Reproductive senescence in laying hens, characterized by a progressive decline in egg production, represents a major challenge for the poultry industry. Although microRNAs (miRNAs) are recognized as important regulators of aging, their specific roles and mechanisms in ovarian aging of hens remain largely unclear. This study was designed to comprehensively analyze miRNA expression patterns during ovarian aging in laying hens. The objectives of this study were to identify key functional miRNAs and to elucidate their molecular regulatory mechanisms. Specifically, this study evaluated ovarian senescence in hens at 350, 500 and 700 days of age, observing a decline in egg production, increased follicular atresia, and p53 upregulation. miRNA sequencing analysis identified 44 differentially expressed miRNAs (DEMs), among which gga-let-7i exhibited the highest abundance and showed progressive upregulation during aging. Functional assays revealed that gga-let-7i induces cell cycle arrest and promotes cellular senescence in ovarian follicle granulosa cells (GCs). Mechanistically, collagen type I alpha 2 chain (COL1A2) was confirmed as a direct target of gga-let-7i, and it has been demonstrated that gga-let-7i accelerates senescence by inhibiting the COL1A2/PI3K/AKT/MDM2 pathway, resulting in p53 accumulation and the downstream cellular senescence signaling pathways activation. These results uncover a novel gga-let-7i/COL1A2 regulatory axis involved in ovarian aging and suggest potential targets for extending reproductive longevity in laying hens.

The development of skeletal muscle are complicated processes involving genes responsible for proper muscle morphology, contractility, cell proliferation, differentiation, interactions, migration, and death. The three-dimensional chromatin architecture of skeletal muscle development has not been studied intensively although dynamic transcriptional regulation during differentiation of muscle cells is one of the most deeply studied processes. The RNA-seq was used to analyze the transcriptome pattern during chicken muscle development across 12 stages. Hi-C was used to build chromatin architectures during four representative stages. ChIP-seq was conducted to identify enhancers and promoters in these four stages, which are occupied by histone H3K27ac and H3K4me3 peaks. Results show that large-scale genome architecture changes are mostly unidirectional, and coupled by complex on/off dynamic patterns of gene expression. Specifically, we observed 258.30 Mb of the genome undergoing A/B compartment switching. Notable alterations (316.57 Mb) of interaction frequencies within TADs were observed. Substantial aging-associated genes exhibited ascending connectivity with the compartment transition from repressive to active status during muscle development. Some muscle-related gene promoters that interacted with active enhancers during development, and some myopathy/aging-associated genes that were activated in aging muscle were founded. These results provide key insights into skeletal muscle development in vivo, and offer a valuable resource that allows in-depth functional characterization of candidate genes.

Ovarian aging in laying hens contributes significantly to the decline in egg production performance during the late laying period, and is accompanied by extensive remodeling of miRNA-mRNA interaction networks, which are critical post-transcriptional regulators during this process. In this study, we investigated the role of miR-30e-3p, which was previously identified as significantly upregulated during ovarian aging in laying hens, in granulosa cell (GC) senescence. To identify its downstream effectors, we performed mRNA transcriptome sequencing and screened for downregulated genes. Among potential targets, the histone methyltransferase complex regulatory subunit dpy-30 (DPY30) was validated as a direct target of miR-30e-3p by luciferase reporter and expression analyses. Functional assays demonstrated that miR-30e-3p significantly suppressed GC proliferation and induced cell cycle arrest, whereas DPY30 exhibited opposite effects. Furthermore, inhibition of miR-30e-3p or overexpression of DPY30 attenuated D-galactose (D-gal) -induced GC senescence. Mechanistically, miR-30e-3p promoted cellular senescence at least in part through downregulation of DPY30. Collectively, our findings indicated that miR-30e-3p promotes GC senescence by targeting DPY30 in chicken ovarian follicles, and suggest that both miR-30e-3p and DPY30 may serve as potential biomarkers for improving reproductive longevity and mitigating ovarian aging-related disorders in poultry.

