Cell Line, Tumor ; Cell Proliferation ; drug effects ; Flow Cytometry ; methods ; Humans ; Point Mutation ; Stomach Neoplasms ; genetics ; pathology ; Tumor Suppressor Protein p53 ; genetics ; pharmacology

Objective To investigate the effect of fluoride on expression of Runx2 mRNA and protein in bone tissue of rats. Methods Fourteen SD rats were randomly divided into two groups: control group(tap water with fluoride ＜ 0.06 mg/L), and fluorosis group(fluoride 50 mg/L in water). After 4 moths, expressions of both mRNA and protein of Runx2 in rat bone tissue were determined by RT-PCR and Western blotting. Results The results showed that the expression of Runx2 mRNA and protein in fluoride-treated bone tissue were 2.287 ± 0.261 and 0.929 ± 0.229, respectively, both of which were significantly higher than those of control group(0.995 ± 0.123,0.317 ± 0.068, t = 11.85,6.78, P ＜ 0.05). Conclusions Fluoride can increase the expression of Runx2 mRNA and protein in bone tissue of rats, and Runx2 may be involved in the pathogenesis of bone injury caused by fluoride.

Objective To observe the expression of osteoprotegerin (OPG), receptor activator of nuclear factor κβ ligand(RANKL) and receptor activator of nuclear factor κβ (RANK) in bone tissue of rats with chronic fluorosis and to explore the relation between OPG/RANKL/RANK system and bone damage in chronic fluoride poisoning rat and the antagonism effects of Danlan Xianpeng capsule. Methods SD rats were randomly divided into six groups according to body weight (equal male and female in each group): fluorosis group, high dose drug group, medium dose drug group, low dose drug group, control group, borax group(positive control), 12 rats in each group. The control group drank tap water and the remaining 5 experimental groups consumed 50 mg/L fluoride water, and high, medium and low doses drug group took Danlan Xianpeng capsule at doses of 0.8,0.4,0.2 g/kg,borax group took borax at dose of 0.8 g/kg. OPG, RANKL, RANK protein in rat tibial metaphysis was detected by immunohistochemistry at the 6 month. Results Compared with the control group(173.79 ± 5.23, 174.17 ± 5.01,155.63 ± 7.11), the expressions of OPG, RANKL were increased and the expression of RANK was decreased in fluorosis group(156.83 ± 5.80, 157.74 ± 6.70, 173.92 ± 4.37), the difference was statistically significant (all P ＜ 0.05). Compared with the fluorosis group, the expression of OPG and RANKL were decreased and the expression of RANK was increased in high-dose drug group, middle-dose drug group(169.67±5.07, 168.08 ± 5.05,162.12 ± 4.24, 170.78 ± 5.01, 168.41 ± 7.19, 166.69 ± 5.78, all P ＜ 0.05). Compared with the borax group (167.27 ± 4.08, 167.85 ± 5.01, 166.14 ± 3.95), the expression of OPG and RANKL was increased in the low-dose drug group (163.40 ± 4.11, 159.49 ± 5.78), the expression of RANK was increased in the high-dose drug group (162.12 ± 4.24) and decreased in the low-dose drug group(171.54 ± 8.06), the difference was statistically significant (all P ＜ 0.05). Conclusions Chronic fluoride poisoning can cause increased bone turnover and enhance the activity of osteoelastic absorption by increasing RANKL. Danlan Xianpeng capsule can affect bone remodeling through the OPG/RANKL/RANK system, and antagonises bone damage caused by fluoride.

