mTORC1/S6K1 signaling promotes sustained oncogenic translation through modulating CRL3IBTK-mediated ubiquitination of eIF4A1 in cancer cells.

Enhanced protein synthesis is a crucial molecular mechanism that allows cancer cells to survive, proliferate, metastasize, and develop resistance to anti-cancer treatments, and often arises as a consequence of increased signaling flux channeled to mRNA-bearing eukaryotic initiation factor 4F (eIF4F). However, the post-translational regulation of eIF4A1, an ATP-dependent RNA helicase and subunit of the eIF4F complex, is still poorly understood. Here, we demonstrate that IBTK, a substrate-binding adaptor of the Cullin 3-RING ubiquitin ligase (CRL3) complex, interacts with eIF4A1. The non-degradative ubiquitination of eIF4A1 catalyzed by the CRL3IBTK complex promotes cap-dependent translational initiation, nascent protein synthesis, oncogene expression, and cervical tumor cell growth both in vivo and in vitro. Moreover, we show that mTORC1 and S6K1, two key regulators of protein synthesis, directly phosphorylate IBTK to augment eIF4A1 ubiquitination and sustained oncogenic translation. This link between the CRL3IBTK complex and the mTORC1/S6K1 signaling pathway, which is frequently dysregulated in cancer, represents a promising target for anti-cancer therapies.

Autophagy initiation triggers p150Glued-AP-2ß interaction on the lysosomes and facilitates their transport.

The endocytic adaptor protein 2 (AP-2) complex binds dynactin as part of its noncanonical function, which is necessary for dynein-driven autophagosome transport along microtubules in neuronal axons. The absence of this AP-2-dependent transport causes neuronal morphology simplification and neurodegeneration. The mechanisms that lead to formation of the AP-2-dynactin complex have not been studied to date. However, the inhibition of mammalian/mechanistic target of rapamycin complex 1 (mTORC1) enhances the transport of newly formed autophagosomes by influencing the biogenesis and protein interactions of Rab-interacting lysosomal protein (RILP), another dynein cargo adaptor. We tested effects of mTORC1 inhibition on interactions between the AP-2 and dynactin complexes, with a focus on their two essential subunits, AP-2ß and p150Glued. We found that the mTORC1 inhibitor rapamycin enhanced p150Glued-AP-2ß complex formation in both neurons and non-neuronal cells. Additional analysis revealed that the p150Glued-AP-2ß interaction was indirect and required integrity of the dynactin complex. In non-neuronal cells rapamycin-driven enhancement of the p150Glued-AP-2ß interaction also required the presence of cytoplasmic linker protein 170 (CLIP-170), the activation of autophagy, and an undisturbed endolysosomal system. The rapamycin-dependent p150Glued-AP-2ß interaction occurred on lysosomal-associated membrane protein 1 (Lamp-1)-positive organelles but without the need for autolysosome formation. Rapamycin treatment also increased the acidification and number of acidic organelles and increased speed of the long-distance retrograde movement of Lamp-1-positive organelles. Altogether, our results indicate that autophagy regulates the p150Glued-AP-2ß interaction, possibly to coordinate sufficient motor-adaptor complex availability for effective lysosome transport.

Effects of asiaticoside on the model of gestational diabetes mellitus in HTR-8/svneo cells via PI3K/AKT pathway.

BACKGROUND: Asiaticoside (AS) has been reported to improve the changes induced by high glucose stimulation, and it may have potential therapeutic effects on gestational diabetes mellitus (GDM). This study aims to explore the effect of AS on the cell model of GDM and the action mechanism of the PI3K/AKT pathway. METHODS: The GDM model was established in HTR-8/Svneo cells with a high glucose (HG) medium. After the cytotoxicity assay of AS, cells were divided into the control group, HG group and HG + AS group to conduct control experiment in cells. The cell proliferation and migration were detected by CCK-8 assay and scratch test, respectively. The mRNA levels of PI3K, AKT2, mTORC1, and GLUT4 in PI3K/AKT signalling pathway were measured by RT-PCR, and the protein expressions of these signalling molecules were monitored by western blot. RESULTS: AS showed a promotion effect on the cell proliferation rate of HTR-8/Svneo cells, and 80 µmol/L AS with a treatment time of 48 h had no cytotoxicity. The cell proliferation rate, migration rate, mRNA levels and protein expressions of PI3K, AKT2, mTORC1, and GLUT4 in the HG group were significantly lower than those in the control group, which were significantly increased in the HG + AS group (p < 0.05). CONCLUSIONS: AS can facilitate the cell proliferation and migration in the cell model of GDM, and might play a role in GDM treatment via PI3K/AKT pathway.

