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1.
Molecules ; 27(21)2022 Oct 27.
Artigo em Inglês | MEDLINE | ID: mdl-36364116

RESUMO

Currently, particular interest among the scientific community is focused on exploring the use of exosomes for several pharmaceutical and biomedical applications. This is due to the identification of the role of exosomes as an excellent intercellular communicator by delivering the requisite cargo comprising of functional proteins, metabolites and nucleic acids. Exosomes are the smallest extracellular vesicles (EV) with sizes ranging from 30-100 nm and are derived from endosomes. Exosomes have similar surface morphology to cells and act as a signal transduction channel between cells. They encompass different biomolecules, such as proteins, nucleic acids and lipids, thus rendering them naturally as an attractive drug delivery vehicle. Like the other advanced drug delivery systems, such as polymeric nanoparticles and liposomes to encapsulate drug substances, exosomes also gained much attention in enhancing therapeutic activity. Exosomes present many advantages, such as compatibility with living tissues, low toxicity, extended blood circulation, capability to pass contents from one cell to another, non-immunogenic and special targeting of various cells, making them an excellent therapeutic carrier. Exosome-based molecules for drug delivery are still in the early stages of research and clinical trials. The problems and clinical transition issues related to exosome-based drugs need to be overcome using advanced tools for better understanding and systemic evaluation of exosomes. In this current review, we summarize the most up-to-date knowledge about the complex biological journey of exosomes from biogenesis and secretion, isolation techniques, characterization, loading methods, pharmaceutical and therapeutic applications, challenges and future perspectives of exosomes.


Assuntos
Exossomos , Vesículas Extracelulares , Ácidos Nucleicos , Exossomos/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Lipossomos/metabolismo , Excipientes , Ácidos Nucleicos/metabolismo
2.
ACS Nano ; 16(11): 17802-17846, 2022 Nov 22.
Artigo em Inglês | MEDLINE | ID: mdl-36354238

RESUMO

Exosomes are a subgroup of nanosized extracellular vesicles enclosed by a lipid bilayer membrane and secreted by most eukaryotic cells. They represent a route of intercellular communication and participate in a wide variety of physiological and pathological processes. The biological roles of exosomes rely on their bioactive cargos, including proteins, nucleic acids, and lipids, which are delivered to target cells. Their distinctive properties─innate stability, low immunogenicity, biocompatibility, and good biomembrane penetration capacity─allow them to function as superior natural nanocarriers for efficient drug delivery. Another notably favorable clinical application of exosomes is in diagnostics. They hold various biomolecules from host cells, which are indicative of pathophysiological conditions; therefore, they are considered vital for biomarker discovery in clinical diagnostics. Here, we use data from the CAS Content Collection and provide a landscape overview of the current state and delineate trends in research advancement on exosome applications in therapeutics and diagnostics across time, geography, composition, cargo loading, and development pipelines. We discuss exosome composition and pathway, from their biogenesis and secretion from host cells to recipient cell uptake. We assess methods for exosome isolation and purification, their clinical applications in therapy and diagnostics, their development pipelines, the exploration goals of the companies, the assortment of diseases they aim to treat, development stages of their research, and publication trends. We hope this review will be useful for understanding the current knowledge in the field of medical applications of exosomes, in an effort to further solve the remaining challenges in fulfilling their potential.


Assuntos
Exossomos , Nanopartículas , Sistemas de Liberação de Medicamentos/métodos , Lipossomos/metabolismo , Exossomos/metabolismo
3.
J Agric Food Chem ; 70(40): 12982-12989, 2022 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-36190122

RESUMO

Lactobacillus S-layer protein (SLP) is a biologically active protein on the cell surface. To further elucidate the structures and functions of SLP in Lactobacillus acidophilus CICC 6074, this study was conducted to identify the functional domains of SLP which is responsible for cell wall anchoring, self-assembly, and adhesion. The gene (slpA) of L. acidophilus CICC 6074 SLP was amplified by polymerase chain reaction and speculated functional domains. Fusion proteins of C-terminal truncations from SLP were exogenously expressed in Escherichia coli BL21 (DE3). FITC-labeling N-terminal truncations of SLP were synthesized. The C-terminal domain was more likely to be the binding region, and the cell wall-anchored receptor of SLP was teichoic acid. Furthermore, N-terminal truncations could self-assemble to milk fat globule membrane polar lipid liposomes observed using a fluorescence microscope. Notably, SAN1 (region 32-55) of N-terminal truncations was mainly responsible for the adhesion of SLP to HT-29 cells. These results showed that SLP played a crucial role in the functions of L. acidophilus CICC 6074, which might be of significant reference value for future studies.


