m6A demethylation of cytidine deaminase APOBEC3B mRNA orchestrates arsenic-induced mutagenesis.

The cytidine deaminase APOBEC3B (A3B) is an endogenous inducer of somatic mutations and causes chromosomal instability by converting cytosine to uracil in single-stranded DNA. Therefore, identification of factors and mechanisms that mediate A3B expression will be helpful for developing therapeutic approaches to decrease DNA mutagenesis. Arsenic (As) is one well-known mutagen and carcinogen, but the mechanisms by which it induces mutations have not been fully elucidated. Herein, we show that A3B is upregulated and required for As-induced DNA damage and mutagenesis. We found that As treatment causes a decrease of N6-methyladenosine (m6A) modification near the stop codon of A3B, consequently increasing the stability of A3B mRNA. We further reveal that the demethylase FTO is responsible for As-reduced m6A modification of A3B, leading to increased A3B expression and DNA mutation rates in a manner dependent on the m6A reader YTHDF2. Our in vivo data also confirm that A3B is a downstream target of FTO in As-exposed lung tissues. In addition, FTO protein is highly expressed and positively correlates with the protein levels of A3B in tumor samples from human non-small cell lung cancer patients. These findings indicate a previously unrecognized role of A3B in As-triggered somatic mutation and might open new avenues to reduce DNA mutagenesis by targeting the FTO/m6A axis.

Stage-specific roles of microbial dysbiosis and metabolic disorders in rheumatoid arthritis.

OBJECTIVE: Rheumatoid arthritis (RA) is a progressive disease including four stages, where gut microbiome is associated with pathogenesis. We aimed to investigate stage-specific roles of microbial dysbiosis and metabolic disorders in RA. METHODS: We investigated stage-based profiles of faecal metagenome and plasma metabolome of 76 individuals with RA grouped into four stages (stages I-IV) according to 2010 RA classification criteria, 19 individuals with osteroarthritis and 27 healthy individuals. To verify bacterial invasion of joint synovial fluid, 16S rRNA gene sequencing, bacterial isolation and scanning electron microscopy were conducted on another validation cohort of 271 patients from four RA stages. RESULTS: First, depletion of Bacteroides uniformis and Bacteroides plebeius weakened glycosaminoglycan metabolism (p<0.001), continuously hurting articular cartilage across four stages. Second, elevation of Escherichia coli enhanced arginine succinyltransferase pathway in the stage II and stage III (p<0.001), which was correlated with the increase of the rheumatoid factor (p=1.35×10-3) and could induce bone loss. Third, abnormally high levels of methoxyacetic acid (p=1.28×10-8) and cysteine-S-sulfate (p=4.66×10-12) inhibited osteoblasts in the stage II and enhanced osteoclasts in the stage III, respectively, promoting bone erosion. Fourth, continuous increase of gut permeability may induce gut microbial invasion of the joint synovial fluid in the stage IV. CONCLUSIONS: Clinical microbial intervention should consider the RA stage, where microbial dysbiosis and metabolic disorders present distinct patterns and played stage-specific roles. Our work provides a new insight in understanding gut-joint axis from a perspective of stages, which opens up new avenues for RA prognosis and therapy.

Chinese mental health workers' family-focused practices: a cross-sectional survey.

BACKGROUND: Mental disorders impose heavy burdens on patients' families and children. It is imperative to provide family-focused services to avoid adverse effects from mental disorders on patients' families and children. However, implementing such services requires a great deal of involvement of mental health workers. This study investigated the attitudes, knowledge, skills, and practices in respect to family-focused practices (FFP) in a sample of Chinese mental health workers. METHODS: A cross-sectional study design was employed to examine the attitudes, knowledge, skills, and practices of a convenience sample of Chinese mental health workers in respect to FFP, using the Chinese version of the Family-Focused Mental Health Practice Questionnaire (FFMHPQ). RESULTS: In total, 515 mental health workers participated in our study, including 213 psychiatrists, 269 psychiatric nurses, and 34 allied mental health professionals (20 clinical psychologists, 9 mental health social workers, and 4 occupational therapists). Compared with psychiatric nurses, psychiatrists and allied mental health professionals provided more support for families and children of patients with mental illness and were more willing to receive further training in FFP. However, there were no significant differences on knowledge, skills, and confidence across different profession types. After adjusting for demographic and occupational variables, previous training in FFP was positively associated with mental health workers' knowledge, skills, and confidence about FFP, but not actual support to families and children. CONCLUSIONS: Professional differences on FFP exist in Chinese mental health workers. Training is needed to engage psychiatrists and other allied workforce in dissemination and implementation of FFP in China.

