Sodium-glucose cotransporter 1 promotes the biofunctions of perivascular preadipocytes mediated by Akt/mTOR/p70S6K signaling pathway.

The influence of SGLT-1 on perivascular preadipocytes (PVPACs) and vascular remodeling is not well understood. This study aimed to elucidate the role and mechanism of SGLT-1-mediated PVPACs bioactivity. PVPACs were cultured in vitro and applied ex vivo to the carotid arteries of mice using a lentivirus-based thermosensitive in situ gel (TISG). The groups were treated with Lv-SGLT1 (lentiviral vector, overexpression), Lv-siSGLT1 (RNA interference, knockdown), or specific signaling pathway inhibitors. Assays were conducted to assess changes in cell proliferation, apoptosis, glucose uptake, adipogenic differentiation, and vascular remodeling in the PVPACs. Protein expression was analyzed by Western blotting, immunocytochemistry, and/or immunohistochemistry. The methyl thiazolyl tetrazolium (MTT) assay and Hoechst 33342 staining indicated that SGLT-1 overexpression significantly promoted PVPACs proliferation and inhibited apoptosis in vitro. Conversely, SGLT-1 knockdown exerted the opposite effect. Oil Red O staining revealed that SGLT-1 overexpression facilitated adipogenic differentiation, while its inhibition mitigated these effects. 3H-labeled glucose uptake experiments demonstrated that SGLT-1 overexpression enhanced glucose uptake by PVPACs, whereas RNA interference-mediated SGLT-1 inhibition had no significant effect on glucose uptake. Moreover, RT-qPCR, Western blotting, and immunofluorescence analyses revealed that SGLT-1 overexpression upregulated FABP4 and VEGF-A levels and activated the Akt/mTOR/p70S6K signaling pathway, whereas SGLT-1 knockdown produced the opposite effects. In vivo studies corroborated these findings and indicated that SGLT-1 overexpression facilitated carotid artery remodeling. Our study demonstrates that SGLT-1 activation of the Akt/mTOR/p70S6K signaling pathway promotes PVPACs proliferation, adipogenesis, glucose uptake, glucolipid metabolism, and vascular remodeling.NEW & NOTEWORTHY SGLT-1 is expressed in PVPACs and can affect preadipocyte glucolipid metabolism and vascular remodeling. SGLT-1 promotes the biofunctions of PVPACs mediated by Akt/mTOR/p70S6K signaling pathway. Compared with caudal vein or intraperitoneal injection, the external application of lentivirus-based thermal gel around the carotid artery is an innovative attempt at vascular remodeling model, it may effectively avoid the transfection of lentiviral vector into the whole body of mice and the adverse effect on experimental results.

Lipoic acid-mediated oral drug delivery system utilizing changes on cell surface thiol expression for the treatment of diabetes and inflammatory diseases.

Lipoic acid (LA), which has good safety and oral absorption, is obtained from various plant-based food sources and needs to be supplemented through human diet. Moreover, substances with a disulfide structure can enter cells through dynamic covalent disulfide exchange with thiol groups on the cell membrane surface. Based on these factors, we constructed LA-modified nanoparticles (LA NPs). Our results showed that LA NPs can be internalized into intestinal epithelial cells through surface thiols, followed by intracellular transcytosis via the endoplasmic reticulum-Golgi pathway. Further mechanistic studies indicated that disulfide bonds within the structure of LA play a critical role in this transport process. In a type I diabetes rat model, the oral administration of insulin-loaded LA NPs exhibited a more potent hypoglycemic effect, with a pharmacokinetic bioavailability of 5.42 ± 0.53%, representing a 1.6 fold enhancement compared to unmodified PEG NPs. Furthermore, a significant upregulation of surface thiols in inflammatory macrophages was reported. Thus, we turned our direction to investigate the uptake behavior of inflammatory macrophages with increased surface thiols towards LA NPs. Inflammatory macrophages showed a 2.6 fold increased uptake of LA NPs compared to non-inflammatory macrophages. Surprisingly, we also discovered that the antioxidant resveratrol facilitates the uptake of LA NPs in a concentration-dependent manner. This is mainly attributed to an increase in glutathione, which is involved in thiol uptake. Consequently, we employed LA NPs loaded with resveratrol for the treatment of colitis and observed a significant alleviation of colitis symptoms. These results suggest that leveraging the variations of thiol expression levels on cell surfaces under both healthy and diseased states through an oral drug delivery system mediated by the small-molecule nutrient LA can be employed for the treatment of diabetes and certain inflammatory diseases.

