Follicle Cells Research Articles

Role of Y box-binding protein 1 (Ybx1) in zebrafish folliculogenesis: Promoting follicle cell proliferation via suppression of p21 (cdkn1a).

Y box-binding protein 1 (YB-1; Ybx1/ybx1) regulates gene expression through DNA/RNA-binding. In zebrafish, Ybx1 is highly abundant in primary growth (PG) follicles in the ovary, but decreases precipitously as the follicles enter the secondary growth (SG). To understand Ybx1 function in folliculogenesis, we created an ybx1 mutant using TALEN and observed disrupted folliculogenesis during the previtellogenic (PV) to early vitellogenic (EV) transition of SG, resulting in underdeveloped ovaries and infertility. Expression and Western blot analyses revealed differential gene expression between ybx1-/- and control ovaries, with significantly increased expression of cdkn1a (p21), a cell cycle inhibitor, in ybx1-/- follicles. While cdkn1a knockout via CRISPR/Cas9 was embryonically lethal, the heterozygote (cdkn1a+/-) displayed advanced follicle activation and maturation, contrasting with the ybx1-/- phenotype. Partial loss of p21 alleviated the ybx1-/-phenotype, restoring folliculogenesis with normal PG-PV and PV-EV transitions in ybx1-/-;cdkn1a+/- mutant. While ybx1-/- mutant follicle cells displayed poor proliferation in vivo and in vitro, the cells from the ybx1-/-;p21+/- follicles resumed normal proliferation. In conclusion, Ybx1 is crucial for early folliculogenesis in zebrafish, potentially by repressing cdkn1a expression, either directly or indirectly.

Landscape transcriptomic analysis of bovine follicular cells during key phases of ovarian follicular development

BackgroundThere are many gaps in our understanding of the mechanisms involved in ovarian follicular development in cattle, particularly regarding follicular deviation, acquisition of ovulatory capacity, and preovulatory changes. Molecular evaluations of ovarian follicular cells during follicular development in cattle, especially serial transcriptomic analyses across key growth phases, have not been reported. This study aims to address this gap by analyzing gene expression using RNA-seq in granulosa and antral cells recovered from ovarian follicular fluid during critical phases of ovarian follicular development in Holstein cows.ResultsIntegrated analysis of gene ontology (GO), gene set enrichment (GSEA), protein–protein interaction (PPI), and gene topology identified that differentially expressed genes (DEGs) in the largest ovarian follicles at deviation (Dev) were primarily involved in FSH-negative feedback, steroidogenesis, cell proliferation, apoptosis, and the prevention of early follicle rupture. In contrast, DEGs in the second largest follicles (DevF2) were mainly related to loss of cell viability, apoptosis, and immune cell invasion. In the dominant (PostDev) and preovulatory (PreOv) follicles, DEGs were associated with vascular changes and inflammatory responses.ConclusionsThe transcriptome of ovarian follicular fluid cells had a predominance of granulosa cells in the dominant follicle at deviation, with upregulation of genes involved in cell viability, steroidogenesis, and apoptosis prevention, whereas in the non-selected follicle there was upregulation of cell death-related transcripts. Immune cell transcripts increased significantly after deviation, particularly in preovulatory follicles, indicating strong intrafollicular chemotactic activity. We inferred that immune cell invasion occurred despite an intact basal lamina, contributing to follicular maturation.Graphical

Immunotherapy response induces divergent tertiary lymphoid structure morphologies in hepatocellular carcinoma.

Tertiary lymphoid structures (TLS) are associated with improved response in solid tumors treated with immune checkpoint blockade, but understanding of the prognostic and predictive value of TLS and the circumstances of their resolution is incomplete. Here we show that in hepatocellular carcinoma treated with neoadjuvant immunotherapy, high intratumoral TLS density at the time of surgery is associated with pathologic response and improved relapse-free survival. In areas of tumor regression, we identify a noncanonical involuted morphology of TLS marked by dispersion of the B cell follicle, persistence of a T cell zone enriched for T cell-mature dendritic cell interactions and increased expression of T cell memory markers. Collectively, these data suggest that TLS can serve as both a prognostic and predictive marker of response to immunotherapy in hepatocellular carcinoma and that late-stage TLS may support T cell memory formation after elimination of a viable tumor.

