NS1-mediated enhancement of MVC transcription and replication promoted by KAT5/H4K12ac.

Histone modifications function in both cellular and viral gene expression. However, the roles of acetyltransferases and histone acetylation in parvoviral infection remain poorly understood. In the current study, we found the histone deacetylase (HDAC) inhibitor, trichostatin A (TSA), promoted the replication and transcription of parvovirus minute virus of canines (MVC). Notably, the expression of host acetyltransferases KAT5, GTF3C4, and KAT2A was increased in MVC infection, as well as H4 acetylation (H4K12ac). KAT5 is not only responsible for H4K12ac but also crucial for viral replication and transcription. The viral nonstructural protein NS1 interacted with KAT5 and enhanced its expression. Further study showed that Y44 in KAT5, which may be tyrosine-phosphorylated, is indispensable for NS1-mediated enhancement of KAT5 and efficient MVC replication. The data demonstrated that NS1 interacted with KAT5, which resulted in an enhanced H4K12ac level to promote viral replication and transcription, implying the epigenetic addition of H4K12ac in viral chromatin-like structure by KAT5 is vital for MVC replication.IMPORTANCEParvoviral genomes are chromatinized with host histones. Therefore, histone acetylation and related acetyltransferases are required for the virus to modify histones and open densely packed chromatin structures. This study illustrated that histone acetylation status is important for MVC replication and transcription and revealed a novel mechanism that the viral nonstructural protein NS1 hijacks the host acetyltransferase KAT5 to enhance histone acetylation of H4K12ac, which relies on a potential tyrosine phosphorylation site, Y44 in KAT5. Other parvoviruses share a similar genome organization and coding potential and may adapt a similar strategy for efficient viral replication and transcription.

Predictive factors that influence the clinical efficacy of umbilical cord-derived mesenchymal stromal cells in the treatment of type 2 diabetes mellitus.

BACKGROUND: Our previous single-center, randomized, double-blinded, placebo-controlled phase 2 study evaluated the safety and effectiveness of human umbilical cord mesenchymal stromal cell (UC-MSC) transfusion for treating patients with type 2 diabetes mellitus (T2DM). Indeed, this potential treatment strategy was able to reduce insulin use by half in a considerable number of patients. However, many other patients' responses to UC-MSC transfusion were insignificant. The selection of patients who might benefit from UC-MSC treatment is crucial from a clinical standpoint. METHODS: In this post hoc analysis, 37 patients who received UC-MSC transfusions were divided into two groups based on whether their glycated hemoglobin (hemoglobin A1c, or HbA1c) level was less than 7% after receiving UC-MSC treatment. The baseline differences between the two groups were summarized, and potential factors influencing efficacy of UC-MSCs for T2DM were analyzed by univariate and multivariate logistic regression. The correlations between the relevant hormone levels and the treatment effect were further analyzed. RESULTS: At the 9-week follow-up, 59.5% of patients achieved their targeted HbA1c level. Male patients with lower baseline HbA1c and greater C-peptide area under the curve (AUCC-pep) values responded favorably to UC-MSC transfusion, according to multivariate analysis. The effectiveness of UC-MSCs transfusion was predicted by AUCC-pep (cutoff value: 14.22 ng/h/mL). Further investigation revealed that AUCC-pep was increased in male patients with greater baseline testosterone levels. CONCLUSIONS: Male patients with T2DM with greater AUCC-pep may be more likely to respond clinically to UC-MSC therapy, and further large-scale multi-ethnic clinical studies should be performed to confirm the conclusion.

N4-acetylcytidine regulates the replication and pathogenicity of enterovirus 71.

Chemical modifications are important for RNA function and metabolism. N4-acetylcytidine (ac4C) is critical for the translation and stability of mRNA. Although ac4C is found in RNA viruses, the detailed mechanisms through which ac4C affects viral replication are unclear. Here, we reported that the 5' untranslated region of the enterovirus 71 (EV71) genome was ac4C modified by the host acetyltransferase NAT10. Inhibition of NAT10 and mutation of the ac4C sites within the internal ribosomal entry site (IRES) suppressed EV71 replication. ac4C enhanced viral RNA translation via selective recruitment of PCBP2 to the IRES and boosted RNA stability. Additionally, ac4C increased the binding of RNA-dependent RNA polymerase (3D) to viral RNA. Notably, ac4C-deficient mutant EV71 showed reduced pathogenicity in vivo. Our findings highlighted the essential role of ac4C in EV71 infection and provided insights into potential antiviral treatments.