Objective To evaluate the effects of perioperative rehabilitation for patients with self-forearm internal arteriovenous fistula.Methods Divided 124 patients with end-stage renal disease into the functional training group (65 cases,A group)and the control group(59 cases,B group) randomly,and then compared certain related indexes between the two groups.Results In the A group,the artery diametcr was obvious widen before functional training without operation,and the vein diameter was obvious widen before functional training without operation too,P ＜ 0.05.After operation,the A group Anastomotic unobstructed was obviously better than B group in one-stage,P ＜ 0.05.A group with thrombosis cases significantly less postoperative than B group,P ＜ 0.05.A group with edema of hands was significantly shorter than B group,P ＜ 0.01.Postoperative fistula time to maturity in the A group was (43.76 ± 2.21) days,which was significantly shorter than that of in the B group,(49.69 ± 5.85) days,P ＜ 0.01.Conclusion Systematic strengthening of perioperative functional exercises can significantly improve preoperative vascular conditions,improve the success rate of operation,reduce risk of thrombosis cases,shorten the time of hand edema and the time of fistula in mature.

Objective To explore the clinical characteristics of Gitelman syndrome in children and the difference between Gitelman syndrome and Bartter syndrome.Methods Clinical date,biochemical tests and therapy of 6 patients diagnosed as Gitelman syndrome in Beijing children′s hospital from Mar.to Dec.2006 were retrospectively analyzed.At the same time,the relative articies of Gitelman syndrome and Bartter syndrome were reviewed.Results The symptoms of 6 patients appeared early.The age of onset of Gitelman syndrome at infancy stage,the main complains were growth delay,weakness,tetany.All patients had normal blood pressure.The biochemical tests showed hypocalemic,hypomagnesium,alkalosis and hyperreninemia.But the concentration of aldosterone was normal or little higher.The manifestations of all patients were relieved after taking both potassium and magnesium.Conclusion Gitelman syndrom and Bartter syndrome have differences at clinical syndrome and machanism of onset.

Objective To detect the osteopontin(OPN)expression in renal tissue of rats with fluorosis and low calcium diet,and study the role of OPN in renal injury of fluorosis.Methods Forty-eight aged 1 month Wistar rats,80-120 g,were randomly divided into 4 groups by 2×2 factorial design(the number of female and male in each group was equal):the control group,high-flluoride group,low-calcium group and low-calcium with high-fluoride group.All rats of the fluorosis groups were fed with feed containing corn exposed to coal-burning from endemic fluorosis areas with high fluoride(100 mg/kg,corn),the other two groups were fed with feed containing coru from nonendemic fluorosis areas(fluoride 5 mg/kg,corn).After 16 weeks,the rats were killed.The change of teeth was examined,and the incidence rate of dental fluorosis was calculated.The expressions of both protein and mRNA of OPN in rat renal tissue were determined by RT-PCR and immunohistochemistry after four-month experimentation.Results The growth of teeth was very well in the control group and the low-calcium group.The two high-fluoride groups showed evident dental fluorosis(100%).The results of immunohistochemistry showed that the OPN protein was localized in renal tubule cytoplasm.The OPN-positive cells from renal tissue were lightly and scatteredly stained in control and low-calcium groups.The OPN-positive cells had deeper color in high-fluoride group and low-calcium with high-fluoride group,widely distributed in the renal tubular epithelial cells.The protein expression of OPN in the two groups exposed to fluoride(168.64±13.21,169.26±8.92)was significantly higher than those of the corresponding control group(145.78±10.26,all P<0.01)and low-calcium group(149.60±16.84,all P<0.01).The mRNA expression of OPN in the two groups exposed to fluoride(1.89±0.37,1.94±0.22)was significantly higher than those of the corresponding control group(1.32±0.26,all P<0.05)and low-calcium group(1.30±0.186,P<0.05),respectively.High fluoride influenced the expression of protein and mRNA of OPN(F=13.821,4.24,all P<0.05).Low calcium did not affect the expression of protein and mRNA of OPN(F=2.164,0.58,all P>0.05).However,high fluoride and low calcium had a cross interaction on the expression of protein and mRNA of OPN(F=6.257,432,all P<0.05).Conclusions Over-dose fluoride enhances the expression of OPN.The higher expression observed in the cases exposed to high fluoride concentration is associated with serious renal injury.OPN may he a potential marker for renal injury in fluorosis.Moreover,over-dose fluoride and low calcium make the renal injury worse,indicating low calcium plays an important part in renal injury by fluoride.