A cellular model for Wilson's disease using patient-derived induced pluripotent stem cells revealed aberrant ß-catenin pathway during osteogenesis.

Wilson's disease (WD) is a rare autosomal recessive disorder of copper metabolism caused by an ATP7B gene mutation. Except for hepatic, neurological symptoms, lower bone mineral density is another most frequent clinical features of WD, but the underlying mechanisms have not been fully understood. This article aims to use induced pluripotent stem cells (iPSCs) to establish cellular osteoblasts model related to WD to identify abnormal osteogenesis and signaling pathways. In this study, we successfully produced functional osteoblasts from normal and WD iPSCs through embryoid bodies (EBs) formation method, and then we found WD osteoblasts may have a lower osteogenesis activity than normal controls by detection of osteogenic marker genes and mineralization ability. Further, through gene expression profiling, detection of ß-catenin in total protein and nuclear protein, and the nuclear localization of ß-catenin, we identified and validated that low osteogenic activity in WD may be due to abnormal ß-catenin pathway. Interestingly, we found SKL2001, a small molecule can reverse decreased osteogenesis of WD. In summery, our results suggested that the low bone density of WD may caused by abnormal ß-catenin signaling pathway, and these may provided a new target for the treatment of WD.

CD109 regulates the inflammatory response and is required for the pathogenesis of rheumatoid arthritis.

OBJECTIVE: The aim of this study was to investigate the role of CD109 in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLSs) and to evaluate its potential as a therapeutic target. METHODS: CD109 expression was examined in synovial tissues and FLSs from RA patients and collagen-induced arthritis (CIA) model mice. CD109-deficient mice were developed to evaluate the severity of CIA. Small interfering RNAs and a neutralising antibody against CD109 (anti-CD109) were designed for functional or treatment studies in RA FLSs and CIA. RESULTS: CD109 was found to be abundantly expressed in the synovial tissues from RA patients and CIA mice. CD109 expression in RA FLSs was upregulated by inflammatory stimuli, such as interleukin-1ß and tumour necrosis factor-α. Silencing of CD109 or anti-CD109 treatment reduced proinflammatory factor production, cell migration, invasion, chemoattractive potential and osteoclast differentiation, thereby reducing the deleterious inflammatory response of RA FLSs in vitro. Mice lacking CD109 were protected against arthritis in the CIA model. Anti-CD109 treatment prevented the onset and ameliorated the severity of CIA lesions. CONCLUSION: Our study uncovers an antiarthritic role for CD109 and suggests that CD109 inhibition might serve as a promising novel therapeutic strategy for RA.

A protocol for isolation and identification and comparative characterization of primary osteoblasts from mouse and rat calvaria.

Calvaria from neonatal mouse and rat is ideal resource for osteoblasts but can be easily contaminated by other cells such as fibroblasts. Here, we established a protocol for isolation and purification of primary osteoblast by enzyme sequential digestion and differential adhesion. In addition, we compared the phenotypic and functional traits of osteoblasts from mouse and rat which are commonly employed in studies. The method applied equally to rat and mouse in osteoblasts isolation and was corroborated its feasibility and validity. The results also provided us evidences for other experiments such as choosing a certain time point to give intervention and do the relevant tests.

Icariin Promotes the Osteogenic Action of BMP2 by Activating the cAMP Signaling Pathway.