Bone mesenchymal stromal cell-derived small extracellular vesicles inhibit inflammation and ameliorate sepsis via delivery of microRNA-21a-5p.

BACKGROUND AIMS: Sepsis is a potentially life-threatening disease that results from a severe systemic inflammatory response due to infection. Mesenchymal stromal cell-derived small extracellular vesicles (MSC sEVs) are able to transfer bioactive molecules and have been demonstrated to play an important role in the pathophysiological process of sepsis. Herein the authors aimed to investigate the potential role and downstream molecular mechanism of MSC sEVs in sepsis. METHODS: MSC sEVs were acquired by ultracentrifugation and then injected into a cecal ligation and puncture mouse model. The efficacy of MSC sEVs in both in vitro and in vivo models of sepsis was evaluated. RESULTS: MSC sEV therapy improved survival, reduced sepsis-induced inflammation, attenuated pulmonary capillary permeability and improved liver and kidney function in septic mice. In addition, the authors found that microRNA-21a-5p (miR-21a-5p) was highly enriched in MSC sEVs, could be transferred to recipient cells, inhibited inflammation and increased survival in septic mice. Furthermore, the authors demonstrated that MSC sEV miR-21a-5p suppressed inflammation by targeting toll-like receptor 4 and programmed cell death 4. The therapeutic efficacy of MSC sEVs was partially abrogated by transfection with miR-21a-5p inhibitors. CONCLUSIONS: Collectively, the authors' data suggest that miR-21a-5p-bearing MSC sEVs may be a prospective and effective sepsis therapeutic strategy.

Human umbilical cord mesenchymal stromal cell small extracellular vesicle transfer of microRNA-223-3p to lung epithelial cells attenuates inflammation in acute lung injury in mice.

BACKGROUND: Acute lung injury (ALI), manifested as strong pulmonary inflammation and alveolar epithelial damage, is a life-threatening disease with high morbidity and mortality. Small extracellular vesicles (sEVs), secreted by multiple types of cells, are critical cellular communication mediators and can inhibit inflammation by transferring bioactive molecules, such as microRNAs (miRNAs). Thus, we hypothesized that sEVs derived from mesenchymal stromal cells (MSC sEVs) could transfer miRNAs to attenuate inflammation of lung epithelial cells during ALI. METHODS: C57BL/6 male mice were intratracheally administered LPS (10 mg/kg). Six hours later, the mice were randomly administered with MSC sEVs (40 µg per mouse in 150 µl of saline), which were collected by ultracentrifugation. Control group received saline administration. After 48 h, the mice were sacrificed to evaluate pulmonary microvascular permeability and inflammatory responses. In vitro, A549 cells and primary human small airway epithelial cells (SAECs) were stimulated with LPS with or without MSC sEVs treatment. RESULTS: In vitro, MSC sEVs could also inhibit the inflammation induced by LPS in A549 cells and SAECs (reducing TNF-α, IL-1ß, IL-6 and MCP-1). Moreover, MSC sEV treatment improved the survival rate, alleviated pulmonary microvascular permeability, and inhibited proinflammatory responses (reducing TNF-α, IL-1ß, IL-6 and JE-1) in ALI mice. Notably, miR-223-3p was found to be served as a critical mediator in MSC sEV-induced regulatory effects through inhibition of poly (adenosine diphosphate-ribose) polymerase-1 (PARP-1) in lung epithelial cells. CONCLUSIONS: Overall, these findings suggest that MSC sEVs may offer a novel promising strategy for ALI.

Human mesenchymal stromal cells small extracellular vesicles attenuate sepsis-induced acute lung injury in a mouse model: the role of oxidative stress and the mitogen-activated protein kinase/nuclear factor kappa B pathway.

