Presented work focuses on the response of osteogenic, osteoblastic and mesenchymal stem cells to real and simulated microgravity. Mesenchymal stem cells are the residents of adult bone marrow stroma. These cells are capable of differentiating towards osteogenic, adipogenic and chondrogenic lineages. The advantage of using MSCs in space-related research is that these cells are relevant tool to develop bone tissue. Osteogenic potential of MSCs allows investigating osteogenesis from the most uncommitted cells while still maintaining all main characteristics of bone differentiation (osteogenic gene expression, osteogenic markers, matrix maturation and mineralization). Recent studies have demonstrated that MSCs are targeted by microgravity and accordingly can be involved in space flight-induced osteopenia. These findings determine the necessity of further study of human MSC biology in real and simulated microgravity conditions.

Mesenchymal stem (stromal) cells (MSCs) are present in a variety of tissues during prenatal and postnatal human development. In adult organism, they are prevalent in bone marrow and supposed to be involved in space-flight induced osteopenia. We studied expression of various genes in human bone marrow MSCs after different terms of simulated microgravity (SMG) provided by Random Positioning Machine. Simulated microgravity induced transient changes in expression level of genes associated with actin cytoskeleton, especially after 48h of SMG. However, after 120h exposure in SMG partial restoration of gene expression levels (relative to the control) was found. Similar results were obtained with bmMSCs subjected to 24h readaptation in static state after 24h in SMG. Analysis of 84 genes related to identification, growth and differentiation of stem cells revealed that expression of nine genes was changed slightly after 48h in SMG. More pronounced changes in gene expression of “stem cells markers” were observed after 120h of simulated microgravity. Among 84 investigated genes, 30 were up-regulated and 24 were down-regulated. Finally, MSCs osteogenesis induced by long-term (10–20 days) simulation of microgravity was accompanied by down-regulation of gene expression of the main osteogenic differentiation markers (ALPL, OMD) and master transcription osteogenic factor of MSCs (Runx2). Thus, our study demonstrated that changes in expression level of some genes associated with actin cytoskeleton and stem cell markers are supposed to be one of the mechanisms, which contribute to precursor's cellular adaptation to the microgravity conditions. These results can clarify genomic mechanisms through which SMG reduces osteogenic differentiation of bmMSCs.

This report is a detailed review of the current data on the mechanic and gravitational sensitivity of osteoblasts and osteogenic precursor cells in vitro. It summarizes the numerous responses of cells with an osteoblastic phenotype and osteogenic precursor cells and especially their responses to the alteration of their mechanic or gravitational surroundings. The review also discusses the osteogenic cell's pathways of signal transduction and the mechanisms of gravitational sensitivity. It was shown that the earliest multipotent stromal precursor cells of an adult organism's bone marrow can sense changes of intensity in a gravitational or mechanic field in model conditions, which may play a certain role in the development of osteopenia in microgravity.

During spaceflight and in simulated microgravity (SMG), cytoskeleton rearrangements were observed in lymphocytes, glial cells and osteoblasts. One potential mechanism for the cytoskeletal gravisensitivity of cells is the disruption of the extracellular matrix and integrin interactions. We investigated the effect of SMG on the structure of the actin cytoskeleton, distribution of cellular vinculin, the expression of some integrin subtypes and cellular adhesion molecules in cultured mesenchymal stem cells (hMSCs) derived from human bone marrow in vitro. Simulated microgravity was produced by desktop RPM equipment (Dutch Space, Netherlands). Cells were exposed to simulated microgravity for 30 min to 120 h. The results showed that the actin cytoskeleton was reorganized very quickly (30 min). Later (6, 24, and 48 h), the number of cells with disrupted actin cytoskeletons was increased; however, after 120 h of exposure, cells partly regained their F-actin structures. RPM exposure augmented the number of cells that express integrin-α2. We also observed a decrease in the number of VCAM-1-positive cells and changes in the expression of ICAM-1. Our findings indicate that SMG induces reversible microfilament reorganization in hMSCs and alters their adhesion properties.

It was shown that proliferative rate of human mesenchymal stem cells (hMSCs) decreased during gravity vector randomization (clinorotation). Clinorotated hMSCs exhibited changes in expression of some external cell markers and adhesion molecules. Marked suppression of osteogenic differentiation of hMSC as to formation of bone nodules and calcium deposits was observed. F-actin stress fibers of cytoskeleton were altered in clinorotated cultures. Prolonged clinorotation of embryoid bodies (EBs) resulted in a delay of embryonic stem cells (ESCs) differentiation into cardiomyocytes. Percentage of beating cardiomyocytes in each EB was significantly reduced. Thus, long-term gravity vector randomization decreases the differentiation processes of precursor cells in vitro.

We studied the effects of long-term (20-day) simulated microgravity (clinostatic exposure) and osteogenic differentiation stimuli on cultured mesenchymal stromal precursor cells isolated from human bone marrow. Clinostatic exposure significantly reduced proliferative activity of mesenchymal stem cells in comparison with the static and dynamic control, increased the number of large flat cells in the culture, and stimulated migration activity of cells. Phenotypic studies of surface antigens (CD90, CD54, CD106, CD105, CD34, CD45, class 1 HLA) during clinostatic exposure of mesenchymal stem cell cultures showed differences in their expression between experimental and control groups. Studies of osteogenesis of precursor cell showed that cell differentiation potential can be directed towards osteogenesis by a combination of clinostatic exposure and differentiation stimuli. The results confirm gravity sensitivity of human bone marrow precursor cells and open new vistas for understanding of the mechanisms of bone tissue loss in humans under conditions of space mission.