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Purpose: Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly, but the therapies are not satisfactory. This study aimed to find AMD specific features through the analysis of high-throughput sequencing., Methods: In this study, we integrated six projects containing single-cell RNA sequencing (scRNA-seq) data to perform a comprehensive analysis for AMD samples in the tissues of retina and retinal pigment epithelium/choroid, and in the positions of macula and periphery. Differentially expressed genes (DEGs) were analyzed and crucial signaling pathways were identified across cell types and between the macula and periphery. The intercellular signaling transduction among cell types were inferred by "CellChat" to build cell-cell communication network under normal and AMD conditions, and verified at the transcriptional level. The CD31+ endothelial cells were obtained to evaluate the enrichment of KEGG pathways in atrophic and neovascular AMD, and GSVA was adopted to discover differential metabolic signals in each AMD type., Results: Thirteen major cell types were identified in the integrated scRNA-seq data. Although no disease-specific cell type or differential cell proportion was found, DEGs and enriched pathways were shown in cell-type- and position-dependent manners. Severe impairment of endothelial cell-mediated cell interactions was found in the signaling transduction network of the macula, and compromised cell interactions were observed in the periphery. Furthermore, distinct signaling pathways and metabolic states were uncovered in atrophic and neovascular AMD. Striking reduction in energy metabolism, lipid metabolism, and oxidative stress was indicated in the atrophic AMD., Conclusion: Conclusively, we discover aberrant signals and metabolic pathways in AMD samples, providing insight into mechanisms and potential therapeutic targets for the AMD treatment.

Somatic cell nuclear transfer (SCNT) has been successfully employed across various mammalian species, yet cloned animals consistently exhibit low pregnancy rates, primarily due to placental abnormalities such as hyperplasia and hypertrophy. This study investigated the involvement of the Hippo signaling pathway in aberrant placental development in SCNT-induced bovine pregnancies. SCNT-derived cattle exhibited placental hypertrophy, including enlarged abdominal circumference and altered placental cotyledon morphology. RNA sequencing analysis indicated significant dysregulation of Hippo signaling pathway genes in SCNT placentas. Co-expression of YAP1 and CCND1 was observed in cloned blastocysts, placental tissues, and bovine placental mesenchymal stem cells (bPMSCs). Manipulation of YAP1 expression demonstrated the capacity to regulate bPMSC proliferation. Experimental assays confirmed the direct binding of YAP1 to CCND1, which subsequently promoted CCND1 expression in bPMSCs. Furthermore, inhibition of CDK6, a downstream target of CCND1, attenuated SCNT bPMSC proliferation. This study identified YAP1 as a key regulatory component within the Hippo signaling pathway that drives placental hyperplasia in cloned cattle through up-regulation of CCND1-CDK6 expression, facilitating cell cycle progression. These findings offer potential avenues for enhancing cloning efficiency, with implications for evolutionary biology and the conservation of valuable germplasm resources.

Feeding behavior is regulated by a complex network of endogenous neuropeptides. In chordates, this role is suggested to be under the control of diverse factors including thyrotropin-releasing hormone (TRH). However, whether this regulatory activity of TRH is functionally conserved in non-chordate metazoans, and to what extent this process is underpinned by interactions of TRH with other neuropeptides such as cholecystokinin (CCK, known as a satiety signal), remain unclear. This study investigated the TRH signaling system in the echinoderm Apostichopus japonicus . Bioinformatic analyses and ligand-binding assays identified a functional TRH receptor (AjTRHR) that activated signaling via the MAPK/ERK1/2 pathways. Experimental administration of TRH significantly reduced feeding activity, while up-regulating CCK expression. RNA interference (RNAi) experiments confirmed that both CCK and TRH are essential components of satiety signaling, working synergistically to mediate feeding inhibition. Evolutionary analysis of TRH-type peptides revealed greater conservation of the short isoform of TRH compared to the long isoform, probably driven by strong selection acting on the functional redundancy. These findings provide compelling evidence of a TRH-mediated signaling system in non-chordate deuterostomes, expanding our understanding of neuropeptide-regulated feeding mechanisms in marine invertebrates.

Pigeons and certain other avian species produce a milk-like secretion in their crop sacs to nourish offspring, yet the detailed processes involved are not fully elucidated. This study investigated the crop sacs of 225-day-old unpaired non-lactating male pigeons (MN) and males initiating lactation on the first day after incubation (ML). Using RNA sequencing, ribosome profiling, and single-cell transcriptome sequencing (scRNA-seq), we identified a significant up-regulation of genes associated with ribosome assembly and protein synthesis in ML compared to MN. Results from scRNA-seq analysis identified 12 distinct cell types and 22 clusters, with secretory epithelial cells (SECs) exhibiting marked expression of plasma cell markers, including IGLL1 and MZB1 . RNA fluorescence in situ hybridization (RNA FISH) and IgY quantification confirmed the critical role of SECs in producing endogenous IgY during lactation. We propose that fibroblast-derived BAFF signals activate SECs, mimicking B cell transformation and enhancing protein production through the unfolded protein response (UPR). These findings shed light on the cellular dynamics of pigeon milk production and contribute to a broader understanding of avian biology.

This study investigates lipid content and expression of genes involved in lipid metabolism in feline cumulus cells and oocytes before and after in vitro maturation (IVM). Domestic cats represent valuable models in reproductive research, yet the efficiency of in vitro embryo production remains suboptimal, with contributing factors still under investigation. We characterized lipid droplets (LDs) in oocytes collected from adult queens, both before and after IVM, using confocal microscopy with Bodipy 493/503 staining. We also quantified the expression of four genes involved in regulation of lipid metabolism. Our findings revealed a substantial accumulation of LDs and a significant upregulation of PLIN2 expression in both cumulus cells and oocytes following IVM. The number and total area of LDs and fluorescence intensity increased markedly in oocytes at the MII stage, while the average LD diameter decreased. Similarly, cumulus cells showed an increase in number and total area of LDs post-IVM, suggesting their involvement in oocyte maturation by modulating lipid homeostasis. This study presents the first detailed characterization of lipid droplet dynamics in feline oocytes and cumulus cells, providing insights into metabolic processes during IVM, which are critical for optimizing assisted reproductive technologies in felids., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Natalia Sowinska reports financial support was provided by Polish National Science Center. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Inc.)

Purpose: Previous studies have reported divergent sexual responses to aging; however, specific variations in gene expression between aging males and females and their potential association with age-related retinal diseases remain unclear. This study collected data from public databases and developed a comprehensive comparison of retina between aging females and males., Methods: Single-cell RNA (scRNA) and bulk RNA sequencing data of the aging retina from females and males in public databases were utilized for integrated analysis to investigate sex-biased expression in retina. Additionally, in vitro experiments were conducted on individuals with retinitis pigmentosa (RP) to validate the sex difference in degenerative retina., Results: Bulk RNA analysis revealed sex-biased expression of specific genes in retina of aging individuals, with immune pathway-related genes exhibiting higher expression in females compared to males. The scRNA analysis demonstrated that sex-biased gene expression was cell-type specific in aging retina. Furthermore, susceptibility genes for age-related macular degeneration and RP exhibited variation across different cell types and sexes. Cell-to-cell communication unveiled an increased interaction associated with TGFB1, CCL7, and VEGFA in Müller glia, microglia, and astrocytes of female retina. Notably, we observed female-biased chemokine expression in microglia contributing to heightened susceptibility to immune inflammation in female retina. Finally, we confirmed a more pronounced inflammatory response during degeneration in female rd10 mouse retina compared to males., Conclusions: This study provides a comprehensive comparison of retina between females and males in healthy aging human retina and highlights the significance of sex as an influential factor in retinal diseases.

Proliferative vitreoretinopathy (PVR) is a multifactorial ocular condition characterized by the development of fibrotic membranes inside the vitreous cavity and on the detached retina, which can result in severe blindness. Semaphorin7A (Sema7a) is involved in axon growth, inflammatory responses, and immune regulation; however, its role in PVR and regulatory mechanisms in retinal pigment epithelium (RPE) cells remains unclear. This study aimed to examine Sema7a in PVR and the underlying mechanisms. Transcriptome sequencing was used to investigate the changes in mRNA expression profiles. Western blotting, immunofluorescence, and real-time polymerase chain reaction (RT-PCR) were utilized to investigate the potential mechanism of Sema7a on epithelial-mesenchymal transition (EMT) in RPE cells. Stimulating RPE cells with transforming growth factor beta-1 (TGF-β1) decreased the levels of epithelial markers but increased those of mesenchymal markers. Based on transcriptome sequencing, many molecules associated with PVR progression were regulated. PVR vitreous fluid proteomics data analysis showed that Sema7a significantly changed at different levels. Silencing Sema7a in RPE cells attenuated TGF-β1-induced EMT and their ability to induce experimental PVR; in contrast, recombinant Sema7a (rSema7a) directly triggered EMT in RPE cells. TGF-β1 induction mechanically activated the PI3k-AKT and MAPK pathways, while Sema7a knockdown by short interfering RNA lowered the phosphorylation of the PI3k-AKT/MAPK signaling pathway. Therefore, Sema7a may be a viable therapeutic target for PVR due to its crucial role in the TGF-β1-induced EMT of RPE cells., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Assessment of the clock genes, Period (Per) 1, Per2, Per3, and Cryptochrome (Cry) 2, Cry3, and Cry4, can help better understand eel spawning ecology. In this study, the circadian rhythm and moonlight effects of these clock genes in the eel retina and hypothalamus were analyzed. We examined clock gene expression patterns under 12 h light:12 h darkness (12L12D), constant darkness (DD), and constant light (LL) conditions; under short photoperiod (SP; 9L15D) and long photoperiod (LP; 15L9D), and during the new moon (NM) and full moon in male eels. Per2 expression increased after sunrise, Cry2, and Cry4 expression increased around sunset, and Per1, Per3, and Cry3 expression increased before sunrise. Under SP conditions, oscillations of retinal Per3 and Cry4, which did not occur under LP conditions, were generated. In addition, retinal Cry4 oscillation was generated under NM conditions. These results suggest that the retina of the eel may play an important role in regulating circadian rhythm, and migration is initiated by the synchronization of clock genes by moonlight, suggesting that photic signals are closely related to the migratory activity of the eel., (© 2024 The Author(s). Journal of Experimental Zoology Part A: Ecological and Integrative Physiology published by Wiley Periodicals LLC.)

