Lysophosphatidic acid (LPA) is a membrane-derived lysophospholipid that exists in the plasma and platelets. It exerts its functions through activation of various LPA receptors (LPARs), which belong to the family of G protein-coupled receptors. Activation of LPARs has important roles in stem cell differentiation. However, how LPA affects human hematopoietic stem cell (HSC) differentiation remains elusive. In our previous studies, we have suggested that LPA receptor 2 (LPA) and LPA receptor 3 (LPA) play opposing roles and may act as a molecular switch during megakaryocytic differentiation in K562 cells. In this study, human CD34 HSCs and zebrafish are adopted to investigate the roles of LPA during megakaryopoiesis/thrombopoiesis in vitro and in vivo. Our results show that LPAR3 mRNA expression level is decreased upon induction by thrombopoietin and stem cell factor in human HSCs. Using pharmacological activators and shRNA knockdown experiments, we demonstrate that activation of LPA inhibits megakaryopoiesis in human HSCs. In addition, pharmacological activation of LPA suppressed thrombopoiesis in zebrafish. Furthermore, blockage of LPA translation by morpholino increased the number of CD41-GFP cells in Tg(CD41:eGFP) zebrafish. Moreover, the mRNA expression level of zCD41 increased significantly in LPA-knockout zebrafish. These results clarify the negative role of LPA during megakaryopoiesis and provide important information for potential treatments of related diseases, such as megakaryopenia.

Insulin-like growth factor (IGF) signaling pathway is an important regulatory mechanism of tumorigenesis and drug resistance in many cancers. The present study explored the potential synergistic effects between IGF receptor (IGFR) inhibition and other molecular targeted agents (MTA) in HCC cells. HCC cell lines (Hep3B, PLC5, and SK-Hep1) and HUVECs were tested. The MTA tested included sorafenib, sunitinib, and the IGFR kinase inhibitor NVP-AEW541. The potential synergistic antitumor effects were tested by median dose effect analysis and apoptosis assay in vitro and by xenograft models in vivo. The activity and functional significance of pertinent signaling pathways and expression of apoptosis-related proteins were measured by RNA interference and Western blotting. We found that IGF can activate IGFR and downstream AKT signaling activities in all the HCC cells tested, but the growth-stimulating effect of IGF was most prominent in Hep3B cells. NVP-AEW541 can abrogate IGF-induced activation of IGFR and AKT signaling in HCC cells. IGF can increase the resistance of HCC cells to sunitinib. The apoptosis-inducing effects of sunitinib, but not sorafenib, were enhanced when IGFR signaling activity was inhibited by NVP-AEW541 or IGFR knockdown. Chk2 kinase activation was found contributory to the synergistic anti-tumor effects between sunitinib and IGFR inhibition. Our data indicate that the apoptosis-potentiating effects of IGFR inhibition for HCC may be drug-specific. Combination therapy of IGFR inhibitors with other MTA may improve the therapeutic efficacy in HCC.

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Lysophosphatidic acid (LPA) is a small bioactive phospholipid enriched in serum and exerts diverse cellular functions through activation of multiple G Protein‐coupled receptors, LPA1‐LPA6. Our previous studies demonstrated that lysophosphatidic acid receptor 3 (LPA3) is critical for megakaryopoiesis and erythropoiesis in zebrafish. However, the roles of LPA3 during the development of other organs of zebrafish remain unclear. Here, we established a zebrafish strain with targeted disruption of lysophosphatidic acid receptor 3 (Lpar3) gene by Transcription Activator‐like Effector Nuclease (TALEN). Paraffin‐section and hematoxylin‐eosin staining were performed for histological analysis to clarify the effects of LPA3 in different organs of adult zebrafish. Histological analysis results showed a shrinking of hematopoietic tissue in kidney marrow of LPA3 deficient fish, implies essential roles of LPA3 in hematopoiesis. In addition, bile duct hyperplasia and mature spermatids loss were also observed in LPA3 deficient fish. Nevertheless, no obvious morphological differences in eyes, thymus, heart, and spleen were observed. Our observation provides tissues profiles of LPA3−/− animal and possible application of LPA3 deficient zebrafish as a disease model in the future.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.