Asiaticoside possesses various pharmacological effects and has been reported to show a beneficial effect on the treatment of diabetes mellitus. This research firstly investigated the effect and mechanism of asiaticoside on gestational diabetes mellitus, and found that asiaticoside could facilitate the cell proliferation and migration of HTR-8/Svneo cells treated with high glucose, and affect the signalling molecules of PI3K/AKT pathway. Therefore, asiaticoside may be a novel useful therapeutic drug in the treatment of gestational diabetes mellitus.

Molecular underpinnings of dedifferentiation and aggressiveness in chromophobe renal cell carcinoma.

Sarcomatoid dedifferentiation is common to multiple renal cell carcinoma (RCC) subtypes, including chromophobe RCC (ChRCC), and is associated with increased aggressiveness, resistance to targeted therapies, and heightened sensitivity to immunotherapy. To study ChRCC dedifferentiation, we performed multiregion integrated paired pathological and genomic analyses. Interestingly, ChRCC dedifferentiates not only into sarcomatoid but also into anaplastic and glandular subtypes, which are similarly associated with increased aggressiveness and metastases. Dedifferentiated ChRCC shows loss of epithelial markers, convergent gene expression, and whole genome duplication from a hypodiploid state characteristic of classic ChRCC. We identified an intermediate state with atypia and increased mitosis but preserved epithelial markers. Our data suggest that dedifferentiation is initiated by hemizygous mutation of TP53, which can be observed in differentiated areas, as well as mutation of PTEN. Notably, these mutations become homozygous with duplication of preexisting monosomes (i.e., chromosomes 17 and 10), which characterizes the transition to dedifferentiated ChRCC. Serving as potential biomarkers, dedifferentiated areas become accentuated by mTORC1 activation (phospho-S6) and p53 stabilization. Notably, dedifferentiated ChRCC share gene enrichment and pathway activation features with other sarcomatoid RCC, suggesting convergent evolutionary trajectories. This study expands our understanding of aggressive ChRCC, provides insight into molecular mechanisms of tumor progression, and informs pathologic classification and diagnostics.

Uncharacterized yeast gene YBR238C, an effector of TORC1 signaling in a mitochondrial feedback loop, accelerates cellular aging via HAP4- and RMD9-dependent mechanisms.

Uncovering the regulators of cellular aging will unravel the complexity of aging biology and identify potential therapeutic interventions to delay the onset and progress of chronic, aging-related diseases. In this work, we systematically compared genesets involved in regulating the lifespan of Saccharomyces cerevisiae (a powerful model organism to study the cellular aging of humans) and those with expression changes under rapamycin treatment. Among the functionally uncharacterized genes in the overlap set, YBR238C stood out as the only one downregulated by rapamycin and with an increased chronological and replicative lifespan upon deletion. We show that YBR238C and its paralog RMD9 oppositely affect mitochondria and aging. YBR238C deletion increases the cellular lifespan by enhancing mitochondrial function. Its overexpression accelerates cellular aging via mitochondrial dysfunction. We find that the phenotypic effect of YBR238C is largely explained by HAP4- and RMD9-dependent mechanisms. Furthermore, we find that genetic- or chemical-based induction of mitochondrial dysfunction increases TORC1 (Target of Rapamycin Complex 1) activity that, subsequently, accelerates cellular aging. Notably, TORC1 inhibition by rapamycin (or deletion of YBR238C) improves the shortened lifespan under these mitochondrial dysfunction conditions in yeast and human cells. The growth of mutant cells (a proxy of TORC1 activity) with enhanced mitochondrial function is sensitive to rapamycin whereas the growth of defective mitochondrial mutants is largely resistant to rapamycin compared to wild type. Our findings demonstrate a feedback loop between TORC1 and mitochondria (the TORC1-MItochondria-TORC1 (TOMITO) signaling process) that regulates cellular aging processes. Hereby, YBR238C is an effector of TORC1 modulating mitochondrial function.

