Mecp2 fine-tunes quiescence exit by targeting nuclear receptors.

Quiescence (G0) maintenance and exit are crucial for tissue homeostasis and regeneration in mammals. Here, we show that methyl-CpG binding protein 2 (Mecp2) expression is cell cycle-dependent and negatively regulates quiescence exit in cultured cells and in an injury-induced liver regeneration mouse model. Specifically, acute reduction of Mecp2 is required for efficient quiescence exit as deletion of Mecp2 accelerates, while overexpression of Mecp2 delays quiescence exit, and forced expression of Mecp2 after Mecp2 conditional knockout rescues cell cycle reentry. The E3 ligase Nedd4 mediates the ubiquitination and degradation of Mecp2, and thus facilitates quiescence exit. A genome-wide study uncovered the dual role of Mecp2 in preventing quiescence exit by transcriptionally activating metabolic genes while repressing proliferation-associated genes. Particularly disruption of two nuclear receptors, Rara or Nr1h3, accelerates quiescence exit, mimicking the Mecp2 depletion phenotype. Our studies unravel a previously unrecognized role for Mecp2 as an essential regulator of quiescence exit and tissue regeneration.

CFD simulation of cannabidiol delivery through microneedle patches.

This study investigates the efficiency and influence of microneedle parameters, specifically Needle Point Angle (a) and Needle Height (h), on the diffusion of Cannabidiol (CBD) across varying skin depths. Utilizing the Latin Hypercube Sampling method, twelve distinct cases were analyzed. Observations reveal a consistent high concentration of CBD delivered via the microneedle patch, with a notable decrease in concentration as the depth increases, displaying a non-linear trend. Multivariate polynomial regression offers a quantitative relationship between the variables, with the third-order bivariate fitting providing the most accurate representation. Compared to other CBD delivery mechanisms, microneedle patches present enhanced CBD concentrations, circumventing challenges faced by other methods such as dosage inaccuracy, systemic absorption issues, and CBD degradation. The results highlight the potential of microneedle patches as a promising avenue for optimized transdermal drug delivery.

ANGPTL4 binds to the leptin receptor to regulate ectopic bone formation.

Leptin protein was thought to be unique to leptin receptor (LepR), but the phenotypes of mice with mutation in LepR [db/db (diabetes)] and leptin [ob/ob (obese)] are not identical, and the cause remains unclear. Here, we show that db/db, but not ob/ob, mice had defect in tenotomy-induced heterotopic ossification (HO), implicating alternative ligand(s) for LepR might be involved. Ligand screening revealed that ANGPTL4 (angiopoietin-like protein 4), a stress and fasting-induced factor, was elicited from brown adipose tissue after tenotomy, bound to LepR on PRRX1+ mesenchymal cells at the HO site, thus promotes chondrogenesis and HO development. Disruption of LepR in PRRX1+ cells, or lineage ablation of LepR+ cells, or deletion of ANGPTL4 impeded chondrogenesis and HO in mice. Together, these findings identify ANGPTL4 as a ligand for LepR to regulate the formation of acquired HO.

Exosome Release Delays Senescence by Disposing of Obsolete Biomolecules.

Accumulation of obsolete biomolecules can accelerate cell senescence and organism aging. The two efficient intracellular systems, namely the ubiquitin-proteasome system and the autophagy-lysosome system, play important roles in dealing with cellular wastes. However, how multicellular organisms orchestrate the processing of obsolete molecules and delay aging remains unclear. Herein, it is shown that prevention of exosome release by GW4869 or Rab27a-/- accelerated senescence in various cells and mice, while stimulating exosome release by nutrient restriction delays aging. Interestingly, exosomes isolate from serum-deprived cells or diet-restricted human plasma, enriched with garbage biomolecules, including misfolded proteins, oxidized lipids, and proteins. These cellular wastes can be englobed by macrophages, eventually, for disintegration in vivo. Inhibition of nutrient-sensing mTORC1 signaling increases exosome release and delays senescence, while constitutive activation of mTORC1 reduces exosome secretion and exacerbates senescence in vitro and in mice. Notably, inhibition of exosome release attenuates nutrient restriction- or rapamycin-delayed senescence, supporting a key role for exosome secretion in this process. This study reveals a potential mechanism by which stimulated exosome release delays aging in multicellular organisms, by orchestrating the harmful biomolecules disposal via exosomes and macrophages.

Tsc1 regulates tight junction independent of mTORC1.

