| Objectives Fluorine is an important trace element in human and animal bodies.Long-term exposure to fluoride in water,air and food will cause fluorosis.Bone tissue is an important target organ for fluoride,and long-term ingestion of excessive fluoride will lead to abnormal metabolism of bone tissue.It is well known that plant anthocyanins have the effects of antioxidation,free radical scavenging,anti-inflammatory and prevention of chronic diseases,but it is still unclear how they affect bone tissue damage and bone metabolism caused by fluorosis.Maize purple plant pigment(MPPP)has the highest content of cornflower anthocyanin,which belongs to anthocyanins and has good biological functions.Its antioxidant capacity is higher than other traditional antioxidants.In this paper,the effect of MPPP on bone metabolism in rats with fluoride poisoning was observed by using an experimental model in vivo.Methods In this experiment,80 healthy weaned SD rats,half male and half female,were selected to be poisoned by drinking tap water.They were divided into a control group(Group C,Tap water without sodium fluoride+ordinary basic pellet feed),a fluorosis group(Group F,tap water with a concentration of 100 mg/L+ordinary basic pellet feed),a fluorosis+low dose MPPP group(Group FA,tap water with a concentration of 100mg/L+MPPP content of 5 g/kg ordinary basic pellet feed),and a fluorosis+high dose MPPP group(Group FAA,tap water with a concentration of 100 mg/L+MPPP content of 10 g/kg ordinary basic pellet feed),A total of 12 weeks of feeding.During this period,observe the changes in the general growth status of rats,and regularly record the occurrence of dental fluorosis.The fluoride ion content in blood,urine,and bone tissue was measured using a fluoride ion selective electrode method to observe the accumulation of fluoride in rats.Biochemical kits were used to detect the changes in calcium,magnesium,and phosphorus in blood and urine,and chemical methods were used to detect the concentrations of oxidation and antioxidant indicators T-AOC,T-SOD,GSH-Px,GSH,and MDA in serum.The tibial bone density and bone mass of rats in each group were measured by bone densitometer,and the microstructure changes at the widest part of the femoral trochanter were observed by bone CT.HE staining was used to observe the effect of MPPP on the trabecular and cortex of fluorosis rats,and ultrastructural observation was conducted to observe the nuclei and bone lacunae in osteoblasts and bone fine particles.Atomic absorption method was used to determine the content of calcium,magnesium,phosphorus,zinc,and iron in bone tissue to detect the effect of corn purple plant pigment on the mineral content of bone tissue in fluorosis rats.Protein immunoblotting was used to determine the protein expression of ALP and BMP-2 in bone tissue of fluorosis rats to verify the effect of MPPP on ALP and BMP-2 protein expression.Results1.Establishment of a rat fluorosis model.(1)General condition of experimental rats.During the 12-week experiment,the appetite of rats in each group was good,and there was no illness or death.The food intake of rats in Group C was higher than that of other experimental groups,but there was no significant difference(P>0.05).The water consumption of rats in each group also increased significantly,but there was no significant difference between the experimental groups(P>0.05).During the experiment,the growth and development of rats in each group were also good,and there was no significant difference between Group C and Group F,as well as between Group F,FA,and FAA groups(P>0.05).(2)Establishment of a successful fluoride model and accumulation of fluoride in rats.Compared with group C,rats in group F,FA,and FAA all had different degrees of dental fluorosis.In the blood,urine,and bone tissues of rats,compared with Group C,other experimental groups showed higher levels of fluoride accumulation(P<0.01).Compared with group F,the fluoride content in bone tissue in FA group and FAA group gradually decreased(P<0.01).2.Effect of MPPP on biochemical indicators in fluorosis rats.(1)Determination of Calcium,Magnesium,and Phosphorus in Blood and Urine of Rats.It can be seen from the blood that there is no significant difference in the content of calcium,magnesium,and phosphorus in the FA and FAA groups compared to the F group(P>0.05).It can be seen from the urine that compared with Group F,the phosphorus/Cr content of FAA group in different doses of MPPP treatment significantly decreased,with a statistical difference(P<0.05),while the calcium/Cr,magnesium/Cr content had no statistical difference(P>0.05).