Estrogen Enhances FDFT1 Expression in Theca Cells of Chicken Hierarchical Ovarian Follicles by Increasing LSD1Ser54p Level Through GSK3β Phosphorylation at 216th Tyrosine

The development of chicken ovarian follicles involves two key stages of primordial follicle recruitment and follicle selection that are tightly regulated by multiple reproductive hormones and cytokines. Our previous study revealed an estrogen-stimulated increase in the phosphorylation level of serine at position 54 of lysine demethylase 1A (LSD1Ser54p) in the theca cells of chicken hierarchical ovarian follicles (Post-TCs). In this study, we further found that the upregulation of LSD1Ser54p by estrogen was performed by glycogen synthase kinase 3 beta (GSK3β) and that GSK3β promoted LSD1Ser54p levels by directly binding to the SWIRM and AOL1 domains of LSD1. Upon estrogen stimulation, the phosphorylation level of tyrosine at position 216 of GSK3β (GSK3βTyr216p) increased, which enhanced the binding between LSD1 and GSK3β. The subsequent transcriptome sequencing on chicken Post-TCs treated with estrogen and CUT&RUN sequencing against the LSD1Ser54p protein revealed that the expression of the farnesyl-diphosphate farnesyltransferase 1 (FDFT1) gene was simultaneously upregulated by estrogen, GSK3β, and LSD1Ser54p. Moreover, the overexpression of FDFT1 further promoted cholesterol biosynthesis in chicken Post-TCs. In short, the findings of this study suggest that estrogen-induced tyrosine phosphorylation at position 216 of GSK3β can upregulate the level of LSD1Ser54p, leading to the activation of FDFT1 expression and subsequently promoting cholesterol biosynthesis in chicken Post-TCs, which may in turn enhance estrogen synthesis.

Ex vivo imaging reveals the spatiotemporal control of ovulation.

During ovulation, an egg is released from an ovarian follicle, ready for fertilization. Ovulation occurs inside the body, impeding direct studies of its progression. Therefore, the exact mechanisms that control ovulation have remained unclear. Here we devised live imaging methods to study the entire process of ovulation in isolated mouse ovarian follicles. We show that ovulation proceeds through three distinct phases, follicle expansion (I), contraction (II) and rupture (III), culminating in the release of the egg. Follicle expansion is driven by hyaluronic acid secretion and an osmotic gradient-directed fluid influx into the follicle. Then, smooth muscle cells in the outer follicle drive follicle contraction. Follicle rupture begins with stigma formation, followed by the exit of follicular fluid and cumulus cells and the rapid release of the egg. These results establish a mechanistic framework for ovulation, a process of fundamental importance for reproduction.

Follicular dendritic cell sarcoma involving the parotid gland with expression of the melanocytic marker PRAME.

Follicular dendritic cell sarcoma is a rare mesenchymal neoplasm arising from follicular dendritic cells (FDC) of lymphoid follicles. While the majority of FDC sarcoma cases arise within lymph nodes, approximately 30% manifest in extranodal sites. Only 4 prior occurrences of intra-parotid FDC sarcomas have been documented. We are reporting a rare case of FDC of the parotid gland in a 65-year-old male with a questionable history of B-cell lymphoma. The patient underwent a right total parotidectomy and bilateral neck dissection. A diagnosis of follicular dendritic cell (FDC) sarcoma was made, with one positive intra-parotid node. The malignant cells expressed the characteristic markers for FDC sarcoma but with positivity of the melanocytic marker PRAME. This is a case of FDC sarcoma with an unusual extranodal localization in the parotid gland. Immunohistochemistry was useful in making a diagnosis although the positivity for the melanocytic marker PRAME was unusual and unreported before.

Blue light inhibits cell viability and proliferation in hair follicle stem cells and dermal papilla cells.

