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The study of secretory protein kinase is an emergent research field in recent years. The secretion phenomenon of type II cGMP‐dependent protein kinase (PKG II) was found in our latest research and our previous study confirmed that PKG II inhibited platelet‐derived growth factor receptor β (PDGFRβ) activation induced by platelet‐derived growth factor BB (PDGF‐BB) within the gastric cancer cells. Thus, this study was designed to investigated effect of secretory PKG II on PDGFRβ. Transwell assay and CCK8 assay indicated that secretory PKG II reversed PDGF‐BB‐induced cell migration, invasion, and proliferation. Immunoprecipitation, GST pull down and Western blotting results showed that secretory PKG II combined with extracellular domains of PDGFRβ and phosphorylated it, and thereby inhibited PDGF‐BB‐induced activation of PDGFRβ, and downstream PI3K/Akt and MAPK/ERK pathways. Mutation at Ser254 of PDGFRβ to alanine abolished the above inhibitory effects of secretory PKG II on PDGFRβ, indicating that Ser254 was the specific site phosphorylated by secretory PKG II. In conclusion, secretory PKG II inhibited PDGFRβ activation via Ser254 site. [ABSTRACT FROM AUTHOR]

Abstract: Previous studies revealed that type II cGMP‐dependent protein kinase G (PKG II) could inhibit the activation of epidermal growth factor receptor (EGFR) which is a widely investigated RTK. PDGFR belongs to family of receptor tyrosine kinases (RTKs) too. However, the effect of PKG II on PDGFR activation is not clear yet. This study investigated potential regulatory effect of PKG II on activation of PDGFRβ and the downstream signaling transductions in gastric cancer. The results from CCK8 assay and Transwell assay indicated that PDGF‐BB induced cell proliferation and migration. Activated PKG II reversed the above variations caused by PDGF‐BB. Immunoprecipitation and Western blotting results showed that PKG II combined with PDGFRβ and phosphorylated this receptor, and thereby inhibited PDGF‐BB induced activation of PDGFRβ, and MAPK/ERK and PI3K/Akt mediated signal transduction pathways. Based on the prediction by phosphorylation site software, Ser643 and Ser712 were mutated to alanine respectively which prevented phosphorylation at these sites. Mutation at Ser712 abolished the inhibitory function of PKG II on PDGFRβ activation but mutation of Ser643 had no such an effect, indicating that Ser712 was PKG II‐specific phosphorylating site of PDGFRβ. In conclusion, PKG II inhibited PDGFRβ activation in gastric cancer via phosphorylating Ser712 of this RTK. [ABSTRACT FROM AUTHOR]

PKG II, cGMP-dependent protein kinase type II; EGF, epidermal growth factor; Ad-LacZ, adenoviral vector containing the -galactosidase gene; Ad-PKG II, adenoviral vector containing the PKG II gene; cGMP, cyclic guanosine monophosphate. PKG II, cGMP-dependent protein kinase type II; Ad-LacZ, adenoviral vector containing the -galactosidase gene; Ad-PKG II, adenoviral vector containing the PKG II gene; 8-pCPT-cGMP, 8-(4-chlorophenylthio)-cGMP; cGMP, cyclic guanosine monophosphate; EGF, epidermal growth factor. [Extracted from the article]

Abstract: The small GTPases regulate many major biological processes in both tumorigenesis and tumor progression such as cell survival, actin cytoskeleton organization, cell polarity and movement. Wnt5a, a non-canonical Wnt family member, is implicated in the activation of small GTPases in breast cancer. We previously demonstrated that Wnt5a signaling stimulates the migration of breast cancer cells MDA-MB-231 via activating RhoA. However, we found here that RhoA activation was not enhanced by Wnt5a in breast cancer cells MCF-7. The conflicting results prompted us to further probe novel small GTPases in response to Wnt5a and investigate the mechanisms whereby cell migration is regulated. We showed here that Wnt5a dose dependently activated Dvl2, Rab35 and Rac1 and subsequently promoted the migration of MCF-7 cells, which was, however, abolished by knocking down Wnt5a expression via small interfering RNA (siRNA) transfection. Dvl2 siRNA significantly decreased background and Wnt5a-induced Rab35/Rac1 activation and, consequently, cell migration. Rab35 short hairpin RNA (shRNA) remarkably inhibited background and Wnt5a-induced Rac1 activation and cell migration. Additionally, blockade of Rac1 activation with Rac1 siRNA suppressed background and Wnt5a-induced cell migration. Co-immunoprecipitation and immunofluorescence assays showed that Dvl2 bound to Rab35 in mammalian cells. Taken together, we demonstrated that Wnt5a promotes breast cancer cell migration via the Dvl2/Rab35/Rac1 signaling pathway. These findings implicate Wnt5a signaling in regulating small GTPases, which could be targeted for manipulating breast cancer cell migration. [Copyright &y& Elsevier]

Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in clinical application because of their potential use in regenerative medicine and tissue engineering. However, the therapeutic application of MSCs still remain limited unless the favorable effect of MSCs for tumor growth in vivo and the long-term safety of the clinical applications of MSCs are better understood. In this study, MSCs derived from fetal bone marrow (FMSCs) and adult MSCs (AMSCs) alone or FMSCs and AMSCs with tumor cell line (F6 or SW480) together were transplanted subcutaneously into BALB/c-nu/nu mice to observe the outgrowth of tumor, and the characteristics of tumor cells were investigated by pathological and immunohistochemical methods, flow cytometry and real-time quantitative PCR. The results showed that both FMSCs and AMSCs could favor tumor growth in vivo. The pathologic examination revealed that tumor tissues had rich vessel distribution, extensive necrosis and invasion surrounding normal tissues, such as muscular tissue and subcutaneous tissue. In the immunohistochemical examination, tumor cells mixed with MSCs transplanted subcutaneously exhibited elevated capability of proliferation, rich angiogenesis in tumor tissues and highly metastatic ability. To understand whether MSCs affected the general properties of the tumor cells in vivo, the expression of some surface antigens and Bmi-1 gene of tumor tissue cells was detected in this study. The results indicated that these parameters were not affected after the interaction of MSCs with tumor cells in vivo. These findings suggested that MSCs could favor tumor growth in vivo. It is necessary to carry out a study for assurance of the long-term safety before MSCs were used as a therapy tools in regenerative medicine and tissue engineering. [Copyright &y& Elsevier]

Abstract: It has previously been implicated that nerve growth factor (NGF) with its high-affinity receptor tyrosine kinase A (TrkA) could play an important role in the growth modulation of human tumor cells, such as glioblastoma multiform cell lines and human breast cancer cell lines. However, the direct mitogenic effects of NGF and TrkA in these tumor cells still remain to be elucidated. Herein we show, by immunofluorescence staining, that NGF was colocalized with γ-tubulin at the centrosomes or the spindle poles throughout the cell cycle and phosphorylated TrkA was colocalized with α-tubulin at mitotic spindle in the glioma cell line U251. The results suggest that NGF concentrated to centrosome can recruit its receptor TrkA there and cause phosphorylation of the latter. The phosphorylated TrkA with the tyrosine kinase activity may phosphorylate the tubulin and promote the mitotic spindle assembly. By these mechanisms, NGF can modulate the mitosis of human glioma cells. [Copyright &y& Elsevier]