Adipose Tissue-derived Mesenchymal Stem Cells Research Articles

Preparation and In Vitro Evaluation of Chitosan-g-Oligolactide Based Films and Macroporous Hydrogels for Tissue Engineering.

In the current study, novel matrices based on chitosan-g-oligo (L,L-/L,D-lactide) copolymers were fabricated. In particular, 2D films were prepared by solvent casting, while 3D macroporous hydrogels were obtained by lyophilization of copolymer solutions. Copolymers of chitosan (Chit) with semi-crystalline oligo (L,L-lactide) (Chit-LL) or amorphous oligo (L,D-lactide) (Chit-LD) were obtained by solid-state mechanochemical synthesis. The structure of the hydrogels was found to be a system of interconnected macropores with an average size of 150 μm. In vitro degradation of these copolymer-based matrices was shown to increase in the case of the Chit-LL-based hydrogel by 34% and decrease for the Chit-LD-based hydrogel by 23% compared to the parameter of the Chit sample. Localization and distribution of mouse fibroblast L929 cells and adipose tissue-derived mesenchymal stromal cells (MSCs) within the hydrogels was studied by confocal laser scanning microscopy (CLSM). Moreover, cellular response, namely cell adhesion, spreading, growth, proliferation, as well as cell differentiation in vitro were also evaluated in the hydrogels for 10-14 days. Both the Chit-LL and Chit-LD matrices were shown to support cell growth and proliferation, while they had improved swelling compared to the Chit matrix. Osteogenic MSCs differentiation on the copolymer-based films was studied by quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR). Maximal expression levels of osteogenesis markers (alkaline phosphatase (ALPL), bone transcription factor (Runx2), and osteopontin (SPP1) were revealed for the Chit-LD films. Thus, osteodifferentiation was demonstrated to depend on the film composition. Both Chit-LL and Chit-LD copolymer-based matrices are promising for tissue engineering.

Mild heat treatment in vitro potentiates human adipose stem cells: delayed aging and improved quality for long term culture

BackgroundMesenchymal stem cells (MSCs) have gained significant attention for diverse biomedical applications, including cell-based therapy. Hence, in vitro expansion of MSCs is critical; however, in vitro MSC culture, especially long-term culture, inevitably leads to significant loss of stemness, growth, and differentiation potential.MethodEffects of mild heat treatment (HT) conditions (temperature, duration, and repetition) on the characteristics of adipose tissue-derived MSCs in vitro were systematically investigated. Characteristics of the MSCs subjected to the predetermined HT conditions (41 or 44ºC, 1 h, and 2X HT) were first analyzed in a single passage using various assays. In addition, the feasibility of HT for long-term MSC culture was studied. The RNA sequencing analyses were performed to elucidate the mechanism of HT effects on MSCs.ResultsA comprehensive exploration of various HT conditions revealed that specific mild HT at 41ºC or 44ºC for 1 h upregulated the expression of heat shock proteins and stemness markers and enhanced differentiation potentials. Furthermore, periodic mild HT extended the maintenance of growth rate and stemness of MSCs up to an additional 10 passages, which substantially retarded their spontaneous aging during subsequent in vitro culture. RNA sequencing analyses unveiled that HT downregulated genes associated with aging and apoptosis.ConclusionOur study successfully demonstrated that mild HT of MSCs has positive effects on their application in various biomedical fields, enhancing their capabilities and slowing down the aging process.Graphical

In vitro Study of Primary Isolation and Culture of Adipose-Derived Stem Cells and Induction of Chondrogenic Differentiation.

