Objective To investigate the significance of osteo-immunology related factor transforming growth factor-β1 (TGF-β1) and interleukin 6 (IL-6) in bone of rats with chronic fluorosis.Methods Thirty-six healthy SD rats were divided to three groups according to body weight with the method of random digits table.The rats of control were fed with tap water(NaF ＜ 1 mg/L) and the experimental rats were exposed to NaF (lower dose group:5 mg/L,higher dose group:50 rmg/L) added to the drinking water to establish the chronic fluorosis model.All rats were killed on the six months and the metaphysic of femoral was collected.Bone fluorine was detected by ashing-fluorin ion selective electrode method.Bone tissues were stained with hematoxylin-eosin and observed under optical microscope.The content of bone alkaline phosphatase (BALP) in rat serum was detected by enzyme-linked immunosorbent assay(ELISA).The expressions of TGF-β1 and IL-6 mRNA and protein in bone were detected by in situ hybridization (ISH) and immunohistochemistry (IHC).Results The contents of bone fluorine were increased gradually in the control,the lower and higher doses fluoride groups[(306.04 ± 12.57),(652.91 ± 51.83),(1 094.11 ± 91.41)mg/kg,F =31.14,P ＜ 0.05].Bone sclerosis could be observed under optical microscope in lower and higher dose groups.The content of BALP in serum increased with the dose of fluoride gradually in the control,the lower and higher doses fluoride groups[(27.78 ± 4.09),(46.59 ± 5.75),(57.45 ± 3.99)U/L],expressions of mRNA (111.84 ± 4.62,123.86 ± 7.46,140.83 ± 5.21) and protein (118.60 ± 7.09,133.17 ± 7.33,145.67 ± 9.61) of TGF-β1 were both increased(F =30.29,73,28,33.65,all P ＜ 0.05).The expressions of mRNA(117.78 ± 7.01,119.90 ± 5.10) and protein(122.79 ± 6.49,123.81 ± 7.99) of IL-6 were both higher than those of the control (106.49 ± 6.76,112.11 ± 5.80,F =15.47、10.83,all P ＜ 0.05).Conclusion The expressions of osteo-immunology related factor TGF-β1 and IL-6 in bone of rats with chronic fluorosis have changed,which indicates that fluoride can impact the increased bone formation by regulating the micro environment of bone.

Objective: To evaluate the malignant potential of oral lichen planus (OLP) by measuring and analyzing the dimensions of spinous cell in mucosal epithelium.Methods:Paraffin-embedded samples of oral mucosa, including normal (N,7 cases), OLP(16 cases), oral leukoplakia(12 cases) with mild and moderate epithelial dysplasia (LK) and well differentiated oral squamous cell carcinoma (SCC,10 cases), were analyzed by MIAS-99 System.The planar and stereoscopic dimensions of spinous cells were measured and compared.Results:The values of morphometric and stereologic parameters in OLP were between those in normal mucosa and in LK, significantly different from those in SCC and not significantly different from those in normal epithelium.Conclusion:Based on the changes of cell morphometric parameter value, OLP is generally a benign lesion with some degree of malignant potentiality that is less than LK.Nucleus/cytoplasm volume ratio and volume density of cell nucleus of spinous cells in mucosal epithelium are of value in distinguishing benign from maliganant lesions.