The objective of this study was to investigate the early follicular apoptosis in canine ovarian follicles by examining the expression of anti-apoptotic BCL-2 and pro-apoptotic BAX proteins throughout the estrous cycle associated with oocyte maturation. Follicular cells from preantral and antral follicles of varying sizes were isolated and grouped based on follicle type and estrous phase. Antral follicles underwent flow cytometry analysis, whereas preantral follicles were subjected to Western blotting. The meiotic capacity of oocytes from these different follicle types was evaluated through in vitro maturation. Statistical analysis included ANOVA and Duncan's test. Results showed fluctuations in BCL-2 and BAX levels across different follicular stages and estrous phases. BCL-2 levels increased (P<0.05) with follicular development in antral follicles, particularly during estrus, while BAX exhibited variations peaking (P<0.05) during estrus. The BCL-2/BAX ratio in antral follicles was higher (P<0.05) in estrus and diestrus compared to anestrus and proestrus. Additionally, BCL-2 and BAX proteins were detected in preantral follicles, with varying expression levels (P<0.05) across estrous phases. The BCL-2/BAX ratio in preantral follicles was highest (P<0.05) during anestrus and estrus and decreased (P<0.05) in proestrus and diestrus. Oocytes from preantral follicles did not reach the MII stage, regardless of the levels of BCL-2/BAX. Conversely, oocytes obtained from large follicles during estrus showed the highest (P<0.05) maturation percentages, which were associated with the highest BCL-2/BAX ratio. These findings provide insights into the dynamic patterns of BCL-2 /BAX in canine follicles across the estrous cycle, shedding light on their potential roles in oocyte development., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 Elsevier B.V. All rights reserved.)

Intermediate filaments (IFs) play a major role in determining and maintaining cell shape and anchoring intracellular organelles in place, in the tissues and organs of several species, starting from the early stages of development. This study was aimed at the immunohistochemical investigation of the presence, cellular localization and temporal distribution of the intermediate filaments keratin 8 (CK8), keratin 18 (CK18), keratin 19 (CK19), vimentin, desmin and laminin, all of which contribute to the formation of the cytoskeleton in the rat mammary gland during pregnancy, lactation and involution. On days 7, 14 and 21 of pregnancy (pregnancy period), on day 7 post-delivery (lactation period) and on day 7 post-weaning (involution period), under ketamine hydrochloride (Ketalar-Pfizer) (90 mg/kg) anesthesia, two mammary glands were fully excised from the abdominal region. It was determined that CK8 showed moderate immunoreactions in the alveolar and ductal epithelia, connective tissue and vascular endothelium of the rat mammary gland throughout pregnancy. On the 7th day of pregnancy, CK18 expression was absent in the alveolar and ductal epithelia, but was observed weakly in some connective tissue cells. Throughout pregnancy, lactation and involution, the alveolar and ductal epithelia of the rat mammary gland were determined to be negative for CK19. Desmin expression predominated in the mammary myoepithelium and vasculature throughout all three of the investigated periods. While vimentin was not expressed in any of the mammary tissue components during pregnancy and lactation, its moderate expression was observed in the alveolar and ductal epithelia during involution. The involution period was also characterized by the vimentin negativity of the myoepithelium, stroma, fat cells and blood vessels of the mammary gland. Throughout all three periods, laminin expression was strong in the alveolar and ductal epithelia, stromal and myoepithelial cells and blood vessels, and did not vary in strength between the investigated periods. These findings demonstrated that intermediate filaments showed cell- and tissue-specific expression patterns in the rat mammary gland under the effects of pregnancy, lactation and involution., (© 2024 The Authors. This is an open access article under the CC BY-NC-ND 4.0 license (https://creativecommons.org/licenses/by-nc-nd/4.0/deed.en), which allows re-users to copy and distribute the material in any medium or format in unadapted form and for noncommercial purposes, and only so long as attribution is given to the creator.)

Pyometra is a common life-threatening inflammatory disease with a complex etiopathogenesis that develops during the diestrus stage and can be observed in elderly intact bitches. The present study evaluated five aquaporin (AQP1, AQP2, AQP3, AQP5, and AQP9) transcript abundances and immunolocalization in the uterine tissue, and investigated their relationship with uterine tissue and blood lipopolysaccharide (LPS) concentration, superoxide dismutase (SOD) and glutathione peroxidase (GPX) activity, and nitric oxide (NO) production in dogs suffering from pyometra. The study sampled 36 client-owned intact bitches from different breeds, of which 24 cases were diagnosed with pyometra. Twelve of these bitches in the diestrus stage that presented for elective ovariohysterectomy were used as the control group. Blood samples were collected into tubes without anticoagulant for serum progesterone, LPS concentration, and antioxidant activities at the time of diagnosis. Bacteriological and tissue samples from the uteri were collected after the ovariohysterectomy. The tissue samples were used to determine antioxidant activity, and hormone and toxin concentrations. Transcript abundance of uterine AQPs were determined by qPCR, and their presence and localization were determined by by immunohistochemistry. For all pyometra samples, the bacteria isolated from the uterine swabs were Escherichia coli. Compared to the control group, AQP1, AQP2, and AQP5 were downregulated more than 2-fold, whereas AQP9 was upregulated nearly 3-fold and AQP3 was upregulated more than 4-fold in the pyometra affected uteri (P<0.05). Uterine AQP1 was moderately negatively correlated with serum LPS concentration (r=-0.568, P<0.01) and tissue NO production (r=-0.407, P<0.05). AQP5 was positively correlated with serum SOD activity (r=0.485, P<0.05) and negatively correlated with serum LPS concentration (r=-0.512, P<0.05). AQP9 was negatively correlated with tissue SOD and serum GPx activity. This is the first study to identify AQP9 transcript abundance and immunolocalization in canine uterine tissue. Uterine AQP1, AQP2, AQP3, AQP5, and AQP9 transcript abundances were altered in spontaneously developed canine pyometra while AQP transcript abundance was negatively related to serum toxin concentration, NO production, and antioxidant enzyme activity. Further studies should be conducted to determine the role of altered abundances of AQPs transcripts in pyometra pathogenesis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Testicular development and spermatogenesis in mice involve complex and dynamic gene regulation and chromatin remodelling. In this study, Real-time fluorescence quantitative PCR (RT-qPCR), Western Blot (WB), Immunofluorescence (IF), transfection and other techniques were used to analyse the expression of Ino80d mRNA and its encoded proteins in mouse testicular tissue and mouse spermatogonial cells, and to further analyse the possible target-regulatory relationship and function of miR-92a-3p and Ino80d. We found that Ino80d mRNA and protein expression was up-regulated in adult mouse testis tissue relative to juvenile mouse testis tissue, whereas miR-92a-3p expression was down-regulated in adult mouse testis tissue. Immunofluorescence results showed that the Ino80d protein was mainly localized in the nucleus of male germ cells. Ino80d protein expression is higher in spermatogonia, spermatid and lower in primary spermatocytes, secondary spermatocytes and sperm. There is a decreasing trend in development from spermatogonia to secondary spermatocytes. The transfection results showed that the expression levels of Ino80d mRNA and protein were down-regulated after overexpression of miR-92a-3p in mouse spermatogonia. Increased miR-92a-3p may be a key factor in inhibiting the expression of Ino80d mRNA and proteins in the miR-92a-3p mimics group of mouse spermatogonial cells, whereas differential expression may be a result of the negative regulation of miR-92a-3p, which regulates testicular development and spermatogenesis in mice., Competing Interests: Declaration of Competing Interest All authors disclosed no relevant relationships., (Copyright © 2024 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier B.V. All rights reserved.)

Increased risk of developing glaucoma has recently been associated with early age of menopause. Here, we examined how age and surgically-induced menopause via ovariectomy (OVX) impacted gene expression in gene pathways previously linked to glaucoma, such as extracellular matrix (ECM) remodeling and TGF-β signaling. Using bulk RNA sequencing, we analyzed changes in young (3-4 months) and middle-aged (9-10 months) Long-Evans rats. We focused on posterior pole tissues (sclera and optic nerve head) but also examined the retina to compare observed changes across different tissue regions. Our results demonstrated that aging and OVX significantly alter gene expression in the sclera and optic nerve head. Generally, OVX triggered the enrichment of immune-related processes. However, OVX in young rats also led to significant enrichment of ECM and TGF-β gene sets. At the same time, these effects were diminished in middle-aged rats, indicating an age dependency of the effects of OVX on matrix-related pathways. Notably, the transcriptional factor Fos was downregulated in the posterior eye and retina in aged and OVX animals. Fos is a major regulator of cell proliferation and survival, and its dysregulation may play an important role in aging and menopause for women. These findings underscore the important role of menopause timing in modulating molecular pathways associated with glaucoma, which is consistent with clinical studies showing that early menopause may heighten the risk of developing this condition. This study also highlights the importance of considering women's health factors, such as menopause, in understanding and managing glaucoma risk., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Ltd.)