Articular cartilage has constrained potential to restore. The mesenchymal stem cellular remedy has presented new treatment possibilities for this circumstance. The experiment aimed to verify the chondrogenic differentiation capacity of rat adipose tissue-derived mesenchymal stem cells (AD-MSCs) in vitroinside the presence or absence of transforming growth factor-beta (TGF-β1). Rat's subcutaneous adipose tissue minced into a small piece (2-3 mm3) was aseptically collected from the subcutaneous fat under anaesthesia and then digested with collagenase type I (1 mg/mL). Spontaneous chondrogenesis occurred in both AD-MSCs pellet cultures and was similar in both TGF-β1 treated. The untreated pellet cultures were collected after 21 days. Histological assessment for evaluating the level of proteoglycan by alcin blue staining and immunohistochemistry for detecting the presence of collagen type II. A monoclonal antibody directed against collagen type II. Adipose-derived stem cells (AD-MSCs) isolated from rats were immunophenotyped for the expression of MSCs cell surface markers and was performed by Flow cytometer, which demonstrated AD-MSCs highly expressed CD73 (99.69±2.6%), CD90 (98.11±0.3%), and week expression CD44 (17.15±0.3%). The result of histological staining showed the presence of extracellular matrix (ECM) in the hyaline cartilage. This staining indicated a deposit of "acid mucopolysaccharides" in the proximity of the cells. Additionally, most cells are rounded cells stained positive for the presence of the cells encompassed by extracellular matrix (ECM), which were like chondrocytes as seen from the magnified view, lightly pink stained nuclei, and nuclear fast red stain. However, the immunohistochemistry method demonstrated that the presence of TGF-β1 decreased the levels of collagen type I and increased the levels of collagen type II. In conclusion, subcutaneous adipose tissue-derived stem cells can be used in cartilage tissue engineering.

Effect of allogeneic adipose tissue-derived mesenchymal stromal cell treatment in chronic ischaemic heart failure with reduced ejection fraction-the SCIENCE trial.

The aim of the SCIENCE trial was to investigate whether a single treatment with direct intramyocardial injections of adipose tissue-derived mesenchymal stromal cells (CSCC_ASCs) was safe and improved cardiac function in patients with chronic ischaemic heart failure with reduced ejection fraction (HFrEF). The study was a European multicentre, double-blind, placebo-controlled phase II trial using allogeneic CSCC_ASCs from healthy donors or placebo (2:1 randomization). Main inclusion criteria were New York Heart Association (NYHA) class II-III, left ventricular ejection fraction (LVEF) <45%, and N-terminal pro-B-type natriuretic peptide (NT-proBNP) levels >300 pg/ml. CSCC_ASCs or placebo (isotonic saline) were injected directly into viable myocardium. The primary endpoint was change in left ventricular end-systolic volume (LVESV) at 6-month follow-up measured by echocardiography. A total of 133 symptomatic HFrEF patients were included. The treatment was safe without any drug-related severe adverse events or difference in cardiac-related adverse events during a 3-year follow-up period. There were no significant differences between groups during follow-up in LVESV (0.3 ± 5.0ml, p=0.945), nor in secondary endpoints of left ventricular end-diastolic volume (-2.0 ± 6.0ml, p=0.736) and LVEF (-1.6 ± 1.0%, p=0.119). The NYHA class improved slightly within the first year in both groups without any difference between groups. There were no changes in 6-min walk test, NT-proBNP, C-reactive protein or quality of life the first year in any groups. The SCIENCE trial demonstrated safety of intramyocardial allogeneic CSCC_ASC therapy in patients with chronic HFrEF. However, it was not possible to improve the pre-defined endpoints and induce restoration of cardiac function or clinical symptoms.

Better isolation, proliferation and differentiation of human adipose-derived mesenchymal stem cells using human serum.

Mesenchymal stem cells have many applications in medicine. Attention to the proliferation and differentiation of stem cell differentiation is an important issue. The aim of this study was to investigate the possibility of optimal isolation, proliferation, and differentiation of adipose tissue-derived mesenchymal stem cells (ADSCs) using human serum. Human serum (HS) was obtained from the venous blood of eight healthy individuals. The rate of proliferation and differentiation of ADSCs and expression of surface markers was assessed by flow cytometry. Bone differentiation was assessed using Alizarin Red staining. Data were analyzed using statistical software. Over time, HS showed more proliferation than fetal bovine serum (FBS) -enriched cells (p <0.05). Differentiation of ADSCs cells ls in HS-enriched medium is faster and more pronounced than differentiation in the control group. The expression of surface markers in the medium containing HS was the same as the medium containing FBS where the expression levels of CD105 and CD95 were found to be positive and the expression of CD34 and CD45 was negative. Due to the better proliferation of adipose tissue-derived mesenchymal cells in the medium containing HS than FBS, it is suggested that human serum be used in future clinical studies. Also, HS is healthier, safer, more accessible, and more affordable than FBS.