(2)Effect of MPPP on oxidation and antioxidant indexes in serum of rats.Compared with Group C,the concentration of MDA in Group F was significantly higher(P<0.05),while the concentrations of T-AOC,T-SOD,GSH,and GSH-Px were not significantly different(P>0.05);Compared with group F,the differences in MDA and GSH-Px in FA group with different doses of MPPP treatment were statistically significant(P<0.05),while the differences in antioxidant indicators in other groups were not statistically significant(P>0.05).3.Effect of MPPP on bone microstructure and mineral metabolism in fluorosis rats.(1)Effects of MPPP on bone mineral density and bone microstructure in fluorosis rats.The effect of MPPP on bone mineral density in fluorosis rats: BMD and BMC in the proximal tibia of rats in Group F were lower than those in Group C,with no significant difference(P>0.05).BMD and BMC in the proximal tibia of rats in Group FA and FAA were higher than those in Group F,with only significant differences between Group FAA and Group F(P<0.05).Three dimensional bone trabecular meshwork: Compared with Group C,Tb.BMD in Group F decreased and there was a significant difference in Tb.BMD(P<0.01),while there was no significant difference in BV/TV,Tb.Th,and Tb.Sp in Group F(P>0.05).Compared with group F,Tb.BMD in the MPPP treatment groups with different doses increased,and there was a significant difference in Tb.BMD in the FAA group(P<0.05).BV/TV in the FA group increased,with significant differences(P<0.05).There was no statistical difference in Tb.Th and Tb.Sp in the FA and FAA groups(P>0.05).Cortical bone three-dimensional: Compared with Group C,the three-dimensional dimensions of TV,BV/TV,TS,and BS in Group F decreased without significant difference(P>0.05).Compared with Group F,the three-dimensional dimensions of TV,BV/TV,TS,and BS in different doses of MPPP treatment groups increased without significant difference(P>0.05).(2)Effects of MPPP on bone trabeculae and cortex in fluorosis rats.Microscopic observation showed that the morphology and thickness of bone trabeculae in Group C were not abnormal and arranged tightly,and the thickness of bone cortex was normal.In Group F,the bone trabeculae became thinner,thinner,and arranged loosely,while the bone cortex became slightly thinner.In different doses of MPPP treatment group,the bone trabeculae became slightly thicker,thickened,and arranged slightly tightly compared to Group F,and the bone cortex became slightly thicker.(3)Effects of MPPP on osteoblasts and osteocytes in fluorosis rats.Normal nuclei can be seen in bone cells and osteocytes of group C,and the size and depth of bone lacunae are consistent.In bone cells and osteoblasts of group F,cell bodies become smaller,nuclei pyknotic,chromatin deepens,intracellular organelles are fewer,and bone lacunae are larger.Compared with group F,the fine cell bodies of different doses of MPPP treatment groups become slightly larger,nuclei expand,and bone lacunae become slightly smaller.(4)Effect of MPPP on bone mineral content in rats.Compared with Group F,the content of iron in FAA group increased and the difference was statistically significant(P<0.05),while the content of magnesium,phosphorus,zinc increased and the content of sodium decreased in FAA group,but the difference was not statistically significant(P>0.05);In the FAA group,the calcium content increased and the sodium content decreased,but the difference was not statistically significant(P>0.05).(5)Effect of MPPP on protein expression in bone tissue of fluorosis rats.Immunohistochemical results showed that compared with group C,the expression of ALP and BMP-2 in group F was not statistically significant(P>0.05),while the expression of ALP in FAA group treated with different doses of MPPP was statistically significant(P<0.01).Compared with group F,the expression of ALP in different doses of MPPP treatment groups increased slightly,while the difference in FAA group was statistically significant(P<0.01).The expression of BMP-2 protein decreased,but the difference was not statistically significant(P>0.05).Conclusions1.Fluoride has a high accumulation in rat bone tissue,and MPPP can reduce oxidative damage in the blood of fluorosis rats.2.MPPP can slow down the abnormal changes of tibial bone mineral density and bone mass in fluorosis rats.3.Fluorosis causes a decrease in bone mineral density in rats,while MPPP can increase both bone mineral density and bone volume fraction.4.MPPP can alleviate the damage of osteoblasts and osteocytes in bone tissue of fluorosis rats,and change the sparsity of bone trabeculae. |
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