Data from: Trophoblast survival signaling during human placentation requires HSP70 activation of MMP2-mediated HBEGF shedding

Survival of trophoblast cells in the low oxygen environment of human
placentation requires metalloproteinase-mediated shedding of HBEGF and
downstream signaling. A matrix metalloproteinase (MMP) antibody array and
quantitative RT-PCR revealed upregulation of MMP2 post-transcriptionally
in human first trimester HTR-8/SVneo trophoblast cells and placental
villous explants exposed to 2% O2. Specific MMP inhibitors established the
requirement for MMP2 in HBEGF shedding and upregulation. Because
α-amanitin inhibited the upregulation of HBEGF, differentially expressed
genes were identified by next-generation sequencing of RNA from
trophoblast cells cultured at 2% O2 for 0, 1, 2 and 4 h. Nine genes, all
containing HIF-response elements, were upregulated at 1 h, but only HSPA6
(HSP70B’) remained elevated at 2–4 h. The HSP70 chaperone inhibitor VER
155008 blocked upregulation of both MMP2 and HBEGF at 2% O2, and increased
apoptosis. However, both HBEGF upregulation and apoptosis were rescued by
exogenous MMP2. Proximity ligation assays demonstrated interactions
between HSP70 and MMP2, and between MMP2 and HBEGF, supporting the concept
that MMP2-mediated shedding of HBEGF, initiated by HSP70, contributes to
trophoblast survival at the low O2 concentrations encountered during the
first trimester, and is essential for successful pregnancy outcomes.
Trophoblast survival during human placentation, when oxygenation is
minimal, required HSP70 activity, which mediated MMP2 accumulation and the
transactivation of anti-apoptotic ERBB signaling by HBEGF shedding.

人类胎盘形成过程中，滋养层细胞在低氧环境下的存活依赖于金属蛋白酶介导的HBEGF脱落及其下游信号传导。基质金属蛋白酶（MMP）抗体阵列与定量逆转录聚合酶链反应（quantitative RT-PCR）结果显示，在暴露于2% O₂的人早孕期HTR-8/SVneo滋养层细胞及胎盘绒毛外植体中，MMP2存在转录后上调。特异性MMP抑制剂实验证实，HBEGF的脱落与上调过程需要MMP2的参与。由于α-鹅膏蕈碱（α-amanitin）可抑制HBEGF的上调，研究人员通过对在2% O₂条件下培养0、1、2及4小时的滋养层细胞RNA进行下一代测序（next-generation sequencing），鉴定出差异表达基因。其中9个均含缺氧诱导因子应答元件（HIF-response elements）的基因在1小时时上调，但仅HSPA6（HSP70B'）在2-4小时仍维持高表达水平。热休克蛋白70（HSP70）分子伴侣抑制剂VER155008可阻断2% O₂条件下MMP2与HBEGF的上调，并增加细胞凋亡。然而，外源性MMP2可逆转HBEGF上调受抑及凋亡增加的状态。邻近连接分析（proximity ligation assays）结果显示HSP70与MMP2、MMP2与HBEGF之间存在相互作用，这支持了以下观点：由HSP70启动的MMP2介导的HBEGF脱落，有助于滋养层细胞在早孕期低氧环境中存活，且对妊娠的成功结局至关重要。在人类胎盘形成过程中（此时氧合作用最弱），滋养层细胞的存活依赖于HSP70的活性——HSP70可介导MMP2的积累，以及通过HBEGF脱落实现抗凋亡ERBB信号通路的反式激活。