BACKGROUND:Programmed cell death plays a critical role in skeletal muscle atrophy. Ferroptosis, an iron-dependent form of programmed cell death driven by lipid peroxidation, has been implicated in various diseases, but its role in skeletal muscle atrophy remains unclear. METHODS:Ferroptosis in skeletal muscle atrophy was investigated using two models: dexamethasone (Dex)-induced atrophy (n = 6 independent cell cultures per group) and simulated microgravity (n = 6 mice per group). Conditional Nestin knockout (KO) mice were generated using CRISPR/Cas9 (n = 6-8 mice per group), with wild-type (WT) controls (n = 6-8). Phenotypic analyses included histopathology (HE staining), functional assessments (muscle strength, weight analysis, treadmill), and dystrophy evaluation (dystrophin staining). Molecular analyses involved flow cytometry, ELISA, transmission electron microscopy, PI staining, and IP/MS to delineate Nestin-regulated ferroptosis pathways in skeletal muscle atrophy. RESULTS:Ferroptosis was significantly activated in both atrophy models, with a 2.5-fold increase in lipid peroxidation (p < 0.01), a 2-fold accumulation of Fe2+ (p < 0.01) and a 50% reduction in Nestin expression (p < 0.001). Nestin KO mice exhibited exacerbated muscle atrophy, showing a 40% decrease in muscle weight (p < 0.01) and a 30% reduction in muscle strength (p < 0.05) compared to WT mice. Nestin overexpression mitigated Dex-induced ferroptosis, reducing lipid peroxidation by 40%, decreasing Fe2+ accumulation by 50% (p < 0.01), and improving muscle function by 30% (p < 0.05). Mechanistically, Nestin interacted with MAP 1LC3B (LC3B) to catalyse LC3B polyubiquitination at lysine-51, reducing LC3B availability for autophagy and inhibiting autophagy flux by 60% (p < 0.01), leading to a 50% reduction in ferroptosis (p < 0.001). CONCLUSIONS:Our study identifies Nestin as a critical regulator of ferroptosis-autophagy crosstalk in skeletal muscle atrophy. Targeting Nestin-LC3B ubiquitination may offer novel therapeutic strategies for preventing muscle wasting in diseases such as cachexia and sarcopenia.

Non-alcoholic fatty liver disease (NAFLD) is a clinical syndrome characterized primarily by hepatocellular steatosis and lipid accumulation, which leads to hepatocyte apoptosis, autophagy, inflammation, and intracellular oxidative stress. NAFLD is recognized as one of the most prevalent and complex chronic liver diseases globally, with its occurrence and associated mortality rates rising swiftly each year. Due to the high similarity between chicken fatty liver syndrome (FLS) and NAFLD, as well as the easy availability of diseased chickens, the chicken is considered an ideal model for studying the pathogenesis of NAFLD. Previous studies have pinpointed several circular RNAs (circRNAs) implicated in the pathogenesis of NAFLD, yet the underlying functions and mechanisms of numerous circRNAs continue to remain elusive. In this experiment, we utilized circRNA sequencing of chicken livers to identify a novel circRNA, named circACACA, and discovered that it disrupts the metabolic homeostasis of lipids within hepatocytes. Consequently, this disruption leads to oxidative stress and the induction of autophagy, ultimately exerting an adverse effect on chicken liver health. Mechanistically, circACACA functions as a molecular sponge for miR-132b-5p and miR-101-2-5p to modulate the expression of the downstream CBFB/PIM1 complex. Consequently, it influenced the activity of the AKT/mTOR and PPAR-γ signaling pathways to perform its physiological functions. Crucially, we noticed substantial sequence similarity of circACACA across diverse species by comprehensively searching databases. Further, our research with a mouse model confirmed that the functional conservation of circACACA across livers of different species. Overall, this study built a mechanistic network for circACACA and confirmed its sequence conservation and functional relevance across various species. Our results not only provide new targets for the prevention and treatment of NAFLD but also present fresh perspectives for progress in healthy production of laying hens.