Assuntos
Lactobacillus acidophilus , Lipossomos , Proteínas de Bactérias/metabolismo , Parede Celular/química , Escherichia coli/genética , Escherichia coli/metabolismo , Fluoresceína-5-Isotiocianato , Lactobacillus acidophilus/metabolismo , Lipídeos/análise , Lipossomos/metabolismo , Glicoproteínas de Membrana
4.
Oxid Med Cell Longev ; 2022: 4942519, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36193089

RESUMO

The macrophage transformation of inflammatory M1 to anti-inflammatory M2 could be promoted by activating PI3K/AKT signaling pathway. In our previous study, it was found that downregulation of lncRNA260 could ameliorate hypoxic cardiomyocyte injury by regulating IL28RA through the activation of PI3K/AKT signaling pathways. It was suggested that lncRNA260 siRNA could promote the macrophages toward M2 polarization by regulating IL28RA. In this study, lncRNA260 siRNA was used to observe its effect on the polarization of murine bone marrow-derived macrophages (BMDM) and investigate its related mechanisms. lncRNA 260 specific siRNA were designed and synthesized which were transfected into murine BMDM with liposomes. The experiment was divided into three groups: Hypoxia group, Hypoxia+lncRNA 260-specific siRNA transfection group, and Normoxia group. The CD206-APC/CD11b-FITC or CD206-FITC/CD107b (Mac-3) double positive proportions were used to compare the M2 polarization proportions in the hypoxia process by using the immunofluorescence staining method. The p-AKT, Arg 1, PI3KCG, IL28RAV1, and IL28RAV2 protein expression changes were observed by using the western blot method. Compared with the Normoxia group, the M2 proportions were significantly decreased in the Hypoxia group (P < 0.05). Compared with the hypoxia group, the M2 proportions were significantly increased in the Hypoxia+lncRNA260 siRNA transfection group (P < 0.05). In the Hypoxia group, the ratios of Arg 1/ß-Actin, p-AKT/ß-Actin, PI3KCG/ß-Actin, and IL28RAV1/ß-Actin were significantly lower than those in the Normoxia group (P < 0.05). After transfection with lncRNA260 siRNA, the ratios of Arg1/ß-Actin, p-AKT/ß-Actin, PI3KCG/ß-Actin, and IL28RAV1/ß-Actin were significantly higher than those in the Hypoxia group (P < 0.05). Compared with the Normoxia group, the IL28RAV2/ß-Actin in the Hypoxia group was significantly increased (P < 0.05). After transfection with lncRNA260 siRNA, the ratio of IL28RAV2/ß-Actin was significantly decreased than that in the Hypoxia group (P < 0.05). lncRNA260 siRNA could promote the M2 polarization of the hypoxia macrophages by reducing the IL28RAV2 alternative splicing variant, which might be related to the activation of the JAK-STAT and PI3K/AKT signaling pathways. It will provide a new strategy for the anti-inflammation, antioxidative stress therapy, and cardiac remodeling after AMI.


Assuntos
Fosfatidilinositol 3-Quinases , RNA Longo não Codificante , Actinas/metabolismo , Processamento Alternativo , Animais , Anti-Inflamatórios/metabolismo , Fluoresceína-5-Isotiocianato/metabolismo , Hipóxia/metabolismo , Lipossomos/metabolismo , Macrófagos/metabolismo , Camundongos , Estresse Oxidativo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo
5.
Biomater Adv ; 142: 213156, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36302330

RESUMO

Bottom-up approaches in creating artificial cells that can mimic natural cells have significant implications for both basic research and translational application. Among various artificial cell models, liposome is one of the most sophisticated systems. By encapsulating proteins and associated biomolecules, they can functionally reconstitute foundational features of biological cells, such as the ability to divide, communicate, and undergo shape deformation. Yet constructing liposome artificial cells from the genetic level, which is central to generate self-sustained systems remains highly challenging. Indeed, many studies have successfully established the expression of gene-coded proteins inside liposomes. Further, recent endeavors to build a direct integration of gene-expressed proteins for reconstituting molecular functions and phenotypes in liposomes have also significantly increased. Thus, this review presents the development of liposome-based artificial cells to demonstrate the process of gene-expressed proteins and their reconstitution to perform desired molecular and cell-like functions. The molecular and cellular phenotypes discussed here include the self-production of membrane phospholipids, division, shape deformation, self-DNA/RNA replication, fusion, and intercellular communication. Together, this review gives a comprehensive overview of gene-expressing liposomes that can stimulate further research of this technology and achieve artificial cells with superior properties in the future.