Hair loss is a prevalent issue worldwide, which, though not life-threatening, can result in psychological problems, low self-esteem, and social anxiety. Previous studies have shown that ultraviolet radiation can have negative effects on hair follicle cells, leading to hair loss, while the impact of blue light on hair and hair follicle has largely been overlooked. This study aimed to examine the effects of blue light on hair follicle stem cells (HFSCs) and primary dermal papilla cells (DPCs), which are essential components of hair follicles. Human HFSCs and primary DPCs were exposed to blue light (457nm) at various intensities (1, 4, 8, and 16 mW/cm2) for 3 days. Subsequently, cell viability, cell proliferation, and intracellular reactive oxygen species (ROS) were assessed. The results showed that blue light (457nm) significantly reduced the cell viability and proliferation of HFSCs and DPCs in vitro, with the inhibition being intensity-dependent. Additionally, blue light triggered the overproduction of ROS in the DPCs. While the exact mechanisms by which blue light affects hair follicle cells remain unclear, these findings suggest that blue light could impede the growth of these cells. This insight may offer a new approach to protecting hair by avoiding exposure to high-intensity blue light.

Regulation of Bone Morphogenetic Protein Receptor Type II Expression by FMR1/Fragile X Mental Retardation Protein in Human Granulosa Cells in the Context of Poor Ovarian Response.

Fragile X mental retardation protein (FMRP) is a translational repressor encoded by FMR1. It targets bone morphogenetic protein receptor type II (BMPR2), which regulates granulosa cell (GC) function and follicle development. However, whether this interaction affects folliculogenesis remains unclear. Therefore, this study investigated the potential effect of FMRP-BMPR2 dysregulation in ovarian reserves and infertility. COV434 cells and patient-derived GCs were used to evaluate FMRP and BMPR2 expression. Similarly, FMR1, BMPR2, LIMK1, and SMAD expression were evaluated in GCs with normal (NOR) and poor (POR) ovarian responses. FMRP and BMPR2 were expressed in both cell types. They were co-localized to the nuclear membrane of COV434 cells and cytoplasm of primary GCs. FMR1 silencing increased the mRNA and protein levels of BMPR2. However, the mRNA levels of FMR1 and BMPR2 were significantly lower in the POR group. FMR1 and BMPR2 levels were strongly positively correlated in the NOR group but weakly correlated in the POR group. Additionally, SMAD9 expression was significantly reduced in the POR group. This study highlights the crucial role of FMR1/FMRP in the regulation of BMPR2 expression and its impact on ovarian function. These findings indicate that the disruption of FMRP-BMPR2 interactions may cause poor ovarian responses and infertility.

The antisense lncRNA of TAB2 that prevents oxidative stress to enhance the follicular growth in mammals

LncRNAs are highly implicated in oxidative stress (OS) during the growth of mammalian follicles. TAK1 binding protein 2 gene (TAB2) has been suggested to involve in the normal apoptosis and proliferation of granulosa cells (GCs), the main supporting cells in ovarian follicles. In this study, we found that TAB2 increased the expressions of SOD1, P50, and P65 to suppress the OS, thereby inhibiting the apoptosis and promoting the proliferation in GCs. Notably, DNMTs appeared to mediate the expression of TAB2 without the changes of DNA methylation at TAB2’s promoter. We identified an antisense lncRNA of TAB2, discovered that DNA methylation regulated the transcription of TAB2-AS in GCs, and found TAB2-AS medicated the follicular growth of ovaries in vivo. Mechanistically, the hypomethylation of the CpG site (−1759/−1760) activated the transcription of TAB2-AS, and the 1–155 nt and 156-241 nt of TAB2-AS were respectively complementary to 4368–4534 nt and 4215–4300 nt of TAB2’s mRNA to increase the expression of TAB2. Moreover, TAB2-AS inhibited the OS and apoptosis of GCs, while promoted the proliferation of GCs to expedite the follicular growth, which was in line with that of TAB2. Collectively, these findings revealed the antisense lncRNA mechanism mediated by DNA methylation, and TAB2-AS might be the target to control OS during follicular growth in mammals.

