凋亡相关基因Survivin、bcl-2和bax表达在高三尖杉酯碱诱导骨髓增生异常综合征MUTZ-1细胞凋亡中的作用

目的：研究骨髓增生异常综合征(MDS)细胞株MUTZ—1凋亡相关基因Survivin、bcl—2和bax的表达及高三尖杉酯碱(HHT)诱导其凋亡的作用机制。方法：用MDS—RAEB细胞系MUTZ—1为体外模型，采用透射电镜和流式细胞仪Annexin-V^FITC/PI双染分析细胞凋亡，PI染色分析细胞周期变化，RT—PCR技术检测抗凋亡基因Survivin、bcl—2和促凋亡基因bax的表达。结果：HHT能诱导MUTZ—1细胞凋亡，凋亡率与作用浓度和时间成正相关，细胞被阻滞在G2期。HHT作用MUTZ—1细胞6小时后细胞内抗凋亡基因Survivin表达明显下降，而抗凋亡基因bcl—2和促凋亡基因bax表达无明显变化。结论：HHT能诱导MUTZ—1细胞凋亡，抗凋亡基因Survivin mRNA下调可能是MUTZ—1细胞凋亡的机制之一。     

Alteration of growth and differentiation factors response by Kirsten and Harvey sarcoma viruses in the IL-3-dependent murine hematopoietic cell line 32D C13(G)

32D C13(G) is an interleukin 3(IL3)-dependent non-tumorigenic murine hematopoietic cell line which undergoes terminal differentiation into granulocytes when exposed to granulocytic colony stimulating factor (G-CSF). Infections of 32D C13(G) cells with either Kirsten rat sarcoma virus or Balb murine sarcoma virus, both containing a v-ras oncogene, generates clones that can permanently grow in G-CSF without differentiation. 32D-Ki-ras cells show a heterogeneous morphology ranging from the promyelocytic to the myelocytic stage of differentiation, and express high levels of both myeloperoxidase (MPO) and lactoferrin (LF) mRNA. 32D-Ha-ras cells show a more immature phenotype and express MPO but no LF mRNA. The apparent differentiation block of both 32D Ki-ras and 32D Ha ras can be reversed by treatment with the chemical inducers retinoic acid, sodium butyrate or dimethylsulphoxide, which leads to terminal differentiation into granulocytes. When 32D-Ki-ras and 32D-Ha-ras cells are cultured in medium containing IL-3 they become adherent and express some monocyte-macrophage markers. Upon prolonged exposure to IL3, 32D-Ki-ras, but not 32D-Ha-ras, resume suspension growth. Both 32D-Ki-ras and 32D-Ha-ras rapidly die if grown in chemically defined medium in the absence of any growth factor and are non-tumorigenic in immunosuppressed mice. These findings indicate that ras activation may interfere with the normal response to growth and differentiation factors in cells of the granulocytic lineage. These alterations may represent a critical, although non-sufficient, step in leukemogenesis.     

bcl-2、bax、p53蛋白表达与外阴鳞癌细胞凋亡的相关性研究

目的　探讨bcl 2、bax、p5 3基因与外阴鳞癌细胞凋亡的关系。方法　应用原位末端标记及免疫组化S P法检测 30份外阴鳞癌石蜡包埋组织中的细胞凋亡API和bcl 2、bax、p5 3的表达情况。 结果　外阴鳞癌组织和正常外阴皮肤中均有细胞凋亡和bcl 2、bax、p5 3的表达。随着外阴鳞癌恶性程度的增高 ,API、bcl 2、bax表达降低 ;p5 3表达增高。在外阴鳞癌的部位凋亡细胞的分布与突变型 p5 3蛋白表达呈负相关 ,p5 3分别与bcl 2及bax的表达呈负相关 (P <0 .0 5 )。在正常外阴皮肤组织中 ,凋亡细胞的分布区域与表达bcl 2及bax蛋白的细胞分布区域一致 ,bcl 2与bax的表达呈正相关 (P <0 .0 5 )。结论　外阴鳞癌的发生、发展与细胞凋亡被抑制有关 ,这一过程可能主要由 p5 3基因调控 ,bcl 2可能仅在早期起作用。     

