Intracellular delivery of nitric oxide enhances the therapeutic efficacy of mesenchymal stem cells for myocardial infarction.

Cell therapy by autologous mesenchymal stem cells (MSCs) is a clinically acceptable strategy for treating various diseases. Unfortunately, the therapeutic efficacy is largely affected by the low quality of MSCs collected from patients. Here, we showed that the gene expression of MSCs from patients with diabetes was differentially regulated compared to that of MSCs from healthy controls. Then, MSCs were genetically engineered to catalyze an NO prodrug to release NO intracellularly. Compared to extracellular NO conversion, intracellular NO delivery effectively prolonged survival and enhanced the paracrine function of MSCs, as demonstrated by in vitro and in vivo assays. The enhanced therapeutic efficacy of engineered MSCs combined with intracellular NO delivery was further confirmed in mouse and rat models of myocardial infarction, and a clinically relevant cell administration paradigm through secondary thoracotomy has been attempted.

Overexpression of Glutathione S-Transferases in Human Diseases: Drug Targets and Therapeutic Implications.

Glutathione S-transferases (GSTs) are a major class of phase II metabolic enzymes. Besides their essential role in detoxification, GSTs also exert diverse biological activities in the occurrence and development of various diseases. In the past few decades, much research interest has been paid to exploring the mechanisms of GST overexpression in tumor drug resistance. Correspondingly, many GST inhibitors have been developed and applied, solely or in combination with chemotherapeutic drugs, for the treatment of multi-drug resistant tumors. Moreover, novel roles of GSTs in other diseases, such as pulmonary fibrosis and neurodegenerative diseases, have been recognized in recent years, although the exact regulatory mechanisms remain to be elucidated. This review, firstly summarizes the roles of GSTs and their overexpression in the above-mentioned diseases with emphasis on the modulation of cell signaling pathways and protein functions. Secondly, specific GST inhibitors currently in pre-clinical development and in clinical stages are inventoried. Lastly, applications of GST inhibitors in targeting cell signaling pathways and intracellular biological processes are discussed, and the potential for disease treatment is prospected. Taken together, this review is expected to provide new insights into the interconnection between GST overexpression and human diseases, which may assist future drug discovery targeting GSTs.

Genetically engineered mesenchymal stem cells as a nitric oxide reservoir for acute kidney injury therapy.

Nitric oxide (NO), as a gaseous therapeutic agent, shows great potential for the treatment of many kinds of diseases. Although various NO delivery systems have emerged, the immunogenicity and long-term toxicity of artificial carriers hinder the potential clinical translation of these gas therapeutics. Mesenchymal stem cells (MSCs), with the capacities of self-renewal, differentiation, and low immunogenicity, have been used as living carriers. However, MSCs as gaseous signaling molecule (GSM) carriers have not been reported. In this study, human MSCs were genetically modified to produce mutant ß-galactosidase (ß-GALH363A). Furthermore, a new NO prodrug, 6-methyl-galactose-benzyl-oxy NONOate (MGP), was designed. MGP can enter cells and selectively trigger NO release from genetically engineered MSCs (eMSCs) in the presence of ß-GALH363A. Moreover, our results revealed that eMSCs can release NO when MGP is systemically administered in a mouse model of acute kidney injury (AKI), which can achieve NO release in a precise spatiotemporal manner and augment the therapeutic efficiency of MSCs. This eMSC and NO prodrug system provides a unique and tunable platform for GSM delivery and holds promise for regenerative therapy by enhancing the therapeutic efficiency of stem cells.

