Alleviating effects of selenium on fluoride-induced testosterone synthesis disorder and reproduction toxicity in rats.

Fluoride (F) exists widely in food, water and other natural resources, and can cause damage to the reproductive system of human and animals. Studies have shown that selenium (Se) is a necessary trace element to maintain the normal male reproductive system. However, it is not clear whether it can alleviate the damage of reproductive system induced by F. Hence, sodium fluoride (NaF) was administered singly in drinking water at 100 mg L-1 alone and co-administered by drinking with sodium selenite (Na2SeO3) at 0.5, 1.0, 2.0 mg L-1 for 10 consecutive weeks. The results demonstrated that the sperm deformity rate were increased significantly by F, however, it was improved significantly after the addition of 2.0 mg L-1 Na2SeO3. The contents of glutathione peroxidase 4 (GPX-4), selenoprotein P (SePP), pregnenolone (PREG), androstenedione (ASD), and testosterone (T) were reduced obviously in the F group, however, it was increased significantly after adding 0.5, 1.0 and 2.0 mg L-1 Na2SeO3. F decreased noticeably the mRNA and protein expression levels of steroidogenic acute regulatory protein (StAR), cytochrome P450 cholesterol side chain lyase (P450scc), 3ß-hydroxysteroid dehydrogenase (3ß-HSD), cytochrome P450 17α-hydroxylase (P450c17) and 17ß-hydroxysteroid dehydrogenase (17ß-HSD), which was increased obviously after the addition of 1.0 and 2.0 mg L-1 Na2SeO3. In summary, 2.0 mg L-1 Na2SeO3 can alleviate testosterone synthesis disorder induced by F via reducing oxidative stress, increasing the level of selenoprotein in testis and regulating the content of related hormones and enzyme activity during testosterone synthesis pathway.

Calcium alleviates fluoride-induced kidney damage via FAS/FASL, TNFR/TNF, DR5/TRAIL pathways in rats.

Long-term excessive intake of fluoride (F) can cause osseous and non-osseous damage. The kidney is the main fluoride excretion organ of the body. This study aimed to explore whether dietary calcium (Ca) supplementation can alleviate kidney damage caused by fluorosis and to further investigate the effects of Ca on the mitigation mechanism of renal cell apoptosis triggered by F. We evaluated the histopathological structure, renal function indicators, and gene and protein expression levels of death receptor-mediated apoptosis pathways in Sprague Dawley (SD) rats treated with sodium fluoride (NaF) and/or calcium carbonate (CaCO3) for 120 days. The results showed that 100 mg/L NaF induced kidney histopathological injury and apoptosis, increased the concentrations of Creatinine (CRE), uric acid (UA), blood urea nitrogen (BUN), potassium (K), phosphorus (P) and F (p < 0.05), and decrease the level of serum magnesium (Mg) (p < 0.05). Moreover, NaF increased the mRNA and protein expression levels of Fas cell surface death receptor (FAS), tumor necrosis factor (TNF), TNF-related apoptosis-inducing ligand (TRAIL), Caspase 8, Caspase 3 and poly ADP-ribose polymerase (PARP) (p < 0.01), which finally activated the death receptor pathway. Inversely, Ca supplementation reversed the decrease of CRE, BUN, UA, F and P levels induced by F, alleviated histopathological damage and apoptosis, and reduced the gene and protein expression levels of death receptor pathway-related markers. In conclusion, 1% Ca alleviates F-induced kidney apoptosis through FAS/FASL, TNFR/TNF, DR5/TRAIL signaling pathways.

Elevated expression of glutaminase confers glucose utilization via glutaminolysis in prostate cancer.