A Less-is-More Strategy for Mitochondria-Targeted Photodynamic Therapy of Rheumatoid Arthritis.

Conventional photodynamic therapy (PDT) of rheumatoid arthritis (RA) faces a dilemma: low-power is insufficient to kill pro-inflammatory cells while high-power exacerbates inflammation. Herein, mitochondrial targeting is introduced in PDT of RA to implement a "less-is-more" strategy, where higher apoptosis in pro-inflammatory cells are achieved with lower laser power. In arthritic rats, chlorine 6-loaded and mitochondria-targeting liposomes (Ce6@M-Lip) passively accumulated in inflamed joints, entered pro-inflammatory macrophages, and actively localized to mitochondria, leading to enhanced mitochondrial dysfunction under laser irradiation. By effectively disrupting mitochondria, pro-inflammatory macrophages are more susceptible to PDT, resulting in increased apoptosis initiation. Additionally, it identifies that high-power irradiation caused cell rupture and release of endogenous danger signals that recruited and activated additional macrophages. In contrast, under low-power irradiation, mitochondria-targeting Ce6@M-Lip not only prevented inflammation but also reduced pro-inflammatory macrophage infiltration and pro-inflammatory cytokine secretion. Overall, targeting mitochondria reconciled therapeutic efficacy and inflammation, thus enabling efficacious yet inflammation-sparing PDT for RA. This highlights the promise of mitochondrial targeting to resolve the dilemma between anti-inflammatory efficacy and inflammatory exacerbation in PDT by implementing a "less-is-more" strategy.

Biomimetic nanoparticle synchronizing pyroptosis induction and mitophagy inhibition for anti-tumor therapy.

Inducing pyroptosis in cancer cells can result in a strong anti-tumor immune response. Our preliminary study indicates that pyroptosis can be temporarily strengthened by disrupting mitochondria, but ultimately diminished by defensive mitophagy. Here, this study reports a nano-system camouflaged with hybrid membranes consisting of homologous cell membrane and corresponding mitochondrial membrane, which is used to deliver a drug complex Ca@GOx consisting of calcium phosphate and glucose oxidase. By taking advantage of the homing effects of cell membrane and the orientated fusion mechanism of subcellular membrane, the nano-system is able to deliver Ca@GOx to mitochondria, induce mitochondrial Ca2+ overload and generate significant levels of ROS, thus leading to pyroptosis. However, it's found that this system exhibits limited anti-tumor effects in vivo due to the compensatory activation of mitophagy serving as negative feedback to pyroptosis. To address this issue, mitophagy-inhibiting chloroquine is loaded into nanoparticles to intensify pyroptosis. As a result, the combination significantly promotes tumor infiltration of CD8+T cells and improves anti-tumor effects. Together, this study establishes a rational combination of targeted mitochondria disruption and mitophagy blockage for effective pyroptosis-based therapy.

Mimicking natural cholesterol assimilation to elevate the oral delivery of liraglutide for type II diabetes therapy.

