Short-term moderate hypothermia stimulates alkaline phosphatase activity and osteocalcin expression in osteoblasts by upregulating Runx2 and osterix in vitro.

Exposure of Normal Human Osteoblast cells (NHOst) to a period of hypothermia may interrupt their cellular functions, lead to changes in bone matrix and disrupt the balance between bone formation and resorption, resulting in bone loss or delayed fracture healing. To investigate this possibility, we exposed NHOst cells to moderate (35 °C) and severe (27 °C) hypothermia for 1, 12, 24 and 72 h. The effects of hypothermia with respect to cell cytoskeleton organization, metabolic activity and the expression of cold shock chaperone proteins, osteoblast transcription factors and functional markers, were examined. Our findings showed that prolonged moderate hypothermia retained the polymerization of the cytoskeletal components. NHOst cell metabolism was affected differently according to hypothermia severity. The osteoblast transcription factors Runx2 and osterix were necessary for the transcription and translation of bone matrix proteins, where alkaline phosphatase (Alp) activity and osteocalcin (OCN) bone protein were over expressed under hypothermic conditions. Consequently, bone mineralization was stimulated after exposure to moderate hypothermia for 1 week, indicating bone function was not impaired. The cold shock chaperone protein Rbm3 was significantly upregulated (p<0.001) during the cellular stress adaption under hypothermic conditions. We suggest that Rbm3 has a dual function: one as a chaperone protein that stabilizes mRNA transcripts and a second one in enhancing the transcription of Alp and Ocn genes. Our studies demonstrated that hypothermia permitted the in vitro maturation of NHOst cells probably through an osterix-dependent pathway. For that reason, we suggest that moderate hypothermia can be clinically applied to counteract heat production at the fracture site that delays fracture healing.

Alteration of cell cytoskeleton and functions of cell recovery of normal human osteoblast cells caused by factors associated with real space flight.

Experiments involving short-term space flight have shown an adverse effect on the physiology, morphology and functions of cells investigated. The causes for this effect on cells are: microgravity, temperature fluctuations, mechanical stress, hypergravity, nutrient restriction and others. However, the extent to which these adverse effects can be repaired by short-term space flown cells when recultured in conditions of normal gravity remains unclear. Therefore this study aimed to investigate the effect of short-term spaceflight on cytoskeleton distribution and recovery of cell functions of normal human osteoblast cells. The ultrastructure was evaluated using ESEM. Fluorescent staining was done using Hoechst, Mito Tracker CMXRos and Tubulin Tracker Green for cytoskeleton. Gene expression of cell functions was quantified using qPCR. As a result, recovered cells did not show any apoptotic markers when compared with control. Tubulin volume density (p<0.001) was decreased significantly when compared to control, while mitochondria volume density was insignificantly elevated. Gene expression for IL-6 (p<0.05) and sVCAM-1 (p<0.001) was significantly decreased while alkaline phosphatase (p<0.001), osteocalcin and sICAM (p<0.05) were significantly increased in the recovered cells compared to the control ones. The changes in gene and protein expression of collagen 1A, osteonectin, osteoprotegerin and beta-actin, caused by short-term spaceflight, were statistically not significant. These data indicate that short term space flight causes morphological changes in osteoblast cells which are consistent with hypertrophy, reduced cell differentiation and increased release of monocyte attracting proteins. The long-term effect of these changes on bone density and remodeling requires more detailed studies.

Real space flight travel is associated with ultrastructural changes, cytoskeletal disruption and premature senescence of HUVEC.

Microgravity, hypergravity, vibration, ionizing radiation and temperature fluctuations are major factors of outer space flight affecting human organs and tissues. There are several reports on the effect of space flight on different human cell types of mesenchymal origin while information regarding changes to vascular endothelial cells is scarce. Ultrastructural and cytophysiological features of macrovascular endothelial cells in outer space flight and their persistence during subsequent culturing were demonstrated in the present investigation. At the end of the space flight, endothelial cells displayed profound changes indicating cytoskeletal lesions and increased cell membrane permeability. Readapted cells of subsequent passages exhibited persisting cytoskeletal changes, decreased metabolism and cell growth indicating cellular senescence.