Assuntos
Células Artificiais , Células Artificiais/metabolismo , Lipossomos/metabolismo , Proteínas/genética , Fenótipo , Expressão Gênica
6.
Int J Pharm ; 628: 122277, 2022 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-36241032

RESUMO

Co-localization of antigens and immunomodulators in the same antigen-presenting cells (APCs) can powerfully activate APCs and enhance immune responses. In this study, the immunomodulator resveratrol (Res) was encapsulated into quaternized chitosan (QCS) - coated liposomes for developing a new nanoparticle delivery system (QCS-Res-LP), and ovalbumin (OVA) was selected as a model antigen and adsorbed on the surface of QCS-Res-LP. The results showed that the particle size of QCS-Res-LP was 96.3 ± 3.52 nm; the PDI value was 0.280 ± 0.010; the Zeta potential was 9.59 ± 0.36 mV. QCS-Res-LP could encapsulate 76.22 ± 1.02 % resveratrol and adsorb 88.2 ± 16.3 % antigen. QCS-Res-LP effectively promoted the co-uptake of antigen and Res by dendritic cells (DCs) with 50-fold greater than resveratrol liposomes (Res-LP). QCS-Res-LP promoted expression levels of CD80, CD86, IL-2, and IL-12 in DCs. QCS-Res-LP did not cause hemolysis. The levels of ovalbumin-specific IgG antibodies and cytokines were significantly increased in mice vaccinated with ovalbumin-absorbed QCS-Res-LP, which induced a mixed Th1/Th2 immune response. In conclusion, these results demonstrated that QCS-coated liposomes enable the co-delivery of antigens and immunomodulators to induce strong and durable immune responses.


Assuntos
Quitosana , Lipossomos , Animais , Camundongos , Lipossomos/metabolismo , Quitosana/metabolismo , Resveratrol , Ovalbumina , Antígenos , Adjuvantes Imunológicos , Imunidade , Células Dendríticas
7.
Int J Pharm ; 628: 122290, 2022 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-36252643

RESUMO

For most external applied antioxidant whitening ingredients, effective stratum corneum breakthrough, epidermal penetration and dermal deposition are necessary premises for inhibition of melanin production and transfer occurring in stratum basale. Herein, xanthan gum was added into vitamin C-containing flexible liposome (Vc-L) suspension. The long polymer chain of xanthan gum string dispersed Vc-L together to gain a lotion (Vc-LX) for external application. In this study, the storage stability experiments demonstrated that the additional xanthan gum could improve the storage stability of Vc liposome suspension. The cumulative in vitro skin penetration and deposition of Vc-LX was found to significantly increase within 24 h in mouse skin, compared with those of the Vc aqueous solution and Vc conventional liposomes treated groups (***p < 0.001). Most importantly, in vivo skin whitening experiments gave that Vc-LX has better skin whitening activity (ΔL*) than marketed products (GARNIER® Vc377), Vc flexible liposomes, and Vc conventional liposomes. Moreover, in vitro cytotoxicity experiments and skin irritation experiments demonstrated that Vc-LX has good biosafety. Therefore, this study suggested that Vc-LX may be a promising local skin delivery system for water-soluble antioxidant ingredients.


Assuntos
Lipossomos , Absorção Cutânea , Camundongos , Animais , Lipossomos/metabolismo , Ácido Ascórbico , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Pele/metabolismo , Vitaminas
8.
J Control Release ; 351: 623-637, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36191673

RESUMO

Idiopathic pulmonary fibrosis (IPF) is a fibrotic interstitial lung disease in which collagen progressively deposits in the supporting framework of the lungs. The pathological collagen creates a recalcitrant barrier in mesenchyme for drug penetration, thus greatly restricting the therapeutical efficacy. On the other hand, this overloaded collagen is gradually exposed to the bloodstream at fibrotic sites because of the vascular hyperpermeability, thus serving as a potential target. Herein, pathological collagen targeting and penetrating liposomes (DP-CC) were constructed to deliver anti-fibrotic dual drugs including pirfenidone (PFD) and dexamethasone (DEX) deep into injured alveoli. The liposomes were co-decorated with collagen binding peptide (CBP) and collagenase (COL). CBP could help vehicle recognize the pathological collagen and target the fibrotic lungs efficiently because of its high affinity to collagen, and COL assisted in breaking through the collagen barrier and delivering vehicle to the center of injured sites. Then, the released dual drugs developed a synergistic anti-fibrotic effect to repair the damaged epithelium and remodel the extracellular matrix (ECM), thus rebuilding the lung architecture. This study provides a promising strategy to deliver drugs deep into pathological collagen accumulated sites for the enhanced treatment of IPF.


Assuntos
Fibrose Pulmonar Idiopática , Humanos , Fibrose Pulmonar Idiopática/tratamento farmacológico , Lipossomos/metabolismo , Colágeno/metabolismo , Pulmão/metabolismo , Matriz Extracelular/metabolismo , Fibrose , Fibroblastos/metabolismo
9.
Int J Biol Sci ; 18(15): 5653-5666, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36263186

RESUMO

Studies have shown that epigenetic enzymes such as histone deacetylase (HDAC) are closely related to cancers and that several HDAC inhibitors exert antitumor effects. Studies have further suggested that class IIa HDAC inhibitors are related to immune functions, including immune responses and the expression of chemokines and complement pathway components. TMP195, a selective class IIa HDAC inhibitor, has been reported to be effective against breast cancer. However, the role and mechanism of TMP195 in colorectal cancer remain unknown. In this study, we found that TMP195 significantly reduced the tumor burden in two mouse models of colitis-associated colorectal cancer (CAC) and subcutaneous tumor. Mechanistically, TMP195 decreased the proportion of total macrophages but increased the proportion of M1 macrophages by promoting polarization, resulting in the increased release of inflammatory cytokines. TMP195 had no direct effect on the proliferation of colorectal cancer cells, and its antitumor effect on the colorectal cancer disappeared when macrophages were partly depleted by clodronate liposomes. In addition, TMP195 enhanced the efficacy of PD-1 blockade. The present study revealed that the combination of TMP195 and PD-1 blockade may provide a therapeutic strategy for colorectal cancer.