A reduction in myometrial contractile activity can lead to inadequate cleaning of the uterine lumen, resulting in persistent endometritis and potentially endometrosis in mares. Oxytocin (OXT) is a key hormonal regulator of myometrial contraction. While epigenetic regulation of myometrial gene expression has been studied in humans, there is limited information on the expression of DNA methyltransferases (DNMTs) and ten-eleven translocation enzymes (TETs) in the myometrium of mares. This study aimed to evaluate the mRNA transcription of these enzymes and the potential role of DNA methylation in the expression of the OXT receptor (OXTR) gene in the myometrium of mares with endometrosis. Myometrial samples were collected post-mortem during the mid-luteal (n = 23) and follicular (n = 20) phases of the estrous cycle and assessed according to Kenney and Doig endometrial category (I, IIA, IIB, III). mRNA transcription of OXTR, DNMT1, -3A, -3B and TET1, -2, -3 were determined using qPCR. DNA methylation analysis at CpG islands of OXTR exons 1 and 2 was performed using bisulfite pyrosequencing. Myometrial OXTR mRNA transcription and DNA methylation in its promoter region showed no significant differences between categories, although increased methylation was observed at CpG island position 6 in exon 2. DNMT1, TET2, and TET3 mRNA transcription was altered in the equine myometrium depending on the phase of the estrous cycle and the severity of endometrosis (P < 0.05). These findings indicate that DNMTs and TETs were expressed in myometrium in a manner specific to the severity of endometrosis and phases of the estrous cycle, suggesting a potential regulatory role in DNA methylation of myometrial gene expression., Competing Interests: Declaration of Competing Interest No., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Purpose: Diabetic retinopathy (DR) is one of the most severe and common complications caused by diabetic mellites. Inhibiting NLRP3 inflammasome activation displays a crucial therapeutic value in DR. Studies have shown that KCNQ1OT1 plays a critical role in regulating NLRP3 inflammasome activation and participates in the pathogenesis of diabetic complications. The present study aims to explore the role, and the potential mechanism of KCNQ1OT1 in regulating the activation of NLRP3 inflammasome in DR., Methods: qRT-PCR was used to detect the expression of KCNQ1OT1, miR-17-5p, TXNIP, NLRP3, ASC, caspase-1 and IL-1β. Western blot was performed to detect the expression of NLRP3, ASC, caspase-1, IL-1β and TXNIP. Immunohistochemistry and immunostaining were performed to detect the expression of caspase-1. The levels of the inflammatory cytokine IL-1β were determined by ELISA assay. FISH was used to detect the subcellular localisation of KCNQ1OT1. Bioinformatic analysis, luciferase reporter assay and in vitro studies were performed to elucidate the mechanism of KCNQ1OT1-mediated dysfunction., Results: The expression of KCNQ1OT1 and the activation of NLRP3 inflammasome were increased in experimental DR models. KCNQ1OT1 knockdown alleviated NLRP3 inflammasome-associated molecules expression. In addition, KCNQ1OT1 was found to be localized mainly in the cytoplasm of Müller cells and facilitated TXNIP expression by acting as a miR-17-5p sponge. KCNQ1OT1 promoted the activation of NLRP3 inflammasome through miR-17-5p/TXNIP axis., Conclusions: In conclusion, it was found in this study that KCNQ1OT1 promoted the activation of NLRP3 inflammasome both in vitro and in vivo , which was mediated by miR-17-5p/TXNIP axis. KCNQ1OT1 might be an effective interference target for the prevention and treatment of DR.

Objective: The choice of a stable reference gene plays a crucial role in gene expression analysis. In most cases, the choice of housekeeping gene (HKG) is quite random. However, the expression of HKGs varies under different experimental conditions and in different tissues., Methods: The study focused on three reference genes in Catla Labeo catla-beta actin (β-actin), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and elongation factor-1 alpha (EF-1α)-under four experimental conditions: bacterial challenge, hypoxia, a combination of bacterial challenge and hypoxia, and control. The fish were subjected to the above-mentioned experimental circumstances, and five tissues (blood, gill, liver, kidney, and spleen) were collected at regular intervals., Result: The investigation of gene expression in the five tissues was performed using real-time (RT) quantitative polymerase chain reaction (qPCR), and the resultant cycle threshold values were subjected to several statistical techniques via the RefFinder tool to determine the most stable HKG. The results were ordered in accordance with the stability of the reference genes. In this study, GAPDH was found to be the least stable gene, while β-actin and EF-1α remained stable in the kidney and liver under various circumstances. Other tissues demonstrated tissue- and treatment-specific stability for the various HKGs., Conclusion: The findings indicate that GAPDH should be avoided in Catla for gene expression studies and that different algorithms should be employed to validate the stability of HKGs in RT-qPCR analysis., (© 2024 American Fisheries Society.)

The proper function of the placenta is essential for the health and growth of the fetus and the mother. The placenta relies on dynamic gene expression for its correct and timely development and function. Although numerous studies have identified genes vital for placental functions, equine placental molecular research has primarily focused on single placental locations, in sharp contrast with the broader approach in human studies. Here, we hypothesized that the molecular differences across different regions of the equine placenta are negligible because of its diffuse placental type with a macroscopic homogenous distribution of villi across the placental surface. We compared the transcriptome and stereological findings of the body, pregnant horn, and non-pregnant horn within the equine chorioallantois. Our transcriptomic analysis indicates that the variation between regions of the placenta within individuals is less than the variation observed between individuals. A low number of differentially expressed genes (DEGs) (n = 8) was identified when comparing pregnant and non-pregnant horns within the same placenta, suggesting a remarkable molecular uniformity. A higher number of DEGs was identified when comparing each horn to the body (193 DEGs comparing pregnant horn with body and 207 DEGs comparing non-pregnant horn with body). Genes with a higher expression in the body were associated with processes such as extracellular matrix synthesis and remodeling, which is relevant for placental maturation and placenta-endometrial separation at term and implies asynchrony of these processes across locations. The stereological analysis showed no differences in microcotyledonary density, and width between the locations. However, we observed a greater chorioallantoic thickness in the body and pregnant horn compared to the non-pregnant horn. Overall, our findings reveal a uniform transcriptomic profile across the placental horns, alongside a more distinct gene expression pattern between the uterine body and horns. These regional differences in gene expression suggest a different pace in the placental maturation and detachment among the placental locations., Competing Interests: Declaration of competing interest The authors have no conflicts of interest to declare., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Overactivation of the classic arm of the renin-angiotensin system (RAS) is one of the main mechanisms involved in obesity-related cardiac remodeling, and a possible relationship between RAS and ER stress in the cardiovascular system have been described. Thus, the aim of this study is to evaluate if activating the protective arm of the RAS by ACE inhibition or aerobic exercise training could overturn diet-induced pathological cardiac hypertrophy by attenuating ER stress. Male C57BL/6 mice were fed a control (SC) or a high-fat diet (HF) for 16 weeks. In the 8th week, HF-fed animals were randomly divided into HF, enalapril treatment (HF-En), and aerobic exercise training (HF-Ex) groups. Body mass (BM), food and energy intake, plasma analyzes, systolic blood pressure (SBP), physical conditioning, and plasma ACE and ACE2 activity were evaluated. Cardiac morphology, and protein expression of hypertrophy, cardiac metabolism, RAS, and ER stress markers were assessed. Data presented as mean ± standard deviation and analyzed by one-way ANOVA with Holm-Sidak post-hoc. HF group had increased BM and SBP, and developed pathological concentric cardiac hypertrophy, with overactivation of the classic arm of the RAS, and higher ER stress. Both interventions reverted the increase in BM, and SBP, and favored the protective arm of the RAS. Enalapril treatment improved pathological cardiac hypertrophy with partial reversal of the concentric pattern, and slightly attenuated cardiac ER stress. In contrast, aerobic exercise training induced physiological eccentric cardiac hypertrophy, and fully diminished ER stress., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Since most yaks have a long postpartum anestrus period, postpartum anestrus is the main factor affecting the reproductive efficiency of yaks. In this study, the third-generation sequencing technology was used to successfully screen differentially expressed genes (DEGs) and differentially expressed transcripts (DETs) in the ovarian tissues of yaks during estrus and anestrus. The functional references of DEGs and DETs were Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and Clusters of Orthologous Genes database. A total of 1149 DEGs and 2294 DETs were successfully identified. These DEGs and DETs were mainly related to biological processes such as "reproduction", "reproductive process", "metabolic process" and "rhythmic process". Kisspeptin-G protein-coupled receptor was found to be involved in regulating the reproductive cycle of yaks. DEGs and DETs were also related to gonadotropin-releasing hormone (GnRH) signaling pathways such as oocyte meiosis, estrogen signaling pathway, and progesterone-mediated induced oocyte maturation. The results showed that SIRT1 , CSNK1A1 , SLIT3 , INHBA , INSL3 , ZP2 , Clock , BMP15 , Bmal1 , KISS1 , and LCHGR regulate the postpartum quiescent state and the reproductive cycle of yaks. This study will help to further clarify the reproductive mechanism of yaks at the molecular level and provide certain assistance for the development of animal husbandry.

