Investigating oral microbiome profiles in patients with cleft lip and palate compared with the healthy control.

BACKGROUND: Patients with cleft lip and palate (CLP) have an oronasal communication differed from the closed state in healthy individuals, leading to a unique oral microbiome. This study aimed to determine if variances in the oral microbiota persist among CLP patients who have received treatments for the closure of these fistulas compared to the microbiota of healthy individuals. METHODS: Saliva samples were collected from a cohort comprising 28 CLP patients (CLP group) and 30 healthy controls (HC group). Utilizing 16S rRNA sequencing on the Illumina NovaSeq platform, we conducted a comprehensive analysis of the diversity and composition of the oral microbiota. RESULTS: The analysis of the microbiota in the saliva samples revealed a total of 23 microbial phyla, 38 classes, 111 orders, 184 families, 327 genera and 612 species. The alpha diversity with microbial abundance and evenness indicated the significant difference between the CLP and HC groups. Principal coordinate analysis (PCoA) and the ADONIS test further supported the presence of distinct microorganisms between the two groups. The CLP group displayed elevated abundances of Neisseria, Haemophilus, Porphyromonas, and Granulicatella, as indicated by LefSe analysis. Conversely, Rothia, Veillonella, and Pauljensenia exhibited significant reductions in abundance in the CLP group. The results of the PICRUSt analysis indicated significant differences in the relative abundance of 25 KEGG pathways within the CLP group. Through Spearman correlation analysis, strong associations between Rothia, Veillonella, and Pauljensenia and 25 functional pathways linked to CLP were identified. CONCLUSION: Findings of this study offer a thorough comprehension of the microbiome profiles of CLP patients after the restoration of oronasal structure and are anticipated to present innovative concepts for the treatment of CLP.

Identification and analysis of key hypoxia- and immune-related genes in hypertrophic cardiomyopathy.

BACKGROUND: Hypertrophic cardiomyopathy (HCM), an autosomal dominant genetic disease, is the main cause of sudden death in adolescents and athletes globally. Hypoxia and immune factors have been revealed to be related to the pathology of HCM. There is growing evidence of a role for hypoxia and inflammation as triggers and enhancers in the pathology in HCM. However, the role of hypoxia- and immune-related genes in HCM have not been reported. METHODS: Firstly, we obtained four HCM-related datasets from the Gene Expression Omnibus (GEO) database for differential expression analysis. Immune cells significantly expressed in normal samples and HCM were then screened by a microenvironmental cell population counter (MCP-counter) algorithm. Next, hypoxia- and immune-related genes were screened by the LASSO + support vector machine recursive feature elimination (SVM-RFE) and weighted gene co-expression network analysis (WGCNA). Single-gene enrichment analysis and expression validation of key genes were then performed. Finally, we constructed a competing endogenous RNA (ceRNA) network of key genes. RESULTS: In this study, 35 differentially expressed hypoxia genes were found. By using LASSO + SVM-RFE analysis, 10 more targets with differentially expressed hypoxia genes were identified. The MCP-count algorithm yielded five differentially expressed immune cells, and after assessing them for WGCNA characteristics, 612 immune genes were discovered. When hypoxia and immune genes were combined for cross-tabulation analysis, three hypoxia- and immune-related genes (ATP2A2, DDAH1, and OMA1) were identified. CONCLUSION: Based on hypoxia characteristic genes, three key genes were identified. These were also significantly related to immune activation, which proves a theoretical basis and reference value for studying the relationship between HCM and hypoxia and immunity.

Atf4 regulates angiogenic differences between alveolar bone and long bone macrophages by regulating M1 polarization, based on single-cell RNA sequencing, RNA-seq and ATAC-seq analysis.