Cancer cells reprogram their metabolism towards aerobic glycolysis and elevated glutaminolysis, which contributes to the aggressive phenotype. Understanding how these metabolic pathways are regulated may provide critical targets for therapeutic intervention. Glutaminase (GLS1) is a key enzyme that converts glutamine to glutamate. In this study, we show the loss of GLS1 function by RNA interference or inhibitor diminished the rates of glucose utilization, growth and invasiveness of prostate cancer cells. We propose that GLS1 positively regulates glucose uptake in addition to glutaminolysis. Further, GLS1 involves the transcriptional repression of thioredoxin interacting protein (TXNIP), which is a potent negative regulator of glucose uptake and aerobic glycolysis. Most importantly, we provided direct evidence that elevated GLS1 expression was highly correlated with the tumor stage and progression in prostate cancer patients. Together, we defined a key role for GLS1 in coupling glutaminolysis of the TCA cycle with elevated glucose uptake and consequently the growth of prostate cancer cells. These data extends the role of GLS1 in regulating cell metabolism and the clinical utility of GLS1 inhibitors in the restriction of essential nutrients.

Threshold microsclerotial inoculum for cotton verticillium wilt determined through wet-sieving and real-time quantitative PCR.

Quantification of Verticillium dahliae microsclerotia is an important component of wilt management on a range of crops. Estimation of microsclerotia by dry or wet sieving and plating of soil samples on semiselective medium is a commonly used technique but this method is resource-intensive. We developed a new molecular quantification method based on Synergy Brands (SYBR) Green real-time quantitative polymerase chain reaction of wet-sieving samples (wet-sieving qPCR). This method can detect V. dahliae microsclerotia as low as 0.5 CFU g(-1) of soil. There was a high correlation (r=0.98) between the estimates of conventional plating analysis and the new wet-sieving qPCR method for 40 soil samples. To estimate the inoculum threshold for cotton wilt, >400 soil samples were taken from the rhizosphere of individual plants with or without visual wilt symptoms in experimental and commercial cotton fields at the boll-forming stage. Wilt inoculum was estimated using the wet-sieving qPCR method and related to wilt development. The estimated inoculum threshold varied with cultivar, ranging from 4.0 and 7.0 CFU g(-1) of soil for susceptible and resistant cultivars, respectively. In addition, there was an overall relationship of wilt incidence with inoculum density across 31 commercial fields where a single composite soil sample was taken at each field, with an estimated inoculum threshold of 11 CFU g(-1) of soil. These results suggest that wilt risk can be predicted from the estimated soil inoculum density using the new wet-sieving qPCR method. We recommend the use of 4.0 and 7.0 CFU g(-1) as an inoculum threshold on susceptible and resistant cultivars, respectively, in practical risk prediction schemes.

[Effects of adenovirus-mediated PTEN on the proliferation of prostate cancer PC-3 cells and expressions of cyclin D1 and p21].

OBJECTIVE: To construct a recombinant adenovirus expression vector containing the anti-oncogene PTEN and to investigate the effects of the PTEN gene on the proliferation of prostate cancer PC-3 cells and the expressions of cyclin D1 and p21 in the PC-3 cells. METHODS: The PTEN gene was amplified from the rat hippocampus by RT-PCR and cloned into the shuttle plasmid pEN-TR2A. The plasmids were constructed and amplified in 293A cells. Prostate cancer PC-3 cells were cultured in vitro and infected with the adenoviral vector carrying the PTEN gene (Ad-PTEN). The up-regulation of the PTEN protein was measured by indirect immuno-fluorescence assay; the expressions of PTEN, cyclin D1 and p21 in the cells infected with Ad-PTEN and Ad-LacZ were determined by RESULTS: The Western blot; and the effect of PTEN on the cell proliferation was detected by MTT assay and plate colony formation. recombinant adenoviral vector Ad-PTEN was successfully constructed. Western blot showed a significantly increased expression of the PTEN protein in the PC-3 cells infected with Ad-PTIEN (0.215 +/-0.065) as compared with that in the control ([0.052 +/-0.009], t = 4. 30, P <0.05) and the Ad-LacZ group ( [0. 056 +/- 0.008 ] , t =4.21, P <0.05). The expression of cyclin D1 was significantly lower in the Ad-PTEN-infected PC-3 cells (0. 256 +/- 0. 072) than in the control ( [0. 502 +/- 0. 087 ], t = 3.77, P < 0.05) and the Ad-LacZ group ([0.498 +/-0.081] , t =3.87, P <0.05), while the expression of p21 remarkably higher in the Ad-PTEN-infected PC-3 cells (0.589 +/-0. 076) than in the control ([0. 146 +/-0.026] , t = 9.55, P<0. 01) and the Ad-LacZ group ([0. 163 +/-0. 024] , t = 9.26, P <0.01). Ad-PTEN significantly inhibited the growth of the PC-3 cells (21.98%) at 48 h (t = 6.80, P <0.01). The colony formation rate of the PC-3 cells was (37.4 +/-4. 18)% in the Ad-PTEN group, significantly lower than (54.9 +/-4.81)% in the control (t =4.76, P<0.01) and (56.5 +/- 5.42)% in the Ad-LacZ group (t=4.83, P<0.01). CONCLUSION: The expression of PTEN induced by Ad-PTEN can significantly inhibit the proliferation of PC-3 cells, down-regulate the expression of cyclin D1, and up-regulate the expression of p21.