The regulation of mammalian ovarian development involves the coordinated processes of autophagy and apoptosis. The autophagy-related gene ATG7 plays a pivotal role in mediating crosstalk between these pathways. Despite its recognized importance, the specific function of ATG7 in ovarian follicular granulosa cells remains poorly understood. This study aimed to explore the effects of ATG7 overexpression on apoptosis and autophagy in porcine ovarian follicular granulosa cells and thereby provide insights into the interplay between these fundamental cellular mechanisms. An ATG7 overexpression vector was introduced into cells, followed by assessment of cell proliferation using the CCK-8 assay, quantification of related gene expression via real-time quantitative PCR and western blotting, and evaluation of apoptosis using TUNEL staining. ATG7 exhibited a predominant cytoplasmic localization and additional nuclear expression in porcine ovarian follicular granulosa cells. The transfection efficiency of the vector was initially verified, indicating that its overexpression notably increased expression of ATG7 protein. Further investigations confirmed that overexpression of ATG7 inhibited cell proliferation, stimulated autophagy, and promoted apoptosis in these cells. In summary, overexpression of ATG7 influences the viability of porcine ovarian follicular granulosa cells by regulating the interplay between autophagy and apoptosis. This study not only broadens the understanding of functional regulation of autophagy and apoptosis by ATG7, but also sheds light on the intricate mechanisms governing ovarian follicular atresia., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of interest in the manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

The development of ovarian follicles in poultry is a key factor affecting the performance of egg production. Ovarian follicle development is regulated via the Wnt/β-catenin signaling pathway, and β-catenin, encoded by CTNNB1, is a core component of this pathway. In this study, using ovary GCs from laying hens, we investigated the regulatory role of CTNNB1 in steroid synthesis. We found that CTNNB1 significantly regulates the expression of StAR and CYP11A1 (key genes related to progesterone synthesis) and the secretion of progesterone (P4). Furthermore, simultaneous overexpression of CTNNB1 and SF1 resulted in significantly higher levels of CYP11A1 and secretion of P4 than in cells overexpressing CTNNB1 or SF1 alone. We also found that in GCs overexpressing SF1 , levels of CYP11A1 and secreted P4 were significantly greater than in controls. Silencing of CYP11A1 resulted in the inhibition of P4 secretion while overexpression of SF1 in CYP11A1- silenced cells restored P4 secretion to normal levels. Together, these results indicate that synergistic cooperation between the β-catenin and SF1 regulates progesterone synthesis in laying hen ovarian hierarchical granulosa cells to promote CYP11A1 expression.

This study analyzed the transcriptional changes in primary human corneal stromal fibroblasts (hCSFs) grown under quiescent (serum-free) and proliferating (serum-supplemented) culture conditions to identify genes, pathways, and protein‒protein interaction networks influencing corneal repair and regeneration. Primary hCSFs were isolated from donor human corneas and maintained in serum-free or serum-laden conditions. RNA was extracted from confluent cultures using Qiagen kit and subjected to RNA sequencing (RNAseq) analysis. Differential gene expression (DGE) and pathway enrichment analyses were conducted using DESeq2 and Gene Set Enrichment Analysis (GSEA), respectively. Protein‒protein interaction (PPI) networks were created exploiting the STRING database and analyzed with Cytoscape and the cytoHubba plugin. RNA-seq revealed 5,181 genes that were significantly differentially expressed/changed among the 18,812 annotated genes (p value ˂0.05). A cutoff value of a log2-fold change of ±1.5 or greater was used to identify 674 significantly upregulated and 771 downregulated genes between quiescent and proliferating hCSFs. Pathway enrichment analysis revealed significant changes in genes linked to cell cycle regulation, inflammatory, and oxidative stress response pathways, such as E2F Targets, G2M Checkpoint, and MYC Targets, TNFA signaling via NF-kB, and oxidative phosphorylation. Protein-protein interaction network analysis highlighted critical hub genes. The FGF22, CD34, ASPN, DPT, LUM, FGF10, PDGFRB, ECM2, DCN, VEGFD, OMD, OGN, ANGPT1, CDH5, and PRELP were upregulated, whereas genes linked to cell cycle regulation and mitotic progression, such as BUB1, TTK, KIF23, KIF11, BUB1B, DLGAP5, NUSAP1, CCNA2, CCNB1, BIRC5, CDK1, KIF20A, AURKB, KIF2C, and CDCA8, were downregulated. The RNA sequences and gene count files have been submitted to the Gene Expression Omnibus (accession # GSE260476). Our study provides a comprehensive information on the transcriptional and molecular changes in hCSFs under quiescent and proliferative conditions and highlights key pathways and hub genes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Ltd.)

Spermatogenesis in eukaryotes is a process that occurs within a very narrow temperature threshold, typically not exceeding 36 °C. SPO11 was isolated from the temperature-sensitive mutant receptor of Saccharomyces cerevisiae and is thought to be the only protein that functions during meiosis. This suggested that SPO11 may be the key protein that influenced the temperature of spermatogenesis not exceeding 36 °C. Elevated temperatures typically damage the spermatogenic cells. Birds have a core body temperature of 41-42 °C, and their testis are located inside their bodies, providing an alternative perspective to investigate the potential impact of temperature threshold on spermatogenesis. The objective of this study was to ascertain whether elevated ambient temperatures affect spermatogenesis in birds and whether SPO11 is the key gene affecting the temperature threshold for spermatogenesis. STRA8, SCP3, SPO11, γ-H2AX, and RAD51 were all crucial components in the process of meiotic initiation, synapsis, DNA double-strand break (DSB) induction, homologous chromosome crossover recombination, and repair of DSB. In this study, 39-day-old Japanese quail were subjected to heat stress (HS) at 38 °C for 8 h per day for 3 (3d HS) and 13 (13d HS) consecutive days and analyzed the expression of meiotic signaling molecules (STRA8, SCP3, SPO11, γ-H2AX, and RAD51) using molecular biology techniques, including Immunohistochemistry (IHC), Western Blot (WB), and Real-time Quantitative Polymerase Chain Reaction (qRT-PCR). We found that spermatogenesis was normal in both groups exposed to HS. Meiotic signaling molecules were expressed normally in the 3d HS group. All detected signaling molecules were normally expressed in the 13d HS group, except for SPO11, which showed a significant increase in expression, indicating that SPO11 was temperature-sensitive. We examined the localized expression of each meiotic signaling molecule in quail testis, explored the temperature sensitivity of SPO11, and determined that quail testis can undergo normal spermatogenesis at ambient temperatures exceeding 36 °C. This study concluded that SPO11 is not the key protein influencing spermatogenesis in birds. These findings enhance our understanding of avian spermatogenesis., Competing Interests: Declaration of competing interest The authors have no competing interests to declare., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Pyometra is a life-threatening disease, the severity of which depends on cervical patency status. This study investigated cervical inflammation status as well as the expression patterns and localization of aquaporin (AQP1, AQP2, AQP3, AQP5, and AQP9), and hormone receptors in cervical tissue that influences canine pyometra. Of the 36 animals enrolled in the study, 24 were diagnosed with pyometra and separated into two groups: open cervix pyometra and close cervix pyometra, while 12 healthy animals presented for elective ovariohysterectomies were allocated into the control group. Surgical treatment was performed for treatment of pyometra. After each operation, cervix samples were collected and analyzed for AQP and hormone receptor expression patterns determined by qPCR and protein expression by means of immunohistochemistry. Blood samples were also collected to determine serum progesterone concentrations. AQP9 expression was downregulated approximately 3-fold while and PGR expression was downregulated more than 2 fold in both pyometra groups compared to the control group. AQP3 and AQP5 gene expression levels were upregulated more than 3 fold in the open-cervix pyometra group than the closed-cervix pyometra group (P < 0.05). This is the first study to describe the expression patterns and immunolocalization of AQPs in canine cervical tissue based on pyometra patency status and to report AQP3 and AQP5 expression in cervical tissue linked to cervical patency., Competing Interests: Declaration of competing interest The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: The long-term evaluation of RPE65 gene augmentation initiated in middle-aged RPE65 mutant dogs previously uncovered notable inter-animal and intra-retinal variations in treatment efficacy. The study aims to gain deeper insights into the status of mutant retinas and assess the treatment impact., Methods: Immunohistochemistry utilizing cell-specific markers and reverse transcription-quantitative PCR (RT-qPCR) analysis were conducted on archival retinal sections from normal and RPE65 mutant dogs., Results: Untreated middle-aged mutant retinas exhibited marked downregulation in the majority of 20 examined genes associated with key retinal pathways. These changes were accompanied by a moderate increase in microglia numbers, altered expression patterns of glial-neuronal transmitter recycling proteins, and gliotic responses in Müller glia. Analysis of advanced-aged mutant dogs revealed mild outer nuclear layer loss in the treated eye compared to moderate loss in the corresponding retinal regions of the untreated control eye. However, persistent Müller glial stress response along with photoreceptor synapse loss were evident in both treated and untreated eyes. Photoreceptor synaptic remodeling, infrequent in treated regions, was observed in all untreated advanced-aged retinas, accompanied by a progressive increase in microglial cells indicative of ongoing inflammation. Interestingly, about half of the examined genes showed similar expression levels between treated and untreated advanced-aged mutant retinas, with some reaching normal levels., Conclusions: Gene expression data suggest a shift from pro-degenerative mechanisms in middle-aged mutant retinas to more compensatory mechanisms in preserved retinal regions at advanced stages, despite ongoing degeneration. Such shift, potentially attributed to a number of surviving resilient cells, may influence disease patterns and treatment outcomes.