Dietary resveratrol improves the flesh quality of Siberian sturgeon (Acipenser baerii) by enhancing myofiber growth, nutrient accumulation and antioxidant capacity.

BACKGROUND: In aquaculture, sturgeons are generally maintained in the confined spaces, which not only hinders sturgeon movement, but also threatens their flesh quality that seriously concerned by aquaculture industry. As a typical antioxidant, resveratrol can improve the flesh quality of livestock and poultry. However, the mechanism of resveratrol's effect on the muscle of Siberian sturgeon is still unclear. RESULTS: In this study, the dietary resveratrol increased the myofiber diameter, the content of the amino acids, antioxidant capacity markers (CAT, LDH and SOD) levels and the expression levels of mTORC1 and MYH9 in muscle of Siberian sturgeon. Further transcriptome analysis displayed that ROS production-related pathways ("Oxidative phosphorylation" and "Chemical carcinogenes-reactive oxygen species") were enriched in KEGG analysis, and the expression levels of genes related to the production of ROS (COX4, COX6A, ATPeF1A, etc.) in mitochondria were significantly down-regulated, while the expression levels of genes related to scavenging ROS (SOD1) were up-regulated. CONCLUSIONS: In summary, this study reveals that resveratrol may promote the flesh quality of Siberian sturgeon probably by enhancing myofiber growth, nutritional value and the antioxidant capacity of muscle, which has certain reference significance for the development of a new type of feed for Siberian sturgeon.

Taenia solium cysticerci's extracellular vesicles Attenuate the AKT/mTORC1 pathway for Alleviating DSS-induced colitis in a murine model.

The excretory-secretory proteome plays a pivotal role in both intercellular communication during disease progression and immune escape mechanisms of various pathogens including cestode parasites like Taenia solium. The cysticerci of T. solium causes infection in the central nervous system known as neurocysticercosis (NCC), which affects a significant population in developing countries. Extracellular vesicles (EVs) are 30-150-nm-sized particles and constitute a significant part of the secretome. However, the role of EV in NCC pathogenesis remains undetermined. Here, for the first time, we report that EV from T. solium larvae is abundant in metabolites that can negatively regulate PI3K/AKT pathway, efficiently internalized by macrophages to induce AKT and mTOR degradation through auto-lysosomal route with a prominent increase in the ubiquitination of both proteins. This results in less ROS production and diminished bacterial killing capability among EV-treated macrophages. Due to this, both macro-autophagy and caspase-linked apoptosis are upregulated, with a reduction of the autophagy substrate sequestome 1. In summary, we report that T. solium EV from viable cysts attenuates the AKT-mTOR pathway thereby promoting apoptosis in macrophages, and this may exert immunosuppression during an early viable stage of the parasite in NCC, which is primarily asymptomatic. Further investigation on EV-mediated immune suppression revealed that the EV can protect the mice from DSS-induced colitis and improve colon architecture. These findings shed light on the previously unknown role of T. solium EV and the therapeutic role of their immune suppression potential.

Timing of TORC1 inhibition dictates Pol III involvement in Caenorhabditis elegans longevity.

Organismal growth and lifespan are inextricably linked. Target of Rapamycin (TOR) signalling regulates protein production for growth and development, but if reduced, extends lifespan across species. Reduction in the enzyme RNA polymerase III, which transcribes tRNAs and 5S rRNA, also extends longevity. Here, we identify a temporal genetic relationship between TOR and Pol III in Caenorhabditis elegans, showing that they collaborate to regulate progeny production and lifespan. Interestingly, the lifespan interaction between Pol III and TOR is only revealed when TOR signaling is reduced, specifically in adulthood, demonstrating the importance of timing to control TOR regulated developmental versus adult programs. In addition, we show that Pol III acts in C. elegans muscle to promote both longevity and healthspan and that reducing Pol III even in late adulthood is sufficient to extend lifespan. This demonstrates the importance of Pol III for lifespan and age-related health in adult C. elegans.

mTORC1 regulates cell survival under glucose starvation through 4EBP1/2-mediated translational reprogramming of fatty acid metabolism.

