The Effectiveness of Exercise-based Rehabilitation in People with Hand Osteoarthritis: A Systematic Review with Meta-analysis.

OBJECTIVE: To investigate the effectiveness of exercise-based rehabilitation programs compared with non-exercise intervention or no intervention for people with hand osteoarthritis (OA). DESIGN: Intervention systematic review with meta-analysis. LITERATURE SEARCH: We searched five databases on 23/07/2023. STUDY SELECTION CRITERIA: We included randomized controlled trials that compared the effectiveness of rehabilitation programs that included an exercise component, with non-exercise intervention or no intervention for people with hand OA. DATA SYNTHESIS: Standardized mean differences (SMD) were pooled using a random effects model. The risk of bias was assessed using the Cochrane Risk of Bias 2.0 tool. The certainty of the evidence was evaluated using the GRADE approach. RESULTS: Fourteen trials were included in the meta-analysis (n = 1341 participants). In the immediate-term (<24 weeks) there was low-certainty evidence of an effect of exercise-based rehabilitation on improving pain (13 trials; SMD -0.65, 95%CI: -1.06, -0.25), function (11 trials; SMD -0.35, 95%CI: -0.54, -0.15), and grip strength (14 trials; SMD 0.21, 95%CI: 0.03, 0.38). There was moderate-certainty evidence of an effect on reducing stiffness (7 trials; SMD -0.33, 95%CI: -0.51, -0.16). There was low-certainty evidence of no effect on improving pinch strength and quality of life. For the long-term ⩾24 weeks), there was low-certainty evidence that exercise-based rehabilitation had no additional effect on improving pain, function, and stiffness. CONCLUSION: Exercise-based rehabilitation improved pain, function, stiffness, and grip strength in people with hand OA in the immediate-term; the benefits were not maintained in the long-term.

Most of the pelvic floor muscle functions in women differ in different body positions, yet others remain similar: systematic review with meta-analysis.

Objectives: This systematic literature review and meta-analysis aimed to determine the effect of body position on the measurement of pelvic floor muscle (PFM) contractility and to analyze the influential factors. Data sources: Five databases (PubMed, Web of Science, EMBASE, Cochrane Library and Scopus) were searched for relevant studies published up to 12nd October 2023. Study selection or eligibility criteria: Included cross-sectional studies had to involve the assessment of pelvic floor muscle function in at least two positions. Study appraisal and synthesis methods: We calculated standardized mean difference (SMD) with 95% confidence intervals (CI) to ascertain the potential effect of body position on outcomes. Results: In total, we included 11 cross-sectional studies to ascertain the potential effect of body position on outcomes. There was no statistical difference in the results of maximum voluntary contraction (MVC) of the pelvic floor muscles when assessed in between supine and standing positions (SMD -0.22; 95% CI -0.72 to 0.28; p = 0.38). The results of the meta-analysis showed significantly larger values of resting voluntary contractions (RVC) measured in the standing position compared to the supine position (SMD -1.76; 95% CI -2.55 to -0.97; p < 0.001). Moreover, pelvic floor muscle movement during pelvic floor muscle contraction in the standing position was significantly better than that measured in the supine position (SMD -0.47; 95% CI -0.73 to 0.20; P < 0.001). Conclusion: The results of this study showed that the RVC and PFM movement varied with the position of the assessment. In contrast, MVC values are independent of the assessment position and can be selected according to clinical needs. Systematic review registration: PROSPERO, identifier CRD42022363734, https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42022363734.

Can we trust computers to assess the cognition of stroke patients? A systematic review.