Icariin (ICA) is the main active flavonoid glucoside from herbs of the genus Epimedium; in traditional Chinese medicine, these herbs have long been prescribed for the treatment of bone fractures and osteoporosis. Several studies have shown that treatment with ICA can increase osteogenic differentiation and reduce bone loss in vivo and in vitro. However, the definite signaling pathway of this osteogenic effect remains unclear. In this study, we selected bone morphogenetic protein 2 (BMP2)-induced osteoblastic differentiation of multipotent mesenchymal progenitor C2C12 cells as a model of osteoblast differentiation. We investigated the effects of ICA on C2C12 cells osteogenic differentiation and the underlying molecular mechanisms. We found that ICA could enhance BMP2-mediated osteoblastic differentiation of C2C12 cells in a dose-dependent manner. Treatment with ICA activated the cAMP/PKA/CREB signaling axis in a time-dependent manner. Blocking cAMP signaling using the PKA selective inhibitor H89 significantly inhibited the stimulatory effect of ICA on osteogenesis. Therefore, the osteoinductive potential and the low cost of ICA indicate that it is a promising alternative treatment or promoter for enhancing the therapeutic effects of BMP2.

Exosomes from C2C12 myoblasts enhance osteogenic differentiation of MC3T3-E1 pre-osteoblasts by delivering miR-27a-3p.

Many regulators have been identified to participate in the cross-talk between muscle and bone, however, most previous studies focus on secreting proteins. In this study, we demonstrated that exosomes from myoblasts C2C12 can promote pre-osteoblasts MC3T3-E1 differentiation to osteoblasts. We revealed that the effect of C2C12 exosomes depended on its miR-27a-3p component, they can increase miR-27a-3p level in the recipient cells, and decrease its direct target adenomatous polyposis coli (APC) expression, thus activating ß-catenin pathway. Furthermore, C2C12 exosomes failed to exert above effects when miR-27a-3p was deprived. These findings indicates exosomal microRNAs can be regarded as a novel type of "myokines" with osteogenesis promoting potential, which would broad our understanding of the muscle-bone interaction under physiological and pathological conditions.

Comparison of the prevalence of respiratory viruses in patients with acute respiratory infections at different hospital settings in North China, 2012-2015.

BACKGROUND: Acute respiratory infections (ARIs) are a great public health challenge globally. The prevalence of respiratory viruses in patients with ARIs attending at different hospital settings is fully undetermined. METHODS: Laboratory-based surveillance for ARIs was conducted at inpatient and outpatient settings of 11 hospitals in North China. The first 2-5 patients with ARIs were recruited in each hospital weekly from 2012 through 2015. The presence of respiratory viruses was screened by PCR assays. The prevalence of respiratory viruses was determined and compared between patients at different hospital settings. RESULTS: A total of 3487 hospitalized cases and 6437 outpatients/Emergency Department (ED) patients were enrolled. The most commonly detected viruses in the hospitalized cases were respiratory syncytial virus (RSV, 33.3%) in children less than two years old, adenoviruses (13.0%) in patients 15-34 years old, and influenza viruses (IFVs, 9.6%) in patients ≥65 years. IFVs were the most common virus in outpatient/ED patients across all age groups (22.7%). After controlling for the confounders caused by other viruses and covariates, adenoviruses (adjusted odds ratio [aOR]: 3.97, 99% confidence interval [99% CI]: 2.19-7.20) and RSV (aOR: 2.04, 99% CI: 1.34-3.11) were independently associated with increased hospitalization in children, as well as adenoviruses in adults (aOR: 2.14, 99% CI: 1.19-3.85). Additionally, co-infection of RSV with IFVs was associated with increased hospitalization in children (aOR: 12.20, 99% CI: 2.65-56.18). CONCLUSIONS: A substantial proportion of ARIs was associated with respiratory viruses in North China. RSV, adenoviruses, and co-infection of RSV and IFVs were more frequent in hospitalized children (or adenoviruses in adults), which might predict the severity of ARIs. Attending clinicians should be more vigilant of these infections.

Isopsoralen Enhanced Osteogenesis by Targeting AhR/ERα.