Energetic stress compels cells to evolve adaptive mechanisms to adjust their metabolism. Inhibition of mTOR kinase complex 1 (mTORC1) is essential for cell survival during glucose starvation. How mTORC1 controls cell viability during glucose starvation is not well understood. Here we show that the mTORC1 effectors eukaryotic initiation factor 4E binding proteins 1/2 (4EBP1/2) confer protection to mammalian cells and budding yeast under glucose starvation. Mechanistically, 4EBP1/2 promote NADPH homeostasis by preventing NADPH-consuming fatty acid synthesis via translational repression of Acetyl-CoA Carboxylase 1 (ACC1), thereby mitigating oxidative stress. This has important relevance for cancer, as oncogene-transformed cells and glioma cells exploit the 4EBP1/2 regulation of ACC1 expression and redox balance to combat energetic stress, thereby supporting transformation and tumorigenicity in vitro and in vivo. Clinically, high EIF4EBP1 expression is associated with poor outcomes in several cancer types. Our data reveal that the mTORC1-4EBP1/2 axis provokes a metabolic switch essential for survival during glucose starvation which is exploited by transformed and tumor cells.

Inhibition of ERK1/2 or CRMP2 Disrupts Alcohol Memory Reconsolidation and Prevents Relapse in Rats.

Relapse to alcohol abuse, often caused by cue-induced alcohol craving, is a major challenge in alcohol addiction treatment. Therefore, disrupting the cue-alcohol memories can suppress relapse. Upon retrieval, memories transiently destabilize before they reconsolidate in a process that requires protein synthesis. Evidence suggests that the mammalian target of rapamycin complex 1 (mTORC1), governing the translation of a subset of dendritic proteins, is crucial for memory reconsolidation. Here, we explored the involvement of two regulatory pathways of mTORC1, phosphoinositide 3-kinase (PI3K)-AKT and extracellular regulated kinase 1/2 (ERK1/2), in the reconsolidation process in a rat (Wistar) model of alcohol self-administration. We found that retrieval of alcohol memories using an odor-taste cue increased ERK1/2 activation in the amygdala, while the PI3K-AKT pathway remained unaffected. Importantly, ERK1/2 inhibition after alcohol memory retrieval impaired alcohol-memory reconsolidation and led to long-lasting relapse suppression. Attenuation of relapse was also induced by post-retrieval administration of lacosamide, an inhibitor of collapsin response mediator protein-2 (CRMP2)-a translational product of mTORC1. Together, our findings indicate the crucial role of ERK1/2 and CRMP2 in the reconsolidation of alcohol memories, with their inhibition as potential treatment targets for relapse prevention.

[Mechanism of Notoginseng Radix et Rhizoma in regulating PI3K/Akt/mTORC1-mitochondrial energy metabolism to treat chronic renal failure in rats with blood stasis syndrome].

This study observed the effects of Notoginseng Radix et Rhizoma on the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/mammalian target of rapamycin complex 1(mTORC1) signaling pathway and mitochondrial energy metabolism in the rat model of adriamycin-induced renal fibrosis with blood stasis syndrome to explore the mechanism of Notoginseng Radix et Rhizoma in protecting the kidney. Thirty male rats with adriamycin-induced renal fibrosis were randomized into model, low-, medium-, and high-dose Notoginseng Radix et Rhizoma, and positive control groups(n=6). Six clean SD male rats were selected into the normal group. The normal group and model group were administrated with normal saline, and other groups with corresponding drugs. After 8 weeks of treatment, the renal function, renal pathology, adenosine triphosphate(ATP) levels, Na~+-K~+-ATPase and Ca~(2+)-Mg~(2+)-ATPase activities, and the protein levels of ATP5B, mTORC1, 70 kDa ribosomal protein S6 kinase(P70S6K), P85, Akt, p-Akt, and SH2-containing inositol phosphatase(SHIP2) in the renal tissue were determined. Compared with the normal group, the model group showed elevated levels of blood urea nitrogen(BUN) and serum creatinine(SCr)(P<0.01). Compared with the model group, Notoginseng Radix et Rhizoma and the positive control lowered the levels of BUN and SCr, which were significant in the medium-and high-dose Noto-ginseng Radix et Rhizoma groups and the positive control group(P<0.05). Compared with the model group, Notoginseng Radix et Rhizoma and the positive control alleviated the pathological changes in the renal tissue, such as vacuolar and fibroid changes, glomerulus atrophy, cystic expansion of renal tubules, and massive infiltration of inflammatory cells. Compared with the normal group, the model group showed decreased mitochondrial ATP content and Na~+-K~+-ATPase and Ca~(2+)-Mg~(2+)-ATPase activities in the renal tissue(P<0.05), and medium-and high-dose Notoginseng Radix et Rhizoma and positive control mitigated such decreases(P<0.05). Compared with the model group, medium-and high-dose Notoginseng Radix et Rhizoma and the positive control up-regulated the protein levels of ATP5B and SHIP2 and down-regulated the protein levels of mTORC1, P70S6K, P85, Akt, and p-Akt(P<0.05 or P<0.01 or P<0.001). Notoginseng Radix et Rhizoma may exert an anti-fibrosis effect by inhibiting the activation of the PI3K/Akt/mTORC1 pathway to restore mitochondrial energy metabolism, thus protecting the kidney.