Spontaneous apoptosis in laryngeal squamous cell carcinoma is independent of bcl-2 and bax protein expression

BACKGROUND: bcl-2 and bax genes are known to be involved in the control of apoptotic cell death, an important mechanism of growth regulation that influences the biologic behavior of tumors. The aim of the current study was to investigate the relationship of bcl-2 and bax expression to the rate of spontaneous apoptosis in laryngeal carcinomas, and to assess its relations to clinicopathologic features of tumors. METHODS: Immunohistochemical analyses for bcl-2 and bax were performed on paraffin embedded tissue sections from 134 primary laryngeal squamous cell carcinomas. To visualize apoptotic cells, the nick end labeling method was used. The proliferative activity of tumors was analyzed by determination of mitotic indices. RESULTS: bcl-2 immunoreactivity was positively correlated with tumor grade (P < 0.00001), high T category (P < 0.02), metastatic involvement of cervical lymph nodes (P < 0.003), and supraglottic or subglottic location of primary tumors (P < 0.00005). An inverse relation was found between bcl-2 and bax expression (P < 0.004). The frequency of spontaneous apoptosis was closely associated with mitotic activity (P < 0.0004) but appeared to be unrelated to protein levels of bcl-2 or bax as well as to bcl-2:bax ratios. CONCLUSIONS: The results of this study point to the significance of cell proliferation as a major determinant of the rate of spontaneous apoptosis in laryngeal carcinomas. The bcl-2:bax expression ratio obviously does not affect the incidence of apoptosis, but it may be considered as a marker of disease progression and poor prognosis.     

胃癌及癌前病变细胞增殖和凋亡与bcl-2/bax表达的关系

目的：观察细胞凋亡和增殖及其调控基因在胃癌及癌前病变发生发展过程中的异常变化,探讨细胞凋亡在胃癌演变中的作用。方法：对１２８例胃癌及癌前病变患者和正常人,采用原位末端标记法（ｔｅｒｍｉｎａｌｄｅｏｘｙｎｕｃｌｅｏｔｉｄｌｙ ｔｒａｎｓｆｅｒａｓｅ ｍｅｄｉａｔｅｄ ｌａｂｅｌｌｉｎｇ,ＴＵＮＥＬ）及免疫组化技术检测胃粘膜上皮细胞凋亡、增殖及ｂｃｌ－２／ｂａｘ表达。结果：正常人、浅表性胃炎、萎缩性胃     

子宫内膜癌中细胞凋亡和bcl-2、bax基因的表达

目的 探讨细胞凋亡及其调控基因ｂｃｌ－２、ｂａｘ在子宫内膜癌变过程中的作用。方法应用免疫组织化学Ｓ－Ｐ法和ＴＵＮＥＬ技术,对子宫内膜腺癌癌变过程各级病变组织中细胞凋亡、ｂｃｌ－２和ｂａｘ蛋白进行测定。结果 正常增生期子宫内膜有偶发凋亡,子宫内膜增生症和子宫内膜腺癌的细胞凋亡率分别为０．６％和２．８％。ｂｃｌ－２蛋白在正常增生期子宫内膜、子宫内膜增生症和子宫内膜腺癌的表达率分别为１００％、７４．４％     

bcl—2基因家族在紫杉醇介导BJAB细胞凋亡中的作用

目的 观察抗微管新药紫杉醇对Ｂ细胞淋巴瘤细胞株ＢＪＡＢ是否具有凋亡诱导作用。并进一步研究ｂｃｌ－２基因家族在此过程中的作用。方法 将不同浓度的紫杉醇作用于ＢＪＡＢ细胞,观察其作用的时间效应及剂量效应;在光镜和电镜下观察其形态变化;用流式细胞仪分析细胞ＤＮＡ含量的改变并做ＤＮＡ片段分析;用免疫组织化学及半定量ＲＴ－ＰＣＲ法观察在紫杉醇作用过程中ｂｃｌ－２基因家族的蛋白及ｍＲＮＡ的变化。结果 紫杉醇能抑制ＢＪＡＢ细胞生长,抑制作用首先表现为Ｇ２／Ｍ期阻滞,一定时间后出现细胞凋亡,并显示剂量和时间效应。在这一过程中,ｂｃｌ－２转录及蛋白表达下降,并出现ｂｃｌ－ｘｓ的转录。结论 紫杉醇可诱导ＢＪＡＢ细胞凋亡,这为其应用于Ｂ细胞淋巴瘤的治疗提供依据。ｂｃｌ－２和ｂｃｌ－ｘｓ参与了紫杉醇介导ＢＪＡＢ细胞凋亡的基因调控。     