An attempt to search for the common cellular mechanism of ovulation across all metazoans: A review.

Ovulation is the process by which a fertilizable oocyte is extruded from the interior of the follicle. Herein, we conducted a literature survey to explore the ovulation patterns of eleven sexually reproducing species belonging to 10 animal phyla. These results indicate a large variety of ovulation patterns. Further comparative biological and evolutionary considerations of these results led us to conclude that most female animals ovulate via follicle rupture. We propose that in all animals that ovulate by follicle rupture, two cellular events may be critically involved in the process: 1) the disintegration of cell junctional systems that lead to intracellular cytoskeleton rearrangement in the follicle cells and 2) the degradation of extracellular matrix (ECM) proteins filling between follicle cells. These events may result in follicular cell deformation and increased motility, both of which are necessary for the formation of a path through which oocytes escape from the follicle. In addition to the requirement of ECM degradation for disintegrating cell junctions, intensive ECM protein degradation at the apical region of the follicle probably became increasingly important in late-evolving animals, such as vertebrates, in which a thick follicle wall containing a large abundance of ECM proteins is formed. We also considered hypothetical scenarios for the evolution of ovulation in these animals. Furthermore, this article discusses the future problems that need to be solved for a more comprehensive understanding of ovulation in the animal kingdom.

Ovarian Puncture Triggers an Inflammatory Response that did not Affect Late Folliculogenesis, Ovulation Rate, and Fertility.

Ovarian puncture has been widely used in assisted reproduction, but there are still gaps about its effects on ovarian morphophysiology, as well as the relationship between inflammation caused by this procedure and the follicular growth and fertility. The aim of this study was to investigate the effects of ovarian puncture on folliculogenesis and fertility. Mice (n = 24) were divided into two groups: (1) SHAM-both ovaries were exposed and repositioned and (2) Punctured-ovaries were exposed, punctured, and repositioned. After 96h of surgery, ovaries were collected for morphofunctional analysis. New females were used for the superovulation (n = 10) and fertility assays (n = 10). Increased volumetric density of inflammatory cells-p = 0.0005, p = 0.0013; hemorrhagic foci-p < 0.0001; and inflammatory exudate-p < 0.0001 could be noticed on the punctured group, compared to SHAM. The percentage of primordial follicles was lower on the punctured ovaries (p = 0.00294). Ovarian puncture has also induced an increase in the proliferation of granulosa cells of primary (p = 0.0321) and antral follicles (p = 0.0395), and an increased apoptotic index of antral follicles (p = 0.0100). There was no influence on expression of some genes related to inflammation, collagen deposition and folliculogenesis progression. The reproductive aspects (oocyte retrieval and number of fetuses per female) were not altered (p > 0.05). Taken together, our findings strongly suggest that ovarian puncture results in a local inflammation that affects follicular growth and atresia. However, it does not affect female fertility, which strengthens the safety of this procedure.

Chicken chemerin alone or in mixture with adiponectin-visfatin impairs progesterone secretion by primary hen granulosa cells

Adipokines including adiponectin (ADIPO), chemerin (CHEM) and visfatin (VISF) are involved in metabolism and reproductive functions. These three adipokines are present in ovarian cells in different preovulatory follicles in hens. We have previously shown that VISF and ADIPO are able to modulate in vitro steroid production by hen granulosa cells (GCs). It is, however, unclear whether CHEM acts on hen ovarian cells. In addition, no study has yet investigated the effect of a mixture of several adipokines such ADIPO, VISF and CHEM on GCs from different preovulatory follicles. In this study, we investigated the effect of CHEM alone and in combination with ADIPO and VISF on cell viability, proliferation and progesterone secretion in cultured granulosa cells (GCs) from the largest follicles F1 and smaller ones (F3/F4) in the presence of gonadotropins (oLH and oFSH) or hIGF-1. First, various concentrations of chemerin were examined (0, 12, 25, 50 and 100 ng/mL) and then we determined the response to CHEM (at 25 ng/mL) in combination with ADIPO (10 µg/mL) and VISF (100 ng/mL). Chemerin exposure did not affect F1 and F3/F4 granulosa cell viability and proliferation whatever the concentation and in the presence of the mixture. However, it reduced progesterone secretion in dose dependent manner in both F1 and F3/F4 follicles. Furthermore, this CHEM inhibitory effect was significantly higher when CHEM was combined with ADIPO and VISF. Furthermore, CHEM reduced significantly oLH and oFSH- induced progesterone secretion in F1 GCs and oFSH and hIGF-1-induced progesterone secretion in F3/F4 GCs. Interestingly, this inhibitory effect of CHEM was similar in F1 GCs when CHEM was in mixture with ADIPO and VISF whereas it was significantly higher in F3/F4 GCs. Taken together, CHEM impairs progesterone secretion in cultured hen GCs and this inhibitory effect can be potentiated when it is in combination with other adipokines.