Animals are made up of cells of different types, with each type of cell specializing on a specific role. But for the body to work properly, the different types of cells must be able to coordinate with each other to respond to internal and external stimuli. This can be achieved through signaling molecules, that is, molecules released by a cell that can elicit a specific response in other cells. There are many types of different molecules, including hormones and signaling proteins. Gases can also be potent signaling molecules, participating in various biological processes. Nitric oxide (NO) is a gas signaling molecule that can freely diffuse through the membranes of cells and has roles in blood vessel constriction and other disease processes, making it a promising therapeutic agent. Unfortunately, using artificial carriers to deliver nitric oxide to the organs and tissues where it is needed can lead to issues, including immune reactions to the carrier and long-term toxicity. One way to avoid these effects is by using cells to deliver nitric oxide to the right place. Huang, Qian, Liu et al. have used mesenchymal stem cells ­ which usually develop to form connective tissues such as bone and muscle ­ to develop a cell-based NO-delivery system. The researchers genetically modified the mesenchymal stem cells to produce a compound called ß-GAL^H363A. On its own ß-GAL^H363A does not do much, but in its presence, a non-toxic, non-reactive compound developed by Huang, Qian, Liu et al., called MGP, can drive the release of NO from cells. To confirm the usefulness of their cells as a delivery system, Huang, Qian, Liu et al. transplanted some of the genetically modified mesenchymal stem cells into the kidneys of mice, and then showed that when these mice were given MGP, the levels of NO increased in the kidneys but not in other organs. This result confirms that the cell-based delivery system provides spatial and temporal control of the production of NO. These findings demonstrate a new delivery system for therapies using gas molecules, which can be controlled spatiotemporally in mice. In the future, these types of systems could be used in the clinic for long-term treatment of conditions where artificial carriers could lead to complications.

Mechanistic Study of Icaritin-Induced Inactivation of Cytochrome P450 2C9.

Icaritin (ICT) is a prenylflavonoid derivative that has been approved by National Medical Products Administration for the treatment of hepatocellular carcinoma. This study aims to evaluate the potential inhibitory effect of ICT against cytochrome P450 (CYP) enzymes and to elucidate the inactivation mechanisms. Results showed that ICT inactivated CYP2C9 in a time-, concentration-, and NADPH-dependent manner with Ki = 1.896 µM, Kinact = 0.02298 minutes-1, and Kinact/Ki = 12 minutes-1 mM-1, whereas the activities of other CYP isozymes was minimally affected. Additionally, the presence of CYP2C9 competitive inhibitor, sulfaphenazole, superoxide dismutase/catalase system, and GSH all protected CYP2C9 from ICT-induced activity loss. Moreover, the activity loss was neither recovered by washing the ICT-CYP2C9 preincubation mixture nor the addition of potassium ferricyanide. These results, collectively, implied the underlying inactivation mechanism involved the covalent binding of ICT to the apoprotein and/or the prosthetic heme of CYP2C9. Furthermore, an ICT-quinone methide (QM)-derived GSH adduct was identified, and human glutathione S-transferases (GST) isozymes GSTA1-1, GSTM1-1, and GSTP1-1 were shown to be substantially involved in the detoxification of ICT-QM. Interestingly, our systematic molecular modeling work predicted that ICT-QM was covalently bound to C216, a cysteine residue located in the F-G loop downstream of substrate recognition site (SRS) 2 in CYP2C9. The sequential molecular dynamics simulation confirmed the binding to C216 induced a conformational change in the active catalytic center of CYP2C9. Lastly, the potential risks of clinical drug-drug interactions triggered by ICT as a perpetrator were extrapolated. In summary, this work confirmed that ICT was an inactivator of CYP2C9. SIGNIFICANCE STATEMENT: This study is the first to report the time-dependent inhibition of CYP2C9 by icaritin (ICT) and the intrinsic molecular mechanism behind it. Experimental data indicated that the inactivation was via irreversible covalent binding of ICT-quinone methide to CYP2C9, while molecular modeling analysis provided additional evidence by predicting C216 as the key binding site which influenced the structural confirmation of CYP2C9's catalytic center. These findings suggest the potential of drug-drug interactions when ICT is co-administered with CYP2C9 substrates clinically.

Single-nucleus profiling unveils a geroprotective role of the FOXO3 in primate skeletal muscle aging.