不同类型子宫内膜腺上皮细胞中bcl-2和bax基因表达的研究

目的:探讨bcl-2、bax在不同类型子宫内膜腺上皮细胞中的表达。方法:采用免疫组化检测子宫内膜腺上皮细胞中二指标的表达。结果:从单纯性、复杂性到不典型增生,bcl-2和bax的表达评分均有逐渐增多的趋势,不典型增生与前二者相比差异均有统计学意义,P<0·01。在内膜样腺癌中,随着分化程度的降低、分期的增加,bcl-2表达评分下降。随着分化程度的降低bax的表达逐渐降低;随着浸润深度的增加bax表达逐渐增加。在增生期和分泌期bcl-2和bax的表达无负相关,但在内膜样腺癌组织中bcl-2表达与bax呈正相关。结论:bcl-2和bax参与了癌前病变的发生,bcl-2、bax表达水平在一定程度上反映了内膜样腺癌的生物学行为。bcl-2和bax在增生期和分泌期内膜变化中的相互作用关系不大,但二者共同作用参与了内膜样腺癌的发生与发展。     

Bcl-2和Bax的表达与乳腺癌组织学分级及预后的关系

目的：探讨bcl-2和bax基因在乳腺癌中的表达与乳腺癌组织学分级及预后的关系。方法：应用SP免疫组化染色方法，检测40例乳腺癌组织中bcl-2、bax的表达情况。结果：组织学I级＋Ⅱ级乳腺癌中bcl-2蛋白阳性表达率与组织学Ⅲ级乳腺癌中bcl-2蛋白阳性表达率有显著差异（P〈0．01），Ⅰ＋Ⅱ级组bax蛋白阳性表达率与Ⅲ级组bax蛋白阳性率表达无显著差异（P〉0．05），Ⅰ级＋Ⅱ级组bcl-2高表达和bax低表达（bcl-2／bax≥1）与Ⅲ级组bcl-2高表达和bax低表达（bcl-2／bax≥1）有显著差异（P〈0．01）。10年以上存活组中bcl-2阳性表达率与10年以下存活组有显著差异（P〈0．01）；10年以上存活组bax阳性表达率与10年以下存活组有显著差异（P〈0．05）；10年以上存活组bcl-2高表达和bax低表达（bcl-2／bax≥1）与10年以下存活组bcl-2高表达和bax低表达（bcl-2／bax≥1）有显著差异（P〈0．05）。结论：bcl-2和bax比值与乳腺癌组织学分级、预后关系密切，bcl-2高表达和bax低表达的乳腺癌患者组织学分级好，预后好。     

Different effects of cyclic AMP and butyrate on eosinophilic differentiation,apoptosis and bcl-2 expression of a human eosinophilic leukemia cell line,EoL-1

A human eosinophilic leukemia cell line, EoL-1, stopped proliferating at the G₁ phase, differentiated into eosinophilic granule-containing cells, and died by apoptosis when stimulated with dibutyryl cyclic AMP (dbcAMP). To clarify the effects of dbcAMP, the effects of butyrate and cAMP-increasing reagents, prostaglandin E₂ (PGE₂) and forskolin, on EoL-1 cellular differentiation and apoptosis were examined and compared. PGE₂ and forskolin but not butyrate induced differentiation to eosinophilic granule-containing cells, suggesting that cAMP played a primary role in eosinophilic differentiation of EoL-1 cells. PGE₂, forskolin and butyrate, when used alone, did not induce apoptosis of EoL-1 cells significantly at the concentrations used, but sequential stimulation of EoL-1 cells with the cAMP-increasing reagents and butyrate showed that butyrate induced further maturation and apoptosis of cAMP-induced eosinophilic granule-containing cells. These results showed that cAMP and butyrate have different effects on eosinophilic differentiation and apoptosis of EoL-1 cells. The cAMP-increasing reagents and butyrate also showed different effects on expression of members of the bcl-2 family; PGE₂ decreased bcl-2 and bax levels, whereas butyrate increased the bcl-2 level. PGE₂ or PGE₂+butyrate, but not butyrate alone, induced bcl-x_S expression. EoL-1 cells constitutively expressed Fas and anti-Fas antibody induced EoL-1 cell death, but the Fas/Fas ligand system was not involved in dbcAMP-induced EoL-1 cell apoptosis. The EoL-1 cell line is thus a useful model in which to examine differentiation and apoptosis of eosinophilic leukemia cells. © 1996 John Wiley & Sons, Ltd.     