Isopsoralen (IPRN), one of the main effective ingredients in Psoralea corylifolia Linn, has a variety of biological effects, including antiosteoporotic effects. In vivo studies show that IPRN can increase bone strength and trabecular bone microstructure in a sex hormone deficiency-induced osteoporosis model. However, the mechanism underlying this osteogenic potential has not been investigated in detail. In the present study, we investigated the molecular mechanism of IPRN-induced osteogenesis in MC3T3-E1 cells. Isopsoralen promoted osteoblast differentiation and mineralization, increased calcium nodule levels and alkaline phosphatase (ALP) activity and upregulated osteoblast markers, including ALP, runt-related transcription factor 2 (RUNX2), and collagen type I alpha 1 chain (COL1A1). Furthermore, IPRN limited the nucleocytoplasmic shuttling of aryl hydrocarbon receptor (AhR) by directly binding to AhR. The AhR target gene cytochrome P450 family 1 subfamily A member 1 (CYP1A1) was also inhibited in vitro and in vivo. This effect was inhibited by the AhR agonists indole-3-carbinol (I3C) and 3-methylcholanthrene (3MC). Moreover, IPRN also increased estrogen receptor alpha (ERα) expression in an AhR-dependent manner. Taken together, these results suggest that IPRN acts as an AhR antagonist and promotes osteoblast differentiation via the AhR/ERα axis.

MiR-5100 promotes osteogenic differentiation by targeting Tob2.

MicroRNAs have emerged as pivotal regulators in various physiological and pathological processes, including osteogenesis. Here we discuss the contribution of miR-5100 to osteoblast differentiation and mineralization. We found that miR-5100 was upregulated during osteoblast differentiation in ST2 and MC3T3-E1 cells. Next, we verified that miR-5100 can promote osteogenic differentiation with gain-of-function and loss-of-function experiments. Target prediction analysis and experimental validation demonstrated that Tob2, which acts as a negative regulator of osteogenesis, was negatively regulated by miR-5100. Furthermore, we confirmed that the important bone-related transcription factor osterix, which can be degraded by binding to Tob2, was influenced by miR-5100 during osteoblast differentiation. Collectively, our results revealed a new molecular mechanism that fine-tunes osteoblast differentiation through miR-5100/Tob2/osterix networks.

Ultra-weak photon emission in healthy subjects and patients with type 2 diabetes: evidence for a non-invasive diagnostic tool.

BACKGROUND: Spontaneous ultra-weak photon emission (UPE) is a common phenomenon in biological systems and has been linked to pathological states. Researchers have always considered ultra-weak photon emission a potential non-invasive diagnostic tool, but its application in the medical field is stagnant due to the lack of relevant data for pathological states. METHODS: Ultra-weak photon signals from five body sites (forehead, neck, heart, stomach, and navel) in fifty patients with type 2 diabetes and sixty age-matched healthy subjects were measured using a moveable whole-body biophoton detection system. Photon signal is measured for 10 min and detected in bins of 50 ms by a photomultiplier with a range of 290-630 nm. Each signal is a time series of 12 000 elements. Various parameters including photon intensity, Q value, squeezed state parameters (|α|, θ, ø, r) and SSI were analyzed. RESULTS AND CONCLUSION: we found significant differences in the abovementioned parameters between groups, and all subjects could be clustered into two groups according to the results obtained by principal component analysis. Methods and results from this study could be useful for constructing a UPE database for a range of diseases, which would promote the application of UPE in clinical diagnosis in the future.

PRELP (proline/arginine-rich end leucine-rich repeat protein) promotes osteoblastic differentiation of preosteoblastic MC3T3-E1 cells by regulating the ß-catenin pathway.

Proline/arginine-rich end leucine-rich repeat protein (PRELP) is a collagen-binding proteoglycan highly expressed in the developing bones. Recent studies indicated that PRELP could inhibit osteoclastogenesis as a NF-κB inhibitor. However, its role during osteoblast differentiation is still unclear. In this study, we confirmed that the expression of PRELP increased with the osteogenesis induction of preosteoblastic MC3T3-E1 cells. Down-regulation of PRELP expression by shRNA reduced ALP activity, mineralization and expression of osteogenic marker gene Runx2. Our microarray analysis data suggested that ß-catenin may act as a hub gene in the PRELP-mediated gene network. We validated furtherly that PRELP knockdown could inhibit the level of connexin43, a key regulator of osteoblast differentiation by affecting ß-catenin protein expression, and its nuclear translocation in MC3T3-E1 preosteoblasts. Therefore, this study established a new role of PRELP in modulating ß-catenin/connexin43 pathway and osteoblast differentiation.