Background: Speckle Type POZ Protein (SPOP), despite its tumor type-dependent role in tumorigenesis, primarily as a tumor suppressor gene is associated with a variety of different cancers. However, its function in pancreatic cancer remains uncertain. Methods: SPOP expression and the association between its expression and patient prognosis and immune function were evaluated using The Cancer Genome Atlas (TCGA), Genotype-Tissue Expression (GTEx), The Tumor Immune Estimation Resource 2.0 (TIMER2.0) database, cBioportal, and various bioinformatic databases. Enrichment analysis of SPOP and the association between SPOP expression with clinical stage and grade were analyzed using the R software package. Then immunohistochemistry (IHC) was used to estimate the correlation between SPOP and tumor-infiltrating lymphocytes (TILs) in patients with pancreatic cancer. Results: As part of our study, we assessed that SPOP was anomalously expressed in kinds of cancers, associated with clinical stage and outcomes. Meanwhile, SPOP also played a crucial role in the tumor microenvironment (TME). The expression level of SPOP was significantly correlated to tumor-infiltrating immune cells (TICs) in pancreatic cancer. Conclusions: Our study uncovered the potential corrections in SPOP with TICs, suggesting that SPOP may act as a biomarker for immunotherapy in pancreatic cancer.

The development of skeletal muscle are complicated processes involving genes responsible for proper muscle morphology, contractility, cell proliferation, differentiation, interactions, migration, and death. The three-dimensional chromatin architecture of skeletal muscle development has not been studied intensively although dynamic transcriptional regulation during differentiation of muscle cells is one of the most deeply studied processes. The RNA-seq was used to analyze the transcriptome pattern during chicken muscle development across 12 stages. Hi-C was used to build a chromatin architectures during four representative stages. ChIP-seq was conducted to identify enhancers in these four stages, which are occupied by histone H3K27ac and H3K4me3 peaks. Results show that large-scale genome architecture changes are mostly unidirectional, and coupled by complex on/off dynamic patterns of gene expression. Specifically, we observed 258.30 Mb of the genome undergoing A/B compartment switching. Notable alterations (316.57 Mb) of interaction frequencies within TADs were observed. Substantial aging-associated genes exhibited ascending connectivity with the compartment transition from repressive to active status during muscle development. Some muscle-related gene promoters that interacted with active enhancers during development, and some myopathy/aging-associated genes that were activated in aging muscle were founded. These results provide key insights into skeletal muscle development in vivo, and offer a valuable resource that allows in-depth functional characterization of candidate genes.

MicroRNAs (miRNAs) play a crucial role as transcription regulators in various aspects of follicular development, including steroidogenesis, ovulation, apoptosis, and gene regulation in poultry. However, there is a paucity of studies examining the specific impact of miRNAs on ovarian granulosa cells (GCs) across multiple grades in laying hens. Consequently, this study aims to investigate the roles of miRNAs in chicken GCs. By constructing miRNA expression profiles of GCs at 10 different time points, encompassing 4 pre-hierarchical, 5 preovulatory, and 1 postovulatory follicles stage, we identified highly expressed miRNAs involved in GC differentiation (miR-148a-3p, miR-143-3p), apoptosis (let7 family, miR-363-3p, miR-30c-5p, etc.), and autophagy (miR-128-3p, miR-21-5p). Furthermore, we discovered 48 developmentally dynamic miRNAs (DDMs) that target 295 dynamic differentially expressed genes (DDGs) associated with follicular development and selection (such as oocyte meiosis, progesterone-mediated oocyte maturation, Wnt signaling pathway, TGF-β signaling pathway) as well as follicular regression (including autophagy and cellular senescence). These findings contribute to a more comprehensive understanding of the intricate mechanisms underlying follicle recruitment, selection, and degeneration, aiming to enhance poultry’s reproductive capacity.