Objective To study the proliferation and apoptosis and investigate the expression of Indian hedgehog (Ihh),parathyroid hormone-related peptide (PTHrp),smoothened (Smo) protein and mRNA in the cultured rat primary chondrocytes exposed to different doses of NaF.Methods The third generation articular chondrocytes of neonate rat were cultured in vitro and treated with 0 (control),5,10,20 and 40 mg/L of fluoride.The proliferation activities of cells at different times (24,48 and 72 h) were tested by Thiazolyl Blue Tetrazolium Bromide (MTT).The apoptosis rate was determined by flow cytometry.The expressions of protein and mRNA of Ihh,Smo and PTHrp at 48 h were determined by Western blotting and semi-quantitative RT-PCR,respectively.Results After exposed to 5 mg/L of fluoride for 24,48 and 72 h,the proliferation rates were significantly increased [(1.17 ± 0.07)％,(1.20 ±0.06)％,(1.16 ± 0.08)％] compared with those of control group [(1.10 ± 0.08)％,(1.13 ± 0.08)％,(1.15 ± 0.08)％],but the proliferation activity at 48 and 72 h in 40 mg/L group [(0.72 ± 0.11)％,(0.68 ± 0.04)％] was significantly lower than those in control group (all P ＜ 0.05).Compared with the control group,apoptosis rate of cartilage cell in fluoride treatment group increased gradually [(1.47 ± 0.05)％,(19.87 ± 3.03)％,(25.30 ± 1.28)％,(45.73 ± 4.63)％,F =123.328,P ＜ 0.01].Western blot analysis and RT-PCR results showed that the Ihh,PTHrp,Smo mRNA and protein expression increased in the fluoride groups at 48 h (Ihh protein:0.77 ± 0.08 vs.0.98 ±-0.07,1.23 ± 0.06,1.37 ±0.07,1.34 ± 0.07;PTHrp protein:0.68 ± 0.04 vs.0.89 ± 0.05,0.83 ± 0.05,1.29 ± 0.05,1.16 ± 0.08;Smo protein:0.37 ± 0.01 vs.0.64 ± 0.06,0.67 ± 0.03,0.96 ± 0.06,0.69 ± 0.06;Ihh mRNA:0.77 ± 0.05 vs.0.98 ± 0.05,1.09 ±0.05,1.27 ± 0.03,1.46 ± 0.06;PTHrp mRNA:0.67 ± 0.07 vs.0.97 ± 0.05,1.07 ± 0.08,1.37 ± 0.05,1.45 ± 0.05;Smo mRNA:0.45 ± 0.03 vs.0.63 ±-0.04,0.71 ± 0.05,0.81 ± 0.01,1.00 ± 0.02,all P ＜ 0.05).Conclusions Low doses of fluoride can promote the proliferation of chondrocytes cultured in vitro,and high doses of fluoride can promote the apoptosis of chondrocytes cultured in vitro.The expression of Ihh signaling pathway RNAs and proteins of the cartilage cells are increased following increased levels of fluoride.The results suggest that fluorine has activated the Ihh signaling pathway in chondrocytes and promoted the proliferation and apoptosis processes which might be involved in chondrocytes injury.

As an important protein molecule for innate immunity,Toll-like receptor 7 (TLR7) is also a bridge between innate immunity and adaptive immunity.When TLR7 is activated,it can exert its effect on liver diseases through different signaling pathways.Studies on the role and application of TLR7 agonist in liver diseases are increasing in recent years.This article reviewed the research on TLR7 agonist in liver diseases.

Objective To investigate the effects of silencing Smo gene on proliferation and apoptosis of rat prima-ry chondrocyte in vitro.Methods The primary chondrocyte was obtained by mechanical-enzyme digestion and identified by Immunohistochemical cells ( ColⅡ) .The animals were divided into control group , control siRNA group and Smo siRNA 1 ~3 group.The siRNA was transfected into chondrocytes by lentivirus vector .After 72 h, the cell viability was detected by MTT, Smo expression was detected by RT-PCR and Western blot, and the apoptosis of chondrocyte was assessed by flow cytometry .Results All types of siRNA were transfected into primary chondrocyte by vectors, the Smo siRNA 1 ~3 may inhibit the expression of Smo mRNA and protein in chondrocytes, and Smo siRNA2 had the highest silencing rate ( the expressions of Smo mRNA and protein were 0.19 ±0.03 and 0.39 ±0.07 ) .The cell viability in Smo siRNA2 group was lowest ( 77.38% ±7.19%) , while the apoptosis rate of Smo siRNA2 was highest ( 21.43%±2.97%) .Conclusions Silencing Smo gene in primary chondrocytes may inhibit proliferation and promote apoptosis , Smo may have a protecting role from apop-tosis of the chondrocyte.