Age-dependent loss of skeletal muscle mass and function is a feature of sarcopenia, and increases the risk of many aging-related metabolic diseases. Here, we report phenotypic and single-nucleus transcriptomic analyses of non-human primate skeletal muscle aging. A higher transcriptional fluctuation was observed in myonuclei relative to other interstitial cell types, indicating a higher susceptibility of skeletal muscle fiber to aging. We found a downregulation of FOXO3 in aged primate skeletal muscle, and identified FOXO3 as a hub transcription factor maintaining skeletal muscle homeostasis. Through the establishment of a complementary experimental pipeline based on a human pluripotent stem cell-derived myotube model, we revealed that silence of FOXO3 accelerates human myotube senescence, whereas genetic activation of endogenous FOXO3 alleviates human myotube aging. Altogether, based on a combination of monkey skeletal muscle and human myotube aging research models, we unraveled the pivotal role of the FOXO3 in safeguarding primate skeletal muscle from aging, providing a comprehensive resource for the development of clinical diagnosis and targeted therapeutic interventions against human skeletal muscle aging and the onset of sarcopenia along with aging-related disorders.

Glutathione S-Transferases Mediate In Vitro and In Vivo Inactivation of Genipin: Implications for an Underlying Detoxification Mechanism.

Genipin (GP), the reactive metabolite of geniposide (GE), is responsible for GE-induced hepatotoxicity. As a potential detoxification pathway, the inactivation of GP by glutathione S-transferases (GSTs) has not yet been characterized. In this study, the thiol-GSH conjugates of GP, M532-1 and M532-2 were first identified and the catalytic activities of GSTs were investigated both in vitro and in vivo. GSTA1-1 and GSTA4-4 showed high activity in the formation of both thiol-GSH conjugates, whereas GSTA4-4 specifically catalyzed M532-2 formation in vitro. The active GST isoforms protect against alkylation of N-acetylcysteine (NAC), a classic model nucleophile. GST inhibition attenuated M532-1 formation in rat bile, confirming the in vivo catalytic role of GSTs. In conclusion, this study demonstrated the inactivation of GP by GSTs and implied that interindividual variability of GSTs may be a risk factor for susceptibility to GE-induced hepatotoxicity.

Phase I Metabolism of Pterostilbene, a Dietary Resveratrol Derivative: Metabolite Identification, Species Differences, Isozyme Contribution, and Further Bioactivation.

Pterostilbene (PTE), a dietary derivative of resveratrol, displayed pleiotropic health-promoting activities. This study aimed to explore the metabolic profiles and species differences of the phase I metabolism of PTE and to investigate subsequent detoxification after PTE bioactivation. PTE was found to be biotransformed to two pharmacologically active metabolites, pinostilbene and 3'-hydroxypterostilbene, in vivo and in vitro with substantial species differences. Human CYP1A2 was proved to be mainly responsible for the demethylation and 3'-hydroxylation of PTE, with its contribution to a demethylation of 94.5% and to a 3'-hydroxylation of 97.9%. An in vitro glutathione trapping experiment revealed the presence of an ortho-quinone intermediate formed by further oxidation of 3'-hydroxypterostilbene. Human glutathione S-transferase isoforms A2, T1, and A1 inactivated the ortho-quinone intermediate by catalyzing glutathione conjugation, implicating a potential protective pathway against PTE bioactivation-derived toxicity. Overall, this study provided a comprehensive view of PTE phase I metabolism and facilitated its further development as a promising nutraceutical.

Subacute oral toxicology and toxicokinetics of pterostilbene, a novel Top1/Tdp1 inhibiting anti-tumor reagent.

This study evaluated the subacute toxicity and toxicokinetics of a potential anti-cancer drug candidate, pterostilbene, in rats. Animals were orally administered at two repeated doses of 200 and 500 mg/kg for 28 days. No mortality was observed during the 28 days of continuous administration of pterostilbene. Body weight and food consumption in each group increased steadily, while no significant difference was found. Liver weight in the 500 mg/kg female, but not male group increased with mild cytoplasmic vacuoles observed in histopathological study. Toxicokinetics was assessed by measuring plasma concentrations of pterostilbene on the first and 28th day of administration using UPLC-MS/MS. Toxicokinetic parameters showed that AUC0-t significantly increased in all animals, while the increase in females was greater than males. System exposure of pterostilbene appeared to be linear within the administrated dose range. In conclusion, our findings suggested a minimal subacute toxicity profile of pterostilbene, which could strongly support further development of this compound as a novel anti-cancer agent.

