Single-cell transcriptomic profiling unveils insights into ovarian fibrosis in obese mice.

BACKGROUND: Adiposity profoundly impacts reproductive health in both humans and animals. However, the precise subpopulations contributing to infertility under obese conditions remain elusive. RESULTS: In this study, we established an obese mouse model through an eighteen-week high-fat diet regimen in adult female mice. Employing single-cell RNA sequencing (scRNA-seq), we constructed a comprehensive single-cell atlas of ovarian tissues from these mice to scrutinize the impact of obesity on the ovarian microenvironment. ScRNA-seq revealed notable alterations in the microenvironment of ovarian tissues in obese mice. Granulosa cells, stromal cells, T cells, and macrophages exhibited functional imbalances compared to the control group. We observed heightened interaction strength in the SPP1-CD44 pairing within lgfbp7+ granulosa cell subtypes and Il1bhigh monocyte subtypes in the ovarian tissues of obese mice. Moreover, the interaction strength between Il1bhigh monocyte subtypes and Pdgfrb+ stromal cell subtypes in the form of TNF - TNFrsf1α interaction was also enhanced subsequently to obesity, potentially contributing to ovarian fibrosis pathogenesis. CONCLUSIONS: We propose a model wherein granulosa cells secrete SPP1 to activate monocytes, subsequently triggering TNF-α secretion by monocytes, thereby activating stromal cells and ultimately leading to the development of ovarian fibrosis. Intervening in this process may represent a promising avenue for improving clinical outcomes in fertility treatments for obese women.

METTL14 downregulation drives S100A4+ monocyte-derived macrophages via MyD88/NF-κB pathway to promote MAFLD progression.

Without intervention, a considerable proportion of patients with metabolism-associated fatty liver disease (MAFLD) will progress from simple steatosis to metabolism-associated steatohepatitis (MASH), liver fibrosis, and even hepatocellular carcinoma. However, the molecular mechanisms that control progressive MAFLD have yet to be fully determined. Here, we unraveled that the expression of the N6-methyladenosine (m6A) methyltransferase METTL14 is remarkably downregulated in the livers of both patients and several murine models of MAFLD, whereas hepatocyte-specific depletion of this methyltransferase aggravated lipid accumulation, liver injury, and fibrosis. Conversely, hepatic Mettl14 overexpression alleviated the above pathophysiological changes in mice fed on a high-fat diet (HFD). Notably, in vivo and in vitro mechanistic studies indicated that METTL14 downregulation decreased the level of GLS2 by affecting the translation efficiency mediated by YTHDF1 in an m6A-depedent manner, which might help to form an oxidative stress microenvironment and accordingly recruit Cx3cr1+Ccr2+ monocyte-derived macrophages (Mo-macs). In detail, Cx3cr1+Ccr2+ Mo-macs can be categorized into M1-like macrophages and S100A4-positive macrophages and then further activate hepatic stellate cells (HSCs) to promote liver fibrosis. Further experiments revealed that CX3CR1 can activate the transcription of S100A4 via CX3CR1/MyD88/NF-κB signaling pathway in Cx3cr1+Ccr2+ Mo-macs. Restoration of METTL14 or GLS2, or interfering with this signal transduction pathway such as inhibiting MyD88 could ameliorate liver injuries and fibrosis. Taken together, these findings indicate potential therapies for the treatment of MAFLD progression.

Dancr-BRG1 regulates Nfatc1 transcription and Pgc1ß-dependent metabolic shifts in osteoclastogenesis.

Long non-coding RNA (lncRNA) serves as a vital regulator of bone metabolism, but its role in pathologically overactive osteoclast differentiation remains elusive. Here, we identify lncRNA Dancr (Differentiation Antagonizing Non-protein Coding RNA) as a critical suppressor of osteoclastogenesis and bone resorption, which is down-regulated in response to estrogen deficiency. Global or osteoclast-specific Dancr Knockout mice display significant trabecular bone deterioration and enhanced osteoclast activity, but minimal alteration of bone formation. Moreover, the bone-targeted delivery of Dancr by Adeno-associated viral remarkably attenuates ovariectomy-induced osteopenia in mice. Mechanistically, Dancr establishes a direct interaction with Brahma-related gene 1 to prevent its binding and preserve H3K27me3 enrichment at the nuclear factor of activated T cells 1 and proliferator-activated receptor gamma coactivator 1-beta promoters, thereby maintaining appropriate expression of osteoclastic genes and metabolic programs during osteoclastogenesis. These results demonstrate that Dancr is a key molecule maintaining proper osteoclast differentiation and bone homeostasis under physiological conditions, and Dancr overexpression constitutes a potential strategy for treating osteoporosis.