Micromotor Based Mini-Tablet for Oral Delivery of Insulin.

Diabetes is a metabolic disorder characterized by hyperglycemia due to defective insulin secretion or its biological dysfunction. However, frequent subcutaneous injection of insulin often results in discomfort and local tissue infection. Herein, we demonstrate the successful fabrication of a mini-tablet system based on self-propelled micromotors with biocompatibility and biodegradability for oral colon administration of insulin. The insulin layer is first constructed onto the surface of a magnesium based micromotor via electrostatic interactions, followed by a tableting process. The resulting mini-tablets are then coated with esterified starch with colonic degradation capability, thus achieving controlled release of the embedded micromotors in the colon region. In the meantime, autonomous movement of the released micromotors with a speed up to 76.22 µm·s-1 further results in enhanced colonic uptake and absorption of insulin, realizing long-term control of blood glucose for more than 5 h. Our micromotor based mini-tablet system can not only broaden the biomedical applications of emerging self-propelled micromotors but also offer an appealing strategy for oral administration of biomacromolecular drugs represented by insulin.

Dietary Calcium Alleviates Fluorine-Induced Liver Injury in Rats by Mitochondrial Apoptosis Pathway.

Excessive fluoride (F) exposure can lead to liver damage; moreover, recent studies found that the addition of appropriate calcium (Ca) can alleviate the symptom of skeletal fluorosis. However, whether Ca can relieve F-induced liver damage through the mitochondrial apoptosis pathway has not been reported yet. Therefore, we assessed the liver morphology, serum transaminase content, liver oxidative stress-related enzymes, and apoptosis-related gene and protein expression in Sprague Dawley (SD) rats treated with 150 mg/L sodium fluoride (NaF) and different concentrations of calcium carbonate (CaCO3) for 120 days. Our results showed that NaF brought out pathological changes in liver morphology, serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels increased, total antioxidant capacity (T-AOC), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) content decreased, and malondialdehyde (MDA) content increased, suggesting that NaF caused hepatotoxicity and oxidative stress. In addition, the results of quantitative real-time PCR (qRT-PCR) and immunohistochemistry showed that NaF exposure upregulated the expression of Bcl-2-associated x protein (Bax), rho-related coiled-coil kinase 1 (ROCK1), cytochrome C (Cyto-C) mRNA and protein (P < 0.01), and downregulated B cell lymphoma 2 (Bcl-2) protein and mRNA (P < 0.01), indicating that excessive F exposure activated mitochondrial-mediated apoptosis in the liver. However, the addition of 1% CaCO3 to the diet significantly increased the expression of anti-apoptotic gene Bcl-2 (P < 0.01), inhibited the activation of the mitochondrial apoptosis pathway, and reduced mitochondrial damage. In summary, supplementing 1% CaCO3 in the diet can alleviate the NaF-induced liver cell damage through the mitochondrial apoptosis pathway.

Mitigation Effects of Selenium Nanoparticles on Depression-Like Behavior Induced by Fluoride in Mice via the JAK2-STAT3 Pathway.