Purpose: To summarize the classification of computerized cognitive assessment (CCA) tools for assessing stroke patients, to clarify their benefits and limitations, and to reveal strategies for future studies on CCA tools. Methods: A literature review was performed using PubMed, Embase, Scopus, JAMA Network, Cochrane Library and PsycINFO databases from January 1st, 2010, to August 1st, 2022. Two authors independently screened the literature following the same criteria, evaluated the study quality, and collected data from the articles. Results: A total of 8,697 papers were acquired from the six databases. A total of 74 potentially eligible articles were selected for review. Of these, 29 articles were not relevant to this research, 3 were reviews, 2 were not written in English, and 1 was on an ongoing trial. By screening the references of the reviews, 3 additional articles were included in this study. Thus, a total of 42 articles met the criteria for the review. In terms of the CCA tools analyzed in these studies, they included five types: virtual reality (VR)-based, robot-based, telephone-based, smartphone-based, and computer-based cognitive assessments. Patients' stages of the disease ranged from the subacute phase and rehabilitation phase to the community phase. A total of 27 studies supported the effectiveness of CCA tools, while 22 out of 42 articles mentioned their benefits and 32 revealed areas for future improvement of CCA tools. Conclusions: Although the use of CCA tools for assessing the cognition of post-stroke patients is becoming popular, there are still some limitations and challenges of using such tools in stroke survivors. More evidence is thus needed to verify the value and specific role of these tools in assessing the cognitive impairment of stroke patients.

HBsAg (-)/HBsAb (-)/HBeAg (-)/HBeAb (+)/HBcAb (+) predicts a high risk of hepatitis B reactivation in patients with B-cell lymphoma receiving rituximab based immunochemotherapy.

Patterns of hepatitis B virus reactivation (HBV-R) in HBsAg (-)/HBcAb (+) patients with B-cell non-Hodgkin lymphoma (NHL) receiving rituximab based immunochemotherapy have not been well described. The retrospective study included 222 HBsAg (-)/HBcAb (+) NHL patients as training cohort and 127 cases as validation cohort. The incidence of HBV-R in HBsAg (-)/HBcAb (+) B-cell NHL patients was 6.3% (14/222), of which that in HBsAg (-)/HBsAb (-)/HBeAg (-)/HBeAb (+)/HBcAb (+) population was 23.7% (9/38). Multivariate analysis showed that HBsAg (-)/HBsAb (-)/HBeAg (-)/HBeAb (+)/HBcAb (+) correlated with a high risk of HBV-R in B-cell lymphoma patients (training phase hazard ratio [HR], 10.123; 95% confidence interval [CI], 3.389-30.239; p < 0.001; validation phase HR, 18.619; 95% CI, 1.684-205.906; p = 0.017; combined HR, 12.264; 95% CI, 4.529-33.207; p < 0.001). In the training cohort, the mortality rate of HBsAg (-)/HBcAb (+) B-cell NHL caused by HBV-R was 14.3% (2/14) while that for HBV reactivated HBsAg (-)/HBsAb (-)/HBeAg (-)/HBeAb (+)/HBcAb (+) population was up to 44.4% (4/9). As a high incidence of HBV-R and high mortality after HBV-R was found in HBsAg (-)/HBsAb (-)/HBcAb (+)/HBeAg (-)/HBeAb (+) patients with B-cell NHL receiving rituximab based immunochemotherapy, prophylactic antiviral therapy is recommended for these patients.

Pyrrolidinium-Based Hyperbranched Anion Exchange Membranes with Controllable Microphase Separated Morphology for Alkaline Fuel Cells.

It is well acknowledged that the microphase-separated morphology of anion exchange membranes (AEMs) is of vital importance for membrane properties utilized in alkaline fuel cells. Herein, a rigid macromolecule poly(methyldiallylamine) (PMDA) is incorporated to regulate the microphase morphology of hyperbranched AEMs. As expected, the hyperbranched poly(vinylbenzyl chloride) (HB-PVBC) is guided to distribute along PMDA chains, and longer PMDA cha leads to a more distinct microphase morphology with interconnected ionic channels. Consequently, high chloride conductivity of 10.49 mS cm-1 at 30 °C and suppressed water swelling ratio lower than 30% at 80 °C are obtained. Furthermore, the ß-H of pyrrolidinium cations in the non-antiperiplanar position increases the energy barrier of ß-H elimination, leading to conformationally disfavored Hofmann elimination and increased alkaline stability. This strategy is anticipated to provide a feasible way for preparing hyperbranched AEMs with clear microphase morphology and good overall properties for alkaline fuel cells.

Stu-miR827-Targeted StWRKY48 Transcription Factor Negatively Regulates Drought Tolerance of Potato by Increasing Leaf Stomatal Density.

