Early pregnancy induced expression of prostaglandin E2 receptors EP2 and EP4 in the ovine endometrium and regulated by interferon tau through multiple cell signaling pathways

Prostaglandin E2 (PGE₂) plays pleiotropic roles at fetal-maternal interface during establishment of pregnancy. The objectives of the study were to: (i) determine regulation of PGE2 receptors EP1, EP2, EP3, and EP4 in the endometrium during the estrous cycle and early pregnancy; and (ii) understand endometrial epithelial and stromal cell-specific hormonal regulation of EP2 and EP4 in sheep. Results indicate that: (i) early pregnancy induces expression of EP2 and EP4 but not EP1 and EP3 proteins in the endometrium on days 12-16 compared to that of estrous cycle; (ii) intrauterine infusion of interferon tau (IFNT) increases expression of EP2 and EP4 proteins in endometrium; and (iii) IFNT activates distinct epithelial and stromal cell-specific JAK, EGFR, ERK1/2, AKT, or JNK signaling module to regulate expression of EP2 and EP4 proteins in the ovine endometrium. Our results indicate a role for EP2 and EP4-mediated PGE₂ signaling in endometrial functions and establishment of pregnancy in ruminants.     

miR-761通过靶向作用CRKL抑制卵巢癌SKOV3细胞增殖研究

目的探讨miR-761对卵巢癌SKOV3细胞增殖的影响以及相关机制。方法采用Lipofectamine 2000将miR-761过表达和沉默质粒载体分别转染卵巢癌SKOV3细胞后,采用Real-Time PCR验证其转染效率;采用CCK8法检测miR-761对卵巢癌SKOV3细胞增殖的影响;采用Real-Time PCR检测卵巢癌SKOV3细胞中CRKL mRNA表达变化。采用双荧光素酶报告基因验证miR-761和CRKL的存在靶向结合并明确其结合位点。将CRKL(+)和CRKL(-)慢病毒载体分别转染至卵巢癌SKOV3细胞;采用CCK8法检测CRKL对卵巢癌SKOV3细胞增殖的影响。将CRKL(+)和CRKL(-)质粒分别转染至已经建立的miR-761过表达和沉默细胞,采用CCK8检测卵巢癌SKOV3细胞增殖变化。结果与对照组比较,miR-761过表达显著抑制了卵巢癌SKOV3细胞的增殖,极大降低了CRKL在SKOV3细胞中的mRNA和蛋白水平;miR-761和CRKL存在靶向结合,并明确了其结合位点;CRKL过表达显著增强了卵巢癌SKOV3细胞增殖能力;CRKL过表达有效阻断了miR-761抑制SKOV3细胞增殖的作用。结论 miR-761能够通过靶向下调CRKL,抑制卵巢癌SKOV3细胞增殖。     

CRKL基因反义寡核苷酸诱导K562细胞的凋亡

背景与目的：近几年研究发现,CRKL（CT10regulator of kinase like)基因编码的蛋白在慢性粒细胞白血病（chronic myeloid leukemia,CML)发病中起重要作用。本实验研究CRKL基因反义寡核苷酸对慢性粒细胞白血病细胞系K562细胞凋亡的影响。方法：以脂质体为载体,用CRKL基因反义寡核苷酸转染K562细胞,通过活细胞计数,透射电镜,流式细胞术,DNALadder测定观察K562细胞形态学,细胞周期,基因表达等的变化。结果：在CRKL基因反义寡核苷酸作用下,K562细胞的生长明显出现抑制;流式细胞仪检测见二倍体峰前出现凋亡峰;在电镜下可见处于凋亡各期的K562细胞;提取其基因组DNA进行琼脂糖凝胶电泳,可见明显的细胞凋亡梯状条带,而对照组K562细胞未观察到上述变化。结论：CRKL基因是Ph染色体阳性CML发病的重要因素。     