Objective To explore the role of Janus kinase/signal transduction and transcriptional activation (JAK/STAT) pathway in rat liver damaged by excessive fluorine.Methods Thirty-six healthy Sprague-Dawley (SD) rats were randomized by weight and divided into three groups (6 males and 6 females per group):a control group (drunk water containing NaF ＜1 mg/L) and two fluorosis groups (drunk water containing NaF of 5 mg/L and 50 mg/ L).After 6 months of experiment treatment,the fluorine contents of urine and bone were detected by fluorine-ion electrode method.The rats liver function was determined by automatic blood chemical analyzer.The protein expression of Janus kinase (JAK1),signal transducer and activator of transcription (STAT3),B-cell lymphoma/ leukemia-2 (Bcl-2) and Bcl-associated x protein (Bax) were detected by immunohistochemistry (IHC) and protein imprinting (Western blotting).The activity of superoxide dismutase (SOD),glutathione peroxidase (GSH-PX) and the content of lipid peroxide (LPO) in liver tissue were determined with oxidative stress kit.Results The fluorine contents in the urine and bone in low-[(1.90 ± 0.12)mg/L,(210.37 ± 15.81)mg/kg] and high-dose [(2.20 ± 0.17)mg/L,(222.84 ± 10.21)mg/kg] fluoride groups were higher than those of control group [(1.74 ± 0.11)mg/L;(165.48 ± 10.37) mg/kg,F =33.840,69.149,P ＜0.05];the activity of serum alanine aminotransferase (ALT) and aspartate transaminase (AST) in high-dose fluorosis group [(69.83 ± 11.18),(167.56 ± 50.85) U/L] was higher than those of control group [(42.67 ± 7.07),(126.31 ± 16.76)U/L,F =32.135,4.984,all P ＜0.05];the protein expression of JAK1,STAT3 and Bax (1.56 ± 0.31,1.49 ± 0.49,1.41 ± 0.55) in high-dose fluorosis group were significantly higher than those of control group(1.01 ± 0.11,1.04 ± 0.15,0.87 ± 0.21,F=10.923,5.361,5.009,all P＜0.05),and Bcl-2 (0.61 ± 0.15) was significantly lower in high-dose fluorosis group than control group (1.04 ± 0.17,F =16.017,P ＜0.05);the activities of SOD and GSH-PX [(7.22 ± 0.88),(7.23 ± 2.47)U/mg prot] were significantly lower in high-dose fluorosis group than control group [(9.52 ± 1.51),(12.01 ± 5.16)U/mg prot,F =11.627,4.824,all P ＜0.05],and the contents of LPO [(9.23 ± 2.24)μmol/g prot] was significantly higher in high-dose fluorosis group than control group [(6.09 ± 1.55)μmol/g prot,F =7.457,P ＜0.05].Conclusion JAK/STAT signaling pathway and the oxidative stress,apoptosis may be the pathogenesis of liver damage in chronic fluorosis.

Objective To investigate the effects of chronic fluorosis on the expressions of matrix metalloproteinase-9 (MMP-9) mRNA and protein and the differentiation and maturation process of bone cell in the osteoclast of bone tissue of rats.Methods According to body weight,thirty-six healthy SD rats(body mass 100-120 g) were divided into three groups by random number table,twelve in each group,half male and half female.The rats of control group were given tap water(NaF ＜ 1 mg/L),and rats of low-fluorine and high-fluorine groups were fed with tap water containing 5 and 50 mg/L NaF to establish chronic fluorosis model.Rats were sacrificed after eight months; the contents of urinary fluoride in 24 hours and bone fluoride were analyzed by fluoride selective electrode.Serum content of tartrate resistant acid phosphatase 5b(TRACP5b)was detected by enzyme-linked immunosorbent assay (ELISA).The paraffin section of bone tissue was stained by hematoxylin-eosin (HE) and pathological morphometry was observed under optical microscope.The protein and mRNA levels of MMP-9 in the osteoclast of bones were detected by immunohistochemistry (IHC) and in situ hybridization (ISH),respectively.Results The differences of fluoride contents of urine and bone in rats were statistically significant between groups(F =400.612,48.229,all P ＜ 0.05).Fluoride contents of urine and bone were increased in lowfluorine and high-fluorine groups[(6.09 + 0.56),(7.69 + 0.64)mg/L,(12.65 ± 3.07),(26.53 + 5.88)mg/kg] compared to the control groups[(1.36 ± 0.51)mg/L,(0.67 ± 0.16)mg/kg,all P ＜ 0.05],and the fluoride contents of urine and bone were gradually increased with increasing fluoride doses(all P ＜ 0.05).The difference of TRACP5b content in serum was statistically significant between groups (F =9.607,P ＜ 0.05),in low-fluorine and high-fluorine groups,the TRACP5b contents[(1.86 ± 0.13),(1.92 ± 0.22)U/L] were higher than that of control group [(1.57 + 0.20)U/L,all P ＜ 0.05].The pathological examination showed osteosclerosis in fluoride exposed groups.The differences of MMP-9 mRNA and protein expressions were statistically significant between groups (F =365.727,331.382,all P ＜ 0.05).Compared to the control groups(97.22 ± 2.24,78.51 ± 1.16),the expressions of MMP-9 protein(108.18 ± 1.97,119.28 ± 1.76) and mRNA(89.44 ± 2.86,102.14 ± 2.39) were increased(all P ＜ 0.05),and the expressions of MMP-9 mRNA and protein were gradually increased with increasing fluoride doses (all P ＜ 0.05).Conclusions Chronic fluorosis might influence osteoclast differentiation and maturation process through regulating the expression levels of MMP-9 protein and mRNA.