Stomata are specialized portals in plant leaves to modulate water loss from plants to the atmosphere by control of the transpiration, thereby determining the water-use efficiency and drought resistance of plants. Despite that the stomata developmental progression is well-understood at the molecular level, the experimental evidence that miRNA regulates stomata development is still lacking, and the underlying mechanism remains elusive. This study demonstrates the involvement of stu-miR827 in regulating the drought tolerance of potato due to its control over the leaf stomatal density. The expression analysis showed that stu-miR827 was obviously repressed by drought stresses and then rapidly increased after rewatering. Suppressing the expression of stu-miR827 transgenic potato lines showed an increase in stomatal density, correlating with a weaker drought resistance compared with wildtype potato lines. In addition, StWRKY48 was identified as the target gene of stu-miR827, and the expression of StWRKY48 was obviously induced by drought stresses and was greatly upregulated in stu-miR827 knockdown transgenic potato lines, suggesting its involvement in the drought stress response. Importantly, the expression of genes associated with stomata development, such as SDD (stomatal density and distribution) and TMM (too many mouths), was seriously suppressed in transgenic lines. Altogether, these observations demonstrated that suppression of stu-miR827 might lead to overexpression of StWRKY48, which may contribute to negatively regulating the drought adaptation of potato by increasing the stomatal density. The results may facilitate functional studies of miRNAs in the process of drought tolerance in plants.

Myotonic dystrophy type 1 presenting with dyspnea: A case report.

BACKGROUND: Myotonic dystrophy type 1 (DM1) is a genetic neuromuscular disease involving multiple systems, especially the cardiopulmonary system. The clinical phenotype of DM1 patients is highly variable, which limits early diagnosis and treatment. In the present study, we reported a 35-year-old female DM1 patient with dyspnea as the primary onset clinical manifestation, analyzed her family's medical history, and reviewed related literature. CASE SUMMARY: A 35-year-old woman was admitted to the hospital with dyspnea of 1 mo duration, and sleep apnea for 3 d. Her respiratory pattern and effort were normal, but limb muscle tension was low. Investigation into the patient's medical history revealed that she might have hereditary neuromuscular disease. Electromyography showed that her myotonia potentials were visible in the resting state of the examined muscles, with decreased motor unit potential time limit and amplitude. Genetic testing for DM1 revealed that the cytosine-thymine-guanine (CTG) repeat number of the DMPK gene exceeded 50, while cytosine-CTG expansion in intron 1 of ZNF9 gene was < 30 repeats. The patient was diagnosed with DM1. CONCLUSION: DM1 is a genetic neuromuscular disease involving multiple systems, and the clinical phenotype in DM1 is extremely variable. Some patients with DM1 may be presented at the respiratory department because of dyspnea, which should be cautioned by the pulmonologists. There may be no obvious or specific symptoms in the early stage of disease, and clinicians should improve their understanding of DM1 and make an early diagnosis, which will improve patients' quality of life.

Sexual Dysfunction After Traumatic Injury Can Be Corrected by Transcranial Magnetic Stimulation: A Case Report.

Neurogenic sexual dysfunction (NSD) is a common problem in patients after spinal and pelvic trauma. New treatment is needed beyond medicine or psychological therapies. A 24-year-old man who fell from a six-floor building suffered from subsequent NSD. Repetitive transcranial magnetic stimulation (rTMS) was the only method used to treat his NSD caused by multiple spinal and pelvic injuries. The therapy lasted for 3 courses. Motor and sensory conduction, as well as sexual function, were evaluated before and after the rTMS intervention. Improvements on patient's nerve conduction and sexual activity were confirmed at a 1-year follow-up. Our findings indicate that rTMS delivered a novel, positive and low-cost modality treatment to the patient with NSD. Clinical efficacy and potential mechanisms by which rTMS regulate NSD need to be investigated by further clinical trials.

Brucine restores sodium nitroprusside-induced chondrocyte dysfunction by suppressing the GSK-3ß/ß-catenin pathway.