Granulosa cells play a pivotal role in growth, development and ovulation of ovarian follicle. Simultaneously, autophagy and apoptosis processes are crucial determinants in the destiny of granulosa cells. Within this context, miR-29-3p, known to regulate a broad spectrum of biological processes and critical for tumor detection, prognosis, and treatment, is poised to clarify its roles in both autophagy and apoptosis. To enhance the understanding of the influence of miR-29-3p on follicular development, our study primarily delved into the realms autophagy and apoptosis. We employed a well-established chicken follicular atrophy model achieved through subcutaneous injection of tamoxifen (TMX) into hens. qPCR analysis revealed a significant decrease in the expression of miR-29-3p within the atrophic follicles. In our in vitro experiments with cultured chicken primary granulosa cells, miR-29-3p emerged as a novel microRNA capable of impeding autophagy and apoptosis when transfected with miR-29-3p mimics and inhibitors. Results from luciferase reporter assays corroborated that PTEN is a legitimate target of miR-29-3p. Unlike miR-29-3p, PTEN appeared to foster autophagy and apoptosis in chicken granulosa cells. Moreover, our findings uncovered that miR-29-3p facilitates the phosphorylation of Akt and mTOR proteins by targeting PTEN in chicken granulosa cells. In conclusion, the findings of this study suggest that miR-29-3p, through its targeting of PTEN via the Akt/mTOR signaling pathway, exerts inhibitory effects on autophagy and apoptosis. These effects may hold significant importance in the context of follicular development.

MicroRNAs (miRNAs) have been demonstrated to control chicken skeletal muscle growth, however, the potential function of the miR-181-5p family in chicken myogenesis remains largely unknown. Here, our study identified the two chicken (Gallus gallus; Gga) miR-181-5p family members widely expressed in various tissues, specifically miR-181a-5p and miR-181b-5p. Besides, the breast muscles of fast-growing broilers expressed higher levels of miR-181a-5p and miR-181b-5p than those of slow-growing layers. Functionally, miR-181a-5p and miR-181b-5p both promote the expression level of myogenic factors including Myogenin (MyoG), Myogenic differentiation 1 (MyoD1), and Myosin heavy chain (MyHC), meanwhile accelerating the myotube formation of skeletal muscle satellite cells (SMSCs). Mechanistically, miR-181a-5p and miR-181b-5p directly bind to the 3' Untranslated region (UTR) of the Transforming growth factor beta receptor 1 (TGFBR1) mRNA, further reducing the expression of TGFBR1. TGFBR1 is a key Transforming growth factor beta (TGF-β) signaling transduction receptor and had a negative function in muscle cell differentiation. Furthermore, knockdown of TGFBR1 facilitated the expression of chicken myogenic factors, boosted myotube formation, and decreased the SMAD family member 2/3 (SMAD2/3) phosphorylation in chicken SMSCs. SMAD2/3 are downstream of TGF-β signaling, and miR-181a-5p and miR-181b-5p could reduce the expression of TGFBR1 to further diminish the SMAD2/3 phosphorylation. Our findings revealed that the miR-181-5p family targets TGFBR1 to break the TGF-β signaling transduction, which resulted in promoting chicken skeletal muscle development.

Ovarian follicle development is an important physiological activity for females and makes great significance in maintaining female health and reproduction performance. The development of ovarian follicle is mainly affected by the granulosa cells (GCs), whose growth is regulated by a variety of factors. Here, we identified a novel circular RNA (circRNA) derived from the Ribosomal protein S19 (RPS19) gene, named circRPS19, which is differentially expressed during chicken ovarian follicle development. Further explorations identified that circRPS19 promotes GCs proliferation and steroid hormone synthesis. Furthermore, circRPS19 was found to target and regulate miR-218-5p through a competitive manner with endogenous RNA (ceRNA). Functionals investigation revealed that miR-218-5p attenuates GCs proliferation and steroidogenesis, which is opposite to that of circRPS19. In addition, we also confirmed that circRPS19 upregulates the expression of Inhibin beta B subunit (INHBB) by binding with miR-218-5p to facilitate GCs proliferation and steroidogenesis. Overall, this study revealed that circRPS19 regulates GCs development by releasing the repression of miR-218-5p on INHBB, which suggests a novel mechanism in respect to circRNA and miRNA regulation in ovarian follicle development.