BACKGROUND AIMS: Acute lung injury (ALI) secondary to sepsis is a complex disease associated with high morbidity and mortality. Mesenchymal stem cells (MSCs) and their conditioned medium have been demonstrated to reduce alveolar inflammation, improve lung endothelial barrier permeability and modulate oxidative stress in vivo and in vitro. Recently, MSCs have been found to release small extracellular vesicles (sEVs) that can deliver functionally active biomolecules into recipient cells. The authors' study was designed to determine whether sEVs released by MSCs would be effective in sepsis-induced ALI mice and to identify the potential mechanisms. METHODS: A total of 6 h after cercal ligation and puncture, the mice received saline, sEV-depleted conditioned medium (sEVD-CM) or MSC sEVs via the tail vein. RESULTS: The administration of MSC sEVs improved pulmonary microvascular permeability and inhibited both histopathological changes and the infiltration of polymorphonuclear neutrophils into lung tissues. In addition, the activities of antioxidant enzymes were significantly increased in the group treated with sEVs compared with the saline and sEVD-CM groups, whereas lipid peroxidation was significantly decreased. Furthermore, sEVs were found to possibly inhibit phosphorylation of the mitogen-activated protein kinase/nuclear factor kappa B (MAPK/NF-κB) pathway and degradation of IκB but increase the activities of nuclear factor erythroid 2-related factor 2 and heme oxygenase 1. CONCLUSIONS: These findings suggest that one of the effective therapeutic mechanisms of sEVs against sepsis-induced ALI may be associated with upregulation of anti-oxidative enzymes and inhibition of MAPK/NF-κB activation.

N6-methyladenosine modification and METTL3 modulate enterovirus 71 replication.

N6-methyladenosine (m6A) constitutes one of the most abundant internal RNA modifications and is critical for RNA metabolism and function. It has been previously reported that viral RNA contains internal m6A modifications; however, only recently the function of m6A modification in viral RNAs has been elucidated during infections of HIV, hepatitis C virus and Zika virus. In the present study, we found that enterovirus 71 (EV71) RNA undergoes m6A modification during viral infection, which alters the expression and localization of the methyltransferase and demethylase of m6A, and its binding proteins. Moreover, knockdown of m6A methyltransferase resulted in decreased EV71 replication, whereas knockdown of the demethylase had the opposite effect. Further study showed that the m6A binding proteins also participate in the regulation of viral replication. In particular, two m6A modification sites were identified in the viral genome, of which mutations resulted in decreased virus replication, suggesting that m6A modification plays an important role in EV71 replication. Notably, we found that METTL3 interacted with viral RNA-dependent RNA polymerase 3D and induced enhanced sumoylation and ubiquitination of the 3D polymerase that boosted viral replication. Taken together, our findings demonstrated that the host m6A modification complex interacts with viral proteins to modulate EV71 replication.

Berberine: A Traditional Natural Product With Novel Biological Activities.

CONTEXT: Having been used for thousands of years to treat gastrointestinal diseases, the natural isoquinoline alkaloid, berberine, has exhibited a wide spectrum of biochemical and pharmacological effects in studies of recent years. OBJECTIVE: The review intended to examine the many novel bioactivities of berberine, including antidiabetic, anticancer, neuroprotective, anti-inflammatory, and anti-atherosclerotic actions. DESIGN: The research team searched the MEDLINE database using PubMed, using different keyword combinations, including berberine AND diabetes, berberine AND cancer, berberine AND (neuron OR brain), berberine AND inflammation, and "berberine AND atherosclerosis to find studies evaluating the various effects exerted berberine. CONCLUSION: Berberine is a promising multipotent agent to combat diabetes, cancer, Alzheimer's disease, and other diseases.

Arsenic trioxide inhibits the differentiation of fibroblasts to myofibroblasts through nuclear factor erythroid 2-like 2 (NFE2L2) protein and the Smad2/3 pathway.

BACKGROUND: Tissue contraction and the extracellular matrix deposition are part of the pathogenesis of hypertrophic scars. The transcriptional factor NFE2L2 inhibits fibroblast differentiation in idiopathic pulmonary fibrosis and promotes myofibroblast dedifferentiation. Our previous study showed that the transcription factor NFE2L2 was strongly induced on treatment with arsenic trioxide (ATO). OBJECTIVE: The present study sought to investigate the effect of ATO on myofibroblast formation to determine its potential role in hypertrophic scar treatment. METHODS: Small interfering RNA against NFE2L2 was used on treatment with ATO in human skin myofibroblasts. The expression levels of fibrosis markers were assessed by reverse transcription polymerase chain reaction, western blot, and immunofluorescence staining. The transforming growth factor-ß1 (TGF-ß1)/Smad2/3 signaling was detected by western blot. A rabbit ear model was used to evaluate the antifibrotic role of ATO. RESULTS: At the cellular level, ATO abolished fibroblast differentiation in response to TGF-ß1. ATO reduced TGF-ß1-induced reactive oxygen species accumulation through increased expression of the antioxidant gene HO-1 in fibroblasts. In addition, ATO promoted the nuclear translocation of NFE2L2 and inhibited the phosphorylation of Smad2/3. In the rabbit ear model, ATO prevented the progression of hypertrophic scar formation. CONCLUSIONS: This study provides the first evidence implying that ATO inhibits the formation of myofibroblasts in vivo and in vitro and provides a possible treatment for hypertrophic scars.