Prognostic significance of p53, bax and bcl-2 gene expression in patients with laryngeal carcinoma.

AIM: This study was designed to examine the prognostic significance of the coexpression of three genes (bax, bcl-2 and p53) which play a critical role in the apoptotic mechanisms in patients with squamous cell laryngeal carcinoma. MATERIALS AND METHODS: The immunohistochemical expression of bcl-2, bax and p53 genes was retrospectively examined in 38 patients with squamous cell laryngeal carcinoma and in five controls (necrotomic tissue). Tissue specimens were obtained both during the diagnostic biopsy and at the time of surgery. Clinicopathological and survival data were correlated with the staining results. RESULTS: Bcl-2 protein expression (P=0.0472), stage (P=0.0087) and lymph-node involvement (P=0.0488) were found to be independent prognostic factors. Increased bcl-2 protein expression correlated with a better 5-year survival (P=0.0472). Patients who were bcl-2(-)/p53(-) (n=25) or bax(+)/bcl-2(-) (n=13) had a significantly worse overall survival (P=0.0305 and P=0.0482, respectively). Similarly, patients who were bax(+)/bcl-2(-)/p53(-) (n=11) also had a worse 5-year survival compared with the rest of the group (P=0.0088). Changes that were noticed in bax and p53 protein expression from the time of biopsy until the time of surgery did not correlate with a significant increase in the overall survival. CONCLUSIONS: The expression of bcl-2 gene appears to be an independent prognostic factor for patients with laryngeal carcinoma. The coexpression of the genes studied can be used to determine aggressive clinical phenotypes.     

bcl-2、bax和p53在鼻咽鳞癌中的表达及其与瘤细胞凋亡的关系

目的探讨bcl-2、bax和p53在鼻咽鳞癌中的表达及其与瘤细胞凋亡指数的关系。方法用免疫组织化学SP法检测48例鼻咽鳞癌中bcl-2、bax和p53的表达,用TUNEL法检测鼻咽鳞癌细胞的凋亡指数。结果48例鼻咽鳞癌中bcl-2、bax和p53阳性率分别为85.00%(41/48)、68.00%(33/48)和77.00%(37/48)。48例鼻咽鳞癌细胞的平均凋亡指数为25.62±25.78/HPF,凋亡指数与bcl-2、bax和p53的表达无相关性。结论鼻咽鳞癌中bcl-2和bax均呈高表达且已达到相对平衡,推测它们可能并不起到介导瘤细胞凋亡的主导作用。鼻咽鳞癌中p53的过表达可能已经失去了对bcl-2和bax表达的调节进而影响细胞凋亡的功能。     

Paclitaxel restores radiation-induced apoptosis in a bcl-2-expressing, radiation-resistant lymphoma cell line