Dynamic analysis of Hashimoto’s Thyroiditis bio-mathematical model using artificial neural network

This article establishes an efficient solution scheme for a mathematical model of Hashimoto’s Thyroiditis (HT) employing artificial neural networks. HT is an auto-immune disorder hostile to the thyroid follicle cells, effectuating hypothyroid or hyperthyroidism. Under this condition, the thyroid-stimulating hormone (TSH) alters incomparably to the free thyroxine (FT4) interrupts the functioning of the hypothalamus-pituitary-thyroid (HPT) axis, implicating the thyroid follicle cells getting destroyed. We primarily focus on utilizing artificial neural network (ANN) to perform numerical simulations for the system of ordinary differential equations describing the dynamics of an existing 4D model of HT. The presented model comprises four time-dependent variables: TSH, FT4, anti-thyroid antibodies (Ab), and size of the thyroid gland (T). We utilize ND-Solver and ANN scheme in the Mathematica software to acquire the computational data and illustrate thus retrieved results with essential performance plots. Further, mean square error has been considered in validating the proposed ANN-based approach accurately. The plot for training and validation loss exhibits the effectiveness of the proposed methodology, and substantiate that the suggested ANN approach is a good fit for the solving the mathematical model of HT.

Ovarian reserve alteration in premenopausal women with systemic sclerosis.

Anti-Muellerian hormone (AMH) is produced by the granulosa cells of ovarian follicles. It serves as a sensitive laboratory parameter for assessing ovarian reserve. A reduced ovarian reserve has been observed in patients with various autoimmune diseases. To compare serum levels of AMH as a surrogate parameter for ovarian reserve in female patients with systemic sclerosis compared to healthy controls and thereby assess fertility. In this single centre study from the University Hospital Tuebingen, Germany, we used serum samples to determine concentrations of AMH via an electrochemiluminescence immunoassay. We analysed 30 premenopausal female patients with systemic sclerosis and 30 age-matched healthy controls from 18 to 40 years. Patients who had received cyclophosphamide were excluded from this study. AMH levels were significantly reduced in patients with systemic sclerosis (955 ng/l versus 1.940 ng/L, p < 0.01). Interestingly, in contrast to healthy controls, we observed no significant correlation between age and AMH levels in patients. For women diagnosed with systemic sclerosis, especially at a younger age, regular assessment of AMH levels should be considered to improve guidance with regard to optimal pregnancy timepoint, fertility preservation and treatment options.

Chemotherapy-Induced Alopecia by Docetaxel: Prevalence, Treatment and Prevention.