Assuntos
Neoplasias Colorretais , Inibidores de Histona Desacetilases , Camundongos , Animais , Inibidores de Histona Desacetilases/farmacologia , Receptor de Morte Celular Programada 1/metabolismo , Ácido Clodrônico/metabolismo , Ácido Clodrônico/farmacologia , Lipossomos/metabolismo , Lipossomos/farmacologia , Macrófagos/metabolismo , Histona Desacetilases/metabolismo , Citocinas/metabolismo , Neoplasias Colorretais/metabolismo , Linhagem Celular Tumoral
10.
Sci Adv ; 8(41): eabn6845, 2022 10 14.
Artigo em Inglês | MEDLINE | ID: mdl-36223470

RESUMO

Membrane proteins are currently investigated after detergent extraction from native cellular membranes and reconstitution into artificial liposomes or nanodiscs, thereby removing them from their physiological environment. However, to truly understand the biophysical properties of membrane proteins in a physiological environment, they must be investigated within living cells. Here, we used a spin-labeled nanobody to interrogate the conformational cycle of the ABC transporter MsbA by double electron-electron resonance. Unexpectedly, the wide inward-open conformation of MsbA, commonly considered a nonphysiological state, was found to be prominently populated in Escherichia coli cells. Molecular dynamics simulations revealed that extensive lateral portal opening is essential to provide access of its large natural substrate core lipid A to the binding cavity. Our work paves the way to investigate the conformational landscape of membrane proteins in cells.


Assuntos
Transportadores de Cassetes de Ligação de ATP , Proteínas de Escherichia coli , Escherichia coli , Transportadores de Cassetes de Ligação de ATP/química , Trifosfato de Adenosina/metabolismo , Proteínas de Bactérias/metabolismo , Detergentes/metabolismo , Escherichia coli/metabolismo , Proteínas de Escherichia coli/química , Lipídeo A , Lipossomos/metabolismo , Proteínas de Membrana/metabolismo , Conformação Proteica
11.
Front Immunol ; 13: 922614, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36159784

RESUMO

Background: Macrophages play important roles in diabetes and sepsis-related intestinal injury. Accumulating evidence suggests that microRNAs (miRNAs) act as the fundamental link between macrophage polarization and tissue injury. However, the underlying mechanisms of miRNAs in regulating macrophage polarization-related intestinal injury under diabetes and sepsis conditions remain unclear. Methods: The cecal ligation and puncture (CLP)-induced sepsis models were established in male wild-type (WT) and diabetic mice. Clodronate liposome was used to deplete macrophage. H&E staining, inflammatory cytokines [tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6], and intestinal mucosal barrier function markers [occludin, ZO-1, lipopolysaccharide (LPS), and intestinal fatty acid binding protein (iFABP)] were used to assess elevated intestinal damage. miRNA array, RNA-seq, and bioinformatic analysis were performed to detect the miRNA and messenger RNA (mRNA) expression and the potential regulation mechanism. In vitro, RAW264.7 cells were cultured in the absence or presence of high glucose and LPS, miR-3061 mimics, and Snail small interfering RNA stimulation, respectively, for further mechanism studies. Luciferase reporter assay was used to confirm the interplay between miRNA and its target genes. Results: Compared with WT CLP mice, the diabetic CLP mice showed severe intestinal damage characterized by significant increases in Chui's scores, expression of inflammatory cytokines (TNF-α, IL-1ß, and IL-6), serum LPS and iFABP concentration, and significant reductions in tight junction protein occludin and ZO-1 levels. Macrophage depletion reversed the intestinal damage caused by CLP. The bioinformatic analysis revealed that miR-3061/Snail1 might be a potential regulation axis of macrophage polarization. Furthermore, high glucose and LPS stimulation increased M1 macrophage and reduced the levels of miR-3061, which was negatively associated with Snail1 in RAW264.7 cells. Mechanistic studies demonstrated that miR-3061 regulated macrophage polarization by targeting the Snail1 mRNA 3'-untranslated region. Moreover, miR-3061 overexpression suppressed Snail1 expression and inhibited M1 macrophage and inflammatory cytokines. Conclusion: This study elucidated that diabetes exacerbated sepsis-induced intestinal injury by promoting M1 macrophage polarization and further demonstrated that the miR-3061/Sani1 axis may be the potential target of macrophage polarization.


