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Objective To explore the influence of fluorine on mRNA and protein expression of the insulin-like growth factor-1 (IGF-1) and its receptor of rat osteoblasts.Methods Osteoblasts were isolated from rat bone by enzyme digestion.Different fluorine concentration [0 (control),10-7,10-6,10-5,10-4,10-3 mol/L] was add to the second generation osteoblasts.The IGF-1 in the culture medium was determined by radioimmunoassay (RIA) at different fluorine concentration and different time (24,48 h).The expression of IGF-1 receptor was measured by the method of fluorescent quantitation PCR and the expression of protein IGF-1 receptor was measured by Western blotting.Results ①With increased dose of fluoride exposure,IGF-1 concentration in the osteoblastic culture medium increased first and then decreased at 24,48 h,respectively.Compared to the control group [(38.83 ± 3.48)ng/L],IGF-1 concentration of the 24 h 10-6 mol/L group[(65.45 ± 4.84)ng/L] was higher,and the difference was statistically significant(P ＜ 0.05).The same result was also shown in the 48 h 10-5 mol/L group [(59.14 ± 1.53)ng/L] to its corresponding control group [(33.79 ± 1.84)ng/L,P ＜ 0.05].②The mRNA expression of IGF-1 receptor of the 24,48 h 10-5 mol/L groups (0.0055 ± 0.0004,0.0262 ± 0.0040) was significantly higher than their corresponding control groups (0.0022 ± 0.0001,0.0073 ± 0.0008,all P ＜ 0.05).③With increased dose of fluoride exposure,the protein expression of IGF-1 receptor increased first and then decreased ;the expression of 24 h 10-5 mol/L group (1.39 ± 0.16) was compared with the corresponding control group (0.86 ±0.12),and the difference was statistically significant (P ＜ 0.05) ; the expression of 48 h every fluorine group was also compared with the corresponding control group,and the difference was not statistically significant(all P＞ 0.05).Conclusions Fluorine can affect the mRNA and protein expression of osteoblastic IGF-1 and its receptor.It indicates that IGFS signal transduction pathways play an important role in fluorine regulation of bone metabolism.
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Objective To determine the impact of fluorine and aluminum,and both action combined on the number of rat osteoclasts and bone resorption cultured in vitro and to explore its mechanisms.Methods The osteoclasts and bone marrow stromal cells (BMSCs) isolated from long bone of new born rats were cultured,respectively,in TC199 medium (containing 10％ fetal bovine serum) with fluoride,aluminum and fluoride combined with aluminum.The osteoclasts were inoculated in 96-well culture plate and ivory slice,BMSCs in 6-well culture plate,and culture medium was changed after 2 hours incubation.The cells were divided into control group,fluoride group,aluminum group and fluoride combined with aluminum group; the doses of sodium fluoride were 0,1.0 × 10-4,0,1.0 × 10-4 mol/L and the doses of aluminum chloride were 0,0,1.0 × 10-5,1.0 × 10-5 mol/L,respectively.Tartrate-resistant acid phosphatase (TRAP) staining positive cells were counted under light microscope after TRAP staining on the 5th day and the pit formed in ivory slices were measured by histomorphometry after staining with toludine blue.The expression of osteoprotegerin(OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL) was detected by real-time fluorescence quantitative PCR in BMSCs after 8 h treatment.Results ① Fluoride,aluminum and the interactive effects of fluoride and aluminum all had impact on the numbers of osteoclasts (F =7.15,6.56 and 7.98,respectively,all P ＜ 0.05).The numbers of osteoclasts in fluoride group,aluminum group and fluoride combined with aluminum group[(136.9 ± 22.99),(135.4 ± 23.5),(163.0 ± 24.4) per well] were higher than that in the control group[(92.5 ± 22.1) per well,all P ＜ 0.05].② Fluoride,aluminum and the interactive effects of fluoride and aluminum all had impact on the resorption pit area on ivory slices(F =10.47,12.64,14.29,respectively,all P ＜ 0.05).The resorption pit area on ivory slices in fluoride group,aluminum group and fluoride combined with aluminum group[(0.242 ± 0.031),(0.293 ± 0.026),(0.333 ± 0.016)mm2 per slice] was higher than that in the control group [(0.088 ± 0.030)mm2 per slice,all P ＜ 0.05].③Fluoride,aluminum and the interactive effects of fluoride and aluminum all had impact on the expression ratios of RANKL/OPG in BMSCs (F =8.15,15.38,23.59,respectively,all P ＜ 0.05).The expression ratios of RANKL/OPG in BMSCs in fluoride group,aluminum group and fluoride combined with aluminum group [(193.98 ± 137.93)％,(326.11 ± 176.78)％,(599.84 ± 275.82)％] were higher than that in the control group[(100.00 ± 56.02)％,all P ＜ 0.05].Conclusions Both fluoride and aluminum can cause increase in the number of osteoclasts in vitro and promote cell differentiation and bone resorption activity,which may be related to increased expression ratio of RANKL/OPG mRNA in BMSCs.The stimulating effects of fluoride on osteoclasts differentiation and bone resorption is enhanced by aluminum.
