In cattle and other species in which the pool of resting, primordial follicles is formed during fetal life, little is known about the regulation of the early stages of ovarian follicular development. We used histological morphometry and a combination of observations in vivo and experiments in vitro to study the timing and regulation of follicle formation and the acquisition of the capacity of primordial follicles to initiate growth in cattle. In vivo, primordial, primary, and secondary follicles were first observed around Days 90, 140, and 210 of gestation, respectively. The long interval between the first appearance of primordial and primary follicles suggests that primordial follicles are not capable of activating when they are first formed, or they are inhibited from activating. This hypothesis was confirmed by the finding that most primordial follicles in pieces of ovarian cortex obtained from fetal ovaries older than 140 days activated (i.e., initiated growth) after 2 days in vitro, whereas follicles in cortical pieces from 90- to 140-day-old fetal ovaries did not. We tested the hypothesis that the oocytes of newly formed primordial follicles are not in meiotic arrest and found that before Day 141, most oocytes ( approximately 73%) were in prediplotene stages of prophase I, whereas after Day 140, the majority of oocytes ( approximately 85%) had arrested at the diplotene stage. This observation was further confirmed by the finding that levels of mRNA for YBX2, a protein associated with meiotic arrest, were 2.3 times higher in ovarian cortical pieces isolated after versus before Day 141. Primordial follicles in cortical pieces from 90- to 140-day-old fetal ovaries did activate during a longer, 10-day culture, but activation could be inhibited by adding estradiol or progesterone, but not dihydrotestosterone (all at 10(-6) M). Fetal ovaries secreted estradiol in vitro, and secretion by ovaries from 83 to 140-day-old fetuses declined precipitously ( approximately 30-fold) with age, consistent with the hypothesis that estradiol inhibits activation of newly formed primordial follicles in vivo. In summary, the results show that newly formed primordial follicles do not activate in vivo or within 2 days in vitro and that capacity to activate is correlated with achievement of meiotic arrest by the oocyte and can be inhibited by estradiol, which fetal ovaries actively produce around the time of follicle formation.
The first objective of this study was to determine whether the death of bovine granulosa cells (GC) isolated from small (= 4 mm), medium (5-8 mm), and large (> 8 mm) follicles during follicular atresia occurs by apoptosis. The second objective was to establish an in vitro model system to elucidate the developmental (GC from follicles of different sizes) and hormonal (FSH and insulin-like growth factor-I [IGF-I]) regulation of bovine GC apoptosis during follicular atresia. Bovine ovaries were obtained from a nearby slaughterhouse. Follicles were classified by morphometric criteria as healthy or atretic. Apoptosis in GC from follicles of different sizes was analyzed by both morphological and biochemical methods. Bovine GC were cultured for 48 h at a density of 5 x 10(6) cells/ml in serum-free media at 39 degrees C to determine the effects of FSH and IGF-I on apoptosis. The results showed that apoptosis occurred in GC from all sizes of follicles. Apoptosis in GC was also detected in some healthy follicles. Degenerate GC displayed the morphological characteristics of apoptosis, including nuclei with marginated chromatin, a single condensed nucleus, multiple nuclear fragments, and/or membrane-bound structures containing variable amounts of chromatin and/or cytoplasm (apoptotic bodies). All GC classified as apoptotic on the basis of their morphology contained fragmented DNA measured by the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) technique. Cells that had undergone apoptosis were observed mainly in GC and in scattered theca cells. Throughout the GC layer, apoptotic cell death was more prevalent among antral GC than among mural GC. Interestingly, morphological results showed that no apoptosis occurred in cumulus cells. A time-dependent, spontaneous onset of apoptosis occurred in GC from small, medium, and large follicles during in vitro serum-free culture. The rate of DNA fragmentation in the culture of GC from small follicles was higher than that from medium and large follicles. FSH attenuated apoptotic cell death in GC from medium follicles more effectively than in those from small follicles. IGF-I also suppressed apoptosis in cultured GC from small follicles. In conclusion, this study showed that 1) GC death during bovine follicular development and atresia occurs by apoptosis; 2) apoptosis occurs in GC and theca cells; however, apoptosis does not occur in cumulus cells even in atretic antral follicles; 3) GC from all small, medium, and large follicles undergo spontaneous onset of apoptosis when cultured under serum-free conditions; and 4) FSH and IGF-I can attenuate apoptosis in cultured bovine GC.