Depression is a mental health problem with typically high levels of distress and dysfunction, and 150 mg/L fluoride (F) can induce depression-like behavior. The development of depression is correlated with neuronal atrophy, insufficient secretion of monoamine neurotransmitters, extreme deviations from the normal microglial activation status, and immune-inflammatory response. Studies found that Se supplementation was related to the improvement of depression. In this study, we applied selenium nanoparticles (SeNPs) for F-induced depression disease mitigation by regulating the histopathology, metabolic index, genes, and protein expression related to the JAK2-STAT3 signaling pathway in vivo. Results showed that F and 2 mg Se/kg BW/day SeNPs lowered the dopamine (DA) content (P < 0.05), altered the microglial morphology, ramification index as well as solidity, and triggered the microglial neuroinflammatory response by increasing the p-STAT3 nuclear translocation (P < 0.01). Furthermore, F reduced the cortical Se content and the number of surviving neurons (P < 0.05), increasing the protein expressions of p-JAK2/JAK2 and p-STAT3/STAT3 of the cortex (P < 0.01), accompanied by the depression-like behavior. Importantly, 1 mg Se/kg BW/day SeNPs alleviated the microglial ramification index as well as solidity changes and decreased the interleukin-1ß secretion induced by F by suppressing the p-STAT3 nuclear translocation (P < 0.01). Likewise, 1 mg Se/kg BW/day SeNPs restored the F-disturbed dopamine and noradrenaline secretion, increased the number of cortical surviving neurons, and reduced the vacuolation area, ultimately suppressing the occurrence of depression-like behavior through inhibiting the JAK2-STAT3 pathway activation. In conclusion, 1 mg Se/kg BW/day SeNPs have mitigation effects on the F-induced depression-like behavior. The mechanism of how SeNPs repair neural functions will benefit depression mitigation. This study also indicates that inhibiting the JAK/STAT pathway can be a promising novel treatment for depressive disorders.

Composition of bacterial communities associated with a plant-parasitic nematode Bursaphelenchus mucronatus.

Bursaphelenchus mucronatus is a plant-parasitic nematode widely existing in Eurasian pine forests. To analyze the diversity and role of bacteria associated with the nematode, culture-dependent and culture-independent methods were used to identify and characterize the composition of bacterial community. A total of 13 bacterial isolates were obtained from B. mucronatus by the culture-dependent method. Sixty-four species of bacteria were identified from two 16S rDNA clone libraries constructed from the nematodes of a Chinese and a Japanese population. These bacteria were clustered into four groups: Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Bacteroidetes. Comparison of the two libraries showed that the Chinese library had a higher diversity than that of the Japanese library, and the dominant group and species in each library were also different. In the Japanese library, Alphaproteobacteria group was obviously dominant (60.3%), and Rhizobium sp. was the most dominant species. Whereas in the Chinese library the proportion of each group was similar (from 19.4 to 23.6%), and Pedobacter sp. was a slightly dominant species. Moreover, 18 operational taxonomic units (OTUs) were obtained from each of the two libraries according to a 97% sequence similarity. Metabolic analysis showed that 61.5 and 38.5% of the bacterial isolates could have protease and lipase activities, respectively. But only one had cellulase activity. Testing of reproductive parameter showed that the wild-type nematodes (bacteria carried) could produce more progeny than the bacterium-free nematodes did. So, we speculated that bacteria could promote the propagation and development of the nematode B. mucronatus.

[Effects of staurosporine on the proliferation and apoptosis of prostate cancer PC-3 cells].

OBJECTIVE: To investigate the effects of staurosporine (ST) on the proliferation and apoptosis of prostate cancer PC-3 cells. METHODS: Prostate cancer PC-3 cells were treated in vitro with ST at 10(-8) mol/L. The expressions of cyclin A and cyclin D1 proteins in the cells were detected by Western blot, the effect of ST on the proliferation of the cells determined by MTT assay and plate colony formation, the apoptosis of the cells examined by flow cytometry, and their morphological changes observed under the light microscope. RESULTS: ST treatment markedly decreased the expressions of cyclin A and cyclin D1 in the PC-3 cells, and significantly inhibited the growth of the PC-3 cells (19.35%) at 48 h. (F = 31.06, P < 0.01). The colony formation rate of the PC-3 cells was (37.10 +/- 3.43) % in the ST group, significantly lower than (64.80 +/- 4.34) % in the control (chi2 = 14.59, P < 0.05) and (62.80 +/- 4.36) % in the DMSO group (chi2 = 12.50, P < 0.05), while the apoptosis rate of the cells was remarkably higher in the ST group ([19.6 +/- 2.20] %) than in the control ([5.33 +/- 1.40] %) and the DMSO group ([5.50 +/- 0.96] %) (F = 104.36, P < 0.01). Under the light microscope, the ST-treated cells were round with indistinct margins as compared with those of the other two groups. CONCLUSION: ST could significantly inhibit the proliferation and induce the apoptosis of PC-3 cells.