The dysfunction of chondrocytes is thought to play a role in the initiation and progression of osteoarthritis (OA). Brucine possesses wide pharmacological activities. But the protective mechanism of the brucine on chondrocytes remains unclear. This study is aimed to determine the therapeutic effects of brucine on the mouse chondrocyte OA model by sodium nitroprusside (SNP). The primary chondrocytes were obtained from the knee articular cartilage of a healthy suckling mouse donor. The cultured chondrocytes were divided into the control group, SNP group, brucine group, brucine-SNP group, brucine-SNP-GSK-3ß antagonist group (brucine-SNP- group), and brucine-SNP-GSK-3ß agonist group (brucine-SNP-GSK-3ß+ group). After 24 h, the chondrocytes from different treated groups were collected to detect chondrocyte proliferation and ultrastructure, regulation factors, apoptosis, oxidative stress, and GSK-3ß/ß-catenin pathway. Compared to the SNP group, chondrocyte proliferation, and regulation factors were promoted, and chondrocyte apoptosis, oxidative stress, and the GSK-3ß/ß-catenin pathway were inhibited by brucine. It indicates that the adverse effect of SNP is reversed by the brucine on the chondrocyte. Compared to the brucine-SNP group, the effect of brucine on the chondrocyte proliferation, regulation factothe apoptosis, and oxidative stress were promoted by the GSK-3ß antagonist. It indicates that the chondrocyte is protected agairucine through buying the GSK-3ß/ß-catenin pathway.

When medical trainees encountering a performance difficulty: evidence from pupillary responses.

BACKGROUND: Medical trainees are required to learn many procedures following instructions to improve their skills. This study aims to investigate the pupillary response of trainees when they encounter moment of performance difficulty (MPD) during skill learning. Detecting the moment of performance difficulty is essential for educators to assist trainees when they need it. METHODS: Eye motions were recorded while trainees practiced the thoracostomy procedure in the simulation model. To make pupillary data comparable among trainees, we proposed the adjusted pupil size (APS) normalizing pupil dilation for each trainee in their entire procedure. APS variables including APS, maxAPS, minAPS, meanAPS, medianAPS, and max interval indices were compared between easy and difficult subtasks; the APSs were compared among the three different performance situations, the moment of normal performance (MNP), MPD, and moment of seeking help (MSH). RESULTS: The mixed ANOVA revealed that the adjusted pupil size variables, such as the maxAPS, the minAPS, the meanAPS, and the medianAPS, had significant differences between performance situations. Compared to MPD and MNP, pupil size was reduced during MSH. Trainees displayed a smaller accumulative frequency of APS during difficult subtask when compared to easy subtasks. CONCLUSIONS: Results from this project suggest that pupil responses can be a good behavioral indicator. This study is a part of our research aiming to create an artificial intelligent system for medical trainees with automatic detection of their performance difficulty and delivering instructional messages using augmented reality technology.

Alantolactone: A sesquiterpene lactone with diverse pharmacological effects.

Alantolactone (Ala) is a sesquiterpene lactone that can be isolated from many herbal plants belonging to Asteraceae. Besides the antimicrobial activities against bacteria, fungi and viruses, Ala has also demonstrated significant anti-inflammatory effects in various models by inhibiting NF-κB and MAPKs to decrease the pro-inflammatory cytokines such as IL-1ß, IL-6 and TNF-α. The antitumor effects of Ala have been demonstrated in vitro and in vivo via inducing intrinsic apoptosis, oxidative stress, ER stress, cell cycle arrest and inhibiting autophagy and STAT3 phosphorylation, which are also involved in its combination or synergy with other antitumor drugs. Ala also has neuroprotective activity through attenuating oxidative stress and inflammation, besides its modulation of glucose and lipid metabolism. This review summarizes the recent advances of the pharmacological effects of Ala, including anti-inflammatory, antitumor, antimicrobial, neuroprotective activities, as well as the underlying mechanisms. Ala might be employed as a potential lead to develop drugs for multiple diseases.

Personal Protective Equipment in COVID-19: Impacts on Health Performance, Work-Related Injuries, and Measures for Prevention.

OBJECTIVE: To assess impact of personal protective equipment (PPE) on healthcare providers (HCPs) in caring for COVID-19 patients. METHODS: A cross-sectional survey was conducted over 50 hospitals in China. Descriptive analyses and Chi-square tests were performed on the collected data. RESULTS: All 104 frontline HCPs report negative impacts of PPE on their clinical performance, 97% of them experienced discomfort and injuries caused by wearing PPE for long hours. Frontline HCPs provided suggestions to alleviate the negative impacts and to enhance communication between healthcare staff and patients. Two hundred eighty two non-frontline HCPs also revealed similar problems; however, we recorded a few discrepancies between answers given by frontline and non-frontline HCPs. CONCLUSIONS: Wearing PPE for long hours degrades health performance. Measures were suggested to improve the design of PPE for protecting HCPs and enhancing their services to COVID patients.