Induction of hepatocytes-derived insulin-producing cells using small molecules and identification of microRNA profiles during this procedure.

The transplantation of insulin-producing cells (IPCs) or pancreatic progenitor cells is a theoretical therapy for diabetes with insulin insufficiency. Isolated hepatocytes from newborn rats (within 24 h after birth) were progressively induced into IPCs using 5-aza-2'-deoxycytidine, Trichostatin A, retinoic acid, insulin-transferrin-selenium, and nicotinamide. We transplanted Pdx1+ pancreatic progenitors into STZ-induced diabetic mice and found the decreased blood glucose and increased insulin level in comparison with diabetic model. The dynamic expression profiles of microRNAs (miRNAs) were identified using microarray. We found 67 miRNAs were decreasingly expressed; 52 miRNAs were increasingly expressed; 27 miRNAs were specially inhibited in Stage 1 cells (multipotent progenitor cells); and 58 miRNAs were specially inhibited in Pdx1+ cells (Stage 2). Further analysis showed these miRNAs' targets were associated with genetic recombination, stem cell pluripotency maintenance, cellular structure reorganization and insulin secretion. Enrichment analysis using KEGG pathway showed the differentiation of IPCs from hepatocytes was massively more likely not mediated by canonical Wnt/ß-catenin signaling. In addition, the BMP/Smad signaling was involved in this progression. We found the dysregulated miRNAs profiles were inconsistent with cell phenotypes and might be responsible for small molecule-mediated cell differentiation during IPCs induction.

Human umbilical cord-derived mesenchymal stem cells elicit macrophages into an anti-inflammatory phenotype to alleviate insulin resistance in type 2 diabetic rats.

Insulin resistance, a major characteristic of type 2 diabetes (T2D), is closely associated with adipose tissue macrophages (ATMs) that induce chronic low-grade inflammation. Recently, mesenchymal stem cells (MSCs) have been identified in alleviation of insulin resistance. However, the underlying mechanism still remains elusive. Thus, we aimed to investigate whether the effect of MSCs on insulin resistance was related to macrophages phenotypes in adipose tissues of T2D rats. In this study, human umbilical cord-derived MSCs (UC-MSCs) infusion produced significantly anti-diabetic effects and promoted insulin sensitivity in T2D rats that were induced by a high-fat diet combined with streptozotocin and directed ATMs into an alternatively activated phenotype (M2, anti-inflammatory). In vitro, MSC-induced M2 macrophages alleviated insulin resistance caused by classically activated macrophages (M1, pro-inflammatory). Further analysis showed that M1 stimulated UC-MSCs to increase expression of interleukin (IL)-6, a molecule which upregulated IL4R expression, promoted phosphorylation of STAT6 in macrophages, and eventually polarized macrophages into M2 phenotype. Moreover, the UC-MSCs effect on macrophages was largely abrogated by small interfering RNA (siRNA) knockdown of IL-6. Together, our results indicate that UC-MSCs can alleviate insulin resistance in part via production of IL-6 that elicits M2 polarization. Additionally, human obesity and insulin resistance were associated with increased pro-inflammatory ATMs infiltration. Thus, MSCs may be a new treatment for obesity-related insulin resistance and T2D concerning macrophage polarized effects.

Umbilical cord-derived mesenchymal stromal cell-conditioned medium exerts in vitro antiaging effects in human fibroblasts.

BACKGROUND AIMS: Chronic wounds are a common complication of diabetes. Fibroblast-myofibroblast differentiation is important for wound repair, which is commonly impaired in non-healing wounds, and the underlying mechanisms need to be further elucidated. METHODS: We used high glucose (HG) to simulated the diabetes microenvironment and explored its effects on the biological features of fibroblasts in vitro. RESULTS: The results showed that prolonged HG induced senescence in fibroblasts through activation of p21 and p16 in a reactive oxygen species (ROS)-dependent manner, further delayed the viability and migration in fibroblasts and also depressed fibroblast differentiation through the TGF-ß/Smad signaling pathway. However, mesenchymal stromal cell-conditioned medium (MSC-CM) counteracts the effects of HG. Treatment of fibroblasts with MSC-CM decreased HG-induced ROS overproduction, ameliorated HG-induced senescence in fibroblasts and reversed the defects in myofibroblast formation. Our results may provide clues for the pathogenesis of chronic wounds and a theoretical basis to develop MSC-CM as an alternative therapeutic method to treatment of chronic wounds.