CRKL基因反义寡核苷酸对K562细胞增殖的影响

目的：通过研究CRKL基因反义寡核苷酸对K562细胞增残活性的影响,探讨CRKL基因在Ph^ 慢性粒细胞性白血病（CML）发病中的作用,方法： 脂质体为载体,CRKL基因反义寡核苷酸转染K562细胞,通过活细胸记数,H^3-TdR掺入,RT－PCR等方法观察K562细胞的增殖活性及基因表达的变化,结果,CRKL反义寡核苷酸对K562细胞的增殖有抑制作用,结论：CRKL基因在Ph^ CML的发病中可能有重要作用,可作为CML治疗的新靶点。     

Novel therapeutic strategies for patients with NSCLC that do not respond to treatment with EGFR inhibitors

Introduction

Treatment with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) yields tumour responses in non-small cell lung cancer (NSCLC) patients harbouring activating EGFR mutations. However, even in long-lasting responses, resistance to EGFR TKIs invariably occurs.

Areas covered

This review examines resistance mechanisms to EGFR TKI treatment, which mainly arise from secondary EGFR mutations. Other resistance-inducing processes include mesenchymal–epithelial transition factor (MET) amplification, epithelial–mesenchymal transformation, phenotypic change from NSCLC to small-cell lung carcinoma, and modifications in parallel signalling pathways. Current therapeutic strategies to overcome these EGFR TKI resistance mechanisms focus on the inhibition or blocking of multiple members of the ErbB family. Several molecules which target multiple ErbB receptors are being investigated in NSCLC and other indications including afatinib, an ErbB Family Blocker, as well as dacomitinib and lapatinib. Novel, non-quinazoline, EGFR inhibitors, that also target EGFR activating and resistance (T790M) mutations, are currently under clinical development. Other therapeutic strategies include inhibition of parallel and downstream pathways, using agents which target heat shock protein (HSP)90 or poly (ADP-ribose) polymerase in addition to mammalian target of rapamycin (mTOR), monoclonal antibodies against the insulin-like growth factor-1 receptor, and fulvestrant-mediated oestrogen receptor regulation.

Conclusion

Improved understanding of mechanisms underlying resistance to EGFR TKIs emphasises the importance of a genotype-guided approach to therapy. Elucidation of resistance mechanisms is indeed crucial to target innovative therapeutic approaches and to improve the efficacy of anticancer regimes in NSCLC.     

CRKL knockdown promotes in vitro proliferation,migration and invasion,in vivo tumor malignancy and lymph node metastasis of murine hepatocarcinoma Hca-P cells

Our previous study (Biomed Pharmacother 2015;69:11) demonstrated that the over-expression of CRKL, a chicken tumor virus number 10 regulator of kinase-like protein, suppresses in vitro proliferation, invasion and migration of murine hepatocarcinoma Hca-P cell, a murine HCC cell with lymph node metastatic (LNM) rate of ∼25%. In current work, we investigated the effects of CRKL knockdown on the in vitro cell proliferation, migration and invasion, and on the in vivo tumor malignancy and LNM rate and level for Hca-P cells. Western blotting assay indicated that CRKL was down-regulated by ∼90% in a monoclonal CrkL-shRNA-transfected Hca-P cells. Compared with Hca-P and unrelated-shRNA-transfected Hca-P cell, the in vitro proliferation, migration and invasion potentials were significantly enhanced following CRKL stable deregulation. CRKL knock-down significantly promoted the tumorigenicity malignancy, LNM rates and level of Hca-P-transplanted mice. Consistent with our previous work, it can be concluded CRKL plays an important role in hepatocarcinoma cell proliferation, invasion and migration as well hepatocarcinoma malignancy and metastasis. It functions as a potential tumor suppressor in hepatocarcinoma.     

CRKL基因对K 562细胞的作用研究

目的：通过研究CRKL基因反义寡核苷酸（AODN）对K562细胞增殖及凋亡的作用。探讨CRKL基因对慢性粒细胞性白血病（CML）发病的影响。方法：应用CRKL基因反义寡核苷酸作用于K562细胞，通过^3H-TdR掺入，流式细胞术，Rt-PCR等方法观察K562细胞形态学，细胞周期，细胞动力学及基因表达等变化。结果：CRKL基因ASODN对K562细胞的增殖有抑制作用。并可诱导K562细胞的凋亡。结论：CRKL基因在CML的发病中可能有重要作用。