Genetic diversity of Venturia inaequalis isolates (Apple scab) in China and U.K. determined by SSR markers.

Apple scab caused by Venturia inaequalis is a serious disease of cultivated apple worldwide. In this study, we collected 132 V. inaequalis isolates from Shaanxi, Gansu, Xinjiang, and the U.K. and analyzed their genetic diversity by using 13 microsatellite markers. Cluster analysis based on population structure and genetic distances suggested high similarity among the four regions. Population differentiation values ranged from 0.044 to 0.155, indicating there is a high level of kinship among the four regions. All isolates could be divided into 5 lineages with a 0.76 similarity coefficient. Among the four regions, Shaanxi had only one lineage, Group II; Gansu had four lineages, Group I, Group II, Group IV, and Group V; Xinjiang had all five lineages, Group I, Group II, Group III, Group IV, and Group V; and the U.K. had three lineages, Group I, Group II and Group IV. High molecular variance was detected for populations in the four regions, with 91% of the variance occurring within the populations and 9% among the populations. Structure analysis there are three common ancestors of these four regions. The results of the present study shed light on the genetic diversity of V. inaequalis in Shaanxi, Gansu and Xinjiang, which will lead to the development of more effective management strategies and new resistant apple cultivars through molecular marker-assisted selection.

Effects of Different Doses of Calcium on the Mitochondrial Apoptotic Pathway and Rho/ROCK Signaling Pathway in the Bone of Fluorosis Rats.

For this study, we investigate more deeply the effect calcium (Ca) develops on the mechanism underlying fluoride-triggered osteocyte apoptosis. We detected the morphology of osteocytes by HE staining, mitochondrial microstructure by using the transmission electron microscope, and the biochemical indexes related to bone metabolism and the expression of apoptosis-related genes. These results showed that NaF brought out the reduced osteocytes and ruptured mitochondrial outer membrane, with a significantly increased StrACP activity by 10.414 IU/L at the 4th week (P < 0.05), markedly upregulating the mRNA expression of Bax, Cyto-C, Apaf-1, caspase-7, ROCK-1, BMP-2, and BGP (P < 0.01), as well as caspase-6 (P < 0.05), while downregulating Bcl-2 by 61.3% (P < 0.01). Through immunohistochemical analysis, we also found that NaF notably increased the protein expression of ROCK-1 (P < 0.05) and Cyto-C, BMP-2, and BGP (P < 0.01), suggesting that NaF triggered the activation of the mitochondrial apoptotic pathway and Rho/ROCK signaling pathway. Nevertheless, 1% Ca supplementation in diet notably enhanced the mRNA expression of Bcl-2 by 39.3% (P < 0.01), thus blocking the increment of the expression of mitochondrial apoptotic pathway-related genes and ROCK-1. Meanwhile, Ca could attenuate the StrACP activity by 10.741 IU/L at the 4th week (P < 0.05) and protect the integrity of the mitochondrial outer membrane. These findings strongly suggest that 1% Ca abated the mitochondrial apoptosis pathway by increasing the anti-apoptotic gene Bcl-2 expression, and effectively inhibited the hyper-activation of ROCK-1, dually protecting the structural integrity of the mitochondrial outer membrane and maintaining normal cellular metabolic function.

[Diversity of bacteria associated with pine wood nematode revealed by metagenome].