Gender dimorphism in hepatocarcinogenesis-DNA methylation modification regulated X-chromosome inactivation escape molecule XIST.

BACKGROUND: Sex disparities constitute a significant issue in hepatocellular carcinoma (HCC). However, the mechanism of gender dimorphism in HCC is still not completely understood. METHODS: 5-Hydroxymethylcytosine (5hmC)-Seal technology was utilised to detect the global 5hmC levels from four female and four male HCC samples. Methylation of XIST was detected by Sequenom MassARRAY methylation profiling between HCC tissues (T) and adjacent normal liver tissues (L). The role of Tet methylcytosine dioxygenase 2 (TET2) was investigated using diethylnitrosamine (DEN)-administered Tet2-/- female mice, which regulated XIST in hepatocarcinogenesis. All statistical analyses were carried out by GraphPad Prism 9.0 and SPSS version 19.0 software. RESULTS: The results demonstrated that the numbers of 5hmC reads in the first exon of XIST from female HCC tissues (T) were remarkably lower than that in female adjacent normal liver tissues (L). Correspondingly, DNA methylation level of XIST first exon region was significantly increased in female T than in L. By contrast, no significant change was observed in male HCC patients. Compared to L, the expression of XIST in T was also significantly downregulated. Female patients with higher XIST in HCC had a higher overall survival (OS) and more extended recurrence-free survival (RFS). Moreover, TET2 can interact with YY1 binding to the promoter region of XIST and maintain the hypomethylation state of XIST. In addition, DEN-administered Tet2-/- mice developed more tumours than controls in female mice. CONCLUSIONS: Our study provided that YY1 and TET2 could interact to form protein complexes binding to the promoter region of XIST, regulating the methylation level of XIST and then affecting the expression of XIST. This research will provide a new clue for studying sex disparities in hepatocarcinogenesis. HIGHLIGHTS: XIST was significantly downregulated in HCC tissues and had gender disparity. Methylation levels in the XIST first exon were higher in female HCC tissues, but no significant change in male HCC patients. The TET2-YY1 complex regulate XIST expression in female hepatocytes. Other ways regulate XIST expression in male hepatocytes.

Circular RNA cFAM210A, degradable by HBx, inhibits HCC tumorigenesis by suppressing YBX1 transactivation.

Hepatitis B protein x (HBx) has been reported to promote tumorigenesis in hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC), but the mechanism awaits further investigation. In this study, we found that cFAM210A (a circular RNA derived from the third exon of transcript NM_001098801 of the FAM210A gene; CircBase ID: hsa_circ_0003979) can be silenced by HBx. cFAM210A expression was downregulated and negatively correlated with tumorigenesis in patients with HBV-related HCC. Furthermore, cFAM210A reduced the proliferation, stemness, and tumorigenicity of HCC cells. Mechanistically, HBx increased the N6-methyladenosine (m6A) level of cFAM210A by promoting the expression of RBM15 (an m6A methyltransferase), thus inducing the degradation of cFAM210A via the YTHDF2-HRSP12-RNase P/MRP pathway. cFAM210A bound to YBX1 and inhibited its phosphorylation, suppressing its transactivation function toward MET. These findings suggest the important role of circular RNAs in HBx-induced hepatocarcinogenesis and identify cFAM210A a potential target in the prevention and treatment of HBV-related HCC.

Roles of RNA m6A modification in nonalcoholic fatty liver disease.

NAFLD is a series of liver disorders, and it has become the most prevalent hepatic disease to date. However, there are no approved and effective pharmaceuticals for NAFLD owing to a poor understanding of its pathological mechanisms. While emerging studies have demonstrated that m6A modification is highly associated with NAFLD. In this review, we summarize the general profile of NAFLD and m6A modification, and the role of m6A regulators including erasers, writers, and readers in NAFLD. Finally, we also highlight the clinical significance of m6A in NAFLD.

FTO promotes liver inflammation by suppressing m6A mRNA methylation of IL-17RA.