Assuntos
Diabetes Mellitus Experimental , MicroRNAs , Sepse , Animais , Ácido Clodrônico , Citocinas/metabolismo , Diabetes Mellitus Experimental/metabolismo , Proteínas de Ligação a Ácido Graxo/metabolismo , Glucose/metabolismo , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/metabolismo , Lipossomos/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Ocludina/metabolismo , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Sepse/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Regiões não Traduzidas
12.
Front Cell Infect Microbiol ; 12: 992198, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36159643

RESUMO

Irgb6 is a member of interferon γ-induced immunity related GTPase (IRG), and one of twenty "effector" IRGs, which coordinately attack parasitophorous vacuole membrane (PVM), causing death of intracellular pathogen. Although Irgb6 plays a pivotal role as a pioneer in the process of PVM disruption, the direct effect of Irgb6 on membrane remained to be elucidated. Here, we utilized artificial lipid membranes to reconstitute Irgb6-membrane interaction in vitro, and revealed that Irgb6 directly deformed the membranes. Liposomes incubated with recombinant Irgb6 were drastically deformed generating massive tubular protrusions in the absence of guanine nucleotide, or with GMP-PNP. Liposome deformation was abolished by incubating with Irgb6-K275A/R371A, point mutations at membrane targeting residues. The membrane tubules generated by Irgb6 were mostly disappeared by the addition of GTP or GDP, which are caused by detachment of Irgb6 from membrane. Binding of Irgb6 to the membrane, which was reconstituted in vitro using lipid monolayer, was stimulated at GTP-bound state. Irgb6 GTPase activity was stimulated by the presence of liposomes more than eightfold. Irgb6 GTPase activity in the absence of membrane was also slightly stimulated, by lowering ionic strength, or by increasing protein concentration, indicating synergistic stimulation of the GTPase activity. These results suggest that membrane targeting of Irgb6 and resulting membrane deformation does not require GTP, but converting into GTP-bound state is crucial for detaching Irgb6 from the membrane, which might coincident with local membrane disruption.


Assuntos
Toxoplasma , GTP Fosfo-Hidrolases/metabolismo , Guanilil Imidodifosfato/metabolismo , Interferon gama/metabolismo , Lipídeos , Lipossomos/metabolismo
13.
Int J Mol Sci ; 23(18)2022 Sep 07.
Artigo em Inglês | MEDLINE | ID: mdl-36142217

RESUMO

Magnetosomes of magnetotactic bacteria consist of magnetic nanocrystals with defined morphologies enclosed in vesicles originated from cytoplasmic membrane invaginations. Although many proteins are involved in creating magnetosomes, a single magnetosome protein, Mms6 from Magnetospirillum magneticum strain AMB-1, can direct the crystallization of magnetite nanoparticles in vitro. The in vivo role of Mms6 in magnetosome formation is debated, and the observation that Mms6 binds Fe3+ more tightly than Fe2+ raises the question of how, in a magnetosome environment dominated by Fe3+, Mms6 promotes the crystallization of magnetite, which contains both Fe3+ and Fe2+. Here we show that Mms6 is a ferric reductase that reduces Fe3+ to Fe2+ using NADH and FAD as electron donor and cofactor, respectively. Reductase activity is elevated when Mms6 is integrated into either liposomes or bicelles. Analysis of Mms6 mutants suggests that the C-terminal domain binds iron and the N-terminal domain contains the catalytic site. Although Mms6 forms multimers that involve C-terminal and N-terminal domain interactions, a fusion protein with ubiquitin remains a monomer and displays reductase activity, which suggests that the catalytic site is fully in the monomer. However, the quaternary structure of Mms6 appears to alter the iron binding characteristics of the C-terminal domain. These results are consistent with a hypothesis that Mms6, a membrane protein, promotes the formation of magnetite in vivo by a mechanism that involves reducing iron.


Assuntos
Magnetossomos , Magnetospirillum , Proteínas de Bactérias/química , FMN Redutase/metabolismo , Óxido Ferroso-Férrico/metabolismo , Flavina-Adenina Dinucleotídeo/metabolismo , Ferro/metabolismo , Lipídeos/análise , Lipossomos/metabolismo , Magnetossomos/metabolismo , Magnetospirillum/metabolismo , Proteínas de Membrana/metabolismo , NAD/metabolismo , Ubiquitinas/metabolismo
14.
Clin Transl Med ; 12(10): e1036, 2022 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-36178087