IL-22 promotes the proliferation of cancer cells in smoking colorectal cancer patients.

Chronic cigarette smoking increases the risk of developing colorectal cancer (CRC) and causes higher mortality of CRC patients. To improve our understanding of the underlying mechanism and devise treatment strategies specifically targeted at chronic smoking CRC patients, we examined the immune system of healthy and CRC patients who are complete nonsmokers or chronic primary smokers. We found that the serum concentrations of CRC nonsmokers and CRC smokers were skewed toward Th17-type cytokines, including interleukin (IL)-17 and IL-22. Notably, smoking CRC subjects had significantly higher levels of IL-22 than nonsmoking CRC patients. We also observed higher percentages of CCR4(+)CCR6(+) Th17 cells in circulating blood and higher secretion of IL-17 and IL-22 by peripheral blood mononuclear cells (PBMCs) of nonsmoking CRC and smoking CRC patients, compared to healthy individuals. Again, we observed elevated IL-17 and IL-22 secretion by CRC smokers than nonsmokers. Since IL-22 has been shown to stimulate tumorigenesis, which was also replicated in our experiments using cancer cell line model, we tested whether CRC patients' cell culture supernatant could also support tumor growth using this model. We found that both HT29 cells and LoVo cells had the highest proliferation in the supernatant from smoking CRC patients. Moreover, the proliferation of LoVo cells in smoking CRC supernatant was significantly higher than that in nonsmoking CRC supernatant. In addition, we found that the IL-22 concentration in normal gut tissue of the smoking CRC patients was significantly increased compared to that in nonsmoking CRC subjects, while no significant differences were observed in tumor tissues. Our results suggest that chronic smokers may have higher risk for CRC and worse prognosis due to dysregulated IL-22 production.

Correlation between the different therapeutic properties of Chinese medicinal herbs and delayed luminescence.

In the practice and principle of Chinese medicine, herbal materials are classified according to their therapeutic properties. 'Cold' and 'heat' are the most important classes of Chinese medicinal herbs according to the theory of traditional Chinese medicine (TCM). In this work, delayed luminescence (DL) was measured for different samples of Chinese medicinal herbs using a sensitive photon multiplier detection system. A comparison of DL parameters, including mean intensity and statistic entropy, was undertaken to discriminate between the 'cold' and 'heat' properties of Chinese medicinal herbs. The results suggest that there are significant differences in mean intensity and statistic entropy and using this method combined with statistical analysis may provide novel parameters for the characterization of Chinese medicinal herbs in relation to their energetic properties.

Classification of Chinese herbs based on the cluster analysis of delayed luminescence.

Traditional Chinese material medica are an important component of the Chinese pharmacopeia. According to the traditional Chinese medicinal concept, Chinese herbal medicines are classified into different categories based on their therapeutic effects, however, the bioactive principles cannot be solely explained by chemical analysis. The aim of this study is to classify different Chinese herbs based on their therapeutic effects by using delayed luminescence (DL). The DL of 56 Chinese herbs was measured using an ultra-sensitive luminescence detection system. The different DL parameters were used to classify Chinese herbs according to a hierarchical cluster analysis. The samples were divided into two groups based on their DL kinetic parameters. Interestingly, the DL classification results were quite consistent with classification according to the Chinese medicinal concepts of 'cold' and 'heat' properties. In this paper, we show for the first time that by using DL technology, it is possible to classify Chinese herbs according to the Chinese medicinal concept and it may even be possible to predict their therapeutic properties.

Identification of prohibitin 1 as a potential prognostic biomarker in human pancreatic carcinoma using modified aqueous two-phase partition system combined with 2D-MALDI-TOF-TOF-MS/MS.

