骨髓增生异常综合征患者N-ras基因突变的连续观察

为连续观察骨髓增生异常综合征(MDS)病程进展过程中N-ras基因的突变情况,取19例MDS患者在不同FAB亚型时的骨髓片,每例患者至少取2次不同FAB亚型的骨髓片,应用PCR和寡核苷酸探针斑点杂交技术进行检测。结果显示:MDS患者中N-ras基因突变率为26.0％,主要为G→A的突变;随着MDS病情进展N-ras突变率逐渐增高,突变组比无突变组更易转化为白血病(P<0.05),突变组生存率明显低于无突变组(X~2=4.0149),P<0.05。结论提示,N-ras基因突变可作为观察MDS病程进展和判断其预后的一个指标。     

原位杂交法检测树舌多糖GF对HepA瘤细胞N—ras基因mRNA表达的影响

目的:探明树舌多糖GF对HepA瘤细胞癌基因N-ras mRNA水平的影响.方法:应用地高辛标记探针原位杂交技术,检测树舌多糖GF对HepA瘤细胞中癌基因N-ras mRNA丰度的作用.结果:树舌多糖组、猪苓多糖组中mRNA表达量均显著低于模型组(P<0.01),树舌多糖组与猪苓多糖组无显著性差异.结论:树舌多糖GF可通过调节N-ras基因mRNA的表达,而抑制HepA瘤细胞的增殖.     

7,12-DMBA-induced rat leukemia: a review with insights into future research

7,12-Dimethylbenz[a]anthracene (DMBA) elicits leukemia in Long–Evans rats (LE). This leukemia is mostly erythroblastic and 30% of leukemias have total and partial trisomy of #2 chromosome and the rest have diploid karyotype. The common duplication site is in 2q26–q34 and N-ras gene is located in 2q34. 7,8,12-Trimethylbenz[a]anthracene (TMBA) also induces similar leukemias. These leukemias reveal a highly specific mutation of N-ras gene as in human leukemias. N-ras mutation is induced 48 h after DMBA treatment. Wild type N-ras allele is frequently lost in diploid leukemias but not in trisomy type. Therefore, a gene dosage problem related to the mutant N-ras gene is involved in development of leukemia. Some secondary genetic rearrangements involving abl and H-ras are also observed in cultured leukemia cells. DMBA-induced chromosome aberrations as well as leukemia are enhanced by erythropoietin and blocked by Sudan III given prior to DMBA treatment. This leukemia will provide an important tool for chemical carcinogenesis and leukemia studies.     

Analysis of ras DNA sequences in rat renal cell carcinoma

The DNA sequences for Ha-, Ki-, and N-ras were determined in six cell lines derived from independent rat hereditary renal cell carcinomas (RCC). Genomic regions encompassing codons 12, 13, and 61 of Ha-ras, Ki-ras, and N-ras, and codon 117 of Ha-ras were PCR amplified and directly sequenced. The DNA sequences of Ha-ras and Ki-ras were normal in all lines tested, as were the codon 12 and 61 sequences of N-ras. However, DNA sequence variations that could code for amino acid substitutions were observed in codons 13, 14, and 18 of N-ras in all the lines. The codon 13 Gly----Val alteration observed was consistent with activating N-ras mutations previously reported. When normal kidney DNA from rats with the hereditary tumor syndrome was sequenced, the same N-ras sequence variations observed in the tumor lines were found. DNA from outbred Long-Evans and inbred Fischer rats also had the altered N-ras sequences. The variant N-ras sequence was not observed in PCR-amplified N-ras cDNA from the RCC lines. Thus, tumor-associated activation of ras oncogene appears to be an infrequent event in spontaneous rat RCC. In addition, these data indicate that rats contain an N-ras DNA polymorphism that appears to be a species-specific anomaly.     

反义N—ras1DNA片段部分抑制人膀胱癌BIU—87细胞VEGF基因的表达

本文报道将载有反义人Ｎ－ｒａｓｌ（ｅｘｏｎｌ）ＤＮＡ片段的重组表达质粒ｆＰＧＶ１－ＭＴ－Ｎｒａｓｌ（Ａ）导入人膀胱移行细胞癌ＢＩＵ－８７细胞系，可以部分抑制ＢＩＵ－８７细胞中ＶＥＧＦｍＲ－ＮＡ的表达。提示ＶＥＧＦ基因的表达可受到反义Ｎ－ｒａｓ１ＤＮＡ片段的调控。本研究不仅增加了从分子水平对ＢＩＵ－８７细胞的认识，有助于进一步确定该细胞的生物学特征，还为以后具有Ｎ－ｒａｓ癌基因及ＶＥＧＦ基因表达的膀胱癌进行反义癌基因治疗提供一定的理论和实验依据。     

