Maki A, Mohammad R M, Smith M, Al-Katib A
Department of Internal Medicine, Wayne State University-School of Medicine, Detroit, MI 48201, USA.
Differentiation. 1996 Mar;60(1):59-66. doi: 10.1046/j.1432-0436.1996.6010059.x.
We have previously demonstrated that the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA), induces differentiation of the acute lymphoblastic leukemia cell line, Reh, to a mature non-dividing state. Associated with this differentiation is the expression of ubiquitin carboxyl terminal hydrolase (UCH-L1). To investigate the role of UCH-L1 in TPA-induced Reh differentiation and apoptosis, molecular and chemical inhibition was used. Molecularly, a sequence-specific antisense oligodeoxynucleotide (AODN) directed against UCH-L1 transcript was used to inhibit the expression of the gene. In addition, its complementary sense oligodeoxynucleotide (SODN) was used to indicate the specificity of AODN action. Chemically, sodium borohydride (NaBH4), an inhibitor of UCH-L, was used to block the transcript product. TPA-induced changes in Reh cell growth and morphology, UCH-L1 protein expression, apoptosis contour, surface phenotype, and enzymatic profile were assessed in the presence or absence of NaBH4, AODN or SODN. As previously reported, TPA induced Reh cells to differentiate into monocytoid B lymphocytes and stimulated the apoptotic pathway. However, adding NaBH4 or AODN inhibited the TPA effect on all parameters measured except apoptosis. The sequence in which NaBH4 or AODN were added in relation to TPA did not affect any of the response variables measured. The use of SODN did not influence any of the parameters measured, indicating the specificity of the action. Thus, we conclude that UCH-L1 is involved in the differentiation process of the lymphoblastic leukemia cell line, Reh. Our data suggest that TPA-induced apoptosis of Reh cells has a separate pathway from that of differentiation or that UCH-L1 expression is independent of the apoptotic pathway.
我们之前已经证明，佛波酯12 - O - 十四烷酰佛波醇13 - 乙酸酯（TPA）可诱导急性淋巴细胞白血病细胞系Reh分化为成熟的非分裂状态。与这种分化相关的是泛素羧基末端水解酶（UCH - L1）的表达。为了研究UCH - L1在TPA诱导的Reh分化和凋亡中的作用，我们采用了分子和化学抑制方法。在分子层面，使用针对UCH - L1转录本的序列特异性反义寡脱氧核苷酸（AODN）来抑制该基因的表达。此外，使用其互补的正义寡脱氧核苷酸（SODN）来表明AODN作用的特异性。在化学层面，使用UCH - L的抑制剂硼氢化钠（NaBH4）来阻断转录产物。在存在或不存在NaBH4、AODN或SODN的情况下，评估TPA诱导的Reh细胞生长和形态变化、UCH - L1蛋白表达、凋亡轮廓、表面表型和酶谱。如先前报道的那样，TPA诱导Reh细胞分化为单核样B淋巴细胞并刺激凋亡途径。然而，添加NaBH4或AODN会抑制TPA对除凋亡外所有测量参数的影响。NaBH4或AODN相对于TPA的添加顺序不影响任何测量的反应变量。使用SODN不影响任何测量参数，表明了作用的特异性。因此，我们得出结论，UCH - L1参与了淋巴细胞白血病细胞系Reh的分化过程。我们的数据表明，TPA诱导的Reh细胞凋亡与分化过程有不同的途径，或者UCH - L1的表达独立于凋亡途径。