RESUMO

BACKGROUND: Emerging evidence provides mechanistic insights into the pathogenesis of pulmonary fibrosis (PF), and rare anti-PF therapeutic method has promising effect in its treatment. Rho-associated coiled-coil kinases (ROCK) inhibition significantly ameliorates bleomycin-induced PF and decreases macrophage infiltration, but the mechanism remains unclear. We established bleomycin and radiation-induced PF to identify the activity of WXWH0265, a newly designed unselective ROCK inhibitor in regulating macrophages. METHODS: Bleomycin-induced PF was induced by intratracheal instillation and radiation-induced PF was induced by bilateral thoracic irradiation. Histopathological techniques (haematoxylin and eosin, Masson's trichrome and immunohistochemistry) and hydroxyproline were used to evaluate PF severity. Western blot, quantitative real-time reverse transcription-polymerase chain reaction and flow cytometry were performed to explore the underlying mechanisms. Bone marrow-derived macrophages (BMDMs) were used to verify their therapeutic effect. Clodronate liposomes were applied to deplete macrophages and to identify the therapeutic effect of WXWH0265. RESULTS: Therapeutic administration of ROCK inhibitor ameliorates bleomycin-induced PF by inhibiting M2 macrophages polarisation. ROCK inhibitor showed no significant anti-fibrotic effect in macrophages-depleted mice. Treatment with WXWH0265 demonstrated superior protection effect in bleomycin-induced PF compared with positive drugs. In radiation-induced PF, ROCK inhibitor effectively ameliorated PF. Fibroblasts co-cultured with supernatant from various M2 macrophages phenotypes revealed that M2 macrophages stimulated by interleukin-4 promoted extracellular matrix production. Polarisation of M2 macrophages was inhibited by ROCK inhibitor treatment in vitro. The p-signal transducer and activator of transcription 3 (STAT3) in lung tissue and BMDMs was significantly decreased in PF in vivo and vitro after treated with ROCK inhibitors. CONCLUSION: Inhibiting ROCK could significantly attenuate bleomycin- and radiation-induced PF by regulating the macrophages polarisation via phosphorylation of STAT3. WXWH0265 is a kind of efficient unselective ROCK inhibitor in ameliorating PF. Furthermore, the results provide empirical evidence that ROCK inhibitor, WXWH0265 is a potential drug to prevent the development of PF.


Assuntos
Fibrose Pulmonar , Fator de Transcrição STAT3 , Quinases Associadas a rho , Animais , Bleomicina/efeitos adversos , Ácido Clodrônico/metabolismo , Interleucina-4/metabolismo , Lipossomos/metabolismo , Macrófagos/metabolismo , Camundongos , Fosforilação , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/patologia , Fator de Transcrição STAT3/metabolismo , Quinases Associadas a rho/antagonistas & inibidores
15.
Diabetes ; 71(12): 2597-2611, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-36125850

RESUMO

Preeclampsia is a pregnancy-specific complication with long-term negative outcomes for offspring, including increased susceptibility to type 2 diabetes (T2D) in adulthood. In a rat reduced uteroplacental perfusion pressure (RUPP) model of chronic placental ischemia, maternal hypertension in conjunction with intrauterine growth restriction mimicked aspects of preeclampsia and resulted in female embryonic day 19 (e19) offspring with reduced ß-cell area and increased ß-cell apoptosis compared with offspring of sham pregnancies. Decreased pancreatic ß-cell area persisted to postnatal day 13 (PD13) in females and could influence whether T2D developed in adulthood. Macrophage changes also occurred in islets in T2D. Therefore, we hypothesized that macrophages are crucial to reduction in pancreatic ß-cell area in female offspring after chronic placental ischemia. Macrophage marker CD68 mRNA expression was significantly elevated in e19 and PD13 islets isolated from female RUPP offspring compared with sham. Postnatal injections of clodronate liposomes into female RUPP and sham offspring on PD2 and PD9 significantly depleted macrophages compared with injections of control liposomes. Depletion of macrophages rescued reduced ß-cell area and increased ß-cell proliferation and size in RUPP offspring. Our studies suggest that the presence of macrophages is important for reduced ß-cell area in female RUPP offspring and changes in macrophages could contribute to development of T2D in adulthood.


Assuntos
Diabetes Mellitus Tipo 2 , Hipertensão , Pré-Eclâmpsia , Humanos , Feminino , Gravidez , Ratos , Animais , Pré-Eclâmpsia/etiologia , Pré-Eclâmpsia/metabolismo , Placenta/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Lipossomos/metabolismo , Útero/metabolismo , Ratos Sprague-Dawley , Isquemia/metabolismo , Macrófagos/metabolismo , Pressão Sanguínea , Modelos Animais de Doenças
16.
J Control Release ; 351: 361-380, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36169040