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Objective To determine the effects of fluoride on osteoclasts's quantity and bone resorption function in vitro and its mechanisms. Methods The osteoclasts and bone marrow stromal cells(BMSCs) isolated from long bone of new born rats were cultured respectively in TC199 medium (containing 10% fetal bovine serum) with fluoride. The osteoclasts were inoculated in 96-well culture plate and ivory slice, BMSCs were inoculated in 6- well culture plate, respectively, medium were changed after 2 hours incubation. They were divided into control group, low-dose fluoride, medium-dose fluoride and high-dose fluoride groups, the doses of sodium fluoride were 0,2.5 × 10-5,5.0 × 10-5,10.0 × 10-5 mol/L, respectively. Tartrate-resistant acid phosphatase(TRAP) staining positive cells were counted under light microscope after TRAP staining on the 2nd and the 5th day and the pit formed in ivory slices were measured by histomorphometry after staining with toludine blue. The expression of receptor activator of NK-κβ ligand(RANKL) and osteoprotegerin(OPC) was detected by real-time fluorescence quantitative (337.5 ± 70.5), (447.5 ± 43.4), (472.9 ± 34.8), (475.3 ± 24.3)/well in the control group, the low-dose, mediumdose and high-dose fluoride groups, respectively. The differences were statistically significant between these groups and the control group (all P < 0.05). After in vitro culture for 5 days, the numbers of osteoclasts were (92.5 ± 22.1), (123.0 ± 26.4), (135.5 ± 22.2), (136.9 ± 23.0) per well in the control group, the low-dose, medium-dose and high-dose fluoride groups, respectively. The differences were statistically significant between these groups and the (0.088 ± 0.030), (0.100 ± 0.018), (0.152 ± 0.015), (0.242 ± 0.031 )mm2 per piece in the control group, the lowdose, medium-dose and high-dose fluoride groups, respectively. The values of medium-dose and high-dose fluoride BMSCs in the control group, the low-dose, medium-dose and high-dose fluoride groups were 100.00 ± 56.02, 144.95 ± 97.21,223.25 ± 184.48,193.98 ± 137.93, respectively. The values of medium-dose and high-dose fluoride groups were significantly higher than that of control group (all P < 0.05). Conclusions Fluoride can cause increase in the number of osteoclasts in vitro and promote their cell differentiation and bone resorption activity, which may be related to increased expression ratio of RANKL/OPG mRNA in BMSCs.