The liver plays a critical role in metabolic activity and is the body's first immune barrier, and maintaining liver health is particularly important for poultry production. MicroRNAs (miRNAs) are involved in a wide range of biological activities due to their capacity as posttranscriptional regulatory elements. A growing body of research indicates that miR-21-5p plays a vital role as a modulator of liver metabolism in various species. However, the effect of miR-21-5p on the chicken liver is unclear. In the current study, we discovered that the fatty liver had high levels of miR-21-5p. Then the qPCR, Western blot, flow cytometry, enzyme-linked immunosorbent assay, dual-luciferase, and immunofluorescence assays were, respectively, used to determine the impact of miR-21-5p in the chicken liver, and it turned out that miR-21-5p enhanced lipogenesis, oxidative stress, and inflammatory responses, which ultimately induced hepatocyte apoptosis. Mechanically, we verified that miR-21-5p can directly target nuclear factor I B (NFIB) and kruppel-like factor 3 (KLF3). Furthermore, our experiments revealed that the suppression of NFIB promoted apoptosis and inflammation, and the KLF3 inhibitor accelerated lipogenesis and enhanced oxidative stress. Furthermore, the cotransfection results suggest that the PI3K/AKT pathway is also involved in the process of miRNA-21-5p-mediate liver metabolism regulation. In summary, our study demonstrated that miRNA-21-5p plays a role in hepatocyte lipogenesis, oxidative stress, inflammation, and apoptosis, via targeting NFIB and KLF3 to suppress the PI3K/AKT signal pathway in chicken.

Within the poultry industry, hens' reproductive performance is of great economic significance. The development and growth of follicles is a key aspect of hen egg production, and ovarian follicle growth and development are closely associated with granulosa cells (GCs) proliferation and the synthesis of steroid hormones. It has been confirmed by numerous studies that microRNAs (miRNAs) play important roles in the steroid hormone synthesis and proliferation of GCs. In this study, we examined the main miRNAs influencing hens' ability to reproduce, identified the miR-223 that is mainly expressed in atretic follicles based on sequencing, and investigated its role in GCs. Then, we used miR-223 mimic and inhibitor to knockdown or overexpress miR-223 expression. The result showed that miR-223 significantly inhibits both the steroid hormone synthesis and the proliferation of GCs. Subsequently, the results of the dual luciferase reporter experiment and bioinformatics prediction demonstrated that cysteine rich transmembrane BMP regulator 1 (CRIM1) was a downstream target gene of miR-223, and overexpression of miR-223 prevented CRIM1 expression. The function of CRIM1 was further investigated, and we observed a significant reduction in the synthesis of steroid hormones and the proliferation of GCs after transfection with CRIM1 siRNA. The opposite function of miR-223 was observed for CRIM1 in our study. Additionally, we demonstrated the involvement of the miR-223/CRIM1 axis in GCs through modulation of the AKT signaling pathway. Our data demonstrate the pivotal role of the miR-223 in the proliferation and steroid hormone synthesis of chicken GCs, which helps to explain how non-coding RNA (ncRNA) affects chicken reproductive function.

Background The reproductive performance of chickens mainly depends on the development of follicles. Abnormal follicle development can lead to decreased reproductive performance and even ovarian disease among chickens. Chicken is the only non-human animal with a high incidence of spontaneous ovarian cancer. In recent years, the involvement of circRNAs in follicle development and atresia regulation has been confirmed. Results In the present study, we used healthy and atretic chicken follicles for circRNA RNC-seq. The results showed differential expression of circRALGPS2. It was then confirmed that circRALGPS2 can translate into a protein, named circRALGPS2-212aa, which has IRES activity. Next, we found that circRALGPS2-212aa promotes apoptosis and autophagy in chicken granulosa cells by forming a complex with PARP1 and HMGB1. Conclusions Our results revealed that circRALGPS2 can regulate chicken granulosa cell apoptosis and autophagy through the circRALGPS2-212aa/PARP1/HMGB1 axis.