Intravenously transplanted mesenchymal stromal cells: a new endocrine reservoir for cardioprotection.

BACKGROUND: Intravenous administration of mesenchymal stromal cells (MSCs) has an acknowledged competence of cardiac repair, despite a lack of systematic description of the underlying biological mechanisms. The lung, but not the heart, is the main trapped site for intravenously transplanted MSCs, which leaves a spatial gap between intravenously transplanted MSCs and the injured myocardium. How lung-trapped MSCs after intravenous transplantation rejuvenate the injured myocardium remains unknown. METHODS: MSCs were isolated from human placenta tissue, and DF-MSCs or Gluc-MSCs were generated by transduced with firefly luciferase (Fluc)/enhanced green fluorescence protein (eGFP) or Gaussia luciferase (Gluc) lactadherin fusion protein. The therapeutic efficiency of intravenously transplanted MSCs was investigated in a murine model of doxorubicin (Dox)-induced cardiotoxicity. Trans-organ communication from the lung to the heart with the delivery of blood was investigated by testing the release of MSC-derived extracellular vesicles (MSC-EVs), and the potential miRNA inner MSC-EVs were screened out and verified. The potential therapeutic miRNA inner MSC-EVs were then upregulated or downregulated to assess the further therapeutic efficiency RESULTS: Dox-induced cardiotoxicity, characterized by cardiac atrophy, left ventricular dysfunction, and injured myocardium, was alleviated by consecutive doses of MSCs. These cardioprotective effects might be attributed to suppressing GRP78 triggering endoplasmic reticulum (ER) stress-induced apoptosis in cardiomyocytes. Our results confirmed that miR-181a-5p from MSCs-derived EVs (MSC-EVs) inhibited GRP78. Intravenous DF-MSCs were trapped in lung vasculature, secreted a certain number of EVs into serum, which could be confirmed by the detection of eGFP+ EVs. GLuc activity was increased in serum EVs from mice administrated with GLuc-MSCs. MiR-181a-5p, inhibiting GRP78 with high efficacy, was highly expressed in serum EVs and myocardium after injecting consecutive doses of MSCs into mice treated with Dox. Finally, upregulation or downregulation of miR-181a-5p levels in MSC-EVs enhanced or weakened therapeutic effects on Dox-induced cardiotoxicity through modulating ER stress-induced apoptosis. CONCLUSIONS: This study identifies intravenously transplanted MSCs, as an endocrine reservoir, to secrete cardioprotective EVs into blood continuously and gradually to confer the trans-organ communication that relieves Dox-induced cardiotoxicity.

Role of prostaglandin E2 in tissue repair and regeneration.

Tissue regeneration following injury from disease or medical treatment still represents a challenge in regeneration medicine. Prostaglandin E2 (PGE2), which involves diverse physiological processes via E-type prostanoid (EP) receptor family, favors the regeneration of various organ systems following injury for its capabilities such as activation of endogenous stem cells, immune regulation, and angiogenesis. Understanding how PGE2 modulates tissue regeneration and then exploring how to elevate the regenerative efficiency of PGE2 will provide key insights into the tissue repair and regeneration processes by PGE2. In this review, we summarized the application of PGE2 to guide the regeneration of different tissues, including skin, heart, liver, kidney, intestine, bone, skeletal muscle, and hematopoietic stem cell regeneration. Moreover, we introduced PGE2-based therapeutic strategies to accelerate the recovery of impaired tissue or organs, including 15-hydroxyprostaglandin dehydrogenase (15-PGDH) inhibitors boosting endogenous PGE2 levels and biomaterial scaffolds to control PGE2 release.

Embryonic stem cell-derived extracellular vesicles promote the recovery of kidney injury.