Docetaxel is a commonly used taxane chemotherapeutic agent in the treatment of a variety of cancers, including breast cancer, ovarian cancer, prostate cancer, non-small cell lung cancer, gastric cancer, and head and neck cancer. Docetaxel exerts its anti-cancer effects through inhibition of the cell cycle and induction of proapoptotic activity. However, docetaxel also impacts rapidly proliferating normal cells in the scalp hair follicles (HFs), rendering the HFs vulnerable to docetaxel-induced cell death and leading to chemotherapy-induced alopecia (CIA). In severe cases, docetaxel causes persistent or permanent CIA (pCIA) when hair does not grow back completely six months after chemotherapy cessation. Hair loss has severe negative impacts on patients' quality of life and may even compromise their compliance with treatment. This review discusses the notable prevalence of docetaxel-induced CIA and pCIA, as well as their prevention and management. At this moment, scalp cooling is the standard of care to prevent CIA. Treatment options to promote hair regrowth include but are not limited to minoxidil, photobiomodulation (PBMT), and platelet-rich plasma (PRP). In addition, a handful of current clinical trials are exploring additional agents to treat or prevent CIA. Research models of CIA, particularly ex vivo human scalp HF organ culture and in vivo mouse models with human scalp xenografts, will help expedite the translation of bench findings of CIA prevention and/or amelioration to the clinic.

Downregulation of TASK-3 Channel Induces Senescence in Granulosa Cells of Bovine Cystic Ovarian Follicles.

Ovarian cysts are linked to hormone imbalances and altered gene expressions, but the connection between cysts and ion channel expression is understudied. This study explored the role of TWIK-related acid-sensitive K+ (TASK) channels in bovine ovarian cyst formation. The ovarian follicles were split into small (5 to 10 mm in diameter) and large (>25 mm in diameter) groups. Among the measured K+, Na+, and Cl- concentrations in follicular fluid (FF) obtained from small-sized follicles (SFs) and large-sized follicles (LFs), the K+ concentration was significantly lower in LFFF. Quantitative PCR, Western blot, and immunocytochemistry data revealed that TASK-3 expression levels significantly decreased by approximately 50% in LFs and granulosa cells obtained from LFs (LFGCs) compared to the corresponding controls. The TASK-3 protein was localized to the plasma membranes of GCs. The diameters of LFGCs were larger than those of SFGCs. The cell swelling response to exposure to a hypotonic solution (200 mOsm/L) was highly reduced in TASK-3-overexpressing cells compared to vector-transfected cells. TASK-3-knockdown cells showed arrested growth. Senescence markers were detected in LFGCs and TASK-3-knockdown cells. These findings suggest that reduced TASK-3 expression in LFs is associated with the inhibition of GC growth, leading to senescence and cyst formation.

Actomyosin contraction in the follicular epithelium provides the major mechanical force for follicle rupture during Drosophila ovulation

Ovulation is critical for sexual reproduction and consists of the process of liberating fertilizable oocytes from their somatic follicle capsules, also known as follicle rupture. The mechanical force for oocyte expulsion is largely unknown in many species. Our previous work demonstrated that Drosophila ovulation, as in mammals, requires the proteolytic degradation of the posterior follicle wall and follicle rupture to release the mature oocyte from a layer of somatic follicle cells. Here, we identified actomyosin contraction in somatic follicle cells as the major mechanical force for follicle rupture. Filamentous actin (F-actin) and nonmuscle myosin II (NMII) are highly enriched in the cortex of follicle cells upon stimulation with octopamine (OA), a monoamine critical for Drosophila ovulation. Pharmacological disruption of F-actin polymerization prevented follicle rupture without interfering with the follicle wall breakdown. In addition, we demonstrated that OA induces Rho1 guanosine triphosphate (GTP)ase activation in the follicle cell cortex, which activates Ras homolog (Rho) kinase to promote actomyosin contraction and follicle rupture. All these results led us to conclude that OA signaling induces actomyosin cortex enrichment and contractility, which generates the mechanical force for follicle rupture during Drosophila ovulation. Due to the conserved nature of actomyosin contraction, this work could shed light on the mechanical force required for follicle rupture in other species including humans.

The Interstitial Gland as a Source of Pro- or Anti-Senescent Cells during Chinchilla Rabbit Ovarian Aging.