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Objective To observe the effects of soybean,selenium and spirulina on hemoglobin(Hb)of rats intoxicated with fluorine and aluminiums.Methods According to body weight,84 SD rats were randomly divided into control group,high aluminum group,high fluorine group,high fluorine-aluminum group,high fluorine-aluminium intoxicated rats strengthened with soybean group,high fluorine-aluminium intoxicated rats strengthened with selenium group and high fluorine-aluminium intoxicated rats strengthened with spirulina group,12 in each group.Rats in the control group and the high aluminum group were fed with feed containing 5.2 mg/kg of fluorine and 6.8 mg/kg of aluminum.In other groups,fluorine wag 106 mg/Kg and aluminum 19.7 mg/kg.Fluorine and aluminum concentration in the drinking water of the control group and the high fluorine group were 0.69 mg/L and 0.20 mg/L,respectively.In other groups' drinking water,these values were 0.69 mg/L and 90.2 mg/L,respectively.Ninety days later,Hb concentration of the whole blood was tested.Results Hb concentration of the control group,the high aluminum group,the high fluorine group,and the high fluorine-aluminum group were (160.8±6.3),(142.2±15.9),(156.1±4.9)and(145.2±6.2)g/L,respectively.Fluorine had an effect on the concentration of Hb(F=29.56,P<0.05).The Hb concentration of the high fluorine-aluminum group,the strengthened with soybean group,the strengthened with selenium group and the strengthened with Spirulina group were(145.2±6.2),(150.7±17.7),(156.8±14.5),(154.5±17.8)g/L,respectively.Though the concentration of Hb had increased,there was no significant difference between the four groups(χ2=3.304,P>0.05).Conclusions High-dose fluorine could cause varied decrease in the concentration of Hb.However,aluminum has neitherantagonistic effect nor synergistic effect on the Hb of fluorotic Rat.Soybean,selenium and Spirulina show a trend to increase fluorotic rat's Hb,but they has no evident antagonistic effect.
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Objective To observe the expressions of bone morphogenetic protein-2(BMP-2)and bone morphogenetie protein-7(BMP-7)in the synovial tissue of fluorosis rats and its correlation with pathogenic mechanism of fluorosis arthritis.Methods Thirty-two SD rats were randomly divided into 4 groups:the control group,low,moderate and high-dose fluoride group.The control group ate commou fodder.The low,moderate and high dose fluoride group were fed with fodder composed of 25%.35%and 68%of corn(containing fluorine of 148.00 mg/kg)in chronic endemic fluorosis region in Guizhou Province.After 140 days,the expressions of BMP-2 and BMP-7 protein were determined by immunohistochemistry and assayed the absorbanee by computer image-pattern analysis system.Light microscope was used to observe the synovial tissue by Hematoxin Eosin,and calculated the pathological integral of synovium according to pathological grade standard.Results The expressions of BMP-2 (32.50±2.73)and BMP-7(38.90±2.56)in the control group was spare.Compared with the control group,the expressions of BMP-2(59.43±5.12,79.82±6.41,101.76±7.56)and BMP-7(55.10±4.82,78.42±5.61,98.46± 6.05)in the synovial tissue was up-regulated in each experimental groups(P＜0.05),especially in the moderate dose and the high-dose groups(P＜0.05).Compared with the control group(0.54±0.21).the pathological integral of synovium increased(P＜0.05)in each experimental groups(1.04±0.98,4.69±1.28,8.60±2.07).The expressions of BMP-2 and BMP-7 in the synovial tissue was found to be positively related with the pathological integral of synovium(r=0.98,0.99,P＜0.05).Conclusion The BMP-2 and BMP-7 play an important role in the development of fluorosis arthritis,probably by affecting osteogenesis.
