Pardo Verónica Gonzalez, Facchinetti Maria Marta, Curino Alejandro, Boland Ricardo, de Boland Ana Russo
Departamento de Biología, Bioquímica & Farmacia, Universidad Nacional del Sur, San Juan 670, Bahia Blanca 8000, Argentina.
Biogerontology. 2007 Feb;8(1):13-24. doi: 10.1007/s10522-006-9031-0. Epub 2006 Jul 20.
In intestinal cells, 1alpha,25(OH)(2)-vitamin D(3) (1alpha,25(OH)(2)D(3)) regulates gene expression via the specific intracellular vitamin D receptor and induces fast non-transcriptional responses involving stimulation of transmembrane signal transduction pathways. In the present study, we analyzed, for the first time, alterations in p38 MAPK response to 1alpha,25(OH)(2)D(3) in rat enterocytes with ageing. In enterocytes from young rats, the hormone increased, in a time- and dose-dependent fashion, the phosphorylation of p38 MAPK, peaking at 3 min (+2-fold). Basal levels of p38 MAPK phosphorylation were lower in enterocytes from old rats and the hormone response was greatly diminished (+0.5-fold at 3 min). p38 MAPK phosphorylation impairment in old animals was not related to significant changes of the kinase protein expression and do not explain the decreased response to 1alpha,25(OH)(2)D(3). Extracellular and intracellular Ca(2+) chelation or c-Src pharmacological inhibition suppressed hormone activation of p38 MAPK in both, young and aged rats, demonstrating that Ca(2+) and the non-receptor tyrosine kinase c-Src are required for full activation of p38 MAPK in cells stimulated with 1alpha,25(OH)(2)D(3). Two other vitamin D(3) metabolites, 25(OH)D(3) and 24,25(OH)(2)D(3, )also enhanced p38 phosphorylation, and to a similar extent than 1alpha,25(OH)(2)D(3), an ability that is lost with ageing. Enterocyte exposure to the hormone also resulted in the rapid induction of c-fos protein (peaking at 5 min, +3-fold) and to a greater extent than that of mRNA induction. With ageing, 1alpha,25(OH)(2)D(3)-dependent increase of c-fos protein level was diminished, but c-fos mRNA expression was not different from young animals. Impairment of 1alpha,25(OH)(2)D(3) activation of p38 MAPK upon ageing and abnormal hormone regulation of the c-fos oncoprotein synthesis may affect intestinal cell function.
在肠道细胞中,1α,25(OH)₂-维生素D₃(1α,25(OH)₂D₃)通过特异性细胞内维生素D受体调节基因表达,并诱导涉及跨膜信号转导途径刺激的快速非转录反应。在本研究中,我们首次分析了衰老大鼠肠上皮细胞中p38丝裂原活化蛋白激酶(p38 MAPK)对1α,25(OH)₂D₃反应的变化。在年轻大鼠的肠上皮细胞中,该激素以时间和剂量依赖性方式增加p38 MAPK的磷酸化,在3分钟时达到峰值(增加2倍)。老年大鼠肠上皮细胞中p38 MAPK磷酸化的基础水平较低,且激素反应大大减弱(3分钟时增加0.5倍)。老年动物中p38 MAPK磷酸化受损与激酶蛋白表达的显著变化无关,也无法解释对1α,25(OH)₂D₃反应的降低。细胞外和细胞内钙螯合或c-Src药理学抑制均抑制了年轻和老年大鼠中p38 MAPK的激素激活,表明钙(Ca²⁺)和非受体酪氨酸激酶c-Src是1α,25(OH)₂D₃刺激细胞中p38 MAPK完全激活所必需的。另外两种维生素D₃代谢物,25(OH)D₃和24,25(OH)₂D₃,也增强了p38磷酸化,且程度与1α,25(OH)₂D₃相似,这种能力会随着衰老而丧失。肠上皮细胞暴露于该激素还导致c-fos蛋白的快速诱导(5分钟时达到峰值,增加3倍),且程度大于mRNA诱导。随着衰老,1α,25(OH)₂D₃依赖性c-fos蛋白水平的增加减弱,但c-fos mRNA表达与年轻动物无异。衰老时1α,25(OH)₂D₃对p38 MAPK的激活受损以及c-fos癌蛋白合成的异常激素调节可能会影响肠道细胞功能。