Kawashima Yohei, Kushida Nobuhiro, Kokubun Shuko, Ogawa Soichiro, Shiomi Homare, Ishibashi Kei, Aikawa Ken, Ikegami Kentaro, Nomiya Masanori, Yamaguchi Osamu
Department of Urology, Fukushima Medical University School of Medicine, Fukushima, Japan.
Division of Bioengineering and LUTD Research, Nihon University College of Engineering, Koriyama, Japan.
Int J Urol. 2015 Aug;22(8):778-84. doi: 10.1111/iju.12799. Epub 2015 Jun 3.
To determine whether lysophosphatidic acid activates the mitogen-activated protein kinase and increases DNA synthesis in human bladder smooth muscle cells, and to examine the involvement of lysophosphatidic acid and lysophosphatidic acid receptor in mechanical stretch-induced mitogen-activated protein kinase activation in cultured human bladder smooth muscle cells.
TaqMan reverse transcription polymerase chain reaction was used to determine the mRNA expression levels of six lysophosphatidic acid receptor subtypes. Mitogen-activated protein kinase activity enhanced by either lysophosphatidic acid or mechanical stretch was measured by western blotting. The effect of lysophosphatidic acid on DNA synthesis was assessed by 5-bromo-2'-deoxy-uridine incorporation assay.
Lysophosphatidic acid 1 subtype mRNA was predominantly expressed (96%). Lysophosphatidic acid activated the mitogen-activated protein kinase in a concentration-dependent manner. C-jun NH2 -terminal kinase showed the highest activity among the three subsets of the mitogen-activated protein kinase family members (c-jun NH2 -terminal kinase, extracellular signal-regulated kinases, p38). Lysophosphatidic acid also increased incorporation of 5-bromo-2'-deoxy-uridine. These responses were suppressed by Ki16425 (lysophosphatidic acid receptor antagonist). Mechanical stretch mainly induced c-jun NH2 -terminal kinase activation. This activation was partially inhibited by Ki16425.
Lysophosphatidic acid might activate the c-jun NH2 -terminal kinase component of the mitogen-activated protein kinase family and DNA synthesis through lysophosphatidic acid receptors (presumably, through lysophosphatidic acid 1) in human bladder smooth muscle cells. The present study also implicates the involvement of lysophosphatidic acid and lysophosphatidic acid receptors in mechanical stretch-induced c-jun NH2 -terminal kinase activation. Lysophosphatidic acid receptor can be partially activated by mechanical stretching through lysophosphatidic acid-dependent or independent mechanism.
确定溶血磷脂酸是否激活丝裂原活化蛋白激酶并增加人膀胱平滑肌细胞中的DNA合成，以及研究溶血磷脂酸和溶血磷脂酸受体在培养的人膀胱平滑肌细胞机械牵张诱导的丝裂原活化蛋白激酶激活中的作用。
采用TaqMan逆转录聚合酶链反应来测定六种溶血磷脂酸受体亚型的mRNA表达水平。通过蛋白质印迹法测定溶血磷脂酸或机械牵张增强的丝裂原活化蛋白激酶活性。通过5-溴-2'-脱氧尿苷掺入试验评估溶血磷脂酸对DNA合成的影响。
溶血磷脂酸1亚型mRNA占主要表达（96%）。溶血磷脂酸以浓度依赖性方式激活丝裂原活化蛋白激酶。在丝裂原活化蛋白激酶家族成员的三个亚组（c-Jun氨基末端激酶、细胞外信号调节激酶、p38）中，c-Jun氨基末端激酶活性最高。溶血磷脂酸还增加了5-溴-2'-脱氧尿苷的掺入。这些反应被Ki16425（溶血磷脂酸受体拮抗剂）抑制。机械牵张主要诱导c-Jun氨基末端激酶激活。这种激活被Ki16425部分抑制。
溶血磷脂酸可能通过人膀胱平滑肌细胞中的溶血磷脂酸受体（可能是通过溶血磷脂酸1）激活丝裂原活化蛋白激酶家族的c-Jun氨基末端激酶成分并促进DNA合成。本研究还表明溶血磷脂酸和溶血磷脂酸受体参与机械牵张诱导的c-Jun氨基末端激酶激活。溶血磷脂酸受体可通过溶血磷脂酸依赖性或非依赖性机制被机械牵张部分激活。