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Objective To study the intervention effect of the fluorosis with the high aluminum plus soybean, selenium powder, spiral algae. Methods Two week-long SD rats were randomly allocated into 5 groups bases on body weight with 8 in each group, respectively being control, the high fluoride aluminum,the high fluoride aluminum plus soybean, the high fluoride aluminum plus selenium and the high fluoride aluminum plus spiral algae group. The control group was fed with corn produced in non-endemic area containing 5.20 mg/L of fluoride and 6.80 mg/L of aluminum and tap water of fluoride 0.70 mg/L and aluminum 0.20 mg/L. Other groups were fed primarily with corn produced in the endemic area with the content of fluoride of 110.00 mg/L and aluminum of 19.70 mg/L and tap water with high aluminum also with fluoride of 0.70 mg/L and aluminum of 90.00 mg/L. The high fluoride aluminum plus soybean group was added soybean in the beginning of the experiment, approximately 30% of the total amount. Selenium (3.00 mg/kg) was given to the high fluoride aluminum plus selenium groups and spiral algae (1000.00 mg/kg) to the high fluoride aluminum plus spiral algae groups after dental fluorosis was formed. The experiment lasted for 22 weeks. In the end of the experiment 24-hour urine and bones of the limbs were collected to test the content of fluorine and aluminum. The ultrastructure of femur spongiosa was observed under electron microscope. Results ① Bone fluoride in high fluoride aluminum group [(204.07 ± 63.78)mg/kg] was higher than that in the control group, high fluoride aluminum plus soybean group, high fluoride aluminum plus selenium group and high fluoride aluminum plus spiral algae group[(30.06 ± 6.11), (54.12 ± 14.56), (30.44 ± 5.02), (105.08 ± 21.07)mg/kg, t = 0.62,0.53,0.62,0.35, all P < 0.01], indicating that it was lower in high fluoride aluminum plus selenium group than high fluoride aluminum plus spiral algae group(t = 0.27, P < 0.01). ② The urinary fluoride levels in high fluoride aluminum group [(4.52 ± 3.09)mg/L] was higher than that in the control group [(0.89± 0.37)mg/L, t = 0.23, P < 0.01] and lower than that in the fluoride aluminum plus selenium group[(10.38 ± 1.58) mg/L, t = 0.34, P < 0.01], it was higher in high fluoride sluminum with selenium group than that in the high fluoride aluminum plus soybean group and high fluoride aluminum plus spiral algae group[(5.56 ± 1.69), (4.38 ± 3.36)mg/L, t = 0.28,0.35, all P < 0.05]. ③ The bone aluminum level in the control group[(3.32 ± 3.02)mg/kg] was lower than that in the high fluoride aluminum group, high fluoride aluminum plus soybean group, high fluoride aluminum plus selenium group and high fluoride aluminum plus spiral algae group [(374.21 ± 56.11), (118.20 ± 23.59), (114.01 ± 22.84), (67.11 ± 11.53)mg/kg, t = 1.42,0.44,0.42,0.24, all P < 0.05], it was higher in the high fluoride aluminum group than high fluoride aluminum plus the soybean and selenium and spiral algae groups (t = 0.56,0.57,0.68, all P < 0.01)lower in high fluoride aluminum plus spiral algae group than the soybean plus selenium group(t = 0.11,0.10, all P < 0.05). ④The urine aluminum level in high fluoride aluminum group [(1.14 ± 0.32)mg/L] was higher than that in the control group and the high fluoride aluminum plus the soybean group [(0.66 ± 0.10) mg/L, t = 1.92,2.24, all P< 0.05] and that in high fluoride aluminum plus the soybean group was lower than aluminum plus the selenium[(1.03 ± 0.22)mg/L] and aluminum plus spiral algae group[(1.10 ± 0.28) mg/L, t = 1.73,2.06, all P < 0.05]. ⑤ Under electron microscope, the collagenous fiber was arranged in disorder in high aluminum fluorine group, the trabeculum of bone merged into patches or mostly vanished, This phenomenon was alleviated in the intervened group compared to the high aluminum fluorine group, in an attenuating order of the selenium group, the spiral algae and the soybean group. Conclusion In this experiment the intervention measure alleviates fluorosis, the selenium is the best, spiral algae the second and soybean the last.