The aging ovary in mammals leads to the reduced production of sex hormones and a deterioration in follicle quality. The interstitial gland originates from the hypertrophy of the theca cells of atretic follicles and represents an accumulative structure of the ovary that may contribute to its aging. Here, reproductive and mature rabbit ovaries are used to determine whether the interstitial gland plays a crucial role in ovarian aging. We demonstrate that, in the mature ovary, interstitial gland cells accumulate lipid droplets and show ultrastructural characteristics of lipophagy. Furthermore, they undergo modifications and present a foamy appearance, do not express the pan-leukocyte CD-45 marker, and express CYP11A1. These cells are the first to present an increase in lipofuscin accumulation. In foamy cells, the expression of p21 remains low, PCNA expression is maintained at mature ages, and their nuclei do not show positivity for H2AX. The interstitial gland shows a significant increase in lipofuscin accumulation compared with the ovaries of younger rabbits, but lipofuscin accumulation remains constant at mature ages. Surprisingly, no accumulation of cells with DNA damage is evident, and an increase in proliferative cells is observed at the age of 36 months. We suggest that the interstitial gland initially uses lipophagy to maintain steroidogenic homeostasis and prevent cellular senescence.

Anti-Müllerian hormone deficiency leads to two distinct ovarian phenotypes with different alterations of sex hormones in gynogenetic carp

Anti-Müllerian hormone (Amh) plays conserved roles in gonadal development and function in vertebrates. Previous studies have demonstrated that amh deficiency results in severe gonadal hypertrophy in teleost. Here, we report a novel phenotype of gonadal atrophy caused by amh knockout in fish. Firstly, it was found that the amh gene had three alleles and was expressed in the cytoplasm of follicle cells in gynogenetic amphiploid gibel carp (Carassius gibelio). Subsequently, we constructed a mutant line of amh and identified two distinct ovarian abnormalities in gibel carp. Most mutants exhibited hypertrophic ovaries with increased proliferation and decreased differentiation of early germ cells, which is consistent with previously reported findings. Intriguingly, a small number of mutants displayed atrophic ovaries with decreased proliferation and premature differentiation of germ cells, which represents a novel phenotype in fish species. Transcriptome analysis revealed that these two distinct ovaries of amh mutants showed different alterations in the sex hormone biosynthesis pathway, when compared with wild-type ovaries. Furthermore, diverse gonadal sex hormone levels were also detected in these two abnormal ovaries, which were consistent with the transcriptional expressions of corresponding genes in the sex hormone synthesis pathway. These findings identify a novel atrophic ovarian phenotype of amh mutants and confirm the amh function in controlling the balance between proliferation and differentiation of germ cells in fish.

Single-cell RNA-seq analysis of rat molars reveals cell identity and driver genes associated with dental mesenchymal cell differentiation

BackgroundThe molecular mechanisms and signaling pathways involved in tooth morphogenesis have been the research focus in the fields of tooth and bone development. However, the cell population in molars at the late bell stage and the mechanisms of hard tissue formation and mineralization remain limited knowledge.ResultsHere, we used the rat mandibular first and second molars as models to perform single-cell RNA sequencing (scRNA-seq) analysis to investigate cell identity and driver genes related to dental mesenchymal cell differentiation during the late bell hard tissue formation stage. We identified seven main cell types and investigated the heterogeneity of mesenchymal cells. Subsequently, we identified novel cell marker genes, including Pclo in dental follicle cells, Wnt10a in pre-odontoblasts, Fst and Igfbp2 in periodontal ligament cells, and validated the expression of Igfbp3 in the apical pulp. The dynamic model revealed three differentiation trajectories within mesenchymal cells, originating from two types of dental follicle cells and apical pulp cells. Apical pulp cell differentiation is associated with the genes Ptn and Satb2, while dental follicle cell differentiation is associated with the genes Tnc, Vim, Slc26a7, and Fgfr1. Cluster-specific regulons were analyzed by pySCENIC. In addition, the odontogenic function of driver gene TNC was verified in the odontoblastic differentiation of human dental pulp stem cells. The expression of osteoclast differentiation factors was found to be increased in macrophages of the mandibular first molar.ConclusionsOur results revealed the cell heterogeneity of molars in the late bell stage and identified driver genes associated with dental mesenchymal cell differentiation. These findings provide potential targets for diagnosing dental hard tissue diseases and tooth regeneration.