Granulocyte-Colony Stimulating Factor (G-CSF) Accelerates Wound Healing in Hemorrhagic Shock Rats by Enhancing Angiogenesis and Attenuating Apoptosis.

BACKGROUND Following severe trauma, treatment of cutaneous injuries is often delayed by inadequate blood supply. The aim of the present study was to determine whether granulocyte-colony stimulating factor (G-CSF) protects endothelial cells (ECs) and enhances angiogenesis in a rat model of hemorrhagic shock (HS) combined with cutaneous injury after resuscitation. MATERIAL AND METHODS The HS rats with full-thickness defects were resuscitated and randomly divided into a G-CSF group (200 µg/kg body weight), a normal saline group, and a blank control group. Histological staining was to used estimate the recovery and apoptosis of skin. Apoptosis- and angiogenesis-related factors were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot (WB). Scratch assay, tube formation, and WB experiments were performed to verify the functional effects of G-CSF on HUVECs in vitro. RESULTS H&E staining and Masson trichrome staining showed earlier inflammation resolution and collagen synthesis in the G-CSF-treated group. Angiogenesis-related factors were elevated at mRNA and protein levels. TUNEL staining suggested fewer apoptotic cells in the G-CSF group. The apoptotic-related factors were down-regulated and anti-apoptotic factors were up-regulated in the G-CSF-treated group. Scratch assay and tube formation experiments revealed that G-CSF facilitated migration ability and angiogenic potential of HUVECs. The angiogenic and anti-apoptotic effects were also enhanced in vitro. CONCLUSIONS Our results suggest that G-CSF after resuscitation attenuates local apoptosis and accelerates angiogenesis. These findings hold great promise for improving therapy for cutaneous injury in severe trauma and ischemia diseases.

Mesenchymal stem cell-conditioned medium accelerates wound healing with fewer scars.

Mesenchymal stem cells (MSCs) derived from umbilical cords (UC-MSCs) have been shown to enhance cutaneous wound healing by means of the paracrine activity. Fibroblasts are the primary cells involved in wound repair. The paracrine effects of UC-MSCs on dermal fibroblasts have not been fully explored in vitro or in vivo. Dermal fibroblasts were treated with conditioned media from UC-MSCs (UC-MSC-CM). In this model, UC-MSC-CM increased the proliferation and migration of dermal fibroblasts. Moreover, adult dermal fibroblasts transitioned into a phenotype with a low myofibroblast formation capacity, a decreased ratio of transforming growth factor-ß1,3 (TGF-ß1/3) and an increased ratio of matrix metalloproteinase/tissue inhibitor of metalloproteinases (MMP/TIMP). Additionally, UC-MSC-CM-treated wounds showed accelerated healing with fewer scars compared with control groups. These observations suggest that UC-MSC-CM may be a feasible strategy to promote cutaneous repair and a potential means to realise scarless healing.

Hypoxia pretreatment of bone marrow-derived mesenchymal stem cells seeded in a collagen-chitosan sponge scaffold promotes skin wound healing in diabetic rats with hindlimb ischemia.

Bone marrow-derived mesenchymal stem cells (BM-MSCs) have properties that make them promising for the treatment of chronic nonhealing wounds. The major challenge is ensuring an efficient, safe, and painless delivery of BM-MSCs. Tissue-engineered skin substitutes have considerable benefits in skin damage resulting from chronic nonhealing wounds. Here, we have constructed a three-dimensional biomimetic scaffold known as collagen-chitosan sponge scaffolds (CCSS) using the cross-linking and freeze-drying method. Scanning electron microscopy images showed that CCSS had an interconnected network pore configuration about 100 µm and exhibited a suitable swelling ratio for maintaining morphological stability and appropriate biodegradability to improve biostability using swelling and degradation assays. Furthermore, BM-MSCs were seeded in CCSS using the two-step seeding method to construct tissue-engineered skin substitutes. In addition, in this three-dimensional biomimetic CCSS, BM-MSCs secreted their own collagen and maintain favorable survival ability and viability. Importantly, BM-MSCs exhibited a significant upregulated expression of proangiogenesis factors, including HIF-1α, VEGF, and PDGF following hypoxia pretreatment. In vivo, hypoxia pretreatment of the skin substitute observably accelerated wound closure via the reduction of inflammation and enhanced angiogenesis in diabetic rats with hindlimb ischemia. Thus, hypoxia pretreatment of the skin substitutes can serve as ideal bioengineering skin substitutes to promote optimal diabetic skin wound healing.

