👤 Tamara Rotshteyn

MicroRNA 193a-3p (miR193a-3p) is a short non-coding RNA with tumor suppressor properties. Breast cancer (BC) progression is governed by active interaction between breast cancer cells, vascular (V)/lymphatic (L) endothelial cells (ECs), and BC secretome. We have recently shown that miR193a-3p, a tumor suppressor miRNA, inhibits MCF-7 BC cell-driven growth of VECs via direct antimitogenic actions and alters MCF-7 secretome. Since LEC-BC cross-talk plays a key role in BC progression, we investigated the effects of miR193a-3p on MCF-7 secretome and estradiol-mediated growth effects in LECs and LEC + MCF-7 spheroids, and delineated the underlying mechanisms. Transfection of LECs with miR193a-3p, as well as secretome from MCF-7 transfected cells, inhibited LEC growth, and these effects were mimicked in LEC + MCF-7 spheroids. Moreover, miR193a-3p inhibited ERK1/2 and Akt phosphorylation in LECs and LEC + MCF-7 spheroids, which are importantly involved in promoting cancer development and metastasis. Treatment of LECs and LEC + MCF-7 spheroids with estradiol (E2)-induced growth, as well as ERK1/2 and Akt phosphorylation, and was abrogated by miR193a-3p and secretome from MCF-7 transfected cells. Gene expression analysis (GEA) in LEC + MCF-7 spheroids transfected with miR193a-3p showed significant upregulation of 54 genes and downregulation of 73 genes. Pathway enrichment analysis of regulated genes showed significant modulation of several pathways, including interferon, interleukin/cytokine-mediated signaling, innate immune system, ERK1/2 cascade, apoptosis, and estrogen receptor signaling. Transcriptomic analysis showed downregulation in interferon and anti-apoptotic and pro-growth molecules, such as IFI6, IFIT1, OSA1/2, IFITM1, HLA-A/B, PSMB8/9, and PARP9, which are known to regulate BC progression. The cytokine proteome array of miR193a-3p transfected MCF secretome and confirmed the upregulation of several growth inhibitory cytokines, including IFNγ, Il-1a, IL-1ra, IL-32, IL-33, IL-24, IL-27, cystatin, C-reactive protein, Fas ligand, MIG, and sTIM3. Moreover, miR193a-3p alters factors in MCF-7 secretome, which represses ERK1/2 and Akt phosphorylation, induces pro-apoptotic protein and apoptosis in LECs, and downregulates interferon-associated proteins known to promote cancer growth and metastasis. In conclusion, miR193a-3p can potentially modify the tumor microenvironment by altering pro-growth BC secretome and inhibiting LEC growth, and may represent a therapeutic molecule to target breast tumors/cancer. Show less

Embryonic developmental studies under microgravity conditions in space are very limited. To study the effects of altered gravity on the embryonic development processes we established an in vitro methodology allowing differentiation of mouse embryonic stem cells (mESCs) under simulated microgravity within a fast-rotating clinostat (clinorotation) and capture of microarray-based gene signatures. The differentiating mESCs were cultured in a 2D pipette clinostat. The microarray and bioinformatics tools were used to capture genes that are deregulated by simulated microgravity and their impact on developmental biological processes. The data analysis demonstrated that differentiation of mESCs in pipettes for 3 days resultet to early germ layer differentiation and then to the different somatic cell types after further 7 days of differentiation in the Petri dishes. Clinorotation influences differentiation as well as non-differentiation related biological processes like cytoskeleton related 19 genes were modulated. Notably, simulated microgravity deregulated genes Cyr61, Thbs1, Parva, Dhrs3, Jun, Tpm1, Fzd2 and Dll1 are involved in heart morphogenesis as an acute response on day 3. If the stem cells were further cultivated under normal gravity conditions (1 g) after clinorotation, the expression of cardiomyocytes specific genes such as Tnnt2, Rbp4, Tnni1, Csrp3, Nppb and Mybpc3 on day 10 was inhibited. This correlated well with a decreasing beating activity of the 10-days old embryoid bodies (EBs). Finally, we captured Gadd45g, Jun, Thbs1, Cyr61and Dll1 genes whose expressions were modulated by simulated microgravity and by real microgravity in various reported studies. Simulated microgravity also deregulated genes belonging to the MAP kinase and focal dhesion signal transduction pathways. One of the most prominent biological processes affected by simulated microgravity was the process of cardiomyogenesis. The most significant simulated microgravity-affected genes, signal transduction pathways, and biological processes which are relevant for mESCs differentiation have been identified and discussed below. Show less