The Effect Of Oxidative Stress Induced By 1,25?OH?₂D Deficiency On Development And Aging Of Tooth And Jaw

1,25(OH)2D plays an important role in the development and mineralization of the bone by regulating metabolism of calcium and phosphorus.Studies have found that the role of 1,25(OH)2D is not only limited to the maintenance of serum calcium and phosphorus levels,but also to regulate biological function directly through combination with its specific receptors.Moreover,1,25(OH)2D plays an important role in many chronic diseases(such as osteoporosis,diabetes,periodontal disease)due to its regulation of oxidative stress levels in vivo.Reactive oxygen species(ROS)are necessary molecular signals in the regulation of normal physiological function of cells,but excessive accumulation of ROS can lead to the increase of oxidative stress level of cells,resulting in DNA damage,which contributes to the development of cardiovascular disease,diabetes,osteoporosis,osteoporosis,hypercholesterolemia,hypercholesterolemia,Osteoarthritis,periodontal disease and other chronic disease and aging.However,the role and mechanism of oxidative stress induced by 1,25(OH)2D deficiency in the development of tooth and jaw formation and mineralization has not been reported by now.To investigate the effects of scrum calcium-phosphorus level and oxidative stress on the formation and mineralization of tooth and jaw in 1,25(OH)2D deficiency mice,we employed 25-hydroxyvitamin D la-hydroxylase knockout[la(OH)ase-/-]mice model in this present study.We maintained 1?(OH)ase-/-mice and their wild-type littermates on a rescue diet(2%calcium,1.25%phosphorous and 20%lactose)from weaning until 18-months of age to exclude possible effects of extracellular calcium and/or phosphorus and/or PTH,while investigating the role of 1,25(OH)2D in development and formation of tooth and jaw by imageology,histochemistry,immunohistochemistry.Based on the findings,to investigate the role of oxidative stress in development and mineralization of tooth and jaw induced by 1,25(OH)2D deficiency,1?(OH)ase-/-mice were maintained on a diet of regular mouse chow containing 1%calcium and 0.67%phosphorus after weaning,or a rescue diet,or a rescue diet with N-acetyl-L-cysteine(NAC)solution(lg/L),and their wild-type littermates were fed on a diet of regular mouse chow from weaning until 10-week-old.The phenotype and the probable mechanism were analyzed by methods of imageology,histomorphology and immunohistochemistry and biochemical markers.Part ?:Effects of 1,25(OH)2D Deficiency on Teeth and Jaw of Aging miceThe biological effect of 1,25(OH)2D includes not only the regulation of serum calcium-phosphorus level together with PTH,but also the influence of cellular function through its direct biological effects.To investigate the effects of1,25(OH)2D on the formation and mineralization of tooth and jaw and whether these effects were only calcium-phosphorus level dependent,or its synergistic direct biological effects,we maintained 1?(OH)ase-/-mice and their wild-type littermates on a rescue diet(2%calcium,1.25%phosphorous and 20%lactose)from weaning until 18-months of age.The phenotypes of tooth and jaw of both groups were observed by imageology,histochemistry,immunohistochemistry.The results showed that:(1)Serum calcium and phosphate levels of 1?(OH)ase-/-mice returned to normal after administration of a rescue diet after weaning,left only serum 1,25(OH)2D level was significantly lower than normal;In the wild-type mice,the levels of serum calcium and phosphorus and 1,25(OH),D were all within the normal range after a rescue diet.(2)18-month-old 1?(OH)ase-/-mice had significantly lower mineralization of cortical bone than that of wild-type mice,the volume of the first molar were significantly decreased,and the thickness of root canal wall was decreased compared with WT littermates.(3)The function of osteoblasts of 1?(OH)ase-/-mice was significantly lower than that of wild-type mice.(4)Biglycan and DSP posi-tive expression of the mandibular first molars in the 18-month-old 1?(OH)ase-/mice were significantly decreased than that of the 18-month-old WT mice.This indicated that 1,25(OH)2D deficiency could have effects on the formation and maturation of predentin in 18-month-old mice.(5)Compared with 18-month-old WT mice,18-month-old la(OH)ase-/-mice had significantly higher DNA damage and cellular senescence in the pulp-dentin-complex and mandibular bone.The results of this study suggested that only a deficiency of 1,25(OH)2D could also lead to phenotypic changes in the teeth and jaw of mice,while 1,25(OH)2D deficiency could cause DNA damage and accelerate cell aging by its direct biological effect.Part ?:The Effect of Oxidative Stress Induced by 1,25(OH)2D Deficiency on Development and Aging of Tooth and JawBased on the results of first part,in this part of study,we investigated the role of calcium-phosphorus level and oxidative stress induced by 1,25(OH)2D deficiency in tooth and jaw.1?(OH)ase-/-mice were maintained on a diet of regular mouse chow containing 1%calcium and 0.67%phosphorus after weaning,or a rescue diet,or a rescue diet with N-acetyl-L-cysteine(NAC)solution(1g/L),and their wild-type littermates were fed on a diet of regular mouse chow.The mice were sacrificed at 10-week-old.The phenotype and the probable mechanism were analyzed by methods of imagcology,histomorphology and immunohistochemistry and biochemical markers detection.The results showed that:(1)10-week-old 1?(OH)ase-/-mice mice with a rescue diet could improve the mineralization of tooth and jaw,but still significantly lower than wild-type;when a rescue diet and NAC water was administrated,tooth and jaw mineralization and tooth volume,bone mass were further increased,but still significantly lower than wild-type level.(2)The result of Osx and Col I immunohistochemistry and TRAP histochemical staining showed that a rescue diet could increase the activity of osteoblasts and decrease the activity of osteoclasts of la(OH)ase-/-mice.A rescue diet and NAC water could further increase the activity of osteoblasts and decrease osteoclast activity,but there were still significant differences between 1?(OH)ase-/-mice and their wild-type littermates.(3)Immunohistochemical staining of Biglycan and DSP showed that a rescue diet could increase the expression of Biglycan and DSP in dentin of la(OH)ase-/-mice,and with help of NAC water,the expression of Biglycan and DSP in dentin could be further increased,but still left significant difference between la(OH)ase-/-mice and their wild type littermates.This result indicated that the lack of 1,25(OH)2D could hinder the formation of predentin and maturation of dentin in 10-week-old mice.(4)DNA damage and cell senescence level in the pulp chamber and mandible bone of 10-week-old la(OH)ase-/-mice were dramatically increased compared with WT mice.Given the rescue diet,DNA damage and cell senescence level could be slightly decreased,and the rescue diet and NAC water could forther decrease positive expression,but still significantly higher that of wild-type mice.(5)The dramatically increased content of H202 and MDA,decreased concentration of T-SOD and GSH-Px were detected in mandibular bone of la(OH)ase-/-mice compared with their wild-type littermates,given the rescue diet the content of H2O2 and MDA could be slightly reduced,and the rescue diet with NAC also could further decreased the content of H2O,and MDA compared with the 1?(OH)ase-/-mice just fed on rescue diet.On the other hand,the significant differences of T-SOD and GSH-Px concentration between each group were not found in this study.These results demonstrated that increased DNA damage induced by decreased calcium-phosphorus level and increased oxidative stress level would play important role in the development and moralization of tooth and jaw of 1,25(OH)2D deficiency mice.