Localization of proliferating cell nuclear antigen in the developing and mature rat heart cell.
Localization of proliferating cell nuclear antigen in the developing and mature rat heart cell.
复制标题
增殖细胞核抗原在发育和成熟大鼠心脏细胞中的定位。
DOI：
10.1002/(sici)1097-0185(199608)245:4
复制
发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Courtney,R
中科院分区：
文献类型：
--
作者：
Marino,TA;Cao,W;Lee,J;Courtney,R
心肌细胞在出生后不久就停止分裂；这个停止之后是一个有限的时期，DNA合成和核分裂发生，没有细胞分裂。这一过程的规律尚不清楚。本研究的目的是探讨可能导致心肌细胞分裂停止的事件。DNA合成所需的一种蛋白质是增殖细胞核抗原（PCNA）。该蛋白是DNA聚合酶δ的辅助蛋白。方法取胎龄18天或出生后0、4、8、12、16天的大鼠。此外，该研究还使用了成人心脏。消化18天大的胎儿大鼠和0天大的新生大鼠的心脏。分离心肌细胞并置于培养中，用三甲基胸腺嘧啶分析DNA合成。从所有年龄的心脏中分离心室肌肉组织，并在液氮中冷冻，用于Northern和Western blot分析。结果胸苷嘧啶分析显示，血清刺激虽然显著增加了标记的胎儿心肌细胞的数量，但对培养的新生儿心肌细胞没有这种影响。Northern blot分析显示，从胎儿第18天到出生后第4天，PCNA mRNA的稳态水平保持不变。稳态水平在产后第二周下降，然后在第16天达到基础水平。成人心脏组织中仍存在PCNA信息。通过间接免疫荧光法和Western blotting检测，PCNA蛋白在出生后2周内可定位于心肌细胞核内。在出生后16天，在细胞质中发现了极少量的蛋白质，但在细胞核中没有发现。在成人心肌的细胞质部分，Western blotting几乎检测不到该蛋白。结论出生后心肌中PCNA信息和蛋白产物有所下降，但在成年心肌中均处于较低水平。然而，在成人心肌细胞中，PCNA蛋白并没有转移到细胞核中。涉及PCNA的事件与细胞分裂和DNA合成在这些细胞中停止的时间密切相关。©1996 Wiley‐Liss, Inc。
BackgroundThe cardiac muscle cell ceases to divide shortly after birth; this cessation is followed by a limited period when DNA synthesis and karyokinesis occur without cytokinesis. The regulation of this process is not known. The purpose of this study is to explore the possible events that could lead to the cessation of cardiac muscle cell division. One protein requisite for DNA synthesis is proliferating cell nuclear antigen (PCNA). This protein is the auxiliary protein of DNA polymerase δ.MethodsRats of fetal age day 18 or days 0, 4, 8, 12, and 16 after birth were obtained. In addition, adult hearts were used for this study. Hearts from the fetal day‐18 rats and the day‐0 neonatal rats were digested. Cardiac myocytes were isolated and placed in culture for an analysis of DNA synthesis by using tridiated thymidine. Ventricular muscle tissue was isolated from hearts of all ages and frozen in liquid nitrogen for Northern and Western blot analyses.ResultsTridiated thymidine analysis revealed that, although serum stimulation significantly increased the number of labeled fetal cardiac muscle cells, it did not have that effect on neonatal cardiac muscle cells in culture. Northern blot analysis revealed that the steady state levels of mRNA for PCNA remained constant from fetal day 18 through day 4 after birth. Steady state levels declined during the second postnatal week and then reached basal levels by day 16. PCNA message was still present in adult heart tissue. By using indirect immunofloursecence and Western blotting, PCNA protein could be located in the nucleus of cardiac muscle cells during the first 2 weeks after birth. At 16 days after birth, the protein was found in the cytoplasm in very low amounts but was not found in the nucleus. The protein was barely detectable by Western blotting in the cytoplasmic fraction from the adult myocardium.ConclusionsThe results of this study suggest that the PCNA message and protein product declined after birth, but both were present at low levels in the adult myocardium. However, the PCNA protein was not translocated to the nucleus in adult myocardial cells. The events involving PCNA correlated closely with the time period when cell division and then DNA synthesis ceased in these cells. © 1996 Wiley‐Liss, Inc.