OBJECTIVE: The pine wood nematode (PWN), Bursaphlenchus xylophilus, which collaborates with its associated bacteria to form ecosystem and has interaction among them, is the pathogen of pine wilt disease. This study focused on revealing the bacterial diversity of ecosystem of pine wood nematode and its associated bacteria. METHODS: The metagenome of ecosystem of bacteria associated with the PWN was analyzed by 16S rRNA gene library and 454 sequencing. RESULTS: The results showed that 25 OTUs (Operational Taxonomic Units) were obtained from the library according to sequences similarity of 97%, which affiliated to Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria and Bacteroidetes. The dominant bacteria were belonged to Gammaproteobacteria, especially Stenotrophomonas maltophilia in this class dominated the library. In terms of dominant bacteria, the results revealed by metagenome were similar to that of 16S rRNA gene library. CONCLUSION: The diversity of bacteria associated with the PWN is high and these bacteria maybe have ecological role to the PWN.

Template-Free Electrochemical Formation of Silicon Nanotubes from Silica.

Silicon, with its elaborate microstructure, plays important roles in energy materials. In operando engineering of microstructure during extraction is an ideal protocol to develop advanced Si-based materials. A template-free electrochemical preparation of silicon nanotubes (Si-NT) is herein achieved by co-electrolysis of SiO2 and AgCl in molten NaCl-CaCl2 at 850 °C. The in situ electrodeposited Ag facilitates the generation of a liquid Ag-Si intermediate, triggering a liquid-solid mechanism to direct the growth of Si-NT. An automatic separation of Ag from Si then occurs in the following cooling process, resulting in Ag deposits on the Ni current collector and recycling of Ag. Such a facile and smart preparation of Si-NT from affordable silica guarantees an enhanced current efficiency of 74%, a decreased energy consumption of 12.1 kW h kgSi -1, and enhanced lithium-storage capability of the electrolytic Si-NT. An in situ coating of Ag over the Si-NT can also be fulfilled by simply introducing soluble AgCl in the melts. The present study provides a template-free preparation and an in situ surface modification of Si-NT.

Calcium relieves fluoride-induced bone damage through the PI3K/AKT pathway.

Bone is the main target of fluorosis, and it has been perfectly elaborated that a moderate dosage of calcium (Ca) can alleviate bone fluorosis. However, whether Ca can alleviate fluorosis through the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT) signaling pathway has not yet been reported. Hence, we evaluated the histopathological structure, the imbalance of the biochemical index of bone metabolism, and the expression levels of PI3K/AKT apoptosis signaling pathway-related genes in rats treated with sodium fluoride (NaF, F) and/or calcium carbonate (CaCO3) for 120 days. Our results suggest that 100 mg L-1 NaF induced histopathological injury as alkaline phosphatase (ALP) and tartrate-resistant acid phosphatase (StrACP) activity increased, with a decrease in the serum Ca levels (p < 0.05). Moreover, the results of qRT-PCR and western blotting showed that F increased the expression levels of transglutaminase 2 (TGM2), focal adhesion kinase (FAK), PI3K, AKT, forkhead box O1 (Foxo1), Bcl-2 interacting mediator of cell death (BIM), Bcl2-associated x protein (Bax) and Caspase 3 (p < 0.05, p < 0.01). It also decreased the expression of AnnexinA5 (Anxa5), 3'-phosphoinositide-dependent kinase 1 (PDK1) and B-cell lymphoma-2 (Bcl-2) (p < 0.05, p < 0.01), which finally activated the PI3K/AKT pathway. On the other hand, CaCO3 supplementation reversed the histopathological injury along with the levels of ALP, StrACP and serum Ca, alleviating the gene expression levels of PI3K/AKT pathway-related markers. Altogether, we can conclude that CaCO3 supplementation mitigated F-induced bone damage via the PI3K/AKT signaling pathway.

[Separation and identification of differential protein in rat's bone with fluorosis and calcium supplementation intervention].