Remote Adipose Tissue-Derived Stromal Cells of Patients with Lung Adenocarcinoma Generate a Similar Malignant Microenvironment of the Lung Stromal Counterpart.

Cancer alters both local and distant tissue by influencing the microenvironment. In this regard, the interplay with the stromal fraction is considered critical as this latter can either foster or hamper the progression of the disease. Accordingly, the modality by which tumors may alter distant niches of stromal cells is still unclear, especially at early stages. In this short report, we attempt to better understand the biology of this cross-talk. In our "autologous stromal experimental setting," we found that remote adipose tissue-derived mesenchymal stem cells (mediastinal AMSC) obtained from patients with lung adenocarcinoma sustain proliferation and clonogenic ability of A549 and human primary lung adenocarcinoma cells similarly to the autologous stromal lung counterpart (LMSC). This effect is not observed in lung benign diseases such as the hamartochondroma. This finding was validated by conditioning benign AMSC with supernatants from LAC for up to 21 days. The new reconditioned media of the stromal fraction so obtained, was able to increase cell proliferation of A549 cells at 14 and 21 days similar to that derived from AMSC of patients with lung adenocarcinoma. The secretome generated by remote AMSC revealed overlapping to the corresponding malignant microenvironment of the autologous local LMSC. Among the plethora of 80 soluble factors analyzed by arrays, a small pool of 5 upregulated molecules including IL1-β, IL-3, MCP-1, TNF-α, and EGF, was commonly shared by both malignant-like autologous A- and L-MSC derived microenvironments vs those benign. The bioinformatics analysis revealed that these proteins were strictly and functionally interconnected to lung fibrosis and proinflammation and that miR-126, 101, 486, and let-7-g were their main targets. Accordingly, we found that in lung cancer tissues and blood samples from the same set of patients here employed, miR-126 and miR-486 displayed the highest expression levels in tissue and blood, respectively. When the miR-126-3p was silenced in A549 treated with AMSC-derived conditioned media from patients with lung adenocarcinoma, cell proliferation decreased compared to control media.

Immortalized Canine Adipose-Derived Mesenchymal Stem Cells as a Novel Candidate Cell Source for Mesenchymal Stem Cell Therapy.

Mesenchymal stem cells are expected to be a cell source for stem cell therapy of various diseases in veterinary medicine. However, donor-dependent cell heterogenicity has been a cause of inconsistent therapeutic efficiency. Therefore, we established immortalized cells from canine adipose tissue-derived mesenchymal stem cells (ADSCs) to minimize cellular heterogeneity by reducing the number of donors, evaluated their properties, and compared them to the primary cells with RNA-sequencing. Immortalized canine ADSCs were established by transduction with combinations of the R24C mutation of human cyclin-dependent kinase 4 (CDKR24C), canine cyclin D1, and canine TERT. The ADSCs transduced with CDK4R24C, cyclin D1, and TERT (ADSC-K4DT) or with CDK4R24C and cyclin D1 (ADSC-K4D) showed a dramatic increase in proliferation (population doubling level >100) without cellular senescence compared to the primary ADSCs. The cell surface markers, except for CD90 of the ADSC-K4DT and ADSC-K4D cells, were similar to those of the primary ADSCs. The ADSC-K4DT and ADSC-K4D cells maintained their trilineage differentiation capacity and chromosome condition, and did not have a tumorigenic development. The ability to inhibit lymphocyte proliferation by the ADSC-K4D cells was enhanced compared with the primary ADSCs and ADSC-K4DT cells. The pathway analysis based on RNA-sequencing revealed changes in the pathways mainly related to the cell cycle and telomerase. The ADSC-K4DT and ADSC-K4D cells had decreased CD90 expression, but there were no obvious defects associated with the decreased CD90 expression in this study. Our results suggest that ADSC-K4DT and ADSC-K4D cells are a potential novel cell source for mesenchymal stem cell therapy.