In the repair of maxillofacial bone defects, autogenous craniofacial bone can often provide superior clinical results over long bone grafts. Most current studies have focused on the osteogenic differences between alveolar bone marrow (ABM) and long bone marrow (LBM), however, studies about the angiogenic differences between the two are currently lacking. We downloaded single-cell RNA sequencing (scRNA-seq) of mouse ABM and LBM respectively from the public database, and the data were processed by using Seurat package. CellphoneDB2 results showed that macrophages had the strongest interaction with mesenchymal stem cells (MSCs) and endothelial cells (ECs). ELISA results confirmed that ABM macrophages secreted a higher level of vascular endothelial growth factor A (Vegfa) compared to LBM macrophages, which further promoted angiogenesis of ECs and MSCs. Using SCENIC package, six key transcription factors (TFs) were identified to regulate the difference between ABM and LBM macrophages, and activating transcription factor 4 (Atf4) was confirmed to be more expressed in ABM macrophages by polymerase chain reaction (PCR) and western blot (WB), with predicted target genes including Vegfa. Besides, the result of scRNA-seq implied ABM macrophages more in M1 status than LBM macrophages, which was confirmed by the following experiments. From the results of another assay for transposase accessible chromatin sequencing (ATAC-seq) and RNA-seq about M1 macrophages, Atf4 was also confirmed to regulate the M1 polarization. So, we suspected that Atf4 regulated the different expression of Vegfa between ABM and LBM macrophages by activating M1 polarization. After knocking down Atf4, the expression of M1 polarization markers and Vegfa were downregulated and vasculogenic differences were eliminated, which were subsequently reversed by the addition of LPS/IFN-γ. Our study might provide a new idea to improve the success rate of autologous bone grafting and treatment of oral diseases.

Associations between mixed urinary phenols and parabens metabolites and bone mineral density: Four statistical models.

Phenols and parabens widely exist in personal care and consumer products and have been proved to be endocrine disrupting chemicals that could disturb bone metabolism. The current studies focusing on the associations between phenols and parabens with bone mineral density (BMD) drew contradictory conclusions. Considering the bias might be due to not considering the effects of mixed exposure, we conducted the first cross-sectional study to investigate the associations of both single and mixed metabolites of phenols and parabens with BMD in three populations by setting up four models: generalized linear regression model (GLM), weighted quantile sum (WQS) regression model, quantile g-computation (qgcomp) model and Bayesian kernel machine regression (BKMR) model, based on the US National Health and Nutrition Examination Survey (NHANES) database. We found that the association between the mixtures and total femur BMD in men was significantly negative. Bisphenol A (BPA) was shown to play the most important role in this negative association in the BKMR model, and this negative association was also confirmed in the GLM model with ß coefficient (95% CI) being -0.02 (-0.04, -0.01). The relationships between the mixtures and femoral neck and trochanter BMD in postmenopausal women were significantly positive. Benzophenone-3 (BP-3) played the most significant role in the positive association with trochanter BMD, as confirmed by the WQS, qgcomp and BKMR models, and this positive association was also verified by the GLM model with ß coefficient (95% CI) being 0.01 (0.00, 0.02). In conclusion, the association between the mixed phenols and parabens and BMD was negative in men while was positive in postmenopausal women, which was gender-specific. This study might provide new ideas for the prevention and treatment of osteoporosis and the control of personal care and consumer products containing phenols and parabens in the future.

Association between mixed aldehydes and bone mineral density based on four statistical models.

Aldehydes as an environmental pollutant may lead to oxidative stress, which is an important mechanism in the development of osteoporosis. This suggests a possible link between aldehyde exposure and osteoporosis. Considering the mixed nature of aldehyde exposure and the interactions between different aldehydes, we explored for the first time the associations between mixed six aldehydes (benzaldehyde, butyraldehyde, heptanal, hexanal, isovaleraldehyde, and propionaldehyde) and BMD in three populations (men, premenopausal women, and postmenopausal women) by applying four statistical models: quantile g-computation (qgcomp) model, Bayesian kernel machine regression (BKMR) model, generalized linear regression model (GLM), and generalized additive model (GAM), based on the National Health and Nutrition Examination Survey (NHANES) 2013-2014. We found that mixed aldehydes could significantly reduce BMD in men, with hexanaldehyde and propanaldehyde having the greatest negative qgcomp model and BKMR model weights, also confirmed by GLM. The associations between isopentanaldehyde and propanaldehyde and femoral BMD in men were non-linear and had threshold effects as derived from the BKMR model and GAM. The associations turned positive when the concentrations of isopentanaldehyde and propanaldehyde exceeded their respective inflection points. To conclude, our study might provide new ideas for the prevention and treatment of osteoporosis, and hexanaldehyde and propanaldehyde should be more regulated to prevent osteoporosis.