The mechanisms controlling the initiation and early stages of follicular growth are poorly understood. Our laboratory developed a serum-free culture system that supports spontaneous and wholesale activation of primordial follicles in pieces of cortex dissected from the ovaries of fetal calves and fetal baboons. However, very few follicles activated in vitro progressed to the secondary stage. To determine whether androgens can promote the primary to secondary follicle transition, pieces of fetal bovine ovarian cortex were cultured in serum-free medium in the absence or presence of testosterone (T, 10(-7) and 10(-6) M) or estradiol (E(2), 10(-6) M) for 10 days. Cortical pieces were then fixed and embedded in plastic for serial sectioning and morphometric analysis; fresh cortical pieces fixed on Day 0 served as uncultured controls. Freshly isolated cortical pieces contained mostly primordial follicles, whereas after 10 days in vitro, most primordial follicles had activated, differentiating into primary follicles as expected. Neither T nor E(2) affected the number of primordial and primary follicles compared with controls (P > 0.05). However, T (10(-7) and 10(-6) M) increased the number of secondary follicles (P < 0.05), whereas E(2) had no effect, suggesting that the effect of T was not due to conversion of T to E(2). In the second experiment, the optimal concentration of T for preantral follicle growth was determined. A range of lower doses of T (10(-10)-10(-7) M) increased the number of secondary follicles in cultured cortical pieces in a dose-dependent manner, with 10(-7) M T being the most effective (P < 0.05). In the third experiment, addition of a specific androgen receptor blocker, flutamide, inhibited the stimulatory effects of T on the primary to secondary follicle transition (P < 0.05), suggesting a receptor-mediated action of T. Localization of androgen receptors by immunohistochemistry revealed immunostaining for the androgen receptor in ovarian stromal cells and increasing immunoreactivity in follicle cells as follicular development progressed from primordial and primary to secondary to antral follicles, suggesting the involvement of the androgen receptor in bovine folliculogenesis. In summary, our results show that T promotes the growth of bovine follicles activated in vitro and suggest that its stimulatory effect is mediated through androgen receptors in the stroma and/or follicular cells.
Little is known about the mechanisms regulating the growth of early preantral follicles, especially in nonrodent mammalian species. To test the hypothesis that vascular endothelial growth factor (VEGF) promotes the primary to secondary follicle transition, pieces of bovine fetal ovarian cortex were cultured with VEGF (0, 1, 10, or 100 ng/ml) for 0 or 10 days, followed by morphometric analysis. On day 0, cortical pieces contained mostly primordial follicles, but after 10 days in vitro most primordial follicles had activated, differentiating into primary follicles. VEGF had no effect on the numbers of primordial or primary follicles, compared with untreated controls, but all doses increased the number of secondary follicles. In the second experiment, a range of lower doses of VEGF (0.1-10 ng/ml) increased the number of secondary follicles in a dose-dependent manner. Analysis of VEGF transcripts by RT-PCR showed expression of mRNA for three isoforms of VEGF (VEGF121, 165, and 189) in fetal bovine ovarian cortex, with VEGF121 and 165 expressed predominantly and levels of mRNA for VEGF121 and 189 increasing after day 211 of gestation, when the first secondary follicles appear. Expression of mRNA for both VEGF receptors (flt-1 and flk-1) was also detected, but did not change with the development of fetal ovaries. Immunohistochemistry revealed positive staining for VEGF in blood vessels and in follicle cells of secondary follicles, consistent with Western blot analyses showing increases in VEGF protein as ovarian development progressed. Taken together, the results provide support for a role for VEGF in the primary to secondary follicle transition.