[Research progress in the regulation of autophagy and mitochondrial homeostasis by AMPK signaling channels].

AMP-activated protein kinase (AMPK) is a widely distributed and evolutionarily conserved serine/threonine protein kinase present in eukaryotic cells. In regulating cellular energy metabolism, AMPK plays an extremely important role as an energy metabolic kinase. When the body is in a low energy state, AMPK is activated in response to changes in intracellular adenine nucleotide levels and is bound to adenosine monophosphate (AMP) or adenosine diphosphate (ADP). Activated AMPK regulates various metabolic processes, including lipid and glucose metabolism and cellular autophagy. AMPK directly promotes autophagy by phosphorylating autophagy-related proteins in the mammalian target of rapamycin complex 1 (mTORC1), serine/threonine protein kinase-dysregulated 51-like kinase 1 (ULK1) and type III phosphatidylinositol 3-kinase-vacuolar protein-sorting 34 (PIK3C3-VPS34) complexes. AMPK also indirectly promotes autophagy by regulating the expression of downstream autophagy-related genes of transcription factors such as forkhead box O3 (FOXO3), lysosomal function transcription factor EB (TFEB) and bromodomain protein 4 (BRD4). AMPK also regulates mitochondrial autophagy, induces the division of damaged mitochondria and promotes the transfer of the autophagic response to damaged mitochondria. Another function of AMPK is to regulate mitochondrial health by stimulating mitochondrial biogenesis and participating in various aspects of mitochondrial homeostasis regulation. This review discusses the specific regulation of mitochondrial biology and internal environmental homeostasis by AMPK signaling channels as central to the cellular response to energy stress and regulation of mitochondria, highlighting the key role of AMPK in regulating cellular autophagy and mitochondrial autophagy, as well as advances in research on the regulation of mitochondrial homeostasis.

Synergistic hyperactivation of both mTORC1 and mTORC2 underlies the neural abnormalities of PTEN-deficient human neurons and cortical organoids.

Mutations in the phosphatase and tensin homolog (PTEN) gene are associated with severe neurodevelopmental disorders. Loss of PTEN leads to hyperactivation of the mechanistic target of rapamycin (mTOR), which functions in two distinct protein complexes, mTORC1 and mTORC2. The downstream signaling mechanisms that contribute to PTEN mutant phenotypes are not well delineated. Here, we show that pluripotent stem cell-derived PTEN mutant human neurons, neural precursors, and cortical organoids recapitulate disease-relevant phenotypes, including hypertrophy, electrical hyperactivity, enhanced proliferation, and structural overgrowth. PTEN loss leads to simultaneous hyperactivation of mTORC1 and mTORC2. We dissect the contribution of mTORC1 and mTORC2 by generating double mutants of PTEN and RPTOR or RICTOR, respectively. Our results reveal that the synergistic hyperactivation of both mTORC1 and mTORC2 is essential for the PTEN mutant human neural phenotypes. Together, our findings provide insights into the molecular mechanisms that underlie PTEN-related neural disorders and highlight novel therapeutic targets.

