Simulated Microgravity Influences VEGF, MAPK, and PAM Signaling in Prostate Cancer Cells

Abstract

Prostate cancer is one of the leading causes of cancer mortality in men worldwide. An unusual but unique environment for studying tumor cell processes is provided by microgravity, either in space or simulated by ground-based devices like a random positioning machine (RPM). In this study, prostate adenocarcinoma-derived PC-3 cells were cultivated on an RPM for time periods of 3 and 5 days. We investigated the genes associated with the cytoskeleton, focal adhesions, extracellular matrix, growth, survival, angiogenesis, and metastasis. The gene expression of signaling factors of the vascular endothelial growth factor (VEGF), mitogen-activated protein kinase (MAPK), and PI3K/AKT/mTOR (PAM) pathways was investigated using qPCR. We performed immunofluorescence to study the cytoskeleton, histological staining to examine the morphology, and a time-resolved immunofluorometric assay to analyze the cell culture supernatants. When PC-3 cells were exposed to simulated microgravity (s-µg), some cells remained growing as adherent cells (AD), while most cells detached from the cell culture flask bottom and formed multicellular spheroids (MCS). After 3-day RPM exposure, PC-3 cells revealed significant downregulation of the VEGF, SRC1, AKT, MTOR, and COL1A1 gene expression in MCS, whereas FLT1, RAF1, MEK1, ERK1, FAK1, RICTOR, ACTB, TUBB, and TLN1 mRNAs were not significantly changed. ERK2 and TLN1 were elevated in AD, and FLK1, LAMA3, COL4A5, FN1, VCL, CDH1, and NGAL mRNAs were significantly upregulated in AD and MCS after 3 days. After a 5-day culture in s-µg, the PC-3 cells showed significant downregulations of VEGF mRNA in AD and MCS, and FN1, CDH1, and LAMA3 in AD and SCR1 in MCS. In addition, we measured significant upregulations in FLT1, AKT, ERK1, ERK2, LCN2, COL1A1, TUBB, and VCL mRNAs in AD and MCS, and increases in FLK1, FN1, and COL4A5 in MCS as well as LAMB2, CDH1, RAF1, MEK1, SRC1, and MTOR mRNAs in AD. FAK1 and RICTOR were not altered by s-µg. In parallel, the secretion rate of VEGFA and NGAL proteins decreased. Cytoskeletal alterations (F-actin) were visible, as well as a deposition of collagen in the MCS. In conclusion, RPM-exposure of PC-3 cells induced changes in their morphology, cytoskeleton, and extracellular matrix protein synthesis, as well as in their focal adhesion complex and growth behavior. The significant upregulation of genes belonging to the PAM pathway indicated their involvement in the cellular changes occurring in microgravity.

Comprehensive Proteomic Profiling of Aqueous Humor Proteins in Proliferative Diabetic Retinopathy

“…13,14 The PI3K-Akt signaling pathway can further upregulate the MAPK and VEGF signaling pathways, which can lead to cell proliferation and angiogenesis. 15,16 Our data showed that integrin alpha 11 significantly increased in the AH of PDR subjects and may initiate the development of PDR.…”

“…15,16,23 HIF-1 responds to hypoxia and upregulates the vascular endothelial growth factor (VEGF) signaling pathway. 15,16 In turn, VEGF initiates the mitogen-activated protein kinase (MAPK)signaling pathway, resulting in cell proliferation and angiogenesis. 15,16 Overall, our pathway analysis and literature review enabled us to generate a novel biological process map for the development of PDR (Fig.…”

Effects of Simulated Microgravity on Ultrastructure and Apoptosis of Choroidal Vascular Endothelial Cells

“…The PI3K/Akt pathway regulates cell survival, proliferation, and motility. The PI3K-Akt pathway has been indicated in participating in eNOs production and inhibition of apoptosis in SMG exposed-cells ( Shi et al, 2012 ; Ferranti et al, 2014 ; Arun et al, 2017 ; Hybel et al, 2020 ). In the present study, it was confirmed that the p-AKT and p-PI3K protein, phosphorylated forms of PI3K and Akt, were enhanced along with the time of exposure to SMG which indicated that the concomitant survival signal was also activated when CVEC apoptosis was triggered under SMG.…”

Human epididymis protein 4 and Lewis y enhance chemotherapeutic resistance in epithelial ovarian cancer through the p38MAPK pathway

“…Therefore, we speculate that Lewis y modi ed HE4 promotes the chemotherapy-resistant biological behaviors of ovarian cancer through the p38MAPK signaling pathway. We then constructed a PPI network, using the MCC and MCODE algorithms, and identi ed four hub genes among the overlapped genes (VEGFA, EGR1, HIF1A, and PTGS2); analyzing the known functions of these genes can provide ideas and breakthroughs for prospective studies on HE4 and Lewis y. VEGF-mediated MAPK/ERK and PI3K/AKT signaling pathways play an important role in regulating the growth, proliferation, migration, and angiogenesis of tumor cells and vascular endothelial cells 36 ; moreover, VEGFA has a known role in regulating tumor cell chemotherapeutic drug sensitivity, In addition to its important function in reproductive health 37,38 . EGR1 is bidirectional regulator of pathological process in malignant tumors; in breast cancer and lung cancer tissues, the EGR1 down-regulation plays a tumor-suppressive role and inhibits the proliferation and migration of tumor cells 39,40 ; whereas, in colorectal and prostate cancers, EGR1 plays an oncogenic role where its signi cant up-regulation promotes the proliferation and migration of tumors 41,42 .…”