Physiological significance of proHB-EGF ectodomain shedding in epideimal development

1)HB-EGF是EGF家族中的一员。为了研究HB-EGF在活体中的作用，我们建立了HE-EGF基因敲除小鼠HB^；；del/del&gt；Hb^&lt；del/del&gt；小鼠出现严重心力衰竭并伴有扩张的室腔，类似于条件性ErbB2基因敲除小鼠。Hb^&lt；del/del&gt；小鼠的心脏瓣膜也变大，类似于EGFR基因敲除小鼠。在Hb^&lt；del/del&gt；心脏中，ErbB2和ErbB4的固有酪氨酸磷酸化显著降低。2)HB-EGF首先以膜锚定形式(proHB-EGF)合成，其可溶性形式(SHB-EGF)是通过胞外脱落FIOM释放出来的。为了检验ProHb-EGF在体内加工的意义，我们通过靶向基因替换产生了突变小鼠，表达该分子的不可切割形式(Hb^&lt；uc&gt；)或跨膜结构域截断形式(Hb^&lt；Δtm&gt；)。Hb^&lt；uc/uc&gt；小鼠出现严重的心力衰竭和e…更多的心脏瓣膜变大，表型与proHB-EGF缺失的小鼠相似。另一方面，接种Hb^lt；Δ的小鼠皮肤和心脏均表现出严重的增生。这些结果表明，ProHb-EGF胞外域的脱落是Hb-EGF在体内发挥作用所必需的，这一过程需要严格控制。3)Hb^；lt；del/del&gt；小鼠在表皮发育过程中没有表现出任何异常。因此，我们研究了HE-EGF在再上皮化中的作用。HB-EGF在成人表皮中的表达在稳态时几乎检测不到，但在全身创伤和局部应用维甲酸(Tra)后，HB-EGF的表达明显增强。在选择性去除HB-EGF表达的突变小鼠(Hb^&lt；del/del&gt；-K5-Cre)、Hb^&lt；UC/UC&gt；小鼠和正常HB-EGF基因敲除小鼠(Hb^&lt；lox/iox&gt；)中，评估了反式反应诱导的角质形成细胞的过度增殖水平和创面愈合的程度。在Hb^&lt；del/del&gt；-K5-Cre小鼠和HBuc/UC小鼠中，反式激活因子诱导的表皮过度增殖和创面愈合活性均低于HBlox/lox小鼠。提示HB-EGF的诱导及其胞外结构域脱落是体内再上皮化过程中的重要事件。较少

1)HB-EGF is a member of the EGF family of giowth factors. To examine the role of HB-EGF in viva, we generated HE-EGF-knockout mice HB^<del/del>). HB^<del/del> mice developed severe heart failure with dilated ventricular chambers, similarly to conditional ErbB2 knockout mice. HB^<del/del> mice developed also enlarged cardiac valves, similarly to EGFR knockout mice. Constitulive tyrosine phosphorylation of both ErbB2 and ErbB4 was significantly reduced in HB^<del/del> hearts. These indicated that HB-EGFactivalion of ErbB is essential for normal heartfunclion.2)HB-EGF is first synthesized as a membrane-anchored form (proHB-EGF), and its soluble form (sHB-EGF) is released by ectodomain shedding fiom proHB-EGF. To examine the significance of proHB-EGF processing in vivo, we generated mutant mice by targeted gene replacement, xpressing either an uncleavable form (HB^<uc>) or a transmembrane domain-truncated form (HB^<Δtm>) of the molecule. HB^<uc/uc> mice developed severe heart failure and e … More nlarged heart valves, phenotypes similar to those in proHB-EGF-null mice. On the other hand, mice canying HB^<Δtm> exhibited severe hyperplasia in both skin and heart These results indicate that ectodomain shedding of proHB-EGF is essential for HB-EGF function in viva, and that this piocess requires strict control.3)HB^<del/del> mice did not show any abnormalities in developmental piocess of epidermis. Thus, we investigated the role of HE-EGF in re-epithelization. HB-EGF expression in adult epidermis was hardly detected at the steady state, but it was strongly induced after full-thickness wounding and topical treatment with retinoic acid (tRA). The level of hyperprolifemtion of keratinocytes induced by tRA-ireatinent and that of wound healing were estimated among the mutant mice selectively ablated HB-EGF expression in basal keratinocytes (HB^<del/del>-K5-Cre),HB^<uc/uc> mice and normal HB-EGF cDNA knock-in mice (HB^<lox/iox>) as an experimenrtal control. In HB^<del/del>-K5-Cre mice and in HBuc/uc mice, both tRA-induced epidermal hyperproliferation and wound healing activity was reduced compared with HBlox/lox mice. These indicated that the induction of HB-EGF and its ectodomain shedding is an important event in the re-epithelization in vivo. Less