In order to explore the mechanisms underlying the calcium alleviating fluorosis at protein level, we made an attempt to establish fluorosis and calcium supplementation rat models to isolate and identify bone differential proteins. The bone proteins of different groups were compared by two-dimensional electrophoresis (2-DE) and mass spectrometry (MALDI-TOF MS), and analyzed by gene ontology annotation, pathway enrichment and interaction networks. The 17 proteins were identified in the fluorosis group (F) and the fluorosis calcium supplement group (F+Ca), including type I collagen (Col1a1), actin (Actb), protein glutamine transferase 2 (Tgm2), compared with the control group (C). These differential proteins are enriched in 38 bone metabolic pathways such as focal adhesion, PI3K-Akt signaling pathway, and AMPK signaling pathway. And the functions of these proteins are mainly related to cytoskeleton, energy metabolism, substance transport, ion channel, and apoptosis. Therefore, it is speculated that calcium may alleviate the fluoride-induced bone damage by regulating the focal adhesion, PI3K-Akt, AMPK and other signaling pathway, but the specific mechanism needs further research.

Calcium Alleviates Fluoride-Induced Bone Damage by Inhibiting Endoplasmic Reticulum Stress and Mitochondrial Dysfunction.

Excessive fluoride mainly causes skeletal lesions. Recently, it has been reported that an appropriate level of calcium can alleviate fluorosis. However, the appropriate concentration and mechanism of calcium addition is unclear. Hence, we evaluated the histopathology and ultrastructure, DNA fragmentation, hormonal imbalances, biomechanical levels, and expression of apoptosis-related genes after treating the rats with 150 mg/L NaF and different concentrations of CaCO3. Our results suggested that NaF induced the histopathological and ultrastructural injury, with a concomitant increase in the DNA fragmentation (P < 0.05) and serum OC (17.5 ± 0.89 pmoL/L) at 120 days. In addition, the qRT-PCR and western blotting results indicated that NaF exposure upregulated the mRNA and protein expression of Bax, Calpain, Caspase 12, Caspase 9, Caspase 7, Caspase 3, CAD, PARP, and AIF while downregulated Bcl-2 (P < 0.01) and decreased the bone ultimate load by 27.1%, the ultimate stress by 10.1%, and the ultimate deformity by 23.3% at 120 days. However, 1% CaCO3 supplementation decreased the serum OC (14.7 ± 0.65 pmoL/L), bone F content (P < 0.01), and fracture and breakage of collagen fibers and changed the expression of endoplasmic reticulum pathway-related genes and proteins at 120 days. Further, 1% CaCO3 supplementation increased the bone ultimate load by 20.9%, the ultimate stress by 4.89%, and the ultimate deformity by 21.6%. In summary, we conclude that 1% CaCO3 supplementation alleviated fluoride-induced bone damage by inhibiting endoplasmic reticulum stress and mitochondrial dysfunction.

GSTO1 acts as a mediator in sodium fluoride-induced alterations of learning and memory related factors expressions in the hippocampus cell line.

The mechanism of GSTO1, as a high-risk factor for neurological damage, in sodium fluoride (NaF)-induced learning and memory impairment remained still unclear. Hence, in this study, we used the siRNA-GSTO1 HT22 model to explore the effect of NaF and siRNA-GSTO1 on the viability, and proliferation rate of HT22 cells, as well as the mRNA and protein expression levels of cyclic adenosine monophosphate (cAMP) response element binding protein (CREB), neural cell adhesion molecule (NCAM), stem cell factor (SCF) and brain-derived neurotrophic factor (BDNF). The results of MTT showed that 10-3, 10-4, and 10-5 moL/L sodium fluoride (NaF) exposure could significantly promote the proliferation of HT22 cells at 24 h, 36 h, and 48 h, respectively. In addition, our results showed that exposure to 10-3, 10-4, and 10-5 moL/l NaF increased GSTO1 mRNA and protein expression, but decreased CREB and BDNF expression levels in a dose and time-dependent manner. The mRNA and protein expressions of GSTO1, CREB and BDNF were significantly decreased in the siRNA-GSTO1 and NaF + siRNA-GSTO1 group (P < 0.05). We have shown that various NaF doses affected the learning and memory ability by down-regulation the expressions of CREB, BDNF, NCAM and SCF. In summary, we concluded that GSTO1 plays a mediator role in NaF-induced neurological damage.