BACKGROUND: Embryonic stem cell-derived extracellular vesicles (ESC-EVs) possess therapeutic potential for a variety of diseases and are considered as an alternative of ES cells. Acute kidney injury (AKI) is a common acute and severe disease in clinical practice, which seriously threatens human life and health. However, the roles and mechanisms of ESC-EVs on AKI remain unclear. METHODS: In this study, we evaluated the effects of ESC-EVs on physiological repair and pathological repair using murine ischemia-reperfusion injury-induced AKI model, the potential mechanisms of which were next investigated. EVs were isolated from ESCs and EVs derived from mouse fibroblasts as therapeutic controls. We then investigated whether ESC-EVs can restore the structure and function of the damaged kidney by promoting physiological repair and inhibiting the pathological repair process after AKI in vivo and in vitro. RESULTS: We found that ESC-EVs significantly promoted the recovery of the structure and function of the damaged kidney. ESC-EVs increased the proliferation of renal tubular epithelial cells, facilitated renal angiogenesis, inhibited the progression of renal fibrosis, and rescued DNA damage caused by ischemia and reperfusion after AKI. Finally, we found that ESC-EVs play a therapeutic effect by activating Sox9+ cells. CONCLUSIONS: ESC-EVs significantly promote the physiological repair and inhibit the pathological repair after AKI, enabling restoration of the structure and function of the damaged kidney. This strategy might emerge as a novel therapeutic strategy for ESC clinical application.

Chitosan hydrogel-loaded MSC-derived extracellular vesicles promote skin rejuvenation by ameliorating the senescence of dermal fibroblasts.

BACKGROUND: The senescence of dermal fibroblasts (DFLs) leads to an imbalance in the synthesis and degradation of extracellular matrix (ECM) proteins, presenting so-called senescence-associated secretory phenotype (SASP), which ultimately leads to skin aging. Recently, mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) have been recognized as a promising cell-free therapy for degenerative diseases, which opens a new avenue for skin aging treatment. METHODS: In this study, we utilized chitosan (CS) hydrogel for effective loading and sustained release of EVs. In vitro, we explored the rejuvenation effects of CS hydrogel-incorporated EVs (CS-EVs) on replicative senescence DFLs through a series of experiments such as senescence-associated ß-galactosidase (SA-ß-gal) staining, RT-PCR, and Western blot analysis. Besides, we employed local multi-site subcutaneous injection to treat skin aging of naturally aged mice with CS-EVs and DiI fluorescent dye was used to label EVs to achieve in vivo real-time tracking. RESULTS: CS-EVs can significantly improve the biological functions of senescent fibroblasts, including promoting their proliferation, enhancing the synthesis of ECM proteins, and inhibiting the overexpression of matrix metalloproteinases (MMPs). Moreover, CS hydrogel could prolong the release of EVs and significantly increase the retention of EVs in vivo. After CS-EVs subcutaneous injection treatment, the aging skin tissues showed a rejuvenation state, manifested explicitly as the enhanced expression of collagen, the decreased expression of SASP-related factors, and the restoration of tissue structures. CONCLUSIONS: CS hydrogel-encapsulated EVs could delay the skin aging processes by ameliorating the function of aging DFLs. Our results also highlight the potential of CS hydrogel-encapsulated EVs as a novel therapeutic strategy for improving aging skin to rejuvenation.

The delivery of hsa-miR-11401 by extracellular vesicles can relieve doxorubicin-induced mesenchymal stem cell apoptosis.

BACKGROUND: Chemotherapy is an effective anti-tumor treatment. Mesenchymal stem cells (MSCs), exerting therapy effect on injured tissues during chemotherapy, may be damaged in the process. The possibility of self-healing through long-range paracrine and the mechanisms are unclear. METHODS: Doxorubicin, a commonly used chemotherapy drug, was to treat human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) for 6 h as an in vitro cell model of chemotherapy-induced damage. Then we use extracellular vesicles derived from placental mesenchymal stem cells (hP-MSCs) to investigate the therapeutic potential of MSCs-EVs for chemotherapy injury. The mechanism was explored using microRNA sequencing. RESULTS: MSC-derived extracellular vesicles significantly alleviated the chemotherapy-induced apoptosis. Using microRNA sequencing, we identified hsa-miR-11401, which was downregulated in the Dox group but upregulated in the EV group. The upregulation of hsa-miR-11401 reduced the expression of SCOTIN, thereby inhibiting p53-dependent cell apoptosis. CONCLUSIONS: Hsa-miR-11401 expressed by MSCs can be transported to chemotherapy-damaged cells by EVs, reducing the high expression of SCOTIN in damaged cells, thereby inhibiting SCOTIN-mediated apoptosis.