Endomucin (EMCN) is a 261 amino acid transmembrane glycoprotein that is highly expressed by venous and capillary endothelial cells where it plays a role in VEGF-mediated angiogenesis and regulation of immune cell recruitment. However, it is better known as a histological marker, where it has become widespread due to the commercial availability of high-quality antibodies that work under a wide range of conditions and in many tissues. The specificity of EMCN staining has been well-validated in retinal vessels, but while it has been used extensively as a marker in other tissues of the eye, including the choroid, the pattern of expression has not been described in detail. Here, in addition to endothelial expression in the choriocapillaris and deeper vascular layers, we characterize a population of EMCN-positive perivascular cells in the mouse choroid that did not co-localize with cells expressing other endothelial markers such as PECAM1 or PODXL. To confirm that these cells represented a new population of EMCN-expressing stromal cells, we then performed single cell RNA sequencing in choroids from adult wild-type mice. Analysis of this new dataset confirmed that, in addition to endothelial cells, Emcn mRNA expression was present in choroidal pericytes and a subset of fibroblasts, but not vascular smooth muscle cells. Besides Emcn, no known endothelial gene expression was detected in these cell populations, confirming that they did not represent endothelial-stromal doublets, a common technical artifact in single cell RNA seq datasets. Instead, choroidal Emcn-expressing fibroblasts exhibited high levels of chemokine and interferon signaling genes, while Emcn-negative fibroblasts were enriched in genes encoding extracellular matrix proteins. Emcn expressing fibroblasts were also detected in published datasets from mouse brain and human choroid, suggesting that stromal Emcn expression was not unique to the choroid and was evolutionarily conserved. Together, these findings highlight unique fibroblast and pericyte populations in the choroid and provide new context for the role of EMCN in the eye., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Thyroid hormones (THs) play important roles in growth, development, morphogenesis, reproduction, and so on. They are mainly meditated by binding to thyroid hormone receptors (TRs) in vertebrates. As important members of the nuclear receptor superfamily, TRs and their ligands are involved in many biological processes. To investigate the potential roles of TRs in the gonadal differentiation and sex change, we cloned and characterized the TRs genes in protogynous rice field eel (Monopterus albus). In this study, three types of TRs were obtained, which were TRαA, TRαB and TRβ, encoding preproproteins of 336-, 409- and 415-amino acids, respectively. Multiple alignments of the three putative TRs protein sequences showed they had a higher similarity. Tissue expression analysis showed that TRαA mainly expressed in the gonad, while TRαB and TRβ in the brain. During female-to-male sex reversal, the expression levels of all the three TRs showed a similar trend of increase followed by a decrease in the gonad. Intraperitoneal injection of triiodothyronine (T3) stimulated the expression of TRαA and TRαB, while it had no significant change on the expression of TRβ in the ovary. Gonadotropin-releasing hormone analogue (GnRHa) injection also significantly upregulated the expression levels of TRαA and TRαB after 6 h, while it had no significant effect on TRβ. These results demonstrated that TRs were involved in the gonadal differentiation and sex reversal, and TRα may play more important roles than TRβ in reproduction by the regulation of GnRHa in rice field eel., (© 2024 Wiley Periodicals LLC.)

Purpose: Limbal stem/progenitor cells (LSCs) continuously proliferate and differentiate to replenish the corneal epithelium and play a vital role in corneal function and normal vision. A previous study revealed that paired box 6 (PAX6) is a master transcription factor involved in determining the fate of corneal epithelial cells (CECs). However, the molecular events downstream of PAX6 remain largely unknown. In this study, we aimed to clarify the regulation network of PAX6 in driving CEC differentiation., Methods: An air-liquid culture system was used to differentiate LSCs into mature CECs. Specific targeting PAX6 short-hairpin RNAs were used to knock down PAX6 in LSC. RNA sequencing (RNA-seq) was used to analyze shPAX6-transfected CECs and CEC differentiation-associated genes to identify the potential downstream targets of PAX6. RNA-seq analysis, quantitative real-time PCR, and immunofluorescence staining were performed to clarify the function of WNK lysine deficient protein kinase 2 (WNK2), a downstream target of PAX6, and its relationship with corneal diseases., Results: WNK2 expression increased during CEC differentiation and decreased upon PAX6 depletion. The distribution of WNK2 was specifically limited to the central corneal epithelium and suprabasal layer of the limbus. Knockdown of WNK2 impaired the expression of CEC-specific markers (KRT12, ALDH3A1, and CLU), disrupted the corneal differentiation process, and activated the terms of keratinization, inflammation, and cell proliferation, consistent with PAX6-depleted CEC and published microbial keratitis. Thus, aberrant expression of WNK2 was linked to corneal ulcers., Conclusions: As a downstream target of PAX6, WNK2 plays an essential role in corneal epithelial cell differentiation and maintenance of corneal homeostasis.

Dormancy is an essential ecological characteristic for the survival of organisms that experience harsh environments. Although factors that initiate dormancy vary, suppression or cessation of feeding activities are common among taxa. To distinguish between extrinsic and intrinsic causes of metabolic reduction, we focused on estivation, which occurs in summer when the feeding activity is generally enhanced. Sand lances (genus Ammodytes) are a unique marine fish with a long estivation period from early summer to late autumn. In the present study, we aimed to elucidate the control mechanisms of estivation in western sand lance (A. japonicus), and firstly examined behavioral changes in 8 months including a transition between active and dormant phases. We found that swimming/feeding behavior gradually decreased from June, and completely disappeared by late August, indicating all individuals had entered estivation. Next, we focused on leptin, known as a feeding suppression hormone in various organisms, and examined leptin-A gene (AjLepA) expression in the brain that may regulate the seasonal behavioral pattern. AjLepA expression decreased after 7 days of fasting, suggesting that leptin has a function to regulate feeding in this species. The monthly expression dynamics of AjLepA during the feeding (active) and non-feeding (estivation) periods showed that the levels gradually increased with the onset of estivation and reached its peak when all the experimental fish had estivated. The present study suggests that the suppression of feeding activity by leptin causes shift in the physiological modes of A. japonicus before estivation., (© 2024 Wiley Periodicals LLC.)

Nerve growth factor (NGF) has long been known as the main ovulation-inducing factor in induced ovulation species, however, recent studies suggested the NGF role also in those with spontaneous ovulation. The first aim of this study was to evaluate the presence and gene expression of NGF and its cognate receptors, high-affinity neurotrophic tyrosine kinase 1 receptor (NTRK1) and low-affinity p75 nerve growth factor receptor (p75NTR), in the ram genital tract. Moreover, the annual trend of NGF seminal plasma values was investigated to evaluate the possible relationship between the NGF production variations and the ram reproductive seasonality. The presence and expression of the NGF/receptors system was evaluated in the testis, epididymis, vas deferens ampullae, seminal vesicles, prostate, and bulbourethral glands through immunohistochemistry and real-time PCR (qPCR), respectively. Genital tract samples were collected from 5 adult rams, regularly slaughtered at a local abattoir. Semen was collected during the whole year weekly, from 5 different adult rams, reared in a breeding facility, with an artificial vagina. NGF seminal plasma values were assessed through the ELISA method. NGF, NTRK1 and p75NTR immunoreactivity was detected in all male organs examined. NGF-positive immunostaining was observed in the spermatozoa of the germinal epithelium, in the epididymis and the cells of the secretory epithelium of annexed glands, NTRK1 receptor showed a localization pattern like that of NGF, whereas p75NTR immunopositivity was localized in the nerve fibers and ganglia. NGF gene transcript was highest (p < 0.01) in the seminal vesicles and lowest (p < 0.01) in the testis than in the other tissues. NTRK1 gene transcript was highest (p < 0.01) in the seminal vesicles and lowest (p < 0.05) in all the other tissues examined. Gene expression of p75NTR was highest (p < 0.01) in the seminal vesicles and lowest (p < 0.01) in the testis and bulbourethral glands. NGF seminal plasma concentration was greater from January to May (p < 0.01) than in the other months. This study highlighted that the NGF system was expressed in the tissues of all the different genital tracts examined, confirming the role of NGF in ram reproduction. Sheep are short-day breeders, with an anestrus that corresponds to the highest seminal plasma NGF levels, thus suggesting the intriguing idea that this factor could participate in an inhibitory mechanism of male reproductive activity, activated during the female anestrus., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

GATA4 plays a pivotal role in the reproductive processes of mammals. However, the research on GATA4 in goat ovary is limited. This study aimed to study the expression and function of GATA4 in goat ovary. Utilizing real-time PCR and western blot analysis, we studied the expression and regulatory mechanisms of GATA4 in goat ovary and granulosa cells (GCs). We found that GATA4 was expressed in all follicle types in the goat ovary, with significantly higher levels in GCs of larger follicles (>3 mm) compared to those in smaller follicles (<3 mm). Additionally, we demonstrated that human chorionic gonadotrophin (hCG) induced GATA4 mRNA expression via the activation of PKA, MEK, p38 MAPK, PKC, and PI3K pathways in vitro. Our study also showed that hCG suppressed the levels of miR-200b and miR-429, which in turn directly target GATA4, thereby modulating the basal and hCG-induced expression of GATA4. Functionally, we examined the effect of siRNA-mediated GATA4 knockdown on cell proliferation and hormone secretion in goat GCs. Our results revealed that knockdown of GATA4, miR-200b, and miR-429 suppressed cell proliferation. Moreover, knockdown of GATA4 decreased estradiol and progesterone production by inhibiting the promoter activities of CYP11A1, CYP19A1, HSD3B, and StAR. Collectively, our findings suggest a critical involvement of GATA4 in regulating goat GC survival and steroidogenesis., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Decreased oocyte quality is a significant contributor to the decline in female fertility that accompanies aging in mammals. Oocytes rely on mRNA stores to support their survival and integrity during the protracted period of transcriptional dormancy as they await ovulation. However, the changes in mRNA levels and interactions that occur during porcine oocyte maturation and aging remain unclear. In this study, the mRNA expression profiles of porcine oocytes during the GV, MII, and aging (24 h after the MII stage) stages were explored by transcriptome sequencing to identify the key genes and pathways that affect oocyte maturation and postovulatory aging. The results showed that 10,929 genes were coexpressed in porcine oocytes during the GV stage, MII stage, and aging stage. In addition, 3037 genes were expressed only in the GV stage, 535 genes were expressed only in the MII stage, and 120 genes were expressed only in the aging stage. The correlation index between the GV and MII stages (0.535) was markedly lower than that between the MII and aging stages (0.942). A total of 3237 genes, which included 1408 upregulated and 1829 downregulated genes, were differentially expressed during porcine oocyte postovulatory aging (aging stage vs. MII stage). Key functional genes, including ATP2A1, ATP2A3, ATP2B2, NDUFS1, NDUFA2, NDUFAF3, SREBF1, CYP11A1, CYP3A29, GPx4, CCP110, STMN1, SPC25, Sirt2, SYCP3, Fascin1/2, PFN1, Cofilin, Tmod3, FLNA, LRKK2, CHEK1/2, DDB1/2, DDIT4L, and TONSL, and key molecular pathways, such as the calcium signaling pathway, MAPK signaling pathway, TGF-β signaling pathway, PI3K/Akt signaling pathway, FoxO signaling pathway, gap junctions, and thermogenesis, were found in abundance during porcine postovulatory aging. These genes are mainly involved in the regulation of many biological processes, such as oxidative stress, calcium homeostasis, mitochondrial function, and lipid peroxidation, during porcine oocyte postovulatory aging. These results contribute to a more in-depth understanding of the biological changes, key regulatory genes and related biological pathways that are involved in oocyte aging and provide a theoretical basis for improving the efficiency of porcine embryo production in vitro and in vivo., Competing Interests: Declaration of competing interest None., (Copyright © 2024. Published by Elsevier Inc.)