Background: Previous studies have demonstrated that inflammation-related interleukin-17 (IL-17) signaling plays a pivotal role in the pathogenesis of non-alcoholic steatohepatitis (NASH)- and alcoholic liver disease (ALD)-induced hepatocellular carcinoma (HCC). However, rare efforts have been intended at implementing the analysis of N6-methyladenosine (m6A) mRNA methylation to elucidate the underpinning function of the IL-17 receptor A (IL-17RA) during the inflammation-carcinogenesis transformation of HCC. Methods: We performed methylated RNA immunoprecipitation sequencing (MeRIP-seq) using normal, HCC tumor and paired tumor adjacent tissues from patients to investigate the dynamic changes of m6A mRNA methylation in the process of HCC. Additionally, murine non-alcoholic fatty liver disease (NAFLD) model and murine chronic liver injury model were utilized to investigate the role of IL-17RA regulated by m6A mRNA modulator fat mass and obesity-associated (FTO) in chronic hepatic inflammation. Results: MeRIP-seq revealed the reduction of m6A mRNA methylation of IL-17RA in tumor adjacent tissues with chronic inflammation, suggesting the potential role of IL-17RA in the inflammation-carcinogenesis transformation of HCC. Besides, we demonstrated that FTO, rather than methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and alkB homolog 5 (ALKBH5) functions as a main modulator for the decrease of m6A mRNA methylation of IL-17RA via knockdown and overexpression of FTO in vitro and in vivo. Conclusions: Overall, we elaborated the underlying mechanisms of the increase of IL-17RA resulting in chronic inflammation via the demethylation of FTO in tumor adjacent tissues and demonstrated that targeting the specific m6A modulator FTO may provide an effective treatment for hepatitis patients to prevent the development of HCC.

Spatial maps of hepatocellular carcinoma transcriptomes reveal spatial expression patterns in tumor immune microenvironment.

Rationale: Hepatocellular carcinoma (HCC) is a highly heterogeneous and malignant disease with the complex immune microenvironment, which ultimately influence clinic outcomes of patients. However, the spatial expression patterns of diverse immune cells among tumor microenvironment remain to be further deciphered. Methods: Spatial transcriptomics sequencing (ST) was implemented on two portions of HCC specimens. Differentially expressed genes, cell cycle phases, epithelial-mesenchymal features, pseudo-time and immune infiltration analysis were applied to demonstrate the intratumor heterogeneity and define the specific immune-related regions, and the results were further validated by a second analysis on another ST study. In vitro and in vivo experiments were conducted to confirm the functional mechanisms of key molecules such as CCL15, CCL19 and CCL21. Clinical tissue samples were used to assess their potential prognostic and therapeutic values. Results: Totally, 7553 spots were categorized into 15 subsets by hierarchical clustering, and malignant subsets with intratumor heterogeneity phenotypes were identified. Spatial heterogeneity from distinct sectors highlights specific chemokines: CCL15 is remarkable in the core region of the carcinoma sector and facilitates the immunosuppressive microenvironment by recruiting and polarizing M2-like macrophages in vitro and in vivo; High expression of CCL15 and CD163 respectively predicts poor prognosis of HCC patients, and the combined application of them has better predictive value. CCL19 and CCL21, sharing similar spatial expression patterns, are highly-correlated and prominent in the immune infiltration enrichment and recruit CD3+ T cells and CD20+ B cells to inhibit the growth of HCC, indicating a good prognosis of HCC patients. Conclusions: Taken together, our studies preliminarily reveal intratumor heterogeneity of HCC based on ST techniques and unravel the previously unexplored spatial expression patterns in the immune microenvironment. We also highlight the clinical significance and spatial discrepancy of key molecules, providing novel insight for further developing therapeutic strategies in HCC.

Dysregulated N6-methyladenosine (m6A) processing in hepatocellular carcinoma.

N6-methyladenosine (m6A) is the most thoroughly studied type of internal RNA modification, as this epigenetic modification is the most abundant in eukaryotic RNAs to date. This modification occurs in various types of RNAs and plays significant roles in dominant RNA-related processes, such as translation, splicing, export and degradation. These processes are catalyzed by three types of prominent enzymes: writers, erasers and readers. Increasing evidence has shown that m6A modification is vital for the regulation of gene expression, carcinogenesis, tumor progression and other abnormal changes, and recent studies have shown that m6A is important in the development of hepatocellular carcinoma (HCC). Herein, we summarize the nature and regulatory mechanisms of m6A modification, including its role in the pathogenesis of HCC and related chronic liver diseases. We also highlight the clinical significance and future strategies involving RNA m6A modifications in HCC.