RESUMO

Upon exhaustive research, the transdermal drug delivery system (TDDS) has appeared as a potential, well-accepted, and popular approach to a novel drug delivery system. Ease of administration, easy handling, minimum systemic exposure, least discomfort, broad flexibility and tunability, controlled release, prolonged therapeutic effect, and many more perks make it a promising approach for effective drug delivery. Although, the primary challenge associated is poor skin permeability. Skin is an intact barrier that serves as a primary defense mechanism to preclude any foreign particle's entry into the body. Owing to the unique anatomical framework, i.e., compact packing of stratum corneum with tight junction and fast anti-inflammatory responses, etc., emerged as a critical physiological barrier for TDDS. Fusion with other novel approaches like nanocarriers, specially designed transdermal delivery devices, permeation enhancers, etc., can overcome the limitations. Utilizing such strategies, some of the products are under clinical trials, and many are under investigation. This review explores all dimensions that overcome poor permeability and allows the drug to attain maximum potential. The article initially compiles fundamental features, components, and design of TDDS, followed by critical aspects and various methods, including in vitro, ex vivo, and in vivo methods of assessing skin permeability. The work primarily aimed to highlight the recent advancement in novel strategies for effective transdermal drug delivery utilizing active methods like iontophoresis, electroporation, sonophoresis, microneedle, needleless jet injection, etc., and passive methods such as the use of liposomes, SLN, NLC, micro/nanoemulsions, dendrimers, transferosomes, and many more nanocarriers. In all, this compilation will provide a recent insight on the novel updates along with basic concepts, the current status of clinical development, and challenges for the clinical translation of TDDS.


Assuntos
Sistemas de Liberação de Medicamentos , Absorção Cutânea , Sistemas de Liberação de Medicamentos/métodos , Administração Cutânea , Pele/metabolismo , Permeabilidade , Lipossomos/metabolismo
17.
J Lipid Res ; 63(10): 100273, 2022 10.
Artigo em Inglês | MEDLINE | ID: mdl-36084713

RESUMO

Liposomes are the most widely used nanocarrier platform for the delivery of therapeutic and diagnostic agents, and a number of liposomes have been approved for use in clinical practice. After systemic administration, most liposomes are cleared by macrophages in the mononuclear phagocyte system, such as the liver and bone marrow (BM). However, the majority of studies have focused on investigating the therapeutic results of liposomal drugs, and too few studies have evaluated the potential side effects of empty nanocarriers on the functions of macrophages in the mononuclear phagocyte system. Here, we evaluate the potential effects of empty liposomes on the functions of BM niche macrophages. Following liposome administration, we observed lipid droplet (LD) accumulation in cultured primary macrophages and BM niche macrophages. We found that these LD-accumulating macrophages, similar to foam cells, exhibited increased expression of inflammatory cytokines, such as IL-1ß and IL-6. We further provided evidence that liposome deposition and degradation induced LD biogenesis on the endoplasmic reticulum membrane and subsequently disturbed endoplasmic reticulum homeostasis and activated the inositol-requiring transmembrane kinase/endoribonuclease 1α/NF-κB signaling pathway, which is responsible for the inflammatory activation of macrophages after liposome engulfment. Finally, we also showed the side effects of dysfunctional BM niche macrophages on hematopoiesis in mice, such as the promotion of myeloid-biased output and impairment of erythropoiesis. This study not only draws attention to the safety of liposomal drugs in clinical practice but also provides new directions for the design of lipid-based drug carriers in preclinical studies.


Assuntos
Medula Óssea , Lipossomos , Camundongos , Animais , Lipossomos/metabolismo , NF-kappa B/metabolismo , Interleucina-6/metabolismo , Macrófagos/metabolismo , Hematopoese , Portadores de Fármacos/metabolismo , Portadores de Fármacos/farmacologia , Citocinas/metabolismo , Endorribonucleases , Inositol/metabolismo , Lipídeos
18.
mBio ; 13(5): e0179022, 2022 10 26.
Artigo em Inglês | MEDLINE | ID: mdl-36135367

RESUMO

Gram-negative bacteria have a robust cell envelope that excludes or expels many antimicrobial agents. However, during infection, host soluble innate immune factors permeabilize the bacterial outer membrane. We identified two small molecules that exploit outer membrane damage to access the bacterial cell. In standard microbiological media, neither compound inhibited bacterial growth nor permeabilized bacterial outer membranes. In contrast, at micromolar concentrations, JAV1 and JAV2 enabled the killing of an intracellular human pathogen, Salmonella enterica serovar Typhimurium. S. Typhimurium is a Gram-negative bacterium that resides within phagosomes of cells from the monocyte lineage. Under broth conditions that destabilized the lipopolysaccharide layer, JAV2 permeabilized the bacterial inner membrane and was rapidly bactericidal. In contrast, JAV1 activity was more subtle: JAV1 increased membrane fluidity, altered reduction potential, and required more time than JAV2 to disrupt the inner membrane barrier and kill bacteria. Both compounds interacted with glycerophospholipids from Escherichia coli total lipid extract-based liposomes. JAV1 preferentially interacted with cardiolipin and partially relied on cardiolipin production for activity, whereas JAV2 generally interacted with lipids and had modest affinity for phosphatidylglycerol. In mammalian cells, neither compound significantly altered mitochondrial membrane potential at concentrations that killed S. Typhimurium. Instead, JAV1 and JAV2 became trapped within acidic compartments, including macrophage phagosomes. Both compounds improved survival of S. Typhimurium-infected Galleria mellonella larvae. Together, these data demonstrate that JAV1 and JAV2 disrupt bacterial inner membranes by distinct mechanisms and highlight how small, lipophilic, amine-substituted molecules can exploit host soluble innate immunity to facilitate the killing of intravesicular pathogens. IMPORTANCE Innovative strategies for developing new antimicrobials are needed. Combining our knowledge of host-pathogen interactions and relevant drug characteristics has the potential to reveal new approaches to treating infection. We identified two compounds with antibacterial activity specific to infection and with limited host cell toxicity. These compounds appeared to exploit host innate immunity to access the bacterium and differentially damage the bacterial inner membrane. Further, both compounds accumulated within Salmonella-containing and other acidic vesicles, a process known as lysosomal trapping, which protects the host and harms the pathogen. The compounds also increased host survival in an insect infection model. This work highlights the ability of host innate immunity to enable small molecules to act as antibiotics and demonstrates the feasibility of antimicrobial targeting of the inner membrane. Additionally, this study features the potential use of lysosomal trapping to enhance the activities of compounds against intravesicular pathogens.