Collagenase IV and clusterin-modified polycaprolactone-polyethylene glycol nanoparticles for penetrating dense tumor tissues.

Purpose: Novel collagenase IV (ColIV) and clusterin (CLU)-modified polycaprolactone-polyethylene glycol (PCL-PEG) nanoparticles that load doxorubicin (DOX) were designed and fully evaluated in vitro and in vivo. Methods: PCL-PEG-ColIV was synthesized by linking PCL-PEG and ColIV through a carbodiimide method. DOX-loaded nanoparticles (DOX-PCL-PEG-ColIV) were self-assembly prepared, followed by noncovalently adsorbing CLU on the DOX-PCL-PEG-ColIV surface to obtain DOX-PCL-PEG-ColIV /CLU nanoparticles, which can penetrate through the tumor extracellular matrix (ECM) and inhibit phagocytosis by macrophage. The physicochemical properties of nanoparticles were characterized. The cellular uptake and antiphagocytosis ability of nanoparticles in MCF-7 tumor cells and RAW264.7 cells were investigated. The penetration ability of nanoparticles was individually evaluated in the two-dimensional (2D) and three-dimensional (3D) ECM models. The tissue distribution and antitumor effect of nanoparticles were evaluated in MCF-7 cell-bearing nude mice. Results: Compared with DOX-PCL-PEG-COOH nanoparticles, DOX-PCL-PEG-ColIV/CLU nanoparticles could effectively overcome the phagocytosis by RAW264.7 and showed excellent cellular uptake in MCF-7 cells. In addition, they showed remarkable penetration ability through the 2D and 3D ECM models. DOX-PCL-PEG-ColIV/CLU nanoparticles significantly reduced the drug distribution in the liver and spleen and enhanced the drug accumulation in tumor tissue compared with DOX-PCL-PEG-COOH or DOX-PCL-PEG-ColIV nanoparticles. DOX-PCL-PEG-ColIV/CLU nanoparticles showed remarkable antitumor effect but did not cause severe pathological damages in the main tissues, including the heart, liver, spleen, lung, and kidney. Conclusion: Novel ColIV and CLU-modified PCL-PEG nanoparticles showed excellent cellular uptake, ECM penetration, antiphagocytosis, and antitumor effects both in vitro and in vivo.

Mesenchymal Stem Cell-Derived Extracellular Vesicles Attenuate Radiation-Induced Lung Injury via miRNA-214-3p.

Aims: Radiotherapy is an effective treatment for thoracic malignancies, but it can cause pulmonary injury and may lead to respiratory failure in a subset of patients. Extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) are now recognized as a new candidate for cell-free treatment of lung diseases. Here, we investigated whether MSC-derived EVs (MSC-EVs) could ameliorate radiation-induced lung injury. Results: We exposed mice to thoracic radiation with a total dose of 15 Gy and assessed the protective effects of MSC-EVs on endothelial cells damage, vascular permeability, inflammation, and fibrosis. We found that MSC-EVs attenuated radiation-induced lung vascular damage, inflammation, and fibrosis. Moreover, MSC-EVs reduced the levels of radiation-induced DNA damage by downregulating ATM/P53/P21 signaling. Our results confirmed that the downregulation of ataxia telangiectasia mutated (ATM) was regulated by miR-214-3p, which was enriched in MSC-EVs. Further analysis demonstrated that MSC-EVs inhibited the senescence-associated secretory phenotype development and attenuated the radiation-induced injury of endothelial cells. Innovation and Conclusion: Our study reveals that MSC-EVs can reduce pulmonary radiation injury through transferring miR-214-3p, providing new avenues to minimize lung injury from radiation therapy. Antioxid. Redox Signal. 35, 849-862.