Dentin Sialophosphoprotein Deletion Leads to Femoral Head Cartilage Attenuation and Subchondral Bone Ill-mineralization.

Dentin sialophosphoprotein (DSPP), which expresses and synthesizes in odontoblasts of dental pulp, is a critical protein for normal teeth mineralization. Originally, DSPP was identified as a dentin-specific protein. In 2010, DSPP was also found in femoral head cartilage, and it is still unclear what roles DSPP play in femoral head cartilage formation, growth, and maintenance. To reveal biological functions of DSPP in the femoral head cartilage, we examined Dspp null mice compared with wild-type (WT) mice to observe DSPP expression as well as localization in WT mice and to uncover differences of femoral head cartilage, bone morphology, and structure between these two kinds of mice. Expression data demonstrated that DSPP had heterogeneous fragments, expressed in each layer of femoral head cartilage and subchondral bone of WT mice. Dspp null mice exhibited a significant reduction in the thickness of femoral head cartilage, with decreases in the amount of proliferating cartilage cells and increases in apoptotic cells. In addition, the subchondral bone mineralization decreased, and the expressions of vessel markers (vascular endothelial growth factor [VEGF] and CD31), osteoblast markers (Osterix and dentin matrix protein 1 [DMP1]), osteocyte marker (sclerostin [SOST]), and osteoclast marker (tartrate-resistant acid phosphatase [TRAP]) were remarkably altered. These indicate that DSPP deletion can affect the proliferation of cartilage cells in the femoral head cartilage and endochondral ossification in subchondral bone. Our data clearly demonstrate that DSPP plays essential roles in the femoral head cartilage growth and maintenance and subchondral biomineralization.

Mechanisms of Transcranial Magnetic Stimulation Treating on Post-stroke Depression.

Post-stroke depression (PSD) is a neuropsychiatric affective disorder that can develop after stroke. Patients with PSD show poorer functional and recovery outcomes than patients with stroke who do not suffer from depression. The risk of suicide is also higher in patients with PSD. PSD appears to be associated with complex pathophysiological mechanisms involving both psychological and psychiatric problems that are associated with functional deficits and neurochemical changes secondary to brain damage. Transcranial magnetic stimulation (TMS) is a non-invasive way to investigate cortical excitability via magnetic stimulation of the brain. TMS is currently a valuable tool that can help us understand the pathophysiology of PSD. Although repetitive TMS (rTMS) is an effective treatment for patients with PSD, its mechanism of action remains unknown. Here, we review the known mechanisms underlying rTMS as a tool for better understanding PSD pathophysiology. It should be helpful when considering using rTMS as a therapeutic strategy for PSD.

Cultural adaptation of an evidence-based lifestyle intervention for diabetes prevention in Chinese women at risk for diabetes: results of a randomized trial.

Background: The prevalence of both obesity and type 2 diabetes has been on the rise in China. This randomized controlled trial was conducted to test the feasibility and effectiveness of an evidence-based diabetes prevention program in Yuci, Shanxi Province, China from 2012 to 2014. Methods: Women with pre-diabetes, ages 25-65 y, were assigned randomly to a comparison (n=75) or 6-mo lifestyle intervention condition (n=109). Weight, fasting glucose, hemoglobin A1c and self-reported diet and physical activity were measured at baseline, 6 mo and 12 mo. Results: All measures except fasting glucose improved favorably in both comparison and intervention participants at the 6- and 12-mo follow-ups. Participants in the intervention group lost more weight (-0.91 kg, p<0.05) and had a lower body mass index (-0.39 kg/m2, p<0.05) than the comparison group at follow-up. A total of 31.6% (31/98) and 16.2% (11/68) of the participants in the intervention and comparison groups, respectively, achieved the weight loss goal of 5% at follow-up. There was no significant group difference in outcome measures at the 12-mo follow-up. Participants in the intervention group also showed favorable changes in self-reported diet and physical activity measures. Conclusions: A lifestyle intervention to prevent diabetes in at-risk women in community health centers in China is feasible and acceptable but effect sizes were small.

