Xiaohan Cao

Age-related thymic atrophy, a hallmark of immunosenescence linked to age-related diseases, involves gonadal and adrenal steroid hormones, but their distinct roles and mechanisms in this process remain unclear. Through biochemical, histological, and RNA-seq analyses, we comprehensively explored the mechanisms underpinning age-related thymic atrophy in response to ovariectomy (OVX) versus adrenalectomy (ADX) in female mice. Compared to the sham controls, OVX overtly ameliorated age-related thymic atrophy, as evidenced by increased thymus mass, a larger gross thymus area, and denser cortex cellularity. In contrast, ADX evidently accelerated age-related thymic atrophy, characterized by increased adipose infiltration and decreased cortex/medulla ratio, obscure cortico-medullary junctions, and sparser thymic cortical cells. Unexpectedly, combined OVX and ADX displayed a more pronounced effect than OVX alone in ameliorating age-related thymic atrophy. Mechanistically, OVX decreased while ADX increased the circulating 17β-estradiol levels in female mice, which drove these opposing outcomes potentially by promoting Pparg-mediated thymic fat deposition and blocking Cdk1-dependent thymocyte cell cycle progression. Although OVX eliminated gonadal 17β-estradiol production, it appeared to trigger a compensatory adrenal-dependent estrogen biosynthesis, whereas combined OVX and ADX nearly eliminated all estrogen sources, thus leading to a more pronounced effect than OVX alone in ameliorating age-related thymic atrophy in female mice. Notably, OVX increased while ADX decreased serum corticosterone levels, but these alterations exerted minimal impacts on age-related thymic atrophy, highlighting a pivotal role of estrogens over glucocorticoids in accelerating age-related thymic atrophy in females. Undesirably, although OVX ameliorated age-related thymic atrophy, it appeared to simultaneously increase autoimmune susceptibility by downregulating thymic Cd74 expression. Taken together, our results indicate that OVX ameliorates while ADX accelerates age-related thymic atrophy in females. Estrogens rather than glucocorticoids act as the predominant regulator of this process, potentially via promoting Pparg-dependent fat deposition and blocking Cdk1-dependent thymocyte cycle progression. However, OVX-induced estrogen depletion also elevated autoimmune risk, emphasizing the need to balance benefits and risks in regulating thymic aging.

The Tibetan cashmere goat is a prolific goat breed in China. In sheep breeds, natural mutations have demonstrated that the transforming growth factor beta (TGF-β) super family ligands, such as growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15) and their type I receptor (bone morphogenetic protein receptor (BMPR1B), are essential for ovulation and increasing litter size. In this study, 216 female Tibetan cashmere goats were sampled, and candidate genes with fecundity traits were detected via restriction fragment length polymorphism (RFLP) and sequenced. Four polymorphic loci were found in specific amplification fragments of BMP15 and GDF9. Two SNP sites of the BMP15 gene were discovered, namely G732A and C805G. The G732A mutation did not cause the change in amino acids, and the frequencies of each genotype were 0.695 for the GG type, 0.282 for the GA type and 0.023 for the AA type. The C805G mutation caused amino acids to change from glutamine to glutamate. The genotype frequencies were 0.620 for the CC type, 0.320 for the CG type and 0.320 for the CG type. For the GG type 0.060, the G3 and G4 mutations of the GDF9 gene were all homozygous mutations. Two known SNP sites, C719T and G1189A, were detected in the Tibetan cashmere goat GDF9 gene, of which the C719T mutation caused a change of alanine to valine, with a genotype frequency of 0.944 for the CC type and 0.056 for the CT type, whereas no TT type was found. The G1189A mutation caused valine to become isoleucine, and the frequencies of each genotype were 0.579 for the GG type, 0.305 for the GA type and 0.116 for the AA type; G1, B2, B3, B4, FecX^H, FecX^I, FecX^L, G2, G5, G6, G7, G8, FecG^E, FecTT and FecB mutations were not found in Tibetan cashmere goats. The results of this study provide a data basis for future studies of BMP15, GDF9 and BMPR1B gene mutations in goats.

Inhibins suppress the FSH production in pituitary gonadotrope cells by robustly antagonizing activin signaling by competitively binding to activin type II receptors (ACTR II). The binding of inhibin A to ACTR II requires the presence of its co-receptor, namely, betaglycan. In humans, the critical binding site for betaglycan to inhibin A was identified on the inhibin α subunit. Through conservation analysis, we found that a core 13-amino-acid peptide sequence within the betaglycan-binding epitope on human inhibin α subunit is highly conserved across species. Based on the tandem sequence of such a conserved 13-amino-acid betaglycan-binding epitope (INHα13AA-T), we developed a novel inhibin vaccine and tested its efficacy in promoting female fertility using the female rat as a model. Compared with placebo-immunized controls, INHα13AA-T immunization induced a marked (p < 0.05) antibody generation, enhanced (p < 0.05) ovarian follicle development, and increased ovulation rate and litter sizes. Mechanistically, INHα13AA-T immunization promoted (p < 0.05) pituitary Fshb transcription and increased (p < 0.05) serum FSH and 17β-estradiol concentrations. In summary, active immunization against INHα13AA-T potently increased FSH levels, ovarian follicle development, ovulation rate and litter sizes, thus causing super-fertility in females. Therefore, immunization against INHα13AA is a promising alternative to the conventional approach of multiple ovulation and super-fertility in mammals.