Membrane proteins are an important source of potential targets for anticancer drugs or biomarkers for early diagnosis. In this study, we used a modified aqueous two-phase partition system combined with two-dimensional (2D) matrix-assisted laser desorption ionization (MALDI) time of flight (TOF) mass spectrometry (MS, 2D-MALDI-TOF-TOF-MS/MS) analysis to isolate and identify membrane proteins in PANC-1 pancreatic cancer cells. Using this method, we identified 55 proteins, of which 31 (56.4 %) were membrane proteins, which, according to gene ontology annotation, are associated with various cellular processes including cell signal transduction, differentiation, and apoptosis. Immunohistochemical analysis showed that the expression level of one of the identified mitochondria membrane proteins, prohibitin 1 (PHB1), is correlated with pancreatic carcinoma differentiation; PHB1 is expressed at a higher level in normal pancreatic tissue than in well-differentiated carcinoma tissue. Further studies showed that PHB1 plays a proapoptotic role in human pancreatic cancer cells, which suggests that PHB1 has antitumorigenic properties. In conclusion, we have provided a modified method for isolating and identifying membrane proteins and demonstrated that PHB1 may be a promising biomarker for early diagnosis and therapy of pancreatic (and potentially other) cancers.

Mutational and structural characteristics of four novel heterozygous C-propeptide mutations in the proα1(I) collagen gene in Chinese osteogenesis imperfecta patients.

OBJECTIVE: Osteogenesis imperfecta (OI) with C-propeptide mutations in proα1(I) collagen gene are rarely reported. We report four novel C-propeptide mutations in COL1A1 gene from Chinese OI patients. METHODS: Clinical characteristics and radiographic findings were described for four OI patients with C-propeptide mutations in proα1(I) collagen gene. Mutations were identified by traditional DNA sequencing based on PCR. The locations of mutations were mapped, and in silico prediction was conducted to analyse their effects on protein structure. Histology studies of skin, bone and muscle tissues were performed. RESULTS: All four C-propeptide heterozygous mutations identified were in the COL1A1 gene. Heterozygous mutation of c.4021C>T (p.Q1341X) disrupted the chain recognition sequences and was found in patients with type IV OI. Mutations of c.3893C>A (p.T1298N) and c.3897C>A (p.C1299X) impeded the formation of disulphide bonds and were associated with type IV OI phenotype. Missense mutation of c.3835A>C (p.N1279H) disrupted Ca(2+) binding and led to a severe type III OI phenotype. In silico programs predicted damaging effects for the patients with type III OI and the creation of an exonic splicing enhancer hexamer sequence for the type IV patients. Expansion of the bone marrow cavity and disorganization of osteocyte alignment was evident in bone specimens; and muscle atrophy and enlargement of intramuscular connective tissue were found in muscle specimens. CONCLUSIONS: Four novel C-propeptide mutations in proα1(I) collagen gene were identified in Chinese OI patients, and their clinical severity ranged from moderate type IV to severe type III. In silico prediction of the mutation effect and histological characteristics of tissue specimens was in accordance with the OI phenotypes.

A novel deletion mutation and structural abnormality in the Bruton's tyrosine kinase gene identified in a Chinese patient with X-linked agammaglobulinemia.

BACKGROUND: X-linked agammaglobulinemia (XLA) is a heritable primary immune deficiency disorder caused by mutation of Bruton's tyrosine kinase (BTK) gene. The main clinical characteristics of XLA are recurrent respiratory tract infections and profoundly low serum immunoglobulin levels and B cells. METHODS: The clinical characteristics of a five-year-old Chinese boy with XLA were described. Mutations of BTK genes were identified by traditional DNA sequencing based on PCR. A three-dimensional model of the truncated BTK protein was constructed. RESULTS: Molecular analysis showed a point deletion of an adenine nucleotide at position 1427 (p.Tyr476Ser), which would cause a frameshift and premature termination at codon 484. Three-dimensional analysis showed that the truncated protein had lost the functional region for both ATP and substrate binding such that tyrosine kinase activity would be affected. CONCLUSIONS: The study identified a novel BTK mutation of one Chinese XLA patient. The truncated BTK model identified the loss of a functional domain.