A novel Mn(2+)-doped core/shell quantum dot-based intracellular probe for fluoride anions sensing in MC3T3-E1 osteoblastic cells.

In this paper, 3-aminobenzeneboronic acid functionalized Mn(2+)-doped ZnTe/ZnSe quantum dots (APBA-dQDs) were prepared. The APBA functional groups had strong binding ability with F(-), resulting in the quenchment of dQDs photoluminescence (PL). Under the optimal condition, the fluorescence intensity of APBA-dQDs was related linearly to the concentration of F(-) in the range of 0.25-1.5µmol/L with a detection limit of 0.1µmol/L. The selectivity of fluorescence quenching of APBA-dQDs for F(-) was enhanced. Moreover, the proposed methodology for the sensing of F(-) at EM 560nm in MC3T3-E1 osteoblastic cells was demonstrated and got a satisfactory results. The results indicate that the APBA-dQDs are promising candidates for intracellular in MC3T3-E1 osteoblastic cells. To the best of our knowledge, it was the first report of F(-) sensing by using the quenched fluorescence of APBA-dQDs in non-cancerous cells.

Mechanisms of Intracellular Calcium Homeostasis in MC3T3-E1 Cells and Bone Tissues of Sprague-Dawley Rats Exposed to Fluoride.

Calcium homeostasis of osteoblasts (OBs) has an important role in the physiology and pathology of bone tissue. In order to study the mechanisms of intracellular calcium homeostasis, MC3T3-E1 cells and Sprague-Dawley rats were treated with different concentrations of fluoride. Then, we examined intracellular-free calcium ion ([Ca(2+)]i) in MC3T3-E1 cells as well as mRNA and protein levels of Cav1.2, the main subunit of L-type voltage-dependent calcium channels (VDCCs), Na(+)/Ca(2+) exchange carriers (NCS), and plasma membrane Ca(2+)-ATPase (PMCA), inositol 1,4,5-trisphosphate receptor (IP3R) channels, sarco/endoplasmic reticulum calcium ATPase 2b (SERCA2b)/ATP2A2 in vitro, and rat bone tissues in vivo. Our results showed that [Ca(2+)]i of fluoride-treated OBs increased in a concentration-dependent manner with an increase in the concentration of fluoride. We also found that the low dose of fluoride led to high expression levels of Cav1.2, NCS-1, and PMCA and low expression levels of IP3R and SERCA2b/ATP2A2, while the high dose of fluoride induced an increase in SERCA2b/ATP2A2 levels and decrease in Cav1.2, PMCA, NCS-1, and IP3R levels. These results demonstrate that calcium channels and calcium pumps of plasma and endoplasmic reticulum (ER) membranes keep intracellular calcium homeostasis by regulating Cav1.2, NCS-1, PMCA, IP3R, and SERCA2b/ATP2A2 expression.

Fluoride Affects Calcium Homeostasis by Regulating Parathyroid Hormone, PTH-Related Peptide, and Calcium-Sensing Receptor Expression.

Parathyroid hormone (PTH), PTH-related peptide (PTHrP), and calcium-sensing receptor (CaSR) play important roles in maintaining calcium homeostasis. Here, we study the effect of fluoride on expression of PTH, PTHrP, and CaSR both in vitro and in vivo. MC3T3-E1 cells and Sprague-Dawley rats were treated with different concentrations of fluoride. Then, the free calcium ion concentration in cell culture supernatant and serum were measured by biochemical analyzer. The expression of PTH, PTHrP, and CaSR was analyzed by qRT-PCR and Western blot. We found that the low dose of fluoride increased ionized calcium (i[Ca(2+)]) and the high dose of fluoride decreased i[Ca(2+)] in cell culture supernatant. The low dose of fluoride inhibited the PTH and PTHrP expression in MC3T3-E1 cells. The high dose of fluoride improved the PTHrP expression in MC3T3-E1 cells. Interestingly, we found that NaF decreased serum i[Ca(2+)] in rats. Fluoride increased CaSR expression at both messenger RNA (mRNA) and protein levels in MC3T3-E1 cells and rats. The expression of PTHrP protein was inhibited by fluoride in rats fed regular diet and was increased by fluoride in rats fed low-calcium diet. Fluoride also increased the expression of PTH, NF-kappaB ligand (RANKL), and osteoprotegerin (OPG) in rats. The ratio of RANKL/OPG in rats fed low-calcium food in presence or absence of fluoride was significantly increased. These results indicated that fluoride might be able to affect calcium homeostasis by regulating PTH, PTHrP, and CaSR.