Extracellular vesicles derived from human placental mesenchymal stem cells alleviate experimental colitis in mice by inhibiting inflammation and oxidative stress.

Mesenchymal stem cells (MSCs) are pluripotent cells that can be applied to the treatment of immune disorders, including inflammatory bowel disease (IBD). The therapeutic effects of MSCs have been mostly attributed to the secretion of soluble factors with paracrine actions, such as extracellular vesicles (EVs), which may play a relevant role in the repair of damaged tissues. In the present study, a mouse model of colitis was induced with the use of trinitrobenzene sulfonic acid (TNBS). EVs derived from human placental mesenchymal stem cells (hP­MSCs) were used for the treatment of colitis by in situ injection. Clinical scores were applied to verify the therapeutic effects of EVs on mice with colitis. Inflammation in the colon was evaluated by measuring the levels of various inflammatory cytokines. The content of reactive oxygen species (ROS) was detected by the use of molecular imaging methods for real­time tracking and the therapeutic effects of EVs on mucosal healing in mice with colitis were evaluated. The results revealed that the injection of EVs regulated the balance of pro­inflammatory and anti­inflammatory cytokines in colon tissue. Treatment with EVs also suppressed oxidative stress by decreasing the activity of myeloperoxidase (MPO) and ROS. Histological analysis further confirmed that the EVs significantly promoted mucosal healing, as reflected by the promotion of the proliferation of colonic epithelial cells and the maintenance of tight junctions. Taken together, the findings of the present study demonstrated that EVs derived from hP­MSCs alleviated TNBS­induced colitis by inhibiting inflammation and oxidative stress. These findings may provide a novel theoretical basis for the EV­based treatment of IBD.

IGF-1C domain-modified chitosan hydrogel accelerates cutaneous wound healing by promoting angiogenesis.

Background: Complete regeneration after skin injury remains a critical clinical challenge. Hydrogels, modified with growth factors or mimicking peptides, have been applied for functional tissue regeneration by increasing the bioactivity of engineered matrices. Methodology & results: We synthesized an injectable biological hydrogel, C domain of IGF-1 (IGF-1C)-modified chitosan (CS-IGF-1C) hydrogel. Mouse model of cutaneous wound healing was established to investigate whether this hydrogel could promote wound healing. Our results demonstrated that CS-IGF-1C hydrogel exhibited superior proangiogenic effects, resulting in accelerated wound closure and improved extracellular matrix remodeling. Bioluminescence imaging and histology analysis confirmed the proangiogenic role of CS-IGF-1C hydrogel. Conclusion: CS-IGF-1C hydrogel could accelerate cutaneous wound healing by stimulating angiogenesis.

Enhanced therapeutic effects of MSC-derived extracellular vesicles with an injectable collagen matrix for experimental acute kidney injury treatment.

BACKGROUND: Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) have been shown to have therapeutic potential for ischemic diseases and are considered an alternative to cell therapy. However, the low retention and poor stability of EVs post-transplantation in vivo remain obstacle prior to the clinical application of EVs. METHODS: This study was designed to investigate whether collagen matrix could increase the retention and stability of EVs and further improve the therapeutic effects in murine acute kidney injury (AKI) model. EVs were isolated from human placental MSCs (hP-MSC-EVs) and encapsulated in a collagen matrix. Then, we investigated whether collagen matrix can prolong the retention of EVs in vivo, further enhancing the therapeutic efficiency of EVs in AKI. RESULTS: Our results indicated that collagen matrix could effectively encapsulate EVs, significantly increase the stability of EVs, and promote the sustained release of EVs. Collagen matrix has improved the retention of EVs in the AKI model, which was proved by Gaussia luciferase (Gluc) imaging. The application of collagen matrix remarkably facilitated the proliferation of renal tubular epithelial cells in AKI compared with EVs alone. Moreover, collagen matrix could further augment the therapeutic effects of hP-MSC-EVs as revealed by angiogenesis, fibrosis and apoptosis, and functional analysis. Finally, we found that EVs play a therapeutic role by inhibiting endoplasmic reticulum (ER) stress. CONCLUSIONS: Collagen matrix markedly enhanced the retention of EVs and further augmented the therapeutic effects of EVs for AKI. This strategy for improving the efficacy of EVs therapy provides a new direction for cell-free therapy.