Influence of Calcium Supplementation against Fluoride-Mediated Osteoblast Impairment in Vitro: Involvement of the Canonical Wnt/ß-Catenin Signaling Pathway.

Fluoride (F) is capable of promoting abnormal proliferation and differentiation in primary cultured mouse osteoblasts (OB cells), although the underlying mechanism responsible remains rare. This study aimed to explore the roles of wingless and INT-1 (Wnt) signaling pathways and screen appropriate doses of calcium (Ca2+) to alleviate the sodium fluoride (NaF)-induced OB cell toxicity. For this, we evaluated the effect of dickkopf-related protein 1 (DKK1) and Ca2+ on mRNA levels of wingless/integrated 3a (Wnt3a), low-density lipoprotein receptor-related protein 5 (LRP5), dishevelled 1 (Dv1), glycogen synthase kinase 3ß (GSK3ß), ß-catenin, lymphoid enhancer binding factor 1 (LEF1), and cellular myelocytomatosis oncogene (cMYC), as well as Ccnd1 (Cyclin D1) in OB cells challenged with 10-6 mol/L NaF for 24 h. The demonstrated data showed that F significantly increased the OB cell proliferation rate. Ectogenic 0.5 mg/L DKK1 significantly inhibited the proliferation of OB cells induced by F. The mRNA expression levels of Wnt3a, LRP5, Dv1, LEF1, ß-catenin, cMYC, and Ccnd1 were significantly increased in the F group, while significantly decreased in the 10-6 mol/L NaF + 0.5 mg/L DKK1 (FY) group. The mRNA expression levels of Wnt3a, LRP5, ß-catenin, and cMYC were significantly decreased in the 10-6 mol/L NaF + 2 mmol/L CaCl2 (F+CaII) group. The protein expression levels of Wnt3a, Cyclin D1, cMYC, and ß-catenin were significantly increased in the F group, whereas they were decreased in the F+CaII group. However, the mRNA and protein expression levels of GSK3ß were significantly decreased in the F group while significantly increased in the F+CaII group. In summary, F activated the canonical Wnt/ß-catenin pathway and changed the related gene expression and ß-catenin protein location in OB cells, promoting cell proliferation. Ca2+ supplementation (2 mmol/L) reversed the expression levels of genes and proteins related to the canonical Wnt/ß-catenin pathway.

Population Variation of Apple Scab (Venturia inaequalis) Isolates from Asia and Europe.

Apple scab, caused by Venturia inaequalis, is one of the most of damaging diseases worldwide on apple and currently is managed mainly by scheduled applications of fungicides. Understanding pathogen population structure is important for breeding and deployment of resistant cultivars. Isolates of V. inaequalis were sampled from a number of cultivars in China, India, and the United Kingdom to estimate differences in pathogen populations. Amplified fragment length polymorphism (AFLP) markers were used to genotype isolates, mostly from China and the United Kingdom. The AFLP data indicated that, overall, there were significant differences in V. inaequalis populations from China and the United Kingdom. Within China, there was no significant differentiation associated with their geographical or cultivar origins. In contrast, populations from four cultivars in two U.K. orchards (monoculture of Gala and a mixture orchard of Bramley, Cox, and Worcester) differed significantly. Furthermore, populations from Gala and Worcester were more homogenous than expected but those from Cox were more diverse than expected. In total, 80 isolates were selected randomly from three countries for virulence testing: 20 from the United Kingdom (10 from Gala and 10 from Cox), 30 from China (10 from Gala, 10 from Fuji, and 10 from Qingquan), and 30 from India (10 from Gala, 10 from Golden Delicious, and 10 from Black Ben Davis); of these 80 isolates, 41, 47, and 59 were inoculated against each of these cultivars in the United Kingdom, India, and China, respectively. The two local cultivars from India (Black Ben Davis) and the United Kingdom (Cox) were more resistant against non-indigenous isolates, particularly those from China, than they were against indigenous isolates; the Chinese local cultivar (Qingguan) showed a higher general level of resistance against isolates regardless of their origin.