Wang Gerald J, Brenner-Anantharam Andrea, Vaughan E Darracott, Herzlinger Doris
Departments of Urology and Physiology and Biophysics (ABA, DL), New York-Presbyterian Hospital, Weill Medical College of Cornell University, New York, New York, USA.
J Urol. 2009 Jan;181(1):401-7. doi: 10.1016/j.juro.2008.08.117. Epub 2008 Nov 17.
Congenital ureteropelvic junction obstruction has been associated with aberrant ureteral smooth muscle organization. Recent evidence has shown that BMP4 may be involved in ureteral morphogenesis. We determined whether the disruption of BMP4 signaling results in abnormal smooth muscle investment of the ureter and ureteropelvic junction.
We used a Cre mediated Bmp4 knockout system to conditionally excise the Bmp4 gene in developing mouse embryos. Kidney rudiments were isolated from embryos at varying gestational ages from WT and conditional knockout mice. Metanephric kidney explants were cultured in the presence or absence of the BMP antagonist Noggin. Agarose beads pre-incubated with Gremlin, another BMP antagonist, were used for localized disruption of BMP signaling. Frozen sections and whole metanephric explants were then analyzed by immunofluorescence.
Bmp4 gene excision resulted in a dose dependent loss of ureteral smooth muscle. Antagonism of BMP signaling inhibited ureteral smooth muscle investment in a dose dependent manner and was paralleled by a dose dependent decrease in the immediate downstream targets of BMP signaling, phosphorylated Smad1, 5 and 8. Localized antagonism of BMP resulted in the focal disruption of ureteral smooth muscle investment.
We report that decreased BMP signaling, whether by the loss of BMP4 in vivo or direct antagonism in vitro, results in a gradual reduction of the normal, well organized coat of smooth muscle surrounding the ureter. Our results also suggest that this occurs via a direct Smad dependent pathway. This raises the possibility that abnormalities in BMP4 signaling may have a role in the development of congenital ureteropelvic junction obstruction.
先天性肾盂输尿管连接部梗阻与输尿管平滑肌组织异常有关。最近的证据表明，BMP4可能参与输尿管形态发生。我们确定BMP4信号通路的破坏是否会导致输尿管及肾盂输尿管连接部平滑肌覆盖异常。
我们使用Cre介导的Bmp4基因敲除系统在发育中的小鼠胚胎中条件性切除Bmp4基因。从野生型和条件性敲除小鼠不同胎龄的胚胎中分离出肾原基。在有或没有BMP拮抗剂Noggin的情况下培养后肾肾外植体。用另一种BMP拮抗剂Gremlin预孵育的琼脂糖珠用于局部破坏BMP信号通路。然后通过免疫荧光分析冰冻切片和整个后肾外植体。
Bmp4基因切除导致输尿管平滑肌呈剂量依赖性缺失。BMP信号通路的拮抗以剂量依赖性方式抑制输尿管平滑肌覆盖，同时BMP信号通路的直接下游靶点磷酸化Smad1、5和8呈剂量依赖性减少。BMP的局部拮抗导致输尿管平滑肌覆盖的局灶性破坏。
我们报告，无论是体内BMP4缺失还是体外直接拮抗导致的BMP信号通路减弱，都会导致输尿管周围正常、组织良好的平滑肌包膜逐渐减少。我们的结果还表明，这是通过直接的Smad依赖性途径发生的。这增加了BMP4信号通路异常可能在先天性肾盂输尿管连接部梗阻的发生中起作用的可能性。