Assuntos
Cardiolipinas , Infecções por Salmonella , Animais , Humanos , Cardiolipinas/metabolismo , Lipopolissacarídeos/metabolismo , Lipossomos/metabolismo , Infecções por Salmonella/metabolismo , Salmonella typhimurium/metabolismo , Fagossomos/metabolismo , Antibacterianos/farmacologia , Antibacterianos/metabolismo , Glicerofosfolipídeos/metabolismo , Escherichia coli/metabolismo , Aminas/metabolismo , Mamíferos/metabolismo
19.
Int Immunopharmacol ; 112: 109209, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-36084540

RESUMO

Autoimmune diseases present a significant clinical problem, highlighting the need for the development of novel or improved therapeutic methods. One of the factors that causes autoimmune diseases is a defect in the clearance of apoptotic cells by phagocytes. Thus, improved apoptotic cell processing has been considered as a strategy to treat autoimmune diseases. However, therapeutic strategies focusing on apoptotic cell clearance have not been approved till date. We have reported that liposomes composed of phosphatidylserine (PS liposomes) exhibit anti-inflammatory or immunosuppressive effects in macrophages. A PS liposome display PS on its surface, which plays a crucial role in the phagocytosis of apoptotic cells by marginal zone macrophages (MZMs), a key player in the clearance of apoptotic cells, by recognizing PS exposed on the surface of apoptotic cells. Therefore, we hypothesized that PS liposomes could be used as "antigen delivery vesicles" to act as a substitute for apoptotic cells in the treatment of autoimmune diseases. In this study, we showed that systemically administered PS liposomes accumulated in the marginal zone of the spleen due to recognition of surface-displayed PS by MZMs because it was observed that liposomes without PS did not accumulate in the marginal zone. In conclusion, PS liposomes may be useful vehicles to function as active agents and/or antigens against autoimmune diseases.


Assuntos
Doenças Autoimunes , Fosfatidilserinas , Camundongos , Animais , Fosfatidilserinas/metabolismo , Lipossomos/metabolismo , Apoptose , Macrófagos , Fagocitose , Antígenos , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/metabolismo
20.
Proc Natl Acad Sci U S A ; 119(39): e2204396119, 2022 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-36122218

RESUMO

Membrane contact sites (MCS), close membrane apposition between organelles, are platforms for interorganellar transfer of lipids including cholesterol, regulation of lipid homeostasis, and co-ordination of endocytic trafficking. Sphingosine kinases (SphKs), two isoenzymes that phosphorylate sphingosine to the bioactive sphingosine-1-phosphate (S1P), have been implicated in endocytic trafficking. However, the physiological functions of SphKs in regulation of membrane dynamics, lipid trafficking and MCS are not known. Here, we report that deletion of SphKs decreased S1P with concomitant increases in its precursors sphingosine and ceramide, and markedly reduced endoplasmic reticulum (ER) contacts with late endocytic organelles. Expression of enzymatically active SphK1, but not catalytically inactive, rescued the deficit of these MCS. Although free cholesterol accumulated in late endocytic organelles in SphK null cells, surprisingly however, cholesterol transport to the ER was not reduced. Importantly, deletion of SphKs promoted recruitment of the ER-resident cholesterol transfer protein Aster-B (also called GRAMD1B) to the plasma membrane (PM), consistent with higher accessible cholesterol and ceramide at the PM, to facilitate cholesterol transfer from the PM to the ER. In addition, ceramide enhanced in vitro binding of the Aster-B GRAM domain to phosphatidylserine and cholesterol liposomes. Our study revealed a previously unknown role for SphKs and sphingolipid metabolites in governing diverse MCS between the ER network and late endocytic organelles versus the PM to control the movement of cholesterol between distinct cell membranes.


Assuntos
Fosfatidilserinas , Esfingosina , Ceramidas/metabolismo , Colesterol/metabolismo , Retículo Endoplasmático/metabolismo , Isoenzimas/metabolismo , Lipossomos/metabolismo , Lisofosfolipídeos , Fosfatidilserinas/metabolismo , Esfingolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/metabolismo
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