In vitro expression of endothelin-1 (ET-1) and the ET(A) and ET(B) ET receptors by the prostatic epithelium and stroma
In vitro expression of endothelin-1 (ET-1) and the ET(A) and ET(B) ET receptors by the prostatic epithelium and stroma
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DOI：
10.1210/jc.82.2.508
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发表时间：
1997-02-01
影响因子：
5.8
通讯作者：
Habib, FK
中科院分区：
文献类型：
--
作者：
Grant, ES;Brown, T;Habib, FK
对从前列腺癌细胞系 DU145 和 PC3 以及原代上皮细胞制备的总 RNA 进行 RT-PCR 分析表明存在内皮素-1 (ET-1) 信使 RNA (mRNA)。 LNCaP 细胞系和原代前列腺基质细胞都不具有 ET-1 mRNA 转录本。 72小时后，来自DU145、PC3和原代上皮细胞的72小时条件培养基含有免疫反应性ET浓度，分别相当于0.814+/-0.048、0.330+/-0.050和0.856+/-0.055 fmol/ml/10(6)细胞。 LNCaP（72小时后0.029+/-0.009 fmol/mL/10(6)细胞）和基质细胞（72小时后0.067+/-0.007 fmol/mL/10(6)细胞）表现出基础免疫反应性ET分泌。通过 RT-PCR 检查 ET(A) 和 ET(B) 基因表达表明，在前列腺癌细胞系中几乎完全检测不到 ET 受体 mRNA。 ET(A) 和 ET(B) mRNA 在前列腺上皮和基质的原代培养物中均可检测到。竞争性结合研究表明，原发性良性上皮细胞（解离常数=1.85x10(-10)mol/L；最大结合能力=2.7x10(4)结合位点/细胞）和间质（解离常数=1.93x10(-10)mol/L；最大结合能力=3.7x10(5)结合位点/细胞）中存在单一类别的结合位点。使用选择性ET受体拮抗剂证实体外表达的主要基质受体亚型是ET(B)。该受体似乎不与有丝分裂途径偶联，因为没有观察到对外源 ET-1 的生长反应或 ET-1 与 bFGF 之间的合作。同样，在 4 天的时间内没有观察到 ET-1 或 ET 转换酶抑制剂磷酰胺对良性上皮细胞的影响。
RT-PCR analysis of total RNA prepared from the prostate cancer cell lines DU145 and PC3 and from primary epithelial cells indicated the presence of endothelin-1 (ET-1) messenger RNA (mRNA). Neither the LNCaP cell line nor primary prostatic stromal cells possess ET-1 mRNA transcripts. Seventy-two-hour-conditioned media derived from DU145, PC3, and primary epithelia contain immunoreactive ET concentrations equivalent to 0.814+/-0.048, 0.330+/-0.050, and 0.856+/-0.055 fmol/ml/10(6) cells after 72 h, respectively. Basal immunoreactive ET secretion was exhibited by LNCaP (0.029+/-0.009 fmol/mL/10(6) cells after 72 h) and stromal cells (0.067+/-0.007 fmol/mL/10(6) cells after 72 h). Examination of ET(A) and ET(B) gene expression by RT-PCR demonstrates that ET receptor mRNA is almost completely undetectable in the prostate cancer cell lines. Both ET(A) and ET(B) mRNAs are detectable in primary cultures of prostatic epithelia and stroma. Competitive binding studies demonstrate a single class of binding site in both primary benign epithelia (dissociation constant =1.85x10(-10) mol/L; maximal binding capacity=2.7x10(4) binding sites/cell), and stroma (dissociation constant=1.93x10(-10) mol/L; maximal binding capacity=3.7x10(5) binding sites/cell). Use Of selective ETreceptor antagonists confirmed that the predominant stromal receptor subtype expressed in vitro is ET(B). This receptor seems not to be coupled to mitogenic pathways because no growth response to exogenous ET-1 or cooperation between ET-1 and bFGF could be observed. Similarly, no effect of ET-1 or the ET-converting enzyme inhibitor, phosphoramidon, on benign epithelial cells could be observed over a 4-day period.