Objective:To investigate the compliance with early Warfarin anticoagulation therapy after cardiac mechanical valve replacement(MHVR)and its related factors in elderly patients.Methods:The convenience sampling method was used to prospectively recruit 210 patients undergone MHVR at the Ningbo Medical Center of Lihuili Hospital from January 2017 to October 2019.Six months after discharge, face-to-face interviews or telephone follow-ups were conducted to assess general information, warfarin anticoagulation knowledge, anticoagulant treatment compliance and social support.Results:The overall compliance of early Warfarin anticoagulation therapy was excellent, with 99.5% of patients compliant with medication and 99.0% compliant with INR monitoring, both higher than the rate of compliance with advised lifestyle adjustment(92.1%). Anticoagulation knowledge and age were the main influencing factors for compliance in elderly patients after MHVR.Conclusions:The compliance with early Warfarin anticoagulation therapy after MHVR is good in elderly patients in the Ningbo area.The correlation analysis suggests that medical professionals need to promote education on anticoagulation knowledge and pay more attention to anticoagulation compliance in elderly people.

Objective To investigate the expression and distribution of the downstream substrate of extracellular regulated protein kinase(ERK1/2) pathway, ternary complex factor phospho-Elk-1, in rat brains with chronic fluorosis, and reveal the mechanism of the impaired learning and memory ability caused by chronic fluorosis. Methods Seventy-two SD rats, weighing 100 - 120 g, were randomly divided into 3 groups, 24 in each group (half male and half female). The rats in control group were fed with tap water (fluoride < 0.5 mg/L); low- and high-dose fluoride groups were fed with tap water with different concentrations of NaF(5.0,50.0 mg/L F-, respectively). After 6 months, body weight was weighed, dental fluorosis was determined by observation and urinary fluoride and bone fluoride were detected by fluorine ion-selective electrode; the learning ability of rats was measured by navigation test of Morris water maze, and memory ability by spatial probe test in Morris water maze; the expression and distribution of phospho-Elk-1 in different brain regions were detected by immunohistochemistry method. Results In low- and high-fluoride groups, the body weight of rat[(449.2 ± 77.1), (312.8 ± 89.7)g] was significantly decreased than that of control [(635.5 ± 76.2 )g, all P< 0.05], the varying degrees of dental fluorosis were observed(x2 = 7.83, P<0.05), urinary fluoride[(2.56 ±0.91),(5.73 ±3.14)mg/L] and bone fluoride[(709.2 ± 37.4) ,(1306.3 ± 102.4) mg/kg] were significantly higher than those in controls[(0.92 ± 0.30)mg/L,(348.5 ± 89.2)mg/kg, all P< 0.05]. The escape latency of low- and high-fluoride groups[ (7.4 ± 4.1), (12.2 ± 5.7)s] was longer than that of control [(4.8 ± 2.7 )s, all P < 0.05] and the escape latency in high-fluoride group was significantly longer than that in other groups (all P < 0.05); in spatial probe test, the time of first crossing platform was longer in rats with fluorosis [(4.18 ± 1.10),(5.89 ± 0.56)s] as compared to control[(1.17 ± 0.75)s, all P< 0.05]. Expressions of phospho-Elk-1 in the hippocampus CA1(167.4 ± 8.3,163.2 ± 9.4), CA2(175.7 ± 5.0,183.3 ± 4.2), CA3(165.2 ± 11.6,162.9 ± 4.4), CA4(168.7± 6.9,169.5 ±5.3), fascia dentate (185.2 ±4.0,193.1 ±6.1) and caudate putamen( 181.4 ± 3.8, 179.8 ± 5.5) in low- and high-fluoride groups were higher than those of controls(142.4 ± 8.1,144.9 ± 8.4,143.6 ± 5.8, 116.8 ± 9.1,140.2 ± 7.8,163.1 ± 13.1, all P< 0.05). Conclusion Chronic fluorosis can cause increased expression of phospho-Elk-1 in the hippocampus and caudate putamen region of rat brains, which might be related to the mechanisms of decreased learning and memory ability of rats overexposed to fluoride.