N－ras癌基因突变体分离方法的建立

采用能区分单个碱基变化的ＰＣＲ－ＳＳＣＰ技术扩增Ｎ－ｒａｓ癌基因第１、２外显子，有效地鉴别了１２／１３、６１位点突变。结果表明：在５例ＡＭＬ患者中有３例发生了Ｎ－ｒａｓ癌基因点突变，其中１２／１３位２例，６１位１例。本文使用的方法无需同位素标记，电泳结果也直接经溴化乙锭染色后在紫外检测仪中观察，可以节省试剂和缩短实验时间，是一种检测癌基因突变体的好方法。     

MiR-214 and N-ras regulatory loop suppresses rhabdomyosarcoma cell growth and xenograft tumorigenesis

Rhabdomyosarcoma (RMS) is a childhood malignant soft tissue cancer that is derived from myogenic progenitors trapped in a permanent mode of growth. Here, we report that miR-214 is markedly down-regulated in human RMS cell lines. Although not required for embryogenesis in mice, miR-214 suppresses mouse embryonic fibroblast (MEF) proliferation. When re-introduced into RD cells, a line of human embryonal RMS cells, miR-214 showed inhibition of tumor cell growth, induction of myogenic differentiation and apoptosis, as well as suppression of colony formation and xenograft tumorigenesis. We show that in the absence of miR-214, expression of proto-oncogene N-ras is markedly elevated in miR-214^−/− MEFs, and manipulations of miR-214 levels using microRNA mimics or inhibitor in RD cells reciprocally altered N-ras expression. We further demonstrate that forced expression of N-ras from a cDNA that lacks its 3''-untranslated region neutralized the pro-myogenic and anti-proliferative activities of miR-214. Finally, we show that N-ras is a conserved target of miR-214 in its suppression of xenograft tumor growth, and N-ras expression is up-regulated in xenograft tumor models as well as actual human RMS tissue sections. Taken together, these data indicate that miR-214 is a bona fide suppressor of human RMS tumorigensis.     

Screening of N-ras Gene Mutations in Urothelial Cell Carcinomas of the Urinary Bladder in the Kashmiri Population

Background and Aims: The objective of this study was to assess the frequency of specific-point mutations inN-ras of the RAS gene family in a group of Kashmiri patients with bladder cancer and to observe any associationwith clinicopathological parameters. Methods: Paired tumor and normal tissue specimens of 55 consecutivepatients with urothelial cell carcinoma were screened and DNA was extracted for detection of N-ras activatingmutations in exons 1 and 2. In addition, blood was also collected from all the cases to rule out any germ linemutation. Results: Specific point mutations of activated N-ras were detected in 9% (5 of 55) of the bladdercancer patients, all being missense. The base substitutions identified included three transversions (two G toTand one A to T) and two transitions ( A-G). Sixty % of the mutations were detected in codon 61 and 40% incodon 12. No significant correlations were found between the mutations and clinical features. Conclusion:Although N-ras gene mutation might be one of the mechanisms underlying oncogenesis of urothelial cancer, itseems to be a relatively rare event in Kasmiris, pointing to involvement of different etiological factors in theinduction of bladder tumor in this population     

原发性肝癌中N—ras,c—myc癌基因的表达研究

以点杂交和原位杂交法检测了34例启东地区原发性肝癌手术切除标本N－ras、c－myc基因的表达水平和定位。点杂交结果显示：N－ras、c-myc的检出率分别为647％（22／34）和23．5％（8／34）。前期的实验表明此34例标本HBV－DNA的检出率为588％（20／34），Southernblot实验结果未见病毒DNA的整合。N－ras、c-myc阳性标本中HBV－DNA的检出率分别为54．5％（12／22）和75％（6／8），两者之间无差异，P＞0．05。原位杂交结果显示：肝癌癌组织中N－ras、c－myc基因的表达水平并没有明显增高，相反，肿瘤周围肝组织中的表达水平增高并主要位于慢性肝炎、肝硬变中的再生肝细胞中，阳性细胞呈灶性及成片分布，不伴有毛玻璃样改变（HE染色）。结论：虽然N－ras、c－myc与肝癌的发生密切相关，肝癌患者中常见其表达水平增高，但这很可能只是一种肝细胞损伤后再生时的细胞增殖性调控的表现，此种N－ras、c-myc的高表达是否直接导致肝癌发生，癌基因是否为突变型，有待进一步研究、此外，本实验中c－myc基因相对于N－ras基因的低检出率可能与c－mycMRNA极短的半衰期有关。