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Objective To investigate the effect of excessive fluoride,aluminum on Zn,Fe,Ca,Mg,Cuin rat blood.Methods Forty eight SD rats were randomly divided into 4 groups matched with their weights:control group,high aluminum group,high fluorine group and high fluorine-aluminum group.Aluminum content in their drinking water was 0,90,0,90 mg/L respectively.Fluorine content of their feed was 5.2,5.2,106.0,106.0 mg/kg and aluminum Was 6.8,6.8,19.7,19.7 mg/kg respectively.90 days later,the level of blood Zn,Fe,Ca,Mg, Cu Was detected by the atomic absorption spectrometry.Results Compared among these groups,Zn,Fe,Mg and Cu content of the whole blood had significant difierences(F=46.25,14.74,6.10,2.93,P<0.05),while Ca content of the whole blood did not significantly change(F=2.81.P>0.05).Factorial analysis showed that excessive intake of aluminum could significantly decreased Zn,Fe,Mg content of the blood(F=42.66,5.41,7.04,P<0.05)and excessive intake of fluorine could significantly decreased Zn,Fe,Mg,Cu content of the blood(F=64.50,37.90,9.75,6.74, P<0.05).The coexistence offluorine and Muminum had interaction to the level of Zn(F=31.59,P<0.05)and did not obviously interact with other elements(F=0.91,1.63,1.51.0.00,P>0.05).Compared with the control group [(131.30 ±13.86)μmol/L,(10.24 ±1.02),(1.71 ±0.19)mmol/L,(20.43 ±4.42)μmol/L],Zn content in the high aluminum group[(90.84±9.98)μmol/L]decreased significantly(P<0.05),so did Zn,Fe,Mg content in tlle high fluorine group[(85.85 ±10.92)μmol/L,(8.49 ±0.68),(1.52 ±0.13)mmol/L],the difference being statistically significant(P<0.05)0Zn,Fe,Mg,Cu content in the high fluorine-aluminum group,being(82.82 ±11.00)μmol/L, (8.16±0.45),(1.46±0.09)mmoL/L,(15.69±2.38)μmol/L,respectively,all decreased signitlcarIdy(P<0.05). Compared with the high aluminum group[(9.43±1.09)mmol/L],Fe content of the high fluorine aluminum group[(8.16±0.45)mmol/L]decreased significantly(P<0.05).Conclusions Excessive fluoride can cause blood zn, Fe,Mg,Cu decline,so can excessive aluminum.Combination of excessive fluofine and aiuminum has 8ignificant synergic effect on the level of Zn but have rio influence on Ca.
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Objective To observe the combined poisonous effects of fluoride and aluminum on sex hormone of male rats.Methods Sixteen weaned SD healthy male rats aged two week were selected and divided into control group,aluminum group,fluoride group,fluorine-aluminum group,four rats in each group.All rats in the experimental groups were fed with corn collected from the prevailng areas containing different fluorine contents respectively for 90 days.Serum testosterone(T)and estradiol(E2)were detected.Results Compared separatelv with the control group[(3.317±0.635)μg/L],serum T level of fluorine-aluminum group[(15.994±6.558)μg/L]was higher(P＜0.05),but aluminum[(8.134±3.134)μg/L]and fluorine[(1.868±0.367)μg/L]groups had no significant differences(P＞0.05).Compared separately with the control group[(0.319±0.072)nmol/L],E2 level of the fluorine group[(0.172±0.030)nmol/L]being lower(P＜0.05),and it was not significant differences(P＞0.05)in the control group when compared with aluminum group[(0.282±0.012)nmol/L],and fluorine-aluminum group[(0.265±0.047)nmol/L].Fluorine and aluminum interacted with each other(F=9.82,P＜0.05).Conclusion The combined poisonous effects of fluorine and aluminum may influence sex hormone levels of male rats.
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Objective To study the effects of coal burning related endemic fluorosis on body development and intelligence levels of fluorosic children in Zhijin county.Methods One hundred and seventy-six fluorosic children and 50 healthy(without fluorostic teeth) children aging 7-12 years who were sampled in cluster sampling were examined for urine fluorosis,physical examination and intelligence tests in Zhijin county,with coal burning pollution related fluorosis.Results Physical development of sick children and the children in control group were in low levels. The intelligence levels and the leves of urine fluorosis in study group were lower than those in control group and there was a significant difference(P
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Objective To explore the prevalence satate of children in burning coal endemic fluorosis in Zhijin county of Guizhou province, provide the scientific basis for the prevention of skeletal fluorosis.Methods One thousand six hundred and sixteen children in school under 16 years old that were sampled in cluster sampling were examined with dental fluorosis,X-ray in the type of burning coal pollution fluorosis areas of Zhijin county Guizhou province.Results Total prevalence rate of dental fluorosis was 96.42%,prevalence rate of skeletal fluorosis was 7.49%, constrictive skeletal fluorosis was main type in Zhijin county Guizhou province.Conclusion Prevalence states of dental fluorosis and skeletal fluorosis are still serious, more effectual preventive and control measure shall be used.