Mesenchymal Stem Cell Conditioned Medium Promotes Proliferation and Migration of Alveolar Epithelial Cells under Septic Conditions In Vitro via the JNK-P38 Signaling Pathway.

BACKGROUND/AIMS: Mesenchymal stem cell (MSC) based therapies may be useful for treating acute respiratory distress syndrome (ARDS), but the underlying mechanisms are incompletely understood. We investigated the impact of human umbilical cord Wharton's jelly-derived MSC (hUC-MSC) secreted factors on alveolar epithelial cells under septic conditions and determined the relevant intracellular signaling pathways. METHODS: Human alveolar epithelial cells (AEC) and primary human small airway epithelial cells (SAEC) were subjected to lipopolysaccharide (LPS) with or without the presence of hUC-MSC-conditioned medium (CM). Proliferation and migration of AEC and SAEC were determined via an MTT assay, a wound healing assay and a transwell migration assay (only for AEC). Protein phosphorylation was determined by western blot and the experiments were repeated in presence of small-molecule inhibitors. The hMSC-secretory proteins were identified by LC-MS/MS mass spectrometry. RESULTS: MSC-CM enhanced proliferation and migration. Activation of JNK and P38, but not ERK, was required for the proliferation and migration of AEC and SAEC. Pretreatment of AEC or SAEC with SP600125, an inhibitor of JNK1 or SB200358, an inhibitor of P38, significantly reduced cell proliferation and migration. An array of proteins including TGF-beta receptor type-1, TGF-beta receptor type-2, Ras-related C3 botulinum toxin substrate 1 and Ras-related C3 botulinum toxin substrate 2 which influencing the proliferation and migration of AEC and SAEC were detected in MSC-CM. CONCLUSION: Our data suggest MSC promote epithelial cell repair through releasing a repertoire of paracrine factors via activation of JNK and P38 MAPK.

LPS-preconditioned mesenchymal stromal cells modify macrophage polarization for resolution of chronic inflammation via exosome-shuttled let-7b.

BACKGROUND: Within the last few years, it has become evident that LPS-preconditioned mesenchymal stromal cells (LPS pre-MSCs) show enhanced paracrine effects, including increased trophic support and improved regenerative and repair properties. MSCs may release large amounts of exosomes for cell-to-cell communication and maintain a dynamic and homeostatic microenvironment for tissue repair. The present study assesses the therapeutic efficacy and mechanisms of LPS-preconditioned MSC-derived exosomes (LPS pre-Exo) for chronic inflammation and wound healing. METHODS: We extracted exosomes from the supernatant of LPS pre-MSCs using a gradient centrifugation method. In vitro, THP-1 cells were cultured with high glucose (HG, 30 mM) as an inflammatory model and treated with LPS pre-Exo for 48 h. The expression of inflammation-related cytokines was detected by real-time RT-PCR, and the distribution of macrophage subtype was measured by immunofluorescence. Next, the miRNA expression profiles of LPS pre-Exo were evaluated using miRNA microarray analysis. The molecular signaling pathway responsible for the regenerative potential was identified by western blotting. In vivo, we established a cutaneous wound model in streptozotocin-induced diabetic rats, and LPS pre-Exo were injected dispersively into the wound edge. The curative effects of LPS pre-Exo on inflammation and wound healing were observed and evaluated. RESULTS: LPS pre-Exo have a better ability than untreated MSC-derived exosomes (un-Exo) to modulate the balance of macrophages due to their upregulation of the expression of anti-inflammatory cytokines and promotion of M2 macrophage activation. Microarray analysis of LPS pre-Exo identified the unique expression of let-7b compared with un-Exo, and the let-7b/TLR4 pathway served as potential contributor to macrophage polarization and inflammatory ablation. Further investigation of the mechanisms that control let-7b expression demonstrated that a TLR4/NF-κB/STAT3/AKT regulatory signaling pathway plays a critical role in the regulation of macrophage plasticity. Knockdown of AKT in THP-1 cells similarly abolished the immunomodulatory effect of LPS pre-Exo. In vivo, LPS pre-Exo greatly alleviated inflammation and enhanced diabetic cutaneous wound healing. CONCLUSION: LPS pre-Exo may have improved regulatory abilities for macrophage polarization and resolution of chronic inflammation by shuttling let-7b, and these exosomes carry much immunotherapeutic potential for wound healing.

