Radiosensitivity and transcription factor NF-kappaB inhibition-progress and pitfalls.
Radiosensitivity and transcription factor NF-kappaB inhibition-progress and pitfalls.
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放射敏感性和转录因子 NF-kappaB 抑制 - 进展和陷阱。
DOI：
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发表时间：
1999
期刊：
影响因子：
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通讯作者：
A. Dritschilo
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作者：
A. Dritschilo
细胞对电离辐射的反应包括信号转导级联的激活，这些信号转导级联可能起源于质膜、细胞质或细胞核。细胞在应对辐射“压力”方面的成功决定了它的生存或死亡。转录因子NFkB的激活是电离辐射暴露后的即时早期反应，具有保护细胞免于凋亡（程序性细胞死亡）的功能，但NFkB在有丝分裂细胞死亡中的作用尚未完全确定（1,2）。NF-kB在淋巴样细胞、不朽共济失调毛细血管扩张（AT）成纤维细胞和其他细胞中构成性激活（3-6）。电离辐射后NF-kB的激活需要AT成纤维细胞中完整的ATM基因功能(7)。由于AT成纤维细胞表现出失调的NFkB激活和极端的辐射敏感性，我们有理由问肿瘤细胞的辐射反应是否可以通过抑制NFkB来改变。NF-kB/Rel转录因子被多种不同的信号激活，这些信号聚集在NFkB抑制剂I kb的磷酸化和降解途径上。这导致揭露导致NFkB/Rel二聚体易位到细胞核的核定位信号。五种已知的哺乳动物NFkB/Rel蛋白包括c-Rel、p65 （RelA）、RelB、p50 （NFkB1）和p52 (NFkB2)，它们以同型二聚体或异源二聚体的形式结合DNA(8)。还应该注意的是，p50-p65 /I kB并不是唯一与NF-kB激活相关的复合体。p50-p65异二聚体可以与其他IkB家族成员相互作用，NFkB也可以存在于其他同二聚体或异二聚体复合物中，如p50-p50和p65-p65(8)。此外，IkB家族由三个成员组成：IkB ba、IkB bb和IkB«。此外，p50和p52、p105和p100前体的羧基末端区域也可以作为抑制蛋白。这些抑制剂在体内以细胞类型依赖和刺激依赖的方式对nfkb诱导信号做出不同的反应(9)。在这一期的Journal中，Pajonk等人（10）报道NFkB的激活并不能决定癌细胞固有的放射敏感性。细胞用表达I - kBa （NF-kB抑制剂）突变形式的腺病毒载体转导。转染突变IkBa的细胞克隆原性显著降低至7.4%（对照细胞为29.5%），凋亡指数为90%。因此，随后的辐射存活实验是在相对较小的细胞亚群上进行的，而生化分析是在大多数凋亡细胞上进行的。在《华尔街日报》上发表本质上是负面的研究是不寻常的。然而，如果确定的数据可以为该领域的其他科学家提供方向，那么就有潜在的价值。在临床上，固有的放射敏感性与肿瘤的治愈率有关（11）。操纵辐射敏感性的能力为提高癌症治疗的治疗比率提供了一种潜在的策略。因此，对Pajonk等人（10）的阴性放射生物学结论的实验结果和数据解释进行检查是适当的。1) PC3和HD-MyZ细胞耐辐射吗？Pajonk等人（10）指出，选择这两种细胞系是因为它们具有高组成水平的NFkB，这可能赋予它们相对较高的抗辐射能力。严格地说，辐射抗性是一个不精确的术语，它可能包括细胞对电离辐射反应的各个方面（12）。另一方面，辐射灵敏度由细胞辐射生存曲线终端斜率的陡度来定义（单打多目标模型）。可以确定ADo值（使存活率降低1/ e所需的剂量），以便对辐射敏感性进行相对定量。PC3和HD-MyZ细胞的Do值估计为1.0-1.1 Gy[图3(10)]，对肿瘤细胞系非常敏感，对AT杂合子成纤维细胞的敏感性相当（13）。据报道，对耐药细胞信号转导通路的破坏会导致放射致敏（14），但可能很难使已经敏感的细胞变得更敏感。在PC3和HD-MyZ细胞中观察到的组成性激活的NFkB水平的另一种解释可能与凋亡应激有关，而不是反映潜在的抗性。这与Pajonk等人（10）随后的观察结果一致，即当突变体I kB-a表达时，90%的细胞死于凋亡，抑制了活化的NFkB的保护功能。2) ikba超抑制产物是否能抑制辐射诱导的一小部分未发生凋亡的转导细胞的NFkB激活？Pajonk等人提供的凝胶转移实验（10）显示，在对照细胞的细胞质提取物中存在NF-kB的DNA结合，而在Ad5-I kb转导的细胞中，无论是否给予30 Gy的电离辐射，都不存在NF-kB的DNA结合。这些实验是在细胞转导后48小时进行的，结果表明突变的I kB基因产物抑制了构成型NFkB活性和辐射诱导的激活。对这些数据的另一种解释可能是，从仍然存活的对照细胞中获得的细胞质提取物与从Ad5-I kb转导的细胞中获得的细胞质提取物不同。存活细胞分数的巨大差异使得报道的幸存者的生化比较不可靠。此外，使用凋亡细胞的胞质提取物必须考虑到靶向NFkB成分的caspases可能产生的影响（15）。
Cellular responses to ionizing radiation include activation of signal transduction cascades that may originate at the plasma membrane, cytoplasm, or nucleus. The cell’s success in dealing with radiation “stress” determines its survival or death. Activation of transcription factor NFkB, an immediate early response after exposure to ionizing radiations, functions to protect cells from apoptosis (programmed cell death), but the role of NFkB in mitotic cell death has not been fully defined (1,2). NF-kB is constitutively activated in lymphoid cells, immortal ataxia telangiectasia (AT) fibroblasts, and other cells (3–6).Activation of NF-kB after ionizing radiation requires intact ATM gene function in AT fibroblasts(7). Because AT fibroblasts exhibit dysregulated NFkB activation and extreme radiation sensitivity, it is reasonable to ask if radiation responses of tumor cells can be modified by inhibiting NFkB. NF-kB/Rel transcription factors are activated by a variety of different signals in pathways that converge on the phosphorylation and degradation of I kBs, inhibitors of NFkB. This results in the unmasking of the nuclear localization signals that lead to translocation of NFkB/Rel dimers into the nucleus. Five known mammalian NFkB/Rel proteins include c-Rel, p65 (RelA), RelB, p50 (NFkB1), and p52 (NFkB2) that bind DNA as homodimers or heterodimers (8). It should also be noted that p50–p65/I kB is not the only complex relevant to NF-kB activation. The p50–p65 heterodimer can interact with other IkB family members, and NFkB can also exist in other homodimer or heterodimer complexes, such as p50–p50 and p65–p65 (8). Furthermore, the I kB family consists of three members—I kBa, IkBb, and IkB«. In addition, the carboxylterminal regions of the precursors for p50 and p52, p105 and p100, respectively, can also function as inhibitory proteins. These inhibitors respond differentially to NFkB-inducing signals in a cell type-dependent and