Although anti-Müllerian hormone (AMH) is involved in the regulation of granulosa cell function in female animals, its role in tissues other than ovarian follicles remains poorly understood. It has also been suggested that cows with high circulating AMH concentrations have increased fertility; however, the mechanism has not been elucidated. This study was conducted to identify the presence of the AMH-signaling system and its target cells in the bovine corpus luteum formed from an ovulated follicle. Immunoblotting revealed that the proteolytically cleaved C-terminal region in AMH (AMH C ), a biologically active peptide, was present in trace amounts in the early corpus luteum and significantly increased during the mid to regressed stages. AMH C and cleaved N-terminal region (AMH N ) in AMH generate a noncovalent isoform that improves the activity of AMH signaling. An immunohistochemical analysis revealed that AMH C , AMH N , and type II AMH receptor (AMHR2) were localized to luteal cells during the entire estrous cycle. AMH in the corpus luteum seemed to be newly synthesized since AMH expression was detected. These findings suggest that AMH signaling is involved in the regulation of luteal cell function through an autocrine and post-translational processing mechanism. The level of AMHR2 and mRNA expression of AMHR2 and type I AMH receptors (activin-like kinase 2, 3, and 6) were highest in the mid stage. Thus, AMH signaling in the corpus luteum may also be regulated by changes in the receptor levels. Since the transforming growth factor-beta superfamily, to which AMH belongs, is a multifunctional polypeptide growth factor, further studies are needed to evaluate whether AMH signaling has a role in facilitating or inhibiting luteal cell functions., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Follicular fluid (FF) is rich in extracellular vesicles (EVs). EVs carries a variety of miRNA involved in regulating follicular development, the function of cells in follicles, primordial follicular formation, follicular recruitment and selection, follicular atresia, oocyte communication, granulosa cells (GCs) function and luteinization and other biological processes of follicular development. Previous studies in our laboratory have shown that bovine follicular fluid (bFF) high density-small extracellular vesicles (HD-sEVs)-miRNA was enriched in autophagy-related pathways. However, the mechanism of bFF EVs carrying miRNA regulating GCs autophagy is not clear. Thus, this study carried out a series of studies on the previous HD-sEVs sequencing data and miR-128-3p contained in bFF HD-sEVs. A total of 38 differentially expressed genes were detected by RNA-Seq after overexpression of miR-128-3p in bovine GCs (bGCs). Through cell transfection, Western blot (WB) and Immunofluorescence (IF), it was proved that overexpression of miR-128-3p could promote the expression of LC3 (microtubule-associated protein I light chain 3), inhibit p62, promote the number of autophagosome, promote the formation of autophagy lysosome and autophagy flow, and activate bGCs autophagy. MiR-128-3p inhibitor significantly inhibited the expression of LC3 and monodansylcadaverine (MDC) in bGCs, and promoted the expression of autophagy substrate p62, indicating that HD-sEVs-miR-128-3p could activate bGCs autophagy. In addition, through double luciferase assay, bioinformatics analysis, WB and RT-qPCR, it was concluded that bFF HD-sEVs-miR-128-3p could target TFEB (transcription factor EB) and FoxO4 (Forkhead box O4) and activate GCs autophagy., Competing Interests: Declaration of competing interest The authors have no disclosure of conflicts of interest related to this investigation., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Dielectric barrier discharge (DBD) plasma regulates the levels of reactive oxygen species (ROS), which are critical for sperm quality. MicroRNAs (miRNAs) are non-coding single-stranded RNA molecules encoded by endogenous genes, which regulate post-transcriptional gene expression in animals. At present, it is unknown whether DBD plasma can regulate sperm ROS levels through miRNAs. To further understand the regulatory mechanism of DBD plasma on sperm ROS levels, miRNAs in fresh boar spermatozoa were detected using Illumina deep sequencing technology. We found that 25 known miRNAs and 50 novel miRNAs were significantly upregulated, and 14 known miRNAs and 74 novel miRNAs were significantly downregulated in DBD plasma-treated spermatozoa. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that target genes of differentially expressed miRNAs were involved in many activities and pathways associated with antioxidants. We verified that DBD plasma significantly increased boar sperm quality and reduced ROS levels. These results suggest that DBD plasma can improve sperm quality by regulating ROS levels via miRNAs. Our findings provide a potential strategy to improve sperm quality through miRNA-targeted regulation of ROS, which helps to increase male reproduction and protect cryopreserved semen in clinical practice., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Purpose: This study aims to investigate the relationship among STRA6, circadian rhythm, and choroidal neovascularization (CNV) formation, as well as the regulatory mechanism of STRA6 in CNV under circadian rhythm disturbances., Methods: C57BL/6J male mice (aged 6 weeks) were randomly divided into control and jet lag groups (using a time shift method every 4 days to disrupt the molecular clock's capacity to synchronize with a stable rhythm). A laser-induced CNV model was established in both the control and the jet lag group after 2 weeks of jet lag. The size of CNV lesions and vascular leakage were detected by morphological and imaging examination on the seventh day post laser. STRA6 was screened by full transcriptome sequencing. Bioinformatics analysis was conducted to assess the variation and association of STRA6 in the GSE29801 dataset. The effects of STRA6 were evaluated both in vivo and in vitro. The pathway mechanism was further elucidated and confirmed through immunofluorescence of paraffin sections and Western blotting., Results: The disturbance of circadian rhythm promotes the formation of CNV. Patients with age-related macular degeneration (AMD) exhibited higher levels of STRA6 expression compared to the control group, and STRA6 was enriched in pathways related to angiogenesis. In addition, CLOCK and BMAL1, which are initiators that drive the circadian cycle, had regulatory effects on STRA6. Knocking down STRA6 reversed the promotion of CNV formation caused by circadian rhythm disturbance in vivo, and it also affected the proliferation, migration, and VEGF secretion of RPE cells without circadian rhythm in vitro, as well as impacting endothelial cells. Through activation of the JAK2/STAT3/VEGFA signaling pathway in unsynchronized RPE cells, STRA6 promotes CNV formation., Conclusions: This study suggests that STRA6 reduces CNV production by inhibiting JAK2/STAT3 phosphorylation after circadian rhythm disturbance. The results suggest that STRA6 may be a new direction for the treatment of AMD.

Fragile X Syndrome (FXS), the most common inherited form of human intellectual disability, is a monogenic neurodevelopmental disorder caused by a loss-of-function mutation of the FMR1 gene. FMR1 is encoding the Fragile X Messenger Ribonucleo Protein (FMRP) an RNA-binding protein that regulates the translation of synaptic proteins. The absence of FMRP expression has many important consequences on synaptic plasticity and function, leading to the FXS clinical phenotype. Over the last decade, a visual neurosensorial phenotype had been described in the FXS patients as well as in the murine model (Fmr1 -/y mice), characterized by retinal deficits associated to retinal perception alterations. However, although the transcriptomic profile in the absence of FMRP has been studied in the cerebral part of the central nervous system (CNS), there are no actual data for the retina which is an extension of the CNS. Herein, we investigate the transcriptomic profile of mRNA from whole retinas of Fmr1 -/y mice. Interestingly, we found a specific signature of Fmrp absence on retinal mRNA expression with few common genes compared to other brain studies. Gene Ontology on these retinal specific genes demonstrated an enrichment in retinal development genes as well as in synaptic genes. These alterations could be linked to the reported retinal phenotype of the FXS condition. In conclusion, we describe for the first time, retinal-specific transcriptomic changes in the absence of FMRP., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

In homeostasis, which plays an important role in the proper functioning and maintenance of the internal functioning of the body, kidneys play a key role in being responsible for the proper homeostasis of glucose. Among glucose transporters, sodium-dependent glucose co-transporters (SGLTs) have a major role in the kidney's ability to reabsorb glucose. Although the localization of these transporters has been extensively studied in mammals, there are still gaps in knowledge of the localization of SGLTs in birds of different age groups. The aim of this study was to immunolocalize in kidneys of hen chickens of different ages the sodium-dependent glucose co-transporters SGLT1 and SGLT2, comparing the localization between different age groups. The kidneys derived from 32 hen chickens (Gallus gallus domesticus) were devided equally into four age groups: 3, 7, 14, and 20 day old broilers, 8 indivuals in each group. The polyclonal primary antibodies Rabbit anti- SGLT1 and Rabbit anti-SGLT2 (Abcam, UK) were used together with the corresponding IHC kit (Abcam, UK). The results were visualized photographically using an AxioCam HRc camera (Germany) connected to a Zeiss Axioplan-2 Imaging microscope (Germany). The study revealed similar immunolocalization of SGLT1 and SGLT2 in the apical part of cells of proximal renal tubules in hen chickens' kidneys in all age groups. Strong staining of SGLT2 was noted also in the cytoplasm of epithelial cells of the proximal straight and convoluted tubules. Based on our study, the kidney tissue of newly hatched chickens is ready immediately after hatching for glucose reabsorption and transport, similarily to that of three-week-old chicks., (© 2024 The Authors. This is an open access article under the CC BY-NC-ND 4.0 license (https://creativecommons.org/licenses/by-nc-nd/4.0/deed.en), which allows re-users to copy and distribute the material in any medium or format in unadapted form and for noncommercial purposes, and only so long as attribution is given to the creator.)

