Effects of second messengers on gap junctional intercellular communication of ovine luteal cells throughout the estrous cycle.
Effects of second messengers on gap junctional intercellular communication of ovine luteal cells throughout the estrous cycle.
复制标题
第二信使对整个发情周期绵羊黄体细胞间隙连接细胞间通讯的影响。
DOI：
10.1095/biolreprod65.3.777
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发表时间：
2001
影响因子：
3.6
通讯作者：
Redmer,DA
中科院分区：
文献类型：
--
作者：
Grazul-Bilska,AT;Reynolds,LP;Bilski,JJ;Redmer,DA
将超数排卵后第5、10和15天的黄体（CL）酶促分散，并将一部分细胞淘洗以获得富含小或大黄体细胞的组分。将混合的、小的和大的黄体细胞部分与不处理或与cAMP（dbcAMP或Rp-cAMPS）、蛋白激酶C（PKC; TPA或H-7）或钙（A23187、EGTA或A23187 + EGTA）的激动剂或拮抗剂一起孵育。激光细胞仪检测细胞间接触依赖性缝隙连接通讯（GJIC）的比率。收集用于孕酮（P4）放射免疫测定的培养基，将未经处理的黄体细胞固定用于免疫细胞化学或冷冻用于Western印迹分析。动情周期各阶段的黄体细胞均表现出GJIC。dbcAMP使动情周期各细胞的GJIC增加（P< 0.05）。Rp-cAMPS在第5天降低小黄体细胞的GJIC（P< 0.05），在第10天和第15天降低所有细胞类型的GJIC（P< 0.05）。TPA对动情周期各细胞类型的GJIC均有抑制作用（P< 0.01），而H-7对GJIC无影响。A23187降低了大黄体细胞接触小黄体细胞或仅接触大黄体细胞的GJIC（P< 0.05），而A23187 + EGTA降低了所有细胞类型的GJIC（P< 0.05）。对于混合型和大型黄体细胞，dbcAMP增加（P< 0.05），TPA和A23187 + EGTA减少（P< 0.05），P4分泌减少（P< 0.05）。A23187单独作用可使大部分黄体细胞P4分泌减少（P< 0.05），但对混合黄体细胞P4分泌无明显影响。在不同的细胞类型和不同的细胞周期中，GJIC与P4分泌率呈显著正相关（r= 0.113-0.249;P< 0.01）。免疫荧光和免疫印迹法检测连接蛋白43在培养的黄体细胞。因此，细胞内调节剂，如cAMP，PKC，或钙离子似乎调节GJIC，这可能是一个重要的机制，协调羊CL的功能。
Corpora lutea (CL) from Days 5, 10, and 15 after superovulation were enzymatically dispersed, and a portion of the cells were elutriated to obtain fractions enriched with small or large luteal cells. Mixed, small, and large luteal cell fractions were incubated with no treatment or with agonists or antagonists of cAMP (dbcAMP or Rp-cAMPS), protein kinase C (PKC; TPA or H-7), or calcium (A23187, EGTA, or A23187 + EGTA). The rate of contact-dependent gap junctional intercellular communication (GJIC) was evaluated by laser cytometry. Media were collected for progesterone (P4) radioimmunoassay, and luteal cells cultured with no treatment were fixed for immunocytochemistry or frozen for Western blot analysis. Luteal cells from each stage of the estrous cycle exhibited GJIC. The dbcAMP increased (P< 0.05) GJIC for all cell types across the estrous cycle. The Rp-cAMPS decreased (P< 0.05) GJIC for small luteal cells on Day 5 and for all cell types on Days 10 and 15. The TPA inhibited (P< 0.01), but H-7 did not affect, GJIC for all cell types across the estrous cycle. The A23187 decreased (P< 0.05) GJIC for large luteal cells touching only small or only large luteal cells, whereas A23187 + EGTA decreased (P< 0.05) GJIC for all cell types across the estrous cycle. For the mixed and large luteal cell fractions, dbcAMP increased (P< 0.05), but TPA and A23187 + EGTA decreased (P< 0.05), P4secretion. The A23187 alone decreased (P< 0.05) P4secretion by large, but not by mixed, luteal cells. For all days and cell types, the rate of GJIC and P4secretion were correlated (r= 0.113–0.249;P< 0.01). Connexin 43 was detected in cultured luteal cells by immunofluorescence and Western immunoblotting. Thus, intracellular regulators like cAMP, PKC, or calcium appear to regulate GJIC, which probably is an important mechanism for coordinating function of the ovine CL.