The egg-laying performance of hens holds significant economic importance within the poultry industry. Broody inheritance of the parent stock of chickens can result in poor options for the improvement of egg production, and is a phenomenon influenced by multiple genetic factors. However, few studies have been conducted to delineate the molecular mechanism of ovarian regression in brooding chickens. Here, we explored the pivotal genes responsible for the regulation of ovarian follicles in laying hens, using RNA-sequencing analysis on the small ovarian follicles from broody and laying chickens. Sequencing data analysis revealed the differential expression of 200 genes, with a predominant enrichment in biological processes related to cell activation and metabolism. Among these genes, we focused on solute carrier family 5 member 5 (SLC5A5), which exhibited markedly higher RNA expression levels in follicles from laying compared with broody chickens. Subsequent cellular function studies with knockdown of SLC5A5 in chicken ovarian follicle granulosa cells (GCs) led to the down-regulation of genes associated with cell proliferation and steroid hormone synthesis, and concurrent promotion of gene expression linked to apoptosis. These findings indicated that SLC5A5 deficiency led to the inhibition of proliferation, steroid hormone synthesis and secretion, and promotion of apoptosis in chicken GCs. Our study demonstrated a pivotal role for SLC5A5 in the development and function of chicken GCs, shedding light on its potential significance in the broader context of chicken ovarian follicle development, and providing a prospective target to improve the egg-laying performance of chickens via molecular marker-assisted breeding technology.

Circular RNAs (circRNAs) have been recognized as critical regulators of skeletal muscle development. Myocyte enhancer factor 2A (MEF2A) is an evolutionarily conserved transcriptional factor that regulates myogenesis. However, it remains unclear whether MEF2A produces functional circRNAs. In this study, we identified two evolutionarily conserved circular MEF2A RNAs (circMEF2As), namely circMEF2A1 and circMEF2A2, in chicken and mouse muscle stem cells. Our findings revealed that circMEF2A1 promotes myogenesis by regulating the miR-30a-3p/PPP3CA/NFATC1 axis, whereas circMEF2A2 facilitates myogenic differentiation by targeting the miR-148a-5p/SLIT3/ROBO2/β-catenin signaling pathway. Furthermore, in vivo experiments demonstrated that circMEF2As both promote skeletal muscle growth. We also discovered that the linear MEF2A mRNA-derived MEF2A protein binds to its own promoter region, accelerating the transcription of MEF2A and upregulating the expression of both linear MEF2A and circMEF2As, forming a MEF2A autoregulated positive feedback loop. Moreover, circMEF2As positively regulate the expression of linear MEF2A by adsorbing miR-30a-3p and miR-148a-5p, which directly contribute to the MEF2A autoregulated feedback loop. Importantly, we found that mouse circMEF2As are essential for the myogenic differentiation of C2C12 cells. Collectively, our results demonstrated the evolution, function, and underlying mechanisms of circMEF2As in animal myogenesis, which may provide novel insight for both the farm animal meat industry and human medicine.

Skeletal muscle is important for animal meat production, regulating movements, and maintaining homeostasis. Circular RNAs (circRNAs) have been founded to play vital role in myogenesis. However, the effects of the numerous circRNAs on growth and development of the skeletal muscle are yet to be uncovered. Herein, we identified circLRRFIP1, which is a novel circular RNA that is preferentially expressed in the skeletal muscle. To study the role of circLRRFIP1 in the skeletal muscle, the skeletal muscle satellite cells (SMSCs) was used to silenced or overexpressed circLRRFIP1. The results obtained in this study showed that circLRRFIP1 play a positive role in the proliferation and differentiation of SMSCs. The SMSCs were generated with stable knockdown and overexpression of circLRRFIP1, and the results showed that circLRRFIP1 exerts a stimulatory effect on the proliferation and differentiation of SMSCs. We further generated SMSCs with stable knockdown and overexpression of circLRRFIP1, and the results revealed that circLRRFIP1 exerts a stimulatory effect on the proliferation and differentiation of SMSCs. Mechanistically, circLRRFIP1 targets the myogenic inhibitory factor-miR-15 family to release the suppression of the miR-15 family to AKT3. The knockdown of AKT inhibits SMSC differentiation through the mTOR/p70S6K pathway. Taken together, the results obtained in this present study revealed the important role and the regulatory mechanisms of circLRRFIP1 in the development of chicken skeletal muscle. Therefore, this study provides an attractive target for molecular breeding to enhance meat production in the chicken industry.