stimulus-dependent manner in vivo (9). In this issue of the Journal, Pajonk et al. (10) report that activation of NFkB does not determine the intrinsic radiosensitivity of cancer cells. Cells were transduced with an adenoviral vector expressing a mutant form of I kBa, an inhibitor of NF-kB. The resultant clonogenicity of cells transduced with mutant IkBa was markedly reduced to 7.4% (compared with 29.5% for control cells), and the apoptotic index was 90%. Therefore, subsequent radiation survival experiments were done on relatively small subsets of cells, whereas biochemical assays were done on mostly apoptotic cells. It is unusual to see publication of essentially negative studies in the Journal. However, there is potential value if definitive data can provide direction for other scientists in the field. Clinically, intrinsic radiation sensitivity has been implicated in tumor curability (11).The ability to manipulate radiation sensitivity offers a potential strategy for improving the therapeutic ratio in cancer treatment. Therefore, an examination of the experimental results and data interpretation underlying the negative radiobiologic conclusions of Pajonk et al. (10) are appropriate. 1) Are PC3 and HD-MyZ cells resistant to radiation? Pajonk et al. (10) state that the two cell lines were chosen because they have high constitutive levels of NFkB that might confer relatively high resistance to radiation. Strictly speaking, radiation resistance is an imprecise term that may include various aspects of cellular responses to ionizing radiation (12). On the other hand, radiation sensitivity is defined by the steepness of the terminal slope of the cellular radiation survival curve (single-hit multitarget model). ADo value (the dose required to decrease survival by 1/ e) can be determined for relative quantitation of radiation sensitivities. Do values for PC3 and HD-MyZ cells are estimated to be 1.0–1.1 Gy [Fig. 3 in (10)], which is quite sensitive for tumor cell lines and is comparable in sensitivity to AT heterozygote fibroblasts (13). Disruption of signal transduction pathways of resistant cells has been reported to result in radiosensitization (14), but it may be difficult to make already sensitive cells more sensitive. An alternative interpretation of the constitutively activated NFkB levels observed in PC3 and HD-MyZ cells may be related to apoptotic stresses rather than reflecting potential resistance. This is consistent with subsequent observations of Pajonk et al. (10) that 90% of these cells die of apoptosis when mutant I kB-a is expressed, inhibiting protective functions of activated NFkB. 2) Does the I kBa superrepressor product inhibit radiationinduced NFkB activation in the small subset of transduced cells that do not undergo apoptosis? Gel-shift experiments are offered by Pajonk et al. (10) showing DNA binding by NF-kB to be present in cytosolic extracts from control cells but not from Ad5-I kB-transduced cells that were either given 30 Gy of ionizing radiation or not. These experiments were performed 48 hours after transduction of cells and are interpreted to show that the mutated I kB gene product inhibits constitutive NFkB activity and radiation-induced activation. An alternative interpretation of these data may be that cytosolic extracts obtained from still viable control cells differ from those obtained from the Ad5-I kB-transduced cells that were mostly dead. The substantial disparity of surviving cell fractions makes the reported biochemical comparisons in survivors unreliable. Furthermore, the use of cytosolic extracts from cells undergoing apoptosis must include consideration of possible effects of caspases targeting components of NFkB (15).
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DOI：
10.1073/pnas.96.1.49
发表时间：
1999-01-05
影响因子：
11.1
作者：
Simeonidis, S;Stauber, D;Thanos, D
通讯作者：
Thanos, D
影响因子：
56.9
作者：
JUNG, M;ZHANG, Y;DRITSCHILO, A
通讯作者：
DRITSCHILO, A
对放疗反应异常严重的患者和共济失调性毛细血管扩张杂合子患者的真皮成纤维细胞培养物对分次放疗的反应。
DOI：
--
发表时间：
1995
期刊：
Clinical cancer research : an official journal of the American Association for Cancer Research.
影响因子：
--
作者：
DahlBerg,WK;Little,JB
通讯作者：
Little,JB
DOI：
10.1016/0360-3016(88)90297-0
发表时间：
1988-09-01
影响因子：
7
作者：
WEICHSELBAUM, RR;BECKETT, MA;DRITSCHILO, A
通讯作者：
DRITSCHILO, A