Fluoride affects calcium homeostasis and osteogenic transcription factor expressions through L-type calcium channels in osteoblast cell line.

Osteoblast L-type voltage-dependent calcium channels (VDCC) play important roles in maintaining intracellular homeostasis and influencing multiple cellular processes. In particular, they contribute to the activities and functions of osteoblasts (OBs). In order to study how L-type VDCC modulate calcium ion (Ca(2+)) homeostasis and the expression of osteogenic transcription factors in OBs exposed to fluoride, MC3T3-E1 cells were exposed to a gradient of concentrations of fluoride (0, 2.0, 5.0, 10.0 mg/L) in combination with 10 µM nifedipine, a specific inhibitor of VDCC, for 48 h. We examined messenger RNA (mRNA) and protein levels of Cav1.2, the main subunit of VDCC, and c-fos, c-jun, runt-related transcription factor 2 (Runx2), osterix (OSX), and intracellular free Ca(2+) ([Ca(2+)]i) concentrations in MC3T3-E1 cells. Our results showed that [Ca(2+)]i levels increased in a dose-dependent manner with increase in concentration of fluoride. Meantime, results indicated that lower concentrations of fluoride (less than 5 mg/L, especially 2 mg/L) can lead to high expression of Cav1.2 and enhance osteogenic function, while high concentration of fluoride (10 mg/L) can induce decreased Cav1.2 and osteogenic transcriptional factors in MC3T3E1 cells exposed to fluoride. However, the levels of [Ca(2+)]i, Cav1.2, c-fos, c-jun, Runx2, and OSX induced by fluoride were significantly altered and even reversed in the presence of nifedipine. These results demonstrate that L-type calcium channels play a crucial role in Ca(2+) homeostasis and they affect the expression of osteogenic transcription factors in fluoride-treated osteoblasts.

Expression of core-binding factor α1 and osteocalcin in fluoride-treated fibroblasts and osteoblasts.

To study the effects and importance of fluoride on FBs in the development of extraperiosteal calcification and the ossification of skeletal fluorosis, the presence of the osteogenic phenotype, which is indicated by the expression of core-binding factor α1 (Cbfa1) and osteocalcin (OCN), in an FB cell line (L929) and in osteoblasts (OBs) exposed to fluoride was determined. Fibroblasts and osteoblasts were exposed to different concentrations of fluoride (0, 0.0001, 0.001, 0.1, 1.0, 10.0 and 20.0 mg/L F(-)). By using RT-PCR and ELISA, the mRNA levels of Cbfa1 and OCN were measured at 48 h, and the protein levels of Cbfa1 and OCN were measured at 2, 4, 24, 48 and 72 h. The data demonstrated the following: (1) The Cbfa1 protein level in fluoride-treated fibroblasts clearly increased at 48 h in the groups treated with 0.0001, 0.001, 0.1, 1.0 and 20.0 mg/L F(-). The Cbfa1 protein level of the group treated with 10 mg/L F(-) at 72 h was higher than that of the control group. The level of Cbfa1 mRNA in the fibroblasts was much higher at 48 h in the group treated with 10.0 mg/L F(-) than in the control group. (2) The OCN protein level in fluoride-treated fibroblasts was significantly higher than that of the control group in the 0.0001, 0.1, 1.0, 10.0 and 20.0 mg/L F(-) groups at 2 h, and in the 0.001 and 0.1 F(-) groups at 4 h. A slightly higher level of OCN mRNA in fluoride-treated fibroblasts was also found in the 1.0 and 20.0 mg/L F(-) groups compared to the control group. (3) The expressions of Cbfa1 and OCN in osteoblasts treated with the same experimental conditions as the fibroblasts were up-regulated by fluoride following the same trend as in the fibroblasts. Our results showed an increase in the expression of Cbfa1 and OCN in fibroblasts and osteoblasts exposed to fluoride and suggested that the osteogenic function of fibroblasts induced by fluoride could play an important role in the development of extraperiosteal ossification during skeletal fluorosis.