Enhanced Lysosomal Escape of pH-Responsive Polyethylenimine-Betaine Functionalized Carbon Nanotube for the Codelivery of Survivin Small Interfering RNA and Doxorubicin.

The combination of gene therapy and chemotherapy has recently received considerable attention for cancer treatment. However, low transfection efficiency and poor endosomal escape of genes from nanocarriers strongly limit the success of the clinical use of small interfering RNA (siRNA). In this study, a novel pH-responsive, surface-modified single-walled carbon nanotube (SWCNT) was designed for the codelivery of doxorubicin (DOX) and survivin siRNA. Polyethylenimine (PEI) was covalently conjugated with betaine, and the resulting PEI-betaine (PB) was further synthesized with the oxidized SWCNT to form SWCNT-PB (SPB), which exhibits an excellent pH-responsive lysosomal escape of siRNA. SPB was modified with the targeting and penetrating peptide BR2 (SPBB), thereby achieving considerably higher uptake of siRNA than SWCNT-PEI (SP) or SPB. Furthermore, SPBB-siRNA presented substantially lower survivin expression and higher apoptotic index than Lipofectamine 2000. DOX and survivin siRNA were adsorbed onto SPB to form DOX-SPBB-siRNA, and siRNA/DOX was released into the cytoplasm and nuclei of adenocarcinomic human alveolar basal epithelial (A549) cells without lysosomal retention. Compared with SPBB-siRNA or DOX-SPBB treatment alone, DOX-SPBB-siRNA significantly reduced tumor volume in A549 cell-bearing nude mice, demonstrating the synergistic effects of DOX and survivin siRNA. Pathological analysis also indicated the potential therapeutic effects of DOX-SPBB-siRNA on tumors without distinct damages to normal tissues. In conclusion, the novel functionalized SWCNT loaded with DOX and survivin siRNA was successfully synthesized, and the nanocomplex exhibited effective antitumor effects both in vitro and in vivo, thereby providing an alternative strategy for the codelivery of antitumor drugs and genes.

Effective deactivation of A549 tumor cells in vitro and in vivo by RGD-decorated chitosan-functionalized single-walled carbon nanotube loading docetaxel.

This study aims to construct and evaluate RGD-decorated chitosan (CS)-functionalized pH-responsive single-walled carbon nanotube (SWCNT) carriers using docetaxel (DTX) as a model anticancer drug. DTX was loaded onto SWCNT via π-π stacking interaction (SWCNT-DTX), followed by the non-covalent conjugation of RGD-decorated CS to SWCNT-DTX to prepare RGD-CS-SWCNT-DTX. The RGD-CS-SWCNT-DTX showed significantly higher drug release than the pure drug, giving higher release rate at pH 5.0 (68%) than pH 7.4 (49%). The RGD-CS-SWCNT-DTX could significantly inhibit the growth of A549 tumor cells in vitro, and the uptake amount of A549 cells was obviously higher than that of MCF-7 cells. Meanwhile, the cellular uptake of RGD-CS-SWCNT-DTX was higher than that of CS-SWCNT-DTX in A549 cells, mainly through clathrin and caveolae-mediated endocytosis. The RGD-CS-SWCNT-DTX significantly inhibited tumor growth of A549 cell-bearing nude mice through active tumor-targeting ability. Furthermore, no pathological changes were found in tissues and organs. The result demonstrated that RGD-CS-SWCNT-DTX displayed high drug loading, pH-responsive drug release, remarkable antitumor effect in vitro and in vivo, and also good safety to animal body.