PURPOSE: To restore radiation-induced apoptosis in a bcl-2-expressing, radiation-resistant murine lymphoma cell line (LY-ar) by pretreatment with paclitaxel (Taxol). Because this cell line also has high intracellular levels of glutathione (GSH), reportedly due to the bcl-2 expression and involved in the cell's antioxidant functions, paclitaxel treatment was correlated with GSH levels. METHODS AND MATERIALS: LY-ar cells were pretreated with paclitaxel and then irradiated with 5 Gy. Apoptosis was measured by DNA fragmentation 6 h later. Dose response and time course experiments were performed. Intracellular GSH levels were measured after treatment. Cell survival analysis was performed for various paclitaxel concentrations +/- 5 Gy. RESULTS: LY-ar cells pretreated with 0 nM, 10 nM, 25 nM, and 50 nM paclitaxel for 20 h underwent apoptosis at 2%, 15%, 25%, and 22%, respectively. With the addition of 5-Gy irradiation, LY-ar cell apoptosis increased to 4%, 30%, 49%, and 57%. Maximal apoptosis was detected with a paclitaxel pretreatment time of 20 h. Intracellular GSH levels were reduced by nearly 50% with paclitaxel pretreatment. Surviving fractions (SFs) with 0 nM, 10 nM, 25 nM, and 50 nM paclitaxel and 0 Gy were 1.0, 0.50, 0.08, and 0.05, respectively. SFs with 0 nM, 10 nM, 25 nM, and 50 nM paclitaxel and 5 Gy were 0.009, 0.003, 3 x 10(-5), and 1 x 10(-5), respectively. CONCLUSION: Radiation-induced apoptosis in LY-ar cells was restored by pretreatment with paclitaxel. This correlated with lowered levels of intracellular GSH. Cell survival analysis indicated that the combination of Taxol and radiation on cell killing was greater than additive.     

Involvement of the bcl-2 gene in Hodgkin's disease

A major obstacle to investigations of Hodgkin's disease is the paucity of malignant cells, i.e., Reed-Sternberg cells and their variants, in tissues of patients with this disease. Consequently, the pathogenesis, cell of origin, and clonality of this relatively frequent lymphoma have remained unresolved. Results of recent studies suggest that in some instances Reed-Sternberg cells carry rearranged immunoglobulin heavy-chain joining region (JH) loci as well as chromosomal translocations involving band 14q32. Prompted by these findings, we sought to determine if the t(14;18) (q32;q21) translocation of follicular, non-Hodgkin's B-cell lymphoma was associated with Hodgkin's disease. To detect the possible t(14;18) (q32;q21) translocation within the rare malignant cells of Hodgkin's disease, we amplified sequences created by the t(14;18) translocation using the polymerase chain reaction (PCR). With this approach, DNA sequences carrying the direct fusion of the major breakpoint region of the candidate oncogene, bcl-2, derived from chromosome 18q21, with JH on chromosome 14q32 can be detected in as few as one in 10(5)-10(6) cells. In the present study, joined bcl-2/JH sequences were detected in tissues involved by Hodgkin's disease in 17 of 53 (32%) patients. The frequent association of bcl-2 translocation with Hodgkin's disease suggests that this oncogene has a role in the pathogenesis of Hodgkin's disease. That bcl-2 is involved in a major class of lymphoma in addition to follicular lymphoma implies a role for additional factors responsible for generating the two distinctive clinical and pathologic disease states.     

自发性细胞凋亡及bcl-2和bax基因与鼻咽癌放射敏感性关系的探讨

目的探讨自发性细胞凋亡及其相关基因bcl-2和bax与鼻咽癌放射敏感性的关系。方法采用TdT酶介导的生物素化dUTP缺口末端标记技术(terminal oxynucleotidyl transferase medi-ated dUTP biotin nick end labeling,TUNEL)和免疫组化法,分别检测51例放疗前的鼻咽癌活检组织中自发性凋亡指数(spontaneous apoptotic index,SAI)和bcl-2、bax基因的表达。结果鼻咽癌活检组织均检测到细胞凋亡,SAI平均为31.19±33.83;SAI与bcl-2和bax表达均无明显相关性,但与bcl-2/bax比率呈负相关(P<0.05);SAI与鼻咽癌放射敏感性明显相关,完全消退组比残留组SAI高(P<0.05);bcl-2的表达与鼻咽癌放射敏感性呈负相关(P<0.05),而bax的表达与鼻咽癌放射敏感性无关(P>0.05);bcl-2/bax的比率与鼻咽癌放射敏感性呈负相关(P<0.05)。结论SAI、bcl-2和bcl-2/bax的比率是反映鼻咽癌放射敏感性和预测鼻咽癌放疗效果的重要指标,而bax不能单独作为判断鼻咽癌放射敏感性的指标。