Age-related reduction in spine density, synaptic marker expression, and synaptic efficiency are frequently reported. These changes provide the cellular and molecular basis for the cognitive decline characteristic for old age. Nevertheless, there are several approaches that have the potential to ameliorate these processes and improve cognition, caloric restriction being one of the most promising and widely studied. While lifelong caloric restriction is known for its numerous beneficial effects, including improved cognitive abilities and increased expression of proteins essential for synaptic structure and function, the effects of late-onset and/or short-term CR on synaptic plasticity have yet to be investigated. We have previously documented that the effects of CR are strongly dependent on whether CR is initiated in young or old subjects. With this in mind, we conducted a long-term study in aging Wistar rats to examine changes in the expression of several key synaptic markers under the regimen of CR started at different time points in life. We found a significant increase in the expression of both presynaptic and postsynaptic markers. However, taking into account previously reported changes in the behavior detected in these animals, we consider that this increase cannot represent beneficial effect of CR., (© 2024 International Union of Biochemistry and Molecular Biology.)

The dog retina contains a central macula-like region, and there are reports of central retinal disorders in dogs with shared genetic etiologies with humans. Defining central/peripheral gene expression profiles may provide insight into the suitability of dogs as models for human disorders. We determined central/peripheral posterior eye gene expression profiles in dogs and interrogated inherited retinal and macular disease-associated genes for differential expression between central and peripheral regions. Bulk tissue RNA sequencing was performed on 8 mm samples of the dog central and superior peripheral regions, sampling retina and retinal pigmented epithelium/choroid separately. Reads were mapped to CanFam3.1, read counts were analyzed to determine significantly differentially expressed genes (DEGs). A similar analytic pipeline was used with a published bulk-tissue RNA sequencing human dataset. Pathways and processes involved in significantly DEGs were identified (Database for Annotation, Visualization and Integrated Discovery). Dogs and humans shared the extent and direction of central retinal differential gene expression, with multiple shared biological pathways implicated in differential expression. Many genes implicated in heritable retinal disorders in dogs and humans were differentially expressed between central and periphery. Approximately half of genes associated with human age-related macular degeneration were differentially expressed in human and dog tissues. We have identified similarities and differences in central/peripheral gene expression profiles between dogs and humans which can be applied to further define the relevance of dogs as models for human retinal disorders., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Sexually dimorphic behaviors are often regulated by gonadal steroid hormones. Species diversity in behavioral sex differences may arise as expression of genes mediating steroid action in brain regions controlling these behaviors evolves. The electric communication signals of apteronotid knifefishes are an excellent model for comparatively studying neuroendocrine regulation of sexually dimorphic behavior. These fish produce and detect weak electric organ discharges (EODs) for electrolocation and communication. EOD frequency (EODf), controlled by the medullary pacemaker nucleus (Pn), is sexually dimorphic and regulated by androgens and estrogens in some species, but is sexually monomorphic and unaffected by hormones in other species. We quantified expression of genes for steroid receptors, metabolizing enzymes, and cofactors in the Pn of two species with sexually dimorphic EODf (Apteronotus albifrons and Apteronotus leptorhynchus) and two species with sexually monomorphic EODf ("Apteronotus" bonapartii and Parapteronotus hasemani). The "A." bonapartii Pn expressed lower levels of androgen receptor (AR) genes than the Pn of species with sexually dimorphic EODf. In contrast, the P. hasemani Pn robustly expressed AR genes, but expressed lower levels of genes for 5α-reductases, which convert androgens to more potent metabolites, and higher levels of genes for 17β-hydroxysteroid dehydrogenases that oxidize androgens and estrogens to less potent forms. These findings suggest that sexual monomorphism of EODf arose convergently via two different mechanisms. In "A." bonapartii, reduced Pn expression of ARs likely results in insensitivity of EODf to androgens, whereas in P. hasemani, gonadal steroids may be metabolically inactivated in the Pn, reducing their potential to influence EODf., Competing Interests: Declaration of competing interest The authors have no financial or personal competing interests that could affect the objectivity of this study., (Copyright © 2024 Elsevier Inc. All rights reserved.)

First, significantly higher mate-finding success was found under light condition than under constant darkness condition in Phauda flammans , a typical diurnal moth. We speculate that mate-finding behavior in P. flammans may be influenced by the light-sensitive opsin genes Long wavelength opsin ( PfLW ), Ultraviolet opsin ( PfUV ) and Blue opsin ( PfBL ), which are potentially regulated by both light-cues and endogenous circadian rhythms. Second, the circadian clock genes Period ( PfPer ), Timeless ( PfTim ), Cryptochrome1 ( PfCry1 ), Cryptochrome2 ( PfCRY2 ), Cryptochrome3 ( PfCry-like ), Clock ( PfClk ), Cycle ( PfCyc ), Vrille ( PfVri ), and Slimb ( PfSli ) were identified in P. flammans . Third, circadian rhythms in the relative expression levels of opsin and circadian clock genes were demonstrated via quantitative real-time PCR analysis, with peak expression coinciding with the mate-finding peak. Notably, the relative expression of PfLW in males P. flammans was significantly higher than that in females P. flammans at the mate-finding peaks Zeitgeber time (ZT) 8 and ZT 10 under light, while the expression of the opsin gene PfBL showed a similar pattern at ZT 10 under light. Additionally, the expression of the clock gene PfCry-like was significantly higher in males than in females at ZT 8 and ZT 10 under light, while PfPer , PfTim , PfClk and PfCyc exhibited similar male-biased expression patterns at ZT 10 under light. Conversely, PfCry1 and PfVri expression was significantly higher in females than in male at ZT 8 under light. In conclusion, sex differences were detected in the expression of opsin and circadian clock genes, which indicated that light-mediated regulation of these genes may contribute to the daytime mate-finding behavior of P. flammans .

Leptins and other related genes have been proven to play vital roles in food intake, weight control, and other life activities. While the function of leptins in yellowtail kingfish (Seriola lalandi) has not yet been explored, in the present study, we investigated the structure and preliminary function of four leptin-related genes in S. lalandi. In detail, the sequence of two leptin genes (lepa and lepb), one leptin receptor gene (lepr), and one leptin receptor overlapping transcript (leprot) gene were obtained by homology cloning and RACE methods, in which lepa and lepb have similar structure. Moreover, homologous sequence alignment and evolutionary analysis of all four genes were clustered with Seriola dumerili. The tissue distribution of these four genes in thirteen tissues of yellowtail kingfish was detected by RT-qPCR. Both lepa and leprot were highly expressed in the brain and ovary, while lepb was highly expressed in the pituitary, gill, muscle, and ovary; lepr was highly expressed in the gill, kidney, and ovary. Additionally, these four genes also played roles in embryo development and early growth and development of larvae and juveniles of yellowtail kingfish. Finally, the function of leptin and leptin-related genes was investigated during fasting and re-feeding adaption of yellowtail kingfish. The results showed that these four genes have different regulation functions in five tissues; for example, the mRNA levels of lepa, lepr, and leprot in the brain decreased during fasting and immediately increased after re-feeding, while the mRNA level of lepb did not show significant fluctuation during starvation but significantly lowered after re-feeding. However, lepa and lepb mRNA levels were significantly elevated during fasting and returned to control levels after re-feeding, and there were no significant changes in the expression of lepr and leprot in the liver during fasting and after re-feeding. Moreover, the body mass of fish in the experimental group was measured, and compensatory growth was found after the resumption of feeding. These results suggested that leptin and receptor genes play different functions in different tissues to regulate the physiological state of fish in food deficiency and gain processes., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The ALDH1A1 gene encodes a cytoplasmic member of the aldehyde dehydrogenase 1 family, which plays an important role in regulating animal reproductive performance, including estrus cycle and embryonic development. The aim of this study was to characterize ALDH1A1 activity in ovaries of 3-5 year-old yaks and to determine its effects on cell proliferation, apoptosis, and progesterone secretion in luteal cells (LCs). The coding sequence (CDS) of the ALDH1A1 gene was cloned by reverse transcription-PCR and immunohistochemical analysis was used to confirm localization of the ALDH1A1 protein in the ovary. To assess the activity of ALDH1A1 in regulating progesterone secretion, si-ALDH1A1 was transfected into LCs in vitro and progesterone levels in LC supernatants were measured by ELISA. The interference efficiency was assessed by real-time quantitative PCR (RT-qPCR) and immunofluorescence staining, and cell proliferation and apoptosis were evaluated by EdU and TUNEL staining, respectively. The cloned ALDH1A1 sequence contained 1462 bp, encoding 487 amino acids. Immunohistochemical analysis showed that ALDH1A1 protein expression, which was significantly higher in LCs, was mainly found in antral follicles and the corpus luteum (CL). The expression of ALDH1A1 mRNA in LCs was effectively inhibited by si-ALDH1A1transfection, and progesterone secretion was markedly decreased along with the significant down-regulation of progesterone pathway-related genes, STAR, CYP11A1, CYP19A1, CYP17A1, 3β-HSD, and HSD17B1. Knockdown of ALDH1A1 mRNA expression decreased cell proliferation and increased apoptosis in LCs. The mRNA expression of the proliferation-related genes, PCNA, CCND1, CCNB1 and CDC25A, was significantly down-regulated, while expression of the apoptosis-promoting CASP3 gene was significantly increased. In summary, we characterized the yak ALDH1A1 gene and revealed that ALDH1A1 knockdown promoted apoptosis, repressed cell proliferation, and decreased progesterone secretion by yak LCs, potentially by regulating the mRNA expression of genes related to proliferation, apoptosis, and progesterone synthesis and secretion., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Reducing the dietary crude protein (CP) could effectively reduce pressure on protein ingredient supplies. However, few data have been reported about the extent to which CP can be reduced and whether limiting the use of soybean meal leads to electrolyte imbalance. In this experiment, using the low protein (LP) diet [2% lower than NRC (2012)], seventy-two piglets (35 days old) were randomly divided into 2 groups with 6 replicates of 6 piglets each: CON group (CP = 18.5%) and LP group (CP = 16.5%), to investigate the effect of the LP diet on electrolyte balance, acid-base balance, intestinal structure and amino acid transport in piglets. The results revealed that the LP diet decreased the average daily gain and dietary CP digestibility, and damaged the villi structure of the small intestine. Compared with the CON diet, the potassium content decreased and the chlorine content increased in the LP diet, and similar trends were shown in piglet serum. The arterial pH, pCO 2 , HCO 3 - , and base excess of piglets in the LP group were lower than those in the CON group, while pO 2 was higher than those in the CON group. Interestingly, the LP diet significantly increased the lysine content in piglet serum and significantly decreased the levels of arginine, leucine, and glutamic acid. Furthermore, the LP diet significantly affected the expression of some amino acid transport vectors (B 0 AT1, EAAC1, and y + LAT1). In summary, these findings suggested that the LP diet leads to acid-base imbalance, amino acid transport disorder and amino acids imbalance in piglets, and the dietary electrolyte may be a key factor in the impact of the LP diet on piglet growth performance and intestinal health., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Circular RNAs (circRNAs) are a new class of endogenous RNA regulating gene expression. However, the regulatory mechanisms of lipid metabolism in yaks involved in circRNAs remain poorly understood. The IMF plays a crucial role in the quality of yak meat, to greatly improve the meat quality. In this study, the fatty acid profiles of yak IMF were determined and circRNAs were sequenced. The results showed that the total of polyunsaturated fatty acid (PUFA) content of adult yak muscle was significantly higher than that in yak calves (p < 0.05). A total of 29,021 circRNAs were identified in IMF tissue, notably, 99 differentially expressed (DE) circRNAs were identified, to be associated with fat deposition, the most significant of which were circ&#95;12686, circ&#95;6918, circ&#95;3582, ci&#95;106 and ci&#95;123 (A circRNA composed of exons is labelled 'circRNA' and a circRNA composed of introns is labelled 'ciRNA'). KEGG pathway enrichment analysis showed that the differential circRNAs were enriched in four pathways associated with fat deposition (e.g., the peroxisome proliferator-activated receptor signalling, fatty acid degradation, sphingolipid metabolism and sphingolipid signalling pathways). We also constructed co-expression networks of DE circRNA-miRNA using high-throughput sequencing in IMF deposition, from which revealed that ci&#95;106 target binding of bta-miR-130b, bta-miR-148a, bta-miR-15a, bta-miR-34a, bta-miR-130a, bta-miR-17-5p and ci&#95;123 target binding of bta-miR-150 were involved in adipogenesis. The study revealed the role of the circRNAs in the IMF deposition in yak and its influence on meat quality the findings demonstrated the circRNA differences in the development of IMF with the increase of age, thus providing a theoretical basis for further research on the molecular mechanism of IMF deposition in yaks., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