Animals ; Apoptosis ; Cell Differentiation ; Cell Proliferation ; Fluoride Poisoning ; metabolism ; pathology ; Fluorosis, Dental ; metabolism ; pathology ; Hedgehog Proteins ; genetics ; metabolism ; Humans ; Osteoblasts ; cytology ; metabolism ; Osteoclasts ; cytology ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Signal Transduction ; Stomach Neoplasms ; metabolism ; pathology

This study aimed at exploring the economic operation and developing trend of county-level public TCM hospitals in Guangxi Province,promoting the comprehensive reform of county-level public hospitals with feasible countermeasures and suggestions.In this study,forty-seven county-level public TCM hospitals were involved in the analysis of their economic operations and benefits by constituent ratio analysis,ratio analysis and trend analysis.As a result,the average gross income of the hospitals exceeded the total expenditure with a slight surplus.Drug income contributed the biggest share of the medical income,while the ratio of Chinese drug income to the medical income was increasing year by year.The debt ratio increased from 38.7％ in 2009 up to 44.6％ in 2013,being over 30％ in this period.In conclusion,it was demonstrated that problems were found in the lack of financial assistance,the low efficiency of economic management and the increasing average medical charges,which should be addressed by increasing investments to TCM hospitals,strengthening the performance appraisal and adjusting the charges of TCM medical services.

Objective:To observe the protein and mRNA expressions of microtubule-associated protein 1 light chain 3 (LC3)B, P62 and Beclin1 in the liver of rats with chronic fluorosis, and to explore the role of autophagy in pathogenesis of liver injury induced by fluorosis.Methods:Using a group design, 54 SD rats were divided into 9 groups according to their weight (100 - 120 g) using a random number table method, each group with 6 rats, half male and half female. They were control group (NC group), low fluoride group (LF group), high fluoride group (HF group), NC + rapamycin (RAP) group, LF + RAP group, HF + RAP group, NC + chloroquine (CQ) group, LF + CQ group, and HF + CQ group. The NC group drank tap water (fluoride concentration was 0.5 mg/L), LF group drank fluoride water (fluoride concentration was 5.0 mg/L), HF group drank fluoride water (fluoride concentration was 50.0 mg/L); NC + RAP group, LF + RAP group and HF + RAP group were fed with corresponding drinking water, respectively, for 3 months, and then RAP (1.5 mg/kg) was intraperitoneally administered for 10 d; NC + CQ group, LF + CQ group and HF + CQ group were fed with corresponding drinking water, respectively, for 3 months, and then CQ (60 mg/kg) was intraperitoneally administered for 10 d. Bone and 24-hour urine samples of rats in each group were collected to detect the contents of bone fluoride and urine fluoride; liver histomorphological changes were observed through hematoxylineosin staining; protein and mRNA expressions of LC3B, P62 and Beclin1 in liver were detected by immunohistochemistry and real-time fluorescence quantitative PCR, respectively.Results:Compared with the NC group [(0.03 ± 0.00) mg/kg, (0.34 ± 0.08) mg/L], the contents of bone fluoride [(3.86 ± 0.08) mg/kg] and urine fluoride [(1.11 ± 0.16) mg/L] in HF group were higher ( P < 0.05). In the NC group, the lobule structure of liver tissue was clear, the hepatic cords were arranged in order, and the cell structure was normal. There were different degrees of hepatocyte edema in LF and HF groups. After intraperitoneal injection of RAP, compared with the corresponding fluoride group, the morphology of hepatocytes did not change significantly. After intraperitoneal injection of CQ, compared with the corresponding fluoride group, the liver cells showed obvious edema, and the degree of edema aggravated with the increase of fluoride concentration. Compared with the NC group, the protein expressions of LC3B and Beclin1 in HF group were higher ( P < 0.05), and the protein expression of P62 was lower ( P < 0.05). After intraperitoneal injection of RAP, the protein expressions of LC3B and P62 in LF + RAP group was lower than that in LF group ( P < 0.05); Compared with HF group, the protein expressions of LC3B and Beclin1 in HF + RAP group were lower ( P < 0.05). After intraperitoneal injection of CQ, protein expression of P62 in LF + CQ group was higher than that in LF group ( P < 0.05); Compared with HF group, protein expression of P62 in HF + CQ group was higher ( P < 0.05). Conclusions:Early (3 month) fluoride intake could promote autophagy and induce edema of hepatocytes in rats, and RAP had similar effects. CQ may induce liver injury by inhibiting autophagy of hepatocytes.