The Tibetan cashmere goat is a precious breed in China and its cashmere is widely used in clothing and textiles. The genes IGF-1, FGF5, and KAP 1.4 have been shown to be crucial regulators of cashmere growth. In this study, we examined mRNA expression levels of these three genes and detected IGF-1, FGF5, and KAP 1.4 SNP loci in the Tibetan cashmere goat. After amplification and sequence alignment of the genes IGF-1, FGF5, and KAP 1.4 among 206 Tibetan cashmere goats, two new SNP loci were detected in gene KAP 1.4, while no SNP loci were found in amplified fragments of genes IGF-1 and FGF5. The expression levels of gene IGF-1 in Baingoin and Nyima counties were significantly higher than in other counties (p < 0.05). Moreover, the expression level of gene FGF5 in Gêrzê was significantly higher than in Rutog. The expression levels of mRNA in KAP 1.4 showed significant variation among seven counties. There were no significant differences in mRNA expression levels of IGF-1, FGF5, and KAP 1.4 in Tibetan cashmere goats when analysed by sex. The gene IGF-1 was slightly up-regulated in one to five-year-old cashmere goats, except in those that were 4 years old. The mRNA expression levels of FGF5 in one and two-year-old cashmere goats was lower compared with those in three to five-year-old cashmere goats. KAP 1.4 was up-regulated across one to five-year-old cashmere goats. In this study, SNP detection and mRNA expression analysis of IGF-1, FGF5, and KAP 1.4 genes was able to add data to genetic evolutionary analysis. Further studies should be carried out in SNPs to detect other fragments in genes IGF-1 and FGF5, as well as signal pathways and gene functions in protein levels of genes IGF-1, FGF5, and KAP 1.4 in the Tibetan cashmere goat.

Spexin2 (SPX2), a paralog of SPX1, is a newly identified gene in non-mammalian vertebrates. Limited studies in fish have evidenced its important role in food intake and energy balance modulation. However, little is known about its biological functions in birds. Using the chicken (c-) as a model, we cloned the full-length cDNA of SPX2 by using RACE-PCR. It is 1189 base pair (bp) in length and predicted to generate a protein of 75 amino acids that contains a 14 amino acids mature peptide. Tissue distribution analysis showed that cSPX2 transcripts were detected in a wide array of tissues, with abundant expression in the pituitary, testis, and adrenal gland. cSPX2 was also observed to be ubiquitously expressed in chicken brain regions, with the highest expression in the hypothalamus. Its expression was significantly upregulated in the hypothalamus after 24 or 36 h of food deprivation, and the feeding behavior of chicks was obviously suppressed after peripheral injection with cSPX2. Mechanistically, further studies evidenced that cSPX2 acts as a satiety factor via upregulating cocaine and amphetamine regulated transcript (CART) and downregulating agouti-related neuropeptide (AGRP) in hypothalamus. Using a pGL4-SRE-luciferase reporter system, cSPX2 was demonstrated to effectively activate a chicken galanin II type receptor (cGALR2), a cGALR2-like receptor (cGALR2L), and a galanin III type receptor (cGALR3), with the highest binding affinity for cGALR2L. Collectively, we firstly identified that cSPX2 serves as a novel appetite monitor in chicken. Our findings will help clarify the physiological functions of SPX2 in birds as well as its functional evolution in vertebrates.

Dysfunctions of the ovaries and adrenal glands are both evidenced to cause aberrant adipose tissue (AT) remodeling and resultant metabolic disorders, but their distinct and common roles are poorly understood. In this study, through biochemical, histological and RNA-seq analyses, we comprehensively explored the mechanisms underpinning subcutaneous (SAT) and visceral adipose tissue (VAT) remodeling, in response to ovariectomy (OVX) versus adrenalectomy (ADX) in female mice. OVX promoted adipocyte differentiation and fat accumulation in both SAT and VAT, by potentiating the Pparg signaling, while ADX universally prevented the cell proliferation and extracellular matrix organization in both SAT and VAT, likely by inactivating the Nr3c1 signaling, thus causing lipoatrophy in females. ADX, but not OVX, exerted great effects on the intrinsic difference between SAT and VAT. Specifically, ADX reversed a large cluster of genes differentially expressed between SAT and VAT, by activating 12 key transcription factors, and thereby caused senescent cell accumulation, massive B cell infiltration and the development of selective inflammatory response in SAT. Commonly, both OVX and ADX enhance circadian rhythmicity in VAT, and impair cell proliferation, neurogenesis, tissue morphogenesis, as well as extracellular matrix organization in SAT, thus causing dysfunction of adipose tissues and concomitant metabolic disorders.

A follicle stimulating hormone (FSH) is widely used in the assisted reproduction and a synthetic peptide corresponding to a receptor binding region of the human (h) FSH-β-(34–37) (TRDL) modulated reproduction. Furthermore, a 13-amino acid sequence corresponding to hFSH-β-(37–49) (LVYKDPARPKIQK) was recently identified as the receptor binding site. We hypothesized that the synthetic peptides corresponding to hFSH-β-(37–49) and hFSH-β-(34–49), created by merging hFSH-β-(34–37) and hFSH-β-(37–49), modulate the reproductive functions, with the longer peptide being more biologically active. In male or female prepubertal mice, a single injection of 200 μg/g BW ip of hFSH-β-(37–49) or hFSH-β-(34–49) hastened (p < 0.05) puberty, whereas the same treatments given daily for 4 d promoted (p < 0.05) the gonadal steroidogenesis and gamete formation. In addition of either peptide to the in vitro cell cultures, promoted (p < 0.05) the proliferation of primary murine granulosa cells and the estradiol production by upregulating the expression of Ccnd2 and Cyp19a1, respectively. In adult female mice, 200 μg/g BW ip of either peptide during diestrus antagonized the FSH-stimulated estradiol increase and uterine weight gain during proestrus. Furthermore, hFSH-β-(34–49) was a more potent (p < 0.05) reproductive modulator than hFSH-β-(37–49), both in vivo and in vitro. We concluded that hFSH-β-(37–49) and especially hFSH-β-(34–49), have the potential for reproductive modulation.