Tuberous sclerosis complex 1 (Tsc1) is a tumor suppressor that functions together with Tsc2 to negatively regulate the mechanistic target of rapamycin complex 1 (mTORC1) activity. Here, we show that Tsc1 has a critical role in the tight junction (TJ) formation of epithelium, independent of its role in Tsc2 and mTORC1 regulation. When an epithelial cell establishes contact with neighboring cells, Tsc1, but not Tsc2, migrates from the cytoplasm to junctional membranes, in which it binds myosin 6 to anchor the perijunctional actin cytoskeleton to ß-catenin and ZO-1. In its absence, perijunctional actin cytoskeleton fails to form. In mice, intestine-specific or inducible, whole-body Tsc1 ablation disrupts adherens junction/TJ structures in intestine or skin epithelia, respectively, causing Crohn's disease-like symptoms in the intestine or psoriasis-like phenotypes on the skin. In patients with Crohn's disease or psoriasis, junctional Tsc1 levels in epithelial tissues are markedly reduced, concomitant with the TJ structure impairment, suggesting that Tsc1 deficiency may underlie TJ-related diseases. These findings establish an essential role of Tsc1 in the formation of cell junctions and underpin its association with TJ-related human diseases.

Exosome Release Is Regulated by mTORC1.

Exosomes are small membrane-bound vesicles released into extracellular spaces by many types of cells. These nanovesicles carry proteins, mRNA, and miRNA, and are involved in cell waste management and intercellular communication. In the present study, it is shown that exosome release, which leads to net loss of cellular membrane and protein content, is negatively regulated by mechanistic target of rapamycin complex 1 (mTORC1). It is found that in cells and animal models exosome release is inhibited by sustained activation of mTORC1, leading to intracellular accumulation of CD63-positive exosome precursors. Inhibition of mTORC1 by rapamycin or nutrient and growth factor deprivation stimulates exosome release, which occurs concomitantly with autophagy. The drug-stimulated release is blocked by siRNA-mediated downregulation of small GTPase Rab27A. Analysis of the cargo content in exosomes released from rapamycin-treated cells reveals that inhibition of mTORC1 does not significantly alter its majority protein and miRNA profiles. These observations demonstrate that exosome release, like autophagy, is negatively regulated by mTORC1 in response to changes in nutrient and growth factor conditions.

Colonic epithelial mTORC1 promotes ulcerative colitis through COX-2-mediated Th17 responses.

The functional role of colonic epithelium in the pathogenesis of ulcerative colitis (UC) remains unclear. Here, we reveal a novel mechanism by which colonic epithelia recruit T helper-17 (Th17) cells during the onset of UC. mTOR complex 1 (mTORC1) was hyper-activated in colonic epithelia of UC mice. While colonic epithelial TSC1 (mTORC1 negative regulator) disruption induced constitutive mTORC1 activation in the colon epithelia and aggravated UC, RPTOR (essential mTORC1 component) depletion inactivated mTORC1 and ameliorated UC. TSC1 deficiency enhanced, whereas RPTOR ablation reduced the expression of cyclooxygenase 2 (COX-2), interleukin-1 (IL-1), IL-6, and IL-23, as well as Th17 infiltration in the colon. Importantly, inhibition of COX-2 reversed the elevation in the expression of these proinflammatory mediators induced by TSC1 deficiency, and subsequently reduced the symptoms and pathological characteristics of UC in mouse models. Mechanistically, mTORC1 activates COX-2 transcription via phosphorylating STAT3 and enhancing it's binding to the COX-2 promoter. Consistently, enhanced mTORC1 activity and COX2 expression, as well as strong positive correlation between each other, were observed in colonic epithelial tissues of UC patients. Collectively, our study demonstrates an essential role of epithelial mTORC1 in UC pathogenesis and establishes a novel link between colonic epithelium, Th17 responses, and UC development.

Casticin attenuates liver fibrosis and hepatic stellate cell activation by blocking TGF-ß/Smad signaling pathway.

Although many advances have been made in understanding the pathogenesis of liver fibrosis, few options are available for treatment. Casticin, one of the major flavonoids in Fructus Viticis extracts, has shown hepatoprotective potential, but its effects on liver fibrosis are not clear. In this study, we investigated the antifibrotic activity of casticin and its underlying mechanism in vivo and in vitro. Male mice were injected intraperitoneally with carbon tetrachloride (CCl4) or underwent bile duct ligation (BDL) to induce liver fibrosis, followed by treatment with casticin or vehicle. In addition, transforming growth factor-ß1(TGF-ß1)-activated LX-2 cells were used. In vivo experiments showed that treatment with casticin alone had no toxic effect while significantly attenuating CCl4-or BDL-induced liver fibrosis, as indicated by reductions in the density of fibrosis, hydroxyproline content, expression of α-SMA and collagen α1(I) mRNA. Moreover, casticin inhibited LX2 proliferation, induced apoptosis in a time- and dose-dependent manner in vitro. The underlying molecular mechanisms for the effect of casticin involved inhibition of hepatic stellate cell (HSC) activation and reduced the expression of matrix metalloproteinase (MMP)-2, MMP-9, tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2 resulting from blocking TGF-ß1/Smad signaling, as well as increased the apoptosis of HSCs. The results suggest that casticin has potential benefits in the attenuation and treatment of liver fibrosis.