研究Ｐ５３和ｂｃｌ－２在大肠肿瘤发生中的作用。方法：应用免疫组化Ｓ－Ｐ法分别检测大肠正常、腺瘤及腺癌中Ｐ５３和ｂｃｌ－２表达。结果：大肠腺癌Ｐ５３表达率高于腺瘤组;Ｐ５３表达与大肠癌的临床病理因素无关,ｂｃｌ－２在正常粘膜基底部上皮细胞表达,在腺瘤（７７．５％）和腺（５５％）ｂｃｌ－２表达差异显著,高分化腺癌ｂｃｌ－２表达率（７３．７％）高于差分化癌（４１．２％）,在腺瘤和腺癌中ｂｃｌ－２和     

Nuclear overexpression of bcl-2 oncoprotein during the progression of human stomach cancer.

We unexpectedly observed strong nuclear overexpression of bcl-2 protein in advanced stomach cancer. As far as we know, such expression has not yet been reported. To investigate the significance of nuclear expression of bcl-2 protein in gastric carcinoma, we immunohistochemically analyzed bcl-2 overexpression in a gastric carcinogenic sequence, including 19 tubular adenomas (TA), 20 early carcinomas (EGC), and 20 advanced carcinomas (AGC). While TA displayed a specific granular and supranuclear cytoplasmic staining pattern, adenocarcinomas showed a strong nuclear staining pattern. Nuclear staining of bcl-2 was observed in 50% of AGC, 30% of EGC, and 10% of TA; cytoplasmic staining, on the other hand, was observed in all TA, 5% of EGC, and 10% of AGC. Nuclear bcl-2 overexpression differed according to the histologic type of AGC, occurring in 67% of the diffuse type and 25% of the moderately-to-well differentiated type. In the diffuse type, nuclear bcl-2 positive AGC predominated. In metastatic lesions, the pattern of bcl-2 immunostaining was almost identical to that seen in primary tumor. These results suggest that nuclear expression of bcl-2 may be related to malignant transformation in the stomach and is frequently associated with diffuse type advanced gastric adenocarcinomas.     

bcl-3基因通过cyclin D1及Bax蛋白影响人结肠癌RKO细胞的凋亡

目的:研究bcl-3基因对人结肠癌RKO细胞迁移及凋亡的影响及机制。方法:采用人bcl-3基因的RNA干扰慢病毒载体沉默人结肠癌RKO细胞bcl-3基因的表达后,划痕实验观察bcl-3基因沉默前后RKO细胞迁移能力的变化,Annexin V/PI双染色法检测bcl-3基因沉默前后RKO细胞凋亡率的变化,Western blot法检测bcl-3基因沉默前后细胞周期蛋白cyclin D1及凋亡相关蛋白Bax、Bcl-2的变化。结果:划痕实验显示,划痕后36 h,bcl-3基因沉默前后RKO细胞划痕愈合率分别为84.00%及40.00%,差异具有统计学意义(P0.05)。Annexin V/PI双染色法流式细胞术分析显示,bcl-3基因沉默前后的RKO细胞均经5μmol/L顺铂处理24 h后,沉默前后的RKO细胞凋亡率分别为12.89%及59.67%,差异具有统计学意义(P0.05)。Western blot法检测显示bcl-3基因沉默后cyclin D1蛋白表达显著下降(P0.05),Bax蛋白表达显著上升(P0.05),但Bcl-2表达无明显变化(P0.05)。结论:沉默bcl-3基因后,RKO细胞迁移能力下降,凋亡率增加,并伴细胞周期蛋白cyclin D1及凋亡相关蛋白Bax表达的变化。bcl-3基因可能通过改变cyclin D1及Bax蛋白的表达而影响RKO细胞的凋亡。     

Expression of the bcl-2 gene product in follicular lymphoma.

Expression of bcl-2 protein was analyzed in 140 cases of follicular lymphoma by immunohistologic staining of paraffin-embedded tissue; 85% of cases were positive, the frequency being related to histologic grade (100% for the small-cleaved cell type, 86% for the mixed cell type, and 76% for the large cell group). There was striking heterogeneity of bcl-2 content in a number of cases and the smaller neoplastic cells (i.e., centrocytes) were usually the most strongly labeled. In most cases, bcl-2 protein staining was much weaker in normal lymphoid cells than in the neoplastic cells. In several cases, staining for bcl-2 revealed patterns of neoplastic cell spread into adjacent tissue (e.g., normal follicles, lymphoid sinuses), and bcl-2 protein expression tended to be highest in these migratory cells.