The function of exosomal miRNAs (miRs) in retinal degeneration is largely unclear. We were aimed to investigate the functions of exosomes as well as their miRs derived from retinal pigment epithelial (RPE) cells following exposure to oxidative stress (OS). After the OS by lipopolysaccharide and rotenone on RPE cells, interleukin-1 beta (IL-1β), Interleukin-6 (IL-6), Tumor Necrosis Factor-alpha (TNF-α) were upregulated, along with the decreased mitochondrial membrane potential and upregulated oxidative damage marker 8-OH-dG in RPE cells. RPE-derived exosomes were then isolated, identified, injected into the subretinal space in mice. After subretinal injection, RPE-exosomes after OS not only induced higher ROS level and apoptotic retinal cells, but also elevated IL-1β, IL-6 alongside TNF-α expressions among retina/RPE/choroidal complex. Next, miRs inside the exosomes were sequenced by the next generation sequencing (NGS) technology. NGS revealed that certain miRs were abundant in exosomes, while others were selectively kept by RPE cells. Further, downregulated miRs, like miR-125b-5p, miR-125a-5p, alongside miR-128-3p, and upregulated miR, such as miR-7-5p were validated byRT-qPCR. Finally, Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were used to find the possible target genes of those selective exosomal miRs. Our results proved that the RPE-derived exosomes after OS selectively express certain miRs, providing novel insights into the pathogenesis of age-related macular degeneration (AMD) in future., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024. Published by Elsevier Ltd.)

Irisin is a hormone secreted by muscle in response to exercise. The irisin receptor (IrisinR) is a heterodimer of integrin alpha V (ITGAV) and integrin beta 5 (ITGB5) subunits. Since irisin may mediate some beneficial effects of exercise on animal reproduction, we tested the hypothesis that bovine gonadotrophs express IrisinR and irisin stimulates luteinizing hormone (LH) and follicle stimulating hormone (FSH) secretion by gonadotrophs. Reverse transcription polymerase chain reaction was used to detect the mRNA expression of both ITGAV and ITGB5 in the anterior pituitary glands (APs) of post pubertal heifers and mouse gonadotroph cell line "LβT2." Western blotting was used to detect protein expression in bovine APs. Immunofluorescence microscopy, utilizing the same antibody, visualized IrisinR on the plasma membrane of majority of gonadotrophs. We prepared AP cells from healthy postpubertal heifers, cultured them for 3.5 d, and treated them with increasing concentrations (0, 0.01, 0.1, 1, or 10 nM) of irisin for 5 min before either no treatment or gonadotropin-releasing hormone (GnRH) stimulation. After 2 h, media were harvested for LH and FSH assays. Irisin (0.1-10 nM) stimulated basal LH and FSH secretion, and these stimulatory effects were inhibited by the extracellular signal-regulated kinase or SMAD pathway inhibitors. In the presence of GnRH, irisin at 0.01-1 nM stimulated LH and FSH secretion. A higher dose of irisin (10 nM), however, suppressed the GnRH-induced LH and FSH levels. In conclusion, bovine gonadotrophs expressed IrisinR, and irisin controlled LH and FSH secretion from bovine gonadotrophs., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Kisspeptins are neuropeptides encoded by the Kiss1 gene that was discovered as a metastasis suppressor gene in melanoma and breast cancer. Kisspeptin has pivotal functions for gonadotropin-releasing hormone secretion and plays integrated roles in the hypothalamic-pituitary-gonadal axis. However, little is known about the peripheral expression of kisspeptin in ruminants, especially in the female reproductive tract. Here, the objectives of the current study were to investigate the spatial localization of kisspeptin and mRNA expression of Kiss1 and its receptor (Kiss1r) in the fallopian tubes (FT) and uterus of goats at varied reproductive activity (cyclic versus true anoestrous goats, n=6, each). Specimens of the uterus and FT were collected and fixed using paraformaldehyde to investigate the localizations of kisspeptin in the selected tissues by immunohistochemistry. Another set of samples was snape-frozen to identify the expressions of mRNAs encoding Kiss1 and Kiss1r using real-time PCR. Results revealed immunolocalizations of kisspeptin in the uterus and the FT. The staining of kisspeptin was found mainly in the mucosal epithelium of the uterus the FT, and the endometrial glands. Very intense staining of kisspeptin was found in the uterine and FT specimens in the true anoestrous goats compared to that in cyclic ones. The expression of mRNA encoding Kiss1 gene was significantly higher in the uterine specimen of cyclic goats (1.00±0.09) compared to that in the true anoestrous goats (0.62±0.08) (P ˂0.05), while the expression of mRNA encoding Kiss1r was significantly (P ˂0.001) higher in the uterine tissues of true anoestrous goats (1.78±0.17) compared to that in cyclic ones (1.00±0.11). In conclusion, immunohistochemical localization of kisspeptin and the expression of mRNA encoding Kiss1/Kiss1r revealed spatial changes in the uterus and FT of goats according to the reproductive potential of goats (cyclic versus true anoestrous goats). However, the definitive local role of kisspeptin in the uterus and FT need further investigation., Competing Interests: Declaration of competing interest The authors have declared no conflict of interest for this paper., (Copyright © 2024. Published by Elsevier Inc.)

Optic nerve injuries are severely disrupt the structural and functional integrity of the retina, often leading to visual impairment or blindness. Despite the profound impact of these injuries, the molecular mechanisms involved remain poorly understood. In this study, we performed a comprehensive whole-transcriptome analysis of mouse retina samples after optic nerve crush (ONC) to elucidate changes in gene expression and regulatory networks. Transcriptome analysis revealed a variety of molecular alterations, including 256 mRNAs, 530 lncRNAs, and 37 miRNAs, associated with metabolic, inflammatory, signaling, and biosynthetic pathways in the injured retina. The integrated analysis of co-expression and protein-protein interactions identified an active interconnected module comprising 5 co-expressed proteins (Fga, Serpina1a, Hpd, Slc38a4, and Ahsg) associated with the complement and coagulation cascades. Finally, 5 mRNAs (Fga, Serpinala, Hpd, Slc38a4, and Ahsg), 2 miRNAs (miR-671-5p and miR-3057-5p), and 6 lncRNAs (MSTRG. 1830.1, Gm10814, A530013C23Rik, Gm40634, MSTRG.9514.1, A330023F24Rik) were identified by qPCR in the injured retina, and some of them were validated as critical components of a ceRNA network active in 661W and HEK293T cells through dual-luciferase reporter assays. In conclusion, our study provides comprehensive insight into the complex and dynamic biological mechanisms involved in retinal injury responses and highlights promising potential targets to enhance neuroprotection and restore vision., (Copyright © 2024 Elsevier Ltd. All rights reserved.)