In this study, we examined the effects of Heat Shock Protein 90 (HSP90) on adipocyte proliferation and differentiation in chickens. To achieve this, we constructed RNA interference (RNAi) vectors to target HSP90 and transfected the vectors into primary adipocytes. After transfection, oil red O staining was performed to determine the status of triglyceride accumulation in the cells, whereas the CCK-8 cell kit and 5-Ethynyl-2'-Deoxyuridine (EdU) assays were used to determine cell proliferation. Thereafter, the mRNA and protein expression levels of PPARγ, FAS, SREBP-1c, and HSP90 were determined, and the results showed that after the interference of HSP90, the mRNA and protein expression levels of HSP90 in the chicken adipocytes decreased significantly compared to the control and blank groups (p < 0.05). The decreased mRNA and protein expression of PPARγ, FAS, and SREBP-1c was related to adipocyte differentiation (p < 0.05). However, HSP90 interference had no effect on adipocyte proliferation (p > 0.05). Taken together, the results of this study showed that HSP90 influenced the expression of PPARγ and adipose-differentiation-related genes, thereby regulating triglyceride accumulation and adipocyte differentiation in chickens.

Ovarian follicle development is a complex and well-orchestrated biological process of great economic significance for poultry production. Specifically, understanding the molecular mechanisms underlying follicular development is essential for high-efficiency follicular development can benefit the entire industry. In addition, domestic egg-laying hens often spontaneously develop ovarian cancer, providing an opportunity to study the genetic, biochemical, and environmental risk factors associated with the development of this cancer. Here, we provide high-quality RNA sequencing data for chicken follicular granulosa cells across 10 developmental stages, which resulted in a total of 204.57 Gb of clean sequencing data (6.82 Gb on average per sample). We also performed gene expression, time-series, and functional enrichment analyses across the 10 developmental stages. Our study revealed that SWF (small while follicle), F1 (F1 hierarchical follicles), and POFs (postovulatory follicles) best represent the transcriptional changes associated with the prehierarchical, preovulatory, and postovulatory stages, respectively. We found that the preovulatory stage F1 showed the greatest divergence in gene expression from the POF stage. Our research lays a foundation for further elucidation of egg-laying performance of chicken and human ovarian disease.

The liver is a vital organ in chickens that performs a number of crucial physiological functions, including the storage of hepatic glycogen, protein synthesis, detoxification, and deoxidation. The growth and metabolism of the liver are complex processes influenced by factors such as environment, diet, and genetics. MicroRNAs (miRNAs), as post-transcriptional regulatory molecules, play a role in various biological processes. There is growing evidence that miR-27b-5p plays a key role in the regulation of liver development and metabolism in various species. However, its role in chicken livers has yet to be determined. In our experiment, we found that chickens with fatty livers had significantly higher levels of serum triglyceride (TG) and total cholesterol (TC) compared to the normal chickens, while the control group had significantly higher levels of very low-density lipoprotein (VLDL) and serum hormones. Further research showed that the mRNA of miR-27b-5p was highly expressed in fatty livers. By exploring the function of miR-27b-5p in chicken livers, we discovered that it promotes lipogenesis, oxidative stress, and inflammatory responses, leading to hepatocyte apoptosis. Our study also established the mechanism by which miR-27b-5p interacts with its target gene, and found that miR-27b-5p targets insulin receptor substrate 2 (IRS2) and modulates the PI3K/AKT signaling pathway. Additionally, our investigation of IRS2 in chicken hepatocytes revealed that knocking down IRS2 has the same effects as overexpressing miR-27b-5p. In conclusion, our study revealed that miR-27b-5p directly binds to IRS2, inhibiting the PI3K/AKT signaling pathway and causing steatosis, oxidative stress, inflammation, and apoptosis in chicken liver.