Comparative effectiveness of adipose-derived mesenchymal stromal cells in the management of knee osteoarthritis: A meta-analysis.

Osteoarthritis (OA) is the most common joint disorder, is associated with an increasing socioeconomic impact owing to the ageing population. To analyze and compare the efficacy and safety of bone-marrow-derived mesenchymal stromal cells (BM-MSCs) and adipose tissue-derived MSCs (AD-MSCs) in knee OA management from published randomized controlled trials (RCTs). Independent and duplicate electronic database searches were performed, including PubMed, EMBASE, Web of Science, and Cochrane Library, until August 2021 for RCTs that analyzed the efficacy and safety of AD-MSCs and BM-MSCs in the management of knee OA. The visual analog scale (VAS) score for pain, Western Ontario McMaster Universities Osteoarthritis Index (WOMAC), Lysholm score, Tegner score, magnetic resonance observation of cartilage repair tissue score, knee osteoarthritis outcome score (KOOS), and adverse events were analyzed. Analysis was performed on the R-platform using OpenMeta (Analyst) software. Twenty-one studies, involving 936 patients, were included. Only one study compared the two MSC sources without patient randomization; hence, the results of all included studies from both sources were pooled, and a comparative critical analysis was performed. At six months, both AD-MSCs and BM-MSCs showed significant VAS improvement (P = 0.015, P = 0.012); this was inconsistent at 1 year for BM-MSCs (P < 0.001, P = 0.539), and AD-MSCs outperformed BM-MSCs compared to controls in measures such as WOMAC (P < 0.001, P = 0.541), Lysholm scores (P = 0.006; P = 0.933), and KOOS (P = 0.002; P = 0.012). BM-MSC-related procedures caused significant adverse events (P = 0.003) compared to AD-MSCs (P = 0.673). Adipose tissue is superior to bone marrow because of its safety and consistent efficacy in improving pain and functional outcomes. Future trials are urgently warranted to validate our findings and reach a consensus on the ideal source of MSCs for managing knee OA.

Pathogenic Role of Adipose Tissue-Derived Mesenchymal Stem Cells in Obesity and Obesity-Related Inflammatory Diseases.

Adipose tissue-derived mesenchymal stem cells (ASCs) are adult stem cells, endowed with self-renewal, multipotent capacities, and immunomodulatory properties, as mesenchymal stem cells (MSCs) from other origins. However, in a pathological context, ASCs like MSCs can exhibit pro-inflammatory properties and attract inflammatory immune cells at their neighborhood. Subsequently, this creates an inflammatory microenvironment leading to ASCs' or MSCs' dysfunctions. One such example is given by obesity where adipogenesis is impaired and insulin resistance is initiated. These opposite properties have led to the classification of MSCs into two categories defined as pro-inflammatory ASC1 or anti-inflammatory ASC2, in which plasticity depends on the micro-environmental stimuli. The aim of this review is to (i) highlight the pathogenic role of ASCs during obesity and obesity-related inflammatory diseases, such as rheumatoid arthritis, multiple sclerosis, psoriasis, inflammatory bowel disease, and cancer; and (ii) describe some of the mechanisms leading to ASCs dysfunctions. Thus, the role of soluble factors, adhesion molecules; TLRs, Th17, and Th22 cells; γδ T cells; and immune checkpoint overexpression will be addressed.

Rab37 Promotes Endothelial Differentiation and Accelerates ADSC-Mediated Diabetic Wound Healing through Regulating Secretion of Hsp90α and TIMP1.

Accumulating evidence indicates that adipose tissue-derived mesenchymal stem cells (ADSCs) are an effective treatment for diabetic refractory wounds. However, the application of ADSCs to diabetic wounds is still limited, indicating that we still lack sufficient knowledge regarding regulators/mediators of ADSCs during wound healing. Rab37, a member of RabGTPase, may function as regulator of vesicle trafficