Aims: Studies have observed changes in autophagic flux in the adipose tissue of type 2 diabetes patients with obesity. However, the role of autophagy in obesity-induced insulin resistance is unclear. We propose to confirm the effect of a high-fat diet (HFD) on autophagy and insulin signaling transduction from adipose tissue to clarify whether altered autophagy-mediated HFD induces insulin resistance, and to elucidate the possible mechanisms in autophagy-regulated adipose insulin sensitivity. Methods: Eight-week-old male C57BL/6 mice were fed with HFD to confirm the effect of HFD on autophagy and insulin signaling transduction from adipose tissue. Differentiated 3T3-L1 adipocytes were treated with 1.2 mM fatty acids (FAs) and 50 nM Bafilomycin A1 to determine the autophagic flux. 2.5 mg/kg body weight dose of Chloroquine (CQ) in PBS was locally injected into mouse epididymal adipose (10 and 24 h) and 40 µM of CQ to 3T3-L1 adipocytes for 24 h to evaluate the role of autophagy in insulin signaling transduction. Results: The HFD treatment resulted in a significant increase in SQSTM1/p62, Rubicon expression, and C/EBP homologous protein (CHOP) expression, yet the insulin capability to induce Akt (Ser473) and GSK3β (Ser9) phosphorylation were reduced. PHLPP1 and PTEN remain unchanged after CQ injection. In differentiated 3T3-L1 adipocytes treated with CQ, although the amount of phospho-Akt stimulated by insulin in the CQ-treated group was significantly lower, CHOP expressions and cleaved caspase-3 were increased and bafilomycin A1 induced less accumulation of LC3-II protein. Conclusion: Long-term high-fat diet promotes insulin resistance, late-stage autophagy inhibition, ER stress, and apoptosis in adipose tissue. Autophagy suppression may not affect insulin signaling transduction via phosphatase expression but indirectly causes insulin resistance through ER stress or apoptosis.

In mammalian ovaries, mitochondria are integral sites of energy production and steroidogenesis. While shifts in cellular activities and steroidogenesis are well characterized during the differentiation of large luteal cells in folliculogenesis and luteal formation, mitochondrial dynamics during this process have not been previously evaluated. In this study, we collected ovaries containing primordial follicles, mature follicles, corpus hemorrhagicum, or corpus luteum from goats at specific times in the estrous cycle. Enzyme histochemistry, ultrastructural observations, and 3D structural analysis of serial sections of mitochondria revealed that branched mitochondrial networks were predominant in follicles, while spherical and tubular mitochondria were typical in large luteal cells. Furthermore, the average mitochondrial diameter and volume increased from folliculogenesis to luteal formation. In primordial follicles, the signals of cytochrome c oxidase and ATP synthase were undetectable in most cells, and the large luteal cells from the corpus hemorrhagicum also showed low enzyme signals and content when compared with granulosa cells in mature follicles or large luteal cells from the corpus luteum. Our findings suggest that the mitochondrial enlargement could be an event during folliculogenesis and luteal formation, while the modulation of mitochondrial morphology and respiratory enzyme expressions may be related to tissue remodeling during luteal formation.

White spot syndrome virus (WSSV) has been reported to cause severe economic loss in the shrimp industry. With WSSV being a large virus still under investigation, the 3D structure of its assembly remains unclear. The current study was planned to clarify the 3D structures of WSSV infections in the cell nucleus of red swamp crayfish (Procambarus clarkii). The samples from various tissues were prepared on the seventh day post-infection. The serial sections of the intestinal tissue were obtained for electron tomography after the ultrastructural screening. After 3D reconstruction, the WSSV-associated structures were further visualized, and the expressions of viral proteins were confirmed with immuno-gold labeling. While the pairs of sheet-like structures with unknown functions were observed in the nucleus, the immature virions could be recognized by the core units of nucleocapsids on a piece of the envelope. The maturation of the particle could include the elongation of core units and the filling of empty nucleocapsids with electron-dense materials. Our observations may bring to light a possible order of WSSV maturation in the cell nucleus of the crayfish, while more investigations remain necessary to visualize the detailed viral–host interactions.

