Effect Of Simulated Weightlessness On Apoptosis In Rat Weight-Bearing Bone Tissues

Background:On the earth,locomotor system bear two stimulus in the normal human bo-dy:gravity (passive stimulus)and exercise(active stimulus).A series of physiological and biochemical processes induced by the gravity and exercise combination interacti-on effect maintain bone tissues to adapt this mechanical environment. Microgravity (Weightlessness) is one of the capital environmental characteristic of spaceflight. Wei-ghtlessness which is the problem mankind must to be faced with and overcome in the spaceflight is the object shows only for the quality but not for the weight in the micr- ogravity(10-6-10-9g).Data collected from space flights over the last three decades have demonstrated that exposure to microgravity results in extensive physiological changes in humans,including a cephalic fluid shift, bone loss, negative calcium balance, loss of muscle mass, reduced immune response,and anemia. Researches show that,the creati- on of above-mentioned physiological functions is by the action of weig htlessness se- nsation afferent message change,body fluids cephalad metastasis and gravity carrying capacity disappearance.Osteoporosis which was considerd as bone mass diminution,bone mineral reduction and bone component invariability is a common metabolism osteopathy. Os- teoporosis involve constitutional osteoporosis and secondary osteoporosis. Weightle- ssness-induced osteoporosis is a secondary osteoporosis,it is a special disuse osteopo- rosis caused by the microgravity environment.Research about skeletal system Pathop- hysiology changes induced by weightlessness indicated that weightlessness evoked astronaut to take place a disuse osteoporosis.Disuse osteoporosis induced by wei- ghtlessness mainly manifested ad reduced bone mass,decreased bone mineral, decr- eased bone mineral density osteoporosis,the mechanical properties of bone decreased and a negative calcium balance. Premodinantly system changes between the series of adaptivity changes in the weightlessness flying possess determinate self-limitation,and will reach original equilibrium.However,the regulation of bone metabolism would be disequlibrium and the lost of bone mass might keep developing, so that,the persistent bone mineral loss in long-term weightlessness flying is one of the main serious physi- cological reaction to astronaut,which is also main obstacles in planetary exploration-n.Aerospace Medicine think highly of the research of bone metabolism in microgra- vity condition.The mechanisms by which skeletal unloading cause bone loss are, how- ever incompletely understood.And the valid prevention and cure interventions of bone loss in weight- lessness environment still not found.As the creation of International S- pace Station and the implementation of mars-exploration, mankind will lengthen det- ention time in space. Therefore,strengthens the research of bone metabolism in envir- onment of microgravity,and provides a new thinking prevention and cure in disuse os- teoporosis.Osseous tissue mainly consists of osteogenitor cell,osteocytes,osteoblasts and osteoclasts.Osteoblasts and osteocytes are the most important mechanical receptive cells in bone tissue.A lot of research indicates that,bone loss induced by microgravity was distinguished by bone formation suppression, declining in the number of osteoblasts,bone mineral dyspoiesis and bone delayed maturity.As the result of osteoblastes and osteocytes are short of necessary mechanical force stimulated in microgravity environment,tthe number of osteoblasts and osteocytes declined,finally brought into bone formation decrease.The research about bone loss in space flight is focus on osteoblasts and osteocytes of the microgravity response,signal transduction,gene expression,functional changes and microgravity on osteoblast differentiation process.Objective:Using the rat tail suspension model, we studied the effect of skeletal unloading on osteoblast and bone marrow cells apoptosis.Method:1.Thirty-six male SD rats were randomly assigned to two groups:not suspended (loaded) or suspended by the tail (unloaded) for 7,14 and 21 days.The tails of tail-suspended rats were pasted by ventilated adhesive tape on two sides.The other end of the adhesive tape was suspended by a plastic circle which can move freely along a rod.The head of rat was kept on low position,and hindlimb unloaded without constraint moving.The control rats were untreated.All the rats were fed equivalence of food without limited drinking in the same cages.2.Observation on rat model:To observe the activities,diet,body weight,colour and luster of hair and tail of rats during tail-suspension.After simulated weightlessness, biochemical markers of bone metabolism were observed. Meanwhile, the right tibia metaphysis was processed for histomorphometric analysis.3.Measurement of biochemical markers of bone metabolism:Serum Ca,P,ALP was measured by automatic biochemistry analyzer, BGP measured by radioimmunoassay. 4.The condition of cell apoptosis in bone tissues was detected with in situ death detection.5.The expression of Fas in bone tissues was detected with SP immunohistochemical method.Results:1.During tail-suspension,the rats moved,ate freely and did not bite tails.All rats were alive without ischemai and necrosis of tail.And body weight of tail-suspended rats didn't decreased significantly(P>0.05);2.Measurement of biochemical markers of bone metabolism:In the rats of 7 days,14days and 21 days simulated weightlessness groups,serum bone Gla protein,alkaline phosphatase content were significantly lower than those in corresponding control group (P<0.05), whereas serum calcium,phosphorus concentration were higher than that in corresponding control group:7d simulated weightlessness group markedly higher(P<0.05),14d and 21d unloaded rats were not significant difference with corresponding loaded rat(P>0.05).3.TUNEL examination showed:compared with corresponding control group,tail suspension for 7 to 21 days increased apoptosis of osteoblasts and bone marrow cells,there were significantly difference(P<0.05).With time goes on,the apoptosis of osteoblasts and bone marrow cells in 14days stimulated weightless were significantly lower than those in 7days stimulated weightlessand. While,the apoptosis of osteoblasts and bone marrow cells in 21 days stimulated weightless were significantly lower than those in 14days stimulated weightless.4.The-Fas protein examination demonstrated:compared with corresponding control group,tail suspension for 7 to 21 days increased Fas protein expression,there were significantly difference(P<0.05).Fas protein expression in 14days stimulated weightless was significantly lower than those in 7days stimulated weightlessand. While,the Fas protein expression in 21 days stimulated weightless were significantly lower than those in 14days stimulated weightless.Conclusion:1.The rat which simulates microgravity condition by tail suspended succeeds resulting in headward shift of body fluid and unloading of weight-bearing bones which are two main physiological effects under microgravity with little stress.It supplies a good rat model for studies on simulated microgravity condition on land.2. Simulated microgravity affect serum markers of bone metabolism.3.Simulated microgravity increased Fas protein expression.4.Simulated microgravity induces a increase apoptosis in osteoblast and bone marrow cells.5.The opotosis of osteoblast and bone marrow cells related to the Fas expression,thus,it can be inferred that microgravity promote the Fas expression,then induce the opotosis of osteoblast and bone marrow cells6.With microgravity on a prolonged, Fas expression and apoptosis cells rate has decreased,it shows that early stimulated microgravity affect apoptosis in bone tissue significantly,therefore,osteoporosis induce by microgravity shoud be taken early prevention.