Mesenchymal stem cell-based therapy for nonhealing wounds: today and tomorrow.

Although advancements have been made with traditional therapies, the treatment of chronic nonhealing wounds still remains a tough challenge. In the past two decades, mesenchymal stem cell (MSC)-based therapy has emerged as a promising therapeutic strategy for nonhealing wounds because of their characteristics including self-renewal and a multidirectional differentiation ability and their easy collection and weak immunogenicity. There is a growing body of basic scientific studies that shed light on the functional mechanism of MSCs in modulating nonhealing wounds. Furthermore, critical advances have been achieved using MSC-based therapy in preclinical animal models as well as in clinics trials. In this present review, we summarize the mechanisms of MSCs and highlight the important preclinical and clinical trials of MSC therapy for nonhealing wounds. In particular, the combination of MSCs transplantation and tissue-engineered skin is addressed as a new strategy to optimize the delivery efficiency and therapeutic potential. Additionally, the current drawbacks of MSC therapy and the potential to further optimize the use of MSCs are implied.

[Effects of conditioned media for rat bone marrow-derived mesenchymal stem cells on palmitic acid-induced insulin resistance in HepG2 cells].

OBJECTIVE: To study the effect of conditioned media for rat bone marrow mesenchymal stem cells (BMSCs-CM) on palmitic acid (PA)-induced insulin resistance (IR) in HepG2 cells and its underlying molecular mechanisms. METHODS: HepG2 cells were treated with or without BMSCs-CM and L-DMEM in the presence or absence of PA.Glucose utilization in HepG2 cells were detected with PAS, glucose and glycogen measurements. Western blotting was used to assess the expression of phospho-insulin receptor substrate (p-IRS), phosphatidylinositol 3-kinase (PI3K) and p-AKT. RESULTS: (1) Incubation of HepG2 cells with 0.25 mmol/L PA for 24 hours significantly increased the glucose concentration and decreased the glycogen content (P<0.05) in the media. (2) Treatment with BMSCs-CM significantly ameliorated the glucose and glycogen alteration in cells pretreated with PA (P<0.05), however, no obvious effect of BMSCs-CM on the cell glucose and glycogen production. (3) BMSCs-CM treatment also increased protein expression of p-IRS, PI3K and p-AKT in PA incubated HapG2 cells (P<0.05). The effect of BMSCs-CM on PI3K and p-AKT expression could be mimicked upon addition of 740Y-P, a PI3K agonist, but abolished by LY294002, a PI3K specific inhibitor. CONCLUSIONS: BMSCs-CM could improve the insulin sensitivity in HepG2 cells pretreated with PA through upregulation of insulin signaling component expression.

Applications of Engineered Skin Tissue for Cosmetic Component and Toxicology Detection.

The scale of the cosmetic market is increasing every day. There are many safety risks to cosmetics, but they benefit people at the same time. The skin can become red, swollen, itchy, chronically toxic, and senescent due to the misuse of cosmetics, triggering skin injuries, with contact dermatitis being the most common. Therefore, there is an urgent need for a system that can scientifically and rationally detect the composition and perform a toxicological assessment of cosmetic products. Traditional detection methods rely on instrumentation and method selection, which are less sensitive and more complex to perform. Engineered skin tissue has emerged with the advent of tissue engineering technology as an emerging bioengineering technology. The ideal engineered skin tissue is the basis for building good in vitro structures and physiological functions in this field. This review introduces the existing cosmetic testing and toxicological evaluation methods, the current development status, and the types and characteristics of engineered skin tissue. The application of engineered skin tissue in the field of cosmetic composition detection and toxicological evaluation, as well as the different types of tissue engineering scaffold materials and three-dimensional (3D) organoid preparation approaches, is highlighted in this review to provide methods and ideas for constructing the next engineered skin tissue for cosmetic raw material component analysis and toxicological evaluation.