Study On The Low Frequency Pulse Electromagnetic Fields Promoted Bone Formation By Primary Cilia And NO-cGMP-PKG Pathway

Micro-structure of osteoporosis is a bone tissue is broken,increased bone fragility due to low bone mass and bone brittleness,and easy to fracture of systemic metabolic disorder of systemic disease,is a kind of common disease and frequently-occurring disease,will bring serious trouble to the elderly patients with life.Electromagnetic field is a kind of physical therapy in the treatment of osteoporosis,has simple operation,no wound,non-toxic vice and other obvious advantages.Research in the prophase research is to obtain optimal parameters of the electromagnetic field in the treatment of osteoporosis,but the mechanism remains unclear.This experiment using homemade low frequency pulse electromagnetic field cells processing instrument to study the pulse electromagnetic field to promote the osteoblast mineralization mature relationship with the primary cilia and monoxide signal approach,part to clarify the molecular mechanism of the electromagnetic field in the treatment of osteoporosis.This experiment studied the first low frequency pulse electromagnetic fields of NO/cGMP/PKG signaling pathways.With 50 Hz 0.6 mT of pulse electromagnetic field radiation treatment in the rat calvaria osteoblasts 0h,0.5h,1.5h,1h,2h,determination of the change of the NO content in cell cultures,osteoblast cGMP concentration change and use Western blotting analysis of PI3 K,and AKT,eNOS,iNOS,PKG-1 the change of protein expression.The results show that after pulse electromagnetic field treatment,NO content showed a trend of rising;cGMP content in 0.5h rise to the highest,followed by decline;PI3K and AKT,eNOS,the protein expression of iNOS,PKG-1 quantity rise to different extent after falling.Shows that the pulse electromagnetic field activated NO/cGMP/PKG signaling pathways.Then,we pretreated with nitric oxide synthetase blocker L-NAME ROBs to observe pulse electromagnetic field to promote bone formation by active,the influence of the test indexes including the ALP activity and osteogenesis related proteins(BMP-2,Collagen-1,Osterix and osteoclast adjustment factor Rankl)gene expression changes in quantity and amount of calcium salt deposition.Results show: blockers,ALP activity into osseous gene expression and calcium salt deposition amount were significantly reduced.Showed that pulsed electromagnetic fields to promote bone formation depends on the NO/cGMP/PKG signaling pathways.Thesis in the fourth part studies the RNAi technology interfere with the primary cilia,pulse electromagnetic fields to activate the NO/cGMP/PKG signaling pathways.After primary cilia is interference suppression,electromagnetic field treatment cannot improve the NO content,also cannot make cGMP content increases,iNOS,sGC,eNOS,PKG-1 gene expression quantity rise,NO longer PI3 K,iNOS and eNOS,PKG-1 protein expression cannot rise.After that interfere with the primary cilia,NO signaling pathway is restrained,showed NO signaling pathways activated without the participation of primary cilia.Finally,after studied the primary cilia is interference,the influence of the pulse electromagnetic field to promote the osteoblast mineralization mature.RNAi intervention after the occurrence of primary cilia,we first in the fourth chapter verifies the efficiency of primary cilia interference,in the case of primary cilia interference efficiency is good,we had a follow-up experiment,studies the interference pulse electromagnetic field after primary cilia effect on osteoblast mineralization is mature.Results showed that RNAi interference after primary cilia,50 Hz 0.6 mT PEMFs processing cannot make activities of ALP,calcified nodule number decreases,into osseous differentiation index Osx,Col-1,gene expression of BMP-2 quantity is reduced.Show that primary cilia in the pulse electromagnetic field to promote the osteoblast mineralization mature process.Comprehensive the above results,50 Hz 0.6 mT PEMFs by activating NOS/NO/cGMP/PKG signaling pathways promote osteoblast mineralization is mature,and the process without the participation of primary cilia,suggests that the primary cilia-NOS-NO-sGC-cGMP-PKG signaling pathways to promote bone formation.