The autophagy inhibitor NSC185058 suppresses mTORC1-mediated protein anabolism in cultured skeletal muscle.

The mammalian target of rapamycin (mTOR), and specifically the mTOR complex 1 (mTORC1) is the central regulator of anabolism in skeletal muscle. Among the many functions of this kinase complex is the inhibition of the catabolic process of autophagy; however, less work has been done in investigating the role of autophagy in regulating mTORC1 signaling. Using an in vitro model to better understand the pathways involved, we activated mTORC1 by several different means (growth factors, leucine supplementation, or muscle contraction), alone or with the autophagy inhibitor NSC185058. We found that inhibiting autophagy with NSC185058 suppresses mTORC1 activity, preventing any increase in cellular protein anabolism. These decrements were the direct result of action on the mTORC1 kinase, which we demonstrate, for the first time, cannot function when autophagy is inhibited by NSC185058. Our results indicate that, far from being a matter of unidirectional action, the relationship between mTORC1 and the autophagic cascade is more nuanced, with autophagy serving as an mTORC1 input, and mTORC1 inhibition of autophagy as a form of homeostatic feedback to regulate anabolic signaling. Future studies of cellular metabolism will have to consider this fundamental intertwining of protein anabolism and catabolism, and how it ultimately serves to regulate muscle proteostasis.

Glutaminolysis regulates endometrial fibrosis in intrauterine adhesion via modulating mitochondrial function.

BACKGROUND: Endometrial fibrosis, a significant characteristic of intrauterine adhesion (IUA), is caused by the excessive differentiation and activation of endometrial stromal cells (ESCs). Glutaminolysis is the metabolic process of glutamine (Gln), which has been implicated in multiple types of organ fibrosis. So far, little is known about whether glutaminolysis plays a role in endometrial fibrosis. METHODS: The activation model of ESCs was constructed by TGF-ß1, followed by RNA-sequencing analysis. Changes in glutaminase1 (GLS1) expression at RNA and protein levels in activated ESCs were verified experimentally. Human IUA samples were collected to verify GLS1 expression in endometrial fibrosis. GLS1 inhibitor and glutamine deprivation were applied to ESCs models to investigate the biological functions and mechanisms of glutaminolysis in ESCs activation. The IUA mice model was established to explore the effect of glutaminolysis inhibition on endometrial fibrosis. RESULTS: We found that GLS1 expression was significantly increased in activated ESCs models and fibrotic endometrium. Glutaminolysis inhibition by GLS1 inhibitor bis-2-(5-phenylacetamido-1,2,4-thiadiazol-2-yl) ethyl sulfide (BPTES or glutamine deprivation treatment suppressed the expression of two fibrotic markers, α-SMA and collagen I, as well as the mitochondrial function and mTORC1 signaling in ESCs. Furthermore, inhibition of the mTORC1 signaling pathway by rapamycin suppressed ESCs activation. In IUA mice models, BPTES treatment significantly ameliorated endometrial fibrosis and improved pregnancy outcomes. CONCLUSION: Glutaminolysis and glutaminolysis-associated mTOR signaling play a role in the activation of ESCs and the pathogenesis of endometrial fibrosis through regulating mitochondrial function. Glutaminolysis inhibition suppresses the activation of ESCs, which might be a novel therapeutic strategy for IUA.

Fraxetin pretreatment alleviates cisplatin-induced kidney injury by antagonizing autophagy and apoptosis via mTORC1 activation.

Cisplatin-induced kidney injury (CKI) is a common complication of chemotherapy. Fraxetin, derived from Fraxinus bungeana A. DC. bark, has antioxidant, anti-inflammatory, and anti-fibrotic effects. This study aims to investigate fraxetin's effects on CKI and its underlying mechanism in vivo and in vitro. Tubular epithelial cells (TECs) and mice were exposed to cisplatin with and without fraxetin preconditioning assess fraxetin's role in CKI. TECs autophagy was observed using transmission electron microscopy. Apoptosis levels in animal tissues were measured using TUNEL staining. The protective mechanism of fraxetin was explored through pharmacological and genetic regulation of mTORC1. Molecular docking was used to identify potential binding sites between fraxetin and mTORC1. The results indicated that fraxetin pretreatment reduced cisplatin-induced kidney injury in a time- and concentration-dependent way. Fraxetin also decreased autophagy in TECs, as observed through electron microscopy. Tissue staining confirmed that fraxetin pretreatment significantly reduced cisplatin-induced apoptosis. Inhibition of mTORC1 using rapamycin or siRNA reversed the protective effects of fraxetin on apoptosis and autophagy in cisplatin-treated TECs, while activation of mTORC1 enhanced fraxetin's protective effect. Molecular docking analysis revealed that fraxetin can bind to HEAT-repeats binding site on mTORC1 protein. In  summary, fraxetin pretreatment alleviates CKI by antagonizing autophagy and apoptosis via mTORC1 activation. This provides evidence for the potential therapeutic application of fraxetin in CKI.