Objective To investigate the changes of reactive oxygen species(ROS) level and mitochondria fission-fusion-balance in cortical neurons of rats with chronic fluorosis and reveal the correlation between these two factors. Methods One hundred and twenty rats were randomly divided into 3 groups(control group, low-dose fluorosis group, high-dose fluorosis group) and 40 rats were in each group according to body weight and the experiments were carried out for 3 months or 6 months. The rats were fed with different concentrations of fluoride (NaF) to establish fluorosis models. Controls were fed with tap water( < 0.5 mg/L), experimental animals in low- or high-dose group were fed with water containing NaF 10.0,50.0 mg/L, respectively. The level of ROS and the morphology in mitochondria fission-fusion balance in neurons of the cortex of rat brains prepared with cortical frozen sections were detected with ROS fluorescent probe and MitoTracker RED probe, respectively. Results Significant differences of the level of ROS and the numbers of abnormal mitochondria in morphology in the cortical neurons were found between 3 groups at the experiment period of 3 month and 6 month(F= 3.07,3.06,3.05,3.07, all P < 0.05). As compared with control group(10.43 ± 5.98,4.12 ± 3.86) at the experiment period of 3 month, the level of ROS and the numbers of abnormal mitochondria in morphology in the cortical neurons were obviously increased in high-dose fluorosis group(25.48 ± 6.09,20.47 ± 6.09, all P < 0.05), whereas no significant changes were found in low-dose fluorosis group(11.67 ± 3.49,6.68 ± 3.48, all P> 0.05). Furthermore, the increases in both ROS level and abnormal numbers of mitochondria were significant observed in the cortical neurons of low-dose fluorosis group (63.02 ± 8.15, 49.33 ± 8.61) and high-dose fluorosis group(65.60 ± 7.40,53.10 ± 6.95) as compared with the control group (25.26 ± 6.41,20.26 ± 6.41) at the experimental period of 6 month (all P < 0.05). The abnormal numbers of mitochondria correlated with ROS level(r = 0.93,0.81, all P < 0.05). Conclusions Taking excessive amount of fluoride results in high level of oxidative stress and impaired the balance of mitochondrial fission-fusion,which is dependent on the feeding times and doses of fluoride. The mechanism of the mitochondrial abnormalities might be associated with the high level of oxidative stress induced by chronic fluorosis.

Objective To observe the expression of mitochondrial fission protein locus Fis1 and ultrastructural changes in the renal cells of rats with chronic fluorosis,and to reveal the mechanism in mitochondrial damage of the renal cells.Methods Sixty SD rats were randomly divided into 3 groups according to sex and body mass(20 in each group):control group,lower fluoride group and higher fluoride group.All the rats were fed with different doses of sodium fluoride in drinking water(0,10 and 50 mg/L,respectively).Six-month later,the expression of Fisl in renal cells was determined by real-time fluorenscence quantitative PCR and immunohistochemistry technology,the mitochondrial morphology of renal cells was observed under transmission electron microscopy (TEM).Results As compared with the control group(28.70 ± 12.41),Fis1 mRNA levels(91.48 + 34.83 and 582.09 ± 184.69) in renal cells of the lower fluoride and the higher fluoride groups were increased(all P ＜ 0.05).As compared with the control group(10.49 ± 7.66),Fisl protein levels(16.33 ± 10.26 and 21.50 ± 5.24) in renal cells of the lower fluoride and the higher fluoride groups showed a trend of increasing,the higher fluoride group was higher than that of the control group(P ＜ 0.05).By TEM,mitochondrial crest in renal cells of the lower fluoride and the higher fluoride groups was vague or disappeared,mitochondrial division section appeared.Conclusions Fluoride is a kind of toxicant that can cause damage to mitochondrion of renal cells,induce the expression of Fis1 in transcriptional and protein level,and lead to the obstacles of mitochondrial fusion-fission and ultrastructural abnormality of mitochondrion,which may play an important role in mechanism of mitochondrial damage in the renal cells of rats with chronic fluorosis.