for various fields, such as drug delivery, [1,2] biosensing, [3] detoxification, [4] and environmental remediation. [5] These miniaturized machines can convert local fuel or external energy (e.g., magnetic, ultrasound, or light) to driving force and obtain selfpropulsion in dynamic surroundings. [6] Due to the increasing demands and rigorous requirements of biomedical community, the composition of microrobots has been evolving from rigid and inorganic materials to functional and endogenous cells. [7] Incorporating natural cells with synthetic micromachines creates an elegant platform with synergistic properties of cell biofunctionalities and effective propulsion. The cargo-loading capability of cells is employed by cell-based microrobots to actively deliver drugs, [2,[8][9][10][11] such as red blood cell (RBC)based micromotors loaded with doxorubicin (DOX) and photosensitizer, [10,11] and neutrophil-based microrobots carrying paclitaxel (PTX). [2] However, the utilized chemotherapeutic agents in the robotic system are not specific to cancerous cells and occur drug leakage during transport, inducing unavoidable toxicity to normal regions along with limited therapeutic efficacy and undesired side effects. [12] Such compromised biosafety and antitumor index hinder the practical A unique robotic medical platform is designed by utilizing cell robots as the active "Trojan horse" of oncolytic adenovirus (OA), capable of tumor-selective binding and killing. The OA-loaded cell robots are fabricated by entirely modifying OA-infected 293T cells with cyclic arginine-glycine-aspartic acid tripeptide (cRGD) to specifically bind with bladder cancer cells, followed by asymmetric immobilization of Fe 3 O 4 nanoparticles (NPs) on the cell surface. OA can replicate in host cells and induce cytolysis to release the virus progeny to the surrounding tumor sites for sustainable infection and oncolysis. The asymmetric coating of magnetic NPs bestows the cell robots with effective movement in various media and wireless manipulation with directional migration in a microfluidic device and bladder mold under magnetic control, further enabling steerable movement and prolonged retention of cell robots in the mouse bladder. The biorecognition of cRGD and robust, controllable propulsion of cell robots work synergistically to greatly enhance their tissue penetration and anticancer efficacy in the 3D cancer spheroid and orthotopic mouse bladder tumor model. Overall, this study integrates cell-based microrobots with virotherapy to generate an attractive robotic system with tumor specificity, expanding the operation scope of cell robots in biomedical community.
The gut microbiome plays an important role in a host’s development and adaption to its dietary niche. In this study, a group of bamboo-feeding insects are used to explore the potential role of the gut microbiota in the convergent adaptation to extreme diet specialization. Specifically, using a 16S rRNA marker and an Illumina sequencing platform, we profiled the microbial communities of 76 gut samples collected from nine bamboo-feeding insects, including both hemimetabolous (Orthoptera and Hemiptera) and holometabolous (Coleoptera and Lepidoptera) species, which are specialized in three distinct dietary niches: bamboo leaf, shoot, and sap. The gut microbiota of these insects were dominated by Proteobacteria, Firmicutes, and Bacteroidetes and were clustered into solid (leaf and shoot) and liquid (sap) dietary niches. The gut bacterial communities of insects feeding on solid diet overlapped significantly, even though these insects belong to phylogenetically distant lineages representing different orders. In addition, the presence of cellulolytic bacterial communities within the gut microbiota allows bamboo-feeding insects to adapt to a highly specialized, fiber-rich diet. Although both phylogeny and diet can impact the structure and composition of gut microbiomes, phylogeny is the primary driving force underlying the convergent adaptation to a highly specialized diet, especially when the related insect species harbor similar gut microbiomes and share the same dietary niche over evolutionary timescales. These combined findings lay the foundation for future research on how convergent feeding strategies impact the interplays between hosts and their gut microbiomes and how the gut microbiota may facilitate convergent evolution in phylogenetically distant species in adaptation to the shared diet.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.