Malvidin attenuates trauma-induced heterotopic ossification of tendon in rats by targeting Rheb for degradation via the ubiquitin-proteasome pathway.

The pathogenesis of trauma-induced heterotopic ossification (HO) in the tendon remains unclear, posing a challenging hurdle in treatment. Recognizing inflammation as the root cause of HO, anti-inflammatory agents hold promise for its management. Malvidin (MA), possessing anti-inflammatory properties, emerges as a potential agent to impede HO progression. This study aimed to investigate the effect of MA in treating trauma-induced HO and unravel its underlying mechanisms. Herein, the effectiveness of MA in preventing HO formation was assessed through local injection in a rat model. The potential mechanism underlying MA's treatment was investigated in the tendon-resident progenitor cells of tendon-derived stem cells (TDSCs), exploring its pathway in HO formation. The findings demonstrated that MA effectively hindered the osteogenic differentiation of TDSCs by inhibiting the mTORC1 signalling pathway, consequently impeding the progression of trauma-induced HO of Achilles tendon in rats. Specifically, MA facilitated the degradation of Rheb through the K48-linked ubiquitination-proteasome pathway by modulating USP4 and intercepted the interaction between Rheb and the mTORC1 complex, thus inhibiting the mTORC1 signalling pathway. Hence, MA presents itself as a promising candidate for treating trauma-induced HO in the Achilles tendon, acting by targeting Rheb for degradation through the ubiquitin-proteasome pathway.

mTORC1 - TFEB pathway was involved in sodium arsenite induced lysosomal alteration, oxidative stress and genetic damage in BEAS-2B cells.

The mechanistic target of rapamycin (RAPA) complex 1 (mTORC1) - transcription factor EB (TFEB) pathway plays a crucial role in response to nutritional status, energy and environmental stress for maintaining cellular homeostasis. But there is few reports on its role in the toxic effects of arsenic exposure and the related mechanisms. Here, we show that the exposure of bronchial epithelial cells (BEAS-2B) to sodium arsenite promoted the activation of mTORC1 (p-mTORC1) and the inactivation of TFEB (p-TFEB), the number and activity of lysosomes decreased, the content of reduced glutathione (GSH) and superoxide dismutase (SOD) decreased, the content of malondialdehyde (MDA) increased, the DNA and chromosome damage elevated. Further, when mTORC1 was inhibited with RAPA, p-mTORC1 and p-TFEB down-regulated, GSH and SOD increased, MDA decreased, the DNA and chromosome damage reduced significantly, as compared with the control group. Our data revealed for the first time that mTORC1 - TFEB pathway was involved in sodium arsenite induced lysosomal alteration, oxidative stress and genetic damage in BEAS-2B cells, and it may be a potential intervention target for the toxic effects of arsenic.

[Effect of Erchen Decoction on liver mitochondrial function by inhibiting mTORC1/SREBP1/CAV1 pathway in mice with high-fat diet].