Association of urinary phthalate metabolites with renal function among 9989 US adults.

PURPOSE: This study aimed to investigate the relationship between phthalate metabolites and renal function. METHODS: We analyzed data from 9989 participants who took part in the National Health and Nutrition Examination Survey (NHANES) from 2005 to 2018. Renal function was reflected by estimated glomerular filtration rate (eGFR), urinary albumin-to-creatinine ratio (UACR), and hypertension. We used generalized linear regression to estimate the correlation between covariate-adjusted creatinine-normalized phthalate metabolites and renal function. In addition, subgroup analysis was used to further compare the effect differences between various populations. RESULTS: In the adjusted model, we found differential associations between phthalates and plasticizers metabolites and renal function. We found that Mono-benzyl phthalate, Mono-(3-carboxypropyl) phthalate, and Mono-(2-ethyl-5-oxohexyl) phthalate were positively associated with lower eGFR with odds ratios (95% confidence intervals) of 1.38 (1.14, 1.67), 1.30 (1.09, 1.57), and 1.27 (1.04, 1.53). While Mono-ethyl phthalate, Mono-(2-ethyl)-hexyl phthalate, Mono-isononyl phthalate and Mono-isobutyl phthalate were negatively associated with lower eGFR with OR values of 0.79 (0.69, 0.90), 0.64 (0.52, 0.78), 0.65 (0.51, 0.82) and 0.80 (0.63, 1.00), respectively. In addition, we found that Mono(carboxyoctyl) phthalate and Mono-isobutyl phthalate were negatively associated with hypertension with ORs of 0.86 (0.78, 0.96) and 0.84 (0.72, 0.98). But phthalates and plasticizers metabolites were not associated with UACR. CONCLUSION: This study found differences in the effects of phthalates and plasticizers metabolites on kidney function, which may raise concerns about possible changes in kidney function resulting from exposure to current levels of plasticizers.

Orexin-A Reverse Bone Mass Loss Induced by Chronic Intermittent Hypoxia Through OX1R-Nrf2/HIF-1α Pathway.

Background: Recent studies suggest that there is a potential connection between obstructive sleep apnea (OSA) and osteoporosis through dysregulation of bone metabolism. Orexin-A, a neuroprotective peptide secreted by the hypothalamus, is at a lower level in the plasma of OSA patients, which regulates appetite, energy expenditure and sleep-wake states. However, the protective effect of orexin-A on bone metabolism in OSA is unclear. Purpose: To investigate whether the activation of OX1R by orexin-A can reverse bone mass loss induced by chronic intermittent hypoxia (CIH). Methods: Mice were randomly divided into the normoxia group and CIH group. Within the CIH or normoxia groups, treatment groups were given a subcutaneous injection of either orexin-A or saline vehicle once every day for 4 weeks and then femurs were removed for micro-CT scans. Histology and immunohistochemical staining were performed to observe and calculate the changes in femurs as a result of hypoxia. Cell immunofluorescence and immunohistochemical staining were used to detect the expression of orexin receptors in MC3T3-E1 cells or in bones. CCK-8 assay, ALP assay kit and alizarin red staining were used to detect the viability, alkaline phosphatase (ALP) activity, and capacity of mineralization, respectively. The effect of orexin-A on osteogenic differentiation of MC3T3-E1 cells was evaluated using qRT-PCR, Western blot and cell staining. Results: CIH led to a decrease in the amount and density of trabecular bone, downregulated OCN expression while increasing osteoclast numbers in femurs and inhibited the expression of RUNX2, OSX, OPN and Nrf2 in MC3T3-E1 cells. Orexin-A treatment alleviated these CIH-induced effects by combining to OX1R. The level of HIF-1α was elevated both in CIH and orexin-A treatment groups. Conclusion: CIH environment inhibits osteogenesis and orexin-A can reverse bone mass loss induced by CIH through OX1R-Nrf2/HIF-1α pathway.