Male obesity negative affects gametic function and offspring metabolism. We here describe that (F₀) obesity and weight loss in male mice reversibly alter metabolism and impair adipose mitochondrial function. These metabolic aberrations are transmitted to male offsprings (F₁), which display reduced mitochondrial gene expression. Mechanistically, we identify microRNAs let-7d/e as epigenetic mediators induced in obese F₀ sperm and in F₀/F₁ adipose tissue, where they silence the miRNA processor DICER1 and impair mitochondrial activity. Microinjecting let-7d/e into lean zygotes phenocopies the paternal obesity phenotype, inducing glucose intolerance and mitochondrial gene suppression in sired offspring. Single-cell RNA sequencing of blastomeres reveals that let-7d/e impair oxidative metabolism in early embryos. Furthermore, lifestyle-induced weight loss in males with obesity downregulates human HSA-LET-7D/E in semen, indicating a conserved role for let-7 in transmission of metabolic health. These findings demonstrate that microRNA let-7 in sperm reprograms offspring metabolism by modulating mitochondrial function during early development.

After ovulation, the mitochondrial enzyme CYP11A1 cleavage the cholesterol into pregnenolone for progesterone synthesis, suggesting that mitochondrial dynamics play a vital role in the female reproductive system. The changes in the mitochondria dynamics throughout the ovarian cycle have been reported in literature, but the correlation to its role in the ovarian cycle remains unclear. In this study, mitochondrial fusion promotor, M1, was used to study the impact of mitochondria dynamics in the female reproductive system. Our results showed that M1 treatment in mice can lead to the disruptions of estrous cycles in vagina smears. The decrease in serum LH was recorded in the animal. And the inhibitions of progesterone secretion and ovulations were observed in ovarian culture. Although no significant changes in mitochondrial networks were observed in the ovaries, significant up-regulation of mitochondrial respiratory complexes was revealed in M1 treatments through transcriptomic analysis. In contrast to the estrogen and steroid biosynthesis up-regulated in M1, the molecules of extracellular matrix, remodeling enzymes, and adhesion signalings were decreased. Collectively, our study provides novel targets to regulate the ovarian cycles through the mitochondria. However, more studies are still necessary to provide the functional connections between mitochondria and the female reproductive systems.

Disclosure: Y.P. Budi: None. M. Hsu: None. Y. Lin: None. Y. Lee: None. H. Chiu: None. C. Chiu: None. Y. Jiang: None. In the female reproductive system, mitochondria play a crucial role in steroid hormone synthesis. Literature reports on changes in mitochondrial dynamics throughout the ovarian cycle, but its role in the ovarian cycle is still unknown. In this study, a mitochondrial fusion promotor, M1, was used to study the impact of mitochondrial dynamics in the female reproductive system. The estrus cycle of mice was monitored before, during, and after the M1 treatment to test the impact of M1 on ovarian functions. Our results showed that M1 treatment in mice leads to the disruptions of estrous cycles in vagina smears. To observe the acute effects of M1, serum hormones level was observed, and the decrease in serum LH and progesterone was recorded in the animal. Similarly, our ovarian tissue culture also indicates inhibitions of progesterone secretion and ovulations. Although no significant changes in mitochondrial networks were observed in the ovaries, there was significant up-regulation of mitochondrial respiratory complexes in M1 treatments through transcriptomic analysis. In contrast to the estrogen and steroid biosynthesis up-regulated in M1, the extracellular matrix molecules, remodeling enzymes, and adhesion signaling decreased. Collectively, our study provides novel targets to regulate the ovarian cycles through the mitochondria. However, more studies are still necessary to provide the functional connections between mitochondria and the female reproductive system. Presentation: Saturday, June 17, 2023