This study aims to investigate the effect of Erchen Decoction(ECD) on liver mitochondrial function in mice with a high-fat diet and its possible mechanism. A total of sixty C57BL/6J mice were randomly divided into a normal group, high-fat group, ECD group, mTORC1 activator(MHY) group, ECD+MHY group, and polyene phosphatidyl choline(PPC) group, with 10 rats in each group. The normal group was given a normal diet, and the other groups were fed a high-fat diet for 20 weeks. At the 17th week, the ECD group and ECD+MHY group were given ECD(8.7 g·kg~(-1)) daily, and the PPC group was given PPC(0.18 g·kg~(-1)) daily, while the remaining groups were given normal saline(0.01 mL·g~(-1)) daily for four weeks. In the 19th week, the MHY group and ECD+MHY group were injected intraperitoneally with MHY(5 mg·kg~(-1)) every other day for two weeks. During the experiment, the general conditions of the mice were observed. The contents of triglyceride(TG) and total cholesterol(TC) in serum were measured. Morphological changes in liver tissue were examined through HE and oil red O staining. The content of adenosine triphosphate(ATP) was determined using chemiluminescence, and mitochondrial membrane potential was assessed using a fluorescence probe(JC-1). Western blot was performed to detect the expression of rapamycin target protein complex 1(mTOR1), ribosomal protein S6 kinase B1(S6K), sterol regulatory element binding protein 1(SREBP1), and caveolin 1(CAV1). RESULTS:: revealed that compared with the normal group, the mice in the high-fat group exhibited significant increases in body weight and abdominal circumference(P<0.01). Additionally, there were significant increases in TG and TC levels(P<0.01). HE and oil red O staining showed that the boundaries of hepatic lobules were unclear; hepatocytes were enlarged, round, and irregularly arranged, with obvious lipid droplet deposition and inflammatory cell infiltration. The liver ATP content and mitochondrial membrane potential decreased significantly(P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 increased significantly(P<0.01), while the expression of CAV1 decreased significantly(P<0.01). Compared with the high-fat group, the body weight and TG content of mice in the ECD group and PPC group decreased significantly(P<0.05). Improvements were observed in hepatocyte morphology, lipid deposition, and inflammatory cell infiltration. Furthermore, there were significant increases in ATP content and mitochondrial membrane potential(P<0.05 or P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly in the ECD group(P<0.01), while CAV1 expression increased significantly(P<0.01). However, the indices mentioned above did not show improvement in the MHY group. When the ECD+MHY group was compared with the MHY group, there were significant reductions in body weight and TG contents(P<0.05). The morphological changes of hepatocytes, lipid deposition, and inflammatory cell infiltration were recovered. Moreover, there were significant increases in liver ATP content and mitochondrial membrane potential(P<0.05 or P<0.05). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly(P<0.01), while CAV1 expression increased significantly(P<0.01). In conclusion, ECD can improve mitochondrial function by regulating the mTORC1/SREBP1/CAV1 pathway. This mechanism may be involved in the resolution of phlegm syndrome and the regulation of lipid metabolism.

[High-intensity interval training (HIIT) induces hepatic ketone body production possibly through altering expression of mTORC1, PPARα and FGF21 in mice].

The present study aims to investigate the production of ketone body in the liver of mice after 6 weeks of high-intensity interval training (HIIT) intervention and explore the possible mechanisms. Male C57BL/6J mice (7-week-old) were randomly divided into control and HIIT groups. The control group did not engage in exercise, while the HIIT group underwent a 6-week HIIT (10° slope treadmill exercise). Changes in weight and body composition were recorded, and blood ketone body levels were measured before, immediately after, and 1 h after each HIIT exercise. After 6-week HIIT, the levels of free fatty acids in the liver and serum were detected using reagent kits, and expression levels of regulatory factors and key enzymes of ketone body production in the mouse liver were detected by Western blot and qPCR. The results showed that, the blood ketone body levels in the HIIT group significantly increased immediately after a single HIIT and 1 h after HIIT, compared with that before HIIT. The body weight of the control group gradually increased within 6 weeks, while the HIIT group mice did not show significant weight gain. After 6-week HIIT, compared with the control group, the HIIT group showed decreased body fat ratio, increased lean body weight ratio, and increased free fatty acid levels in liver and serum. Liver carnitine palmitoyl transferase-I (CPT-I), peroxisome proliferator activated receptor α (PPARα), and fibroblast growth factor 21 (FGF21) protein expression levels were up-regulated, whereas mammalian target of rapamycin complex 1 (mTORC1) protein expression level was significantly down-regulated in the HIIT group, compared with those in the control group. These results suggest that HIIT induces hepatic ketone body production through altering mTORC1, PPARα and FGF21 expression in mice.