Glucagon-like peptide-1 receptor agonists (GLP-1 RA) are a series of polypeptides broadly applied in the long-term treatment of type Ⅱ diabetes. However, administration of GLP-RA is mainly through repetitive subcutaneous injection, which may seriously decrease the compliance and safety. Herein, a bio-inspired oral delivery system was designed to enhance the oral absorption of liraglutide (Lira), a kind of GLP-1 RA, by mimicking the natural cholesterol assimilation. 25-hydroxycholesterol (25HC), a cholesterol derivative, was modified on the surfaced of Lira-loaded PLGA nanoparticles (Lira 25HC NPs) and functioned as a "top-down" actuator to facilitate unidirectional transcytosis across the intestinal epithelium. After oral delivery, Lira 25HC NPs displayed improved therapeutic effect as compared with oral free Lira on type Ⅱ diabetes db/db mice, as evidenced by multiple relieved diabetic symptoms including the enhanced glucose tolerance, repressed weight growth, improved liver glucose metabolism, decreased fasting blood glucose, HbA1c, serum lipid, and increased ß cells activity. Surprisingly, the fasting blood glucose, liver glucose metabolism, and HbA1c of oral Lira-loaded 25HC NPs were comparable to subcutaneous injection of free Lira. Further mechanisms revealed that 25HC ligand could mediate the nanoparticles to mimic natural cholesterol absorption by exerting high affinity towards apical Niemann-Pick C1 Like 1 (NPC1L1) and then basolateral ATP binding cassette transporter A1 (ABCA1) overexpressed on the opposite side of intestinal epithelium. This cholesterol assimilation-mimicking strategy achieve the unidirectional transport across the intestinal epithelium, thus improving the oral absorption of liraglutide. In general, this study established a cholesterol simulated platform and provide promising insight for the oral delivery of GLP-1 RA.

Angiopep-2-functionalized nanoparticles enhance transport of protein drugs across intestinal epithelia by self-regulation of targeted receptors.

Ligand-modified nanoparticles (NPs) have been widely used in oral drug delivery systems to promote endocytosis on intestinal epithelia. However, their transcytosis across the intestinal epithelia is still limited. Except for complex intracellular trafficking, recycling again from the apical sides into the intestinal lumen of the endocytosed NPs cannot be ignored. In this study, we modified NP surfaces with angiopep-2 (ANG) that targeted the low-density lipoprotein receptor-related protein 1 (LRP-1) expressed on the intestine to increase both the apical endocytosis and basolateral transcytosis of NPs. Notably, our finding revealed that ANG NPs could increase the apical expression and further basolateral redistribution of LRP-1 on Caco-2 cells, thus generating an apical-to-basolateral absorption pattern. Because of the enhanced transcytosis, insulin loaded ANG NPs possessed much stronger absorption efficiency and induced maximal blood glucose reduction to 61.46% in diabetic rats. Self-regulating the distribution of receptors on polarized intestine cells to promote basolateral transcytosis will provide promising insights for the rational design of oral delivery systems of protein/peptide drugs.

Complying with the physiological functions of Golgi apparatus for secretory exocytosis facilitated oral absorption of protein drugs.

Intestinal epithelial cells are the primary biological barriers for orally administrated nano-formulations and the delivered protein drugs. Thereinto, besides the cellular uptake, intracellular trafficking pathway and the related exocytosis are of great importance to the trans-epithelial transport of drug-loaded NPs. Herein, inspired by the physiological functions of Golgi apparatus for secreting proteins out of cells, Golgi localization-related amino acid l-cysteine (Cys) was modified on the surface of NPs to see whether and how this modification could guide the Golgi pathway-related transport and facilitate the exocytosis of drug-loaded NPs. Meanwhile, cell-penetrating peptide octa-arginine (R8) was co-modified to increase the cellular uptake. The proportion of R8 and Cys modification was explored to get the best effect of endocytosis and exocytosis of NPs. As a result, 25%R8 + 75%Cys NPs with most Cys modification showed efficient transcytosis with the highest transcytosis/endocytosis ratio (0.87). Interestingly, exocytosis mechanism studies indicated that they trafficked through the Golgi secretory pathway and bypassed lysosomes due to Cys modification. The detailed Golgi position mechanism studies further suggested that the thiol group from Cys was important for mediating Golgi transport. In particular, competitive inhibition studies demonstrated that Cys-modified NPs were more conducive to their exocytosis after being transported through the Golgi secretory pathway. We proved that cargos transported via Golgi apparatus tended to be trafficked out of the cells and avoid degradation, which contributed to the transcytosis of 25%R8 + 75%Cys NPs in vitro. Inspiringly, compared with unmodified NPs, 25%R8 + 75%Cys NPs also exhibited promoted intestinal penetration and oral absorption in vivo. Oral delivery of insulin-loaded 25%R8 + 75%Cys NPs showed stronger hypoglycemic effects in diabetic rats. In summary, this work provides a strategy for complying with the physiological functions of Golgi apparatus for secreting to facilitate the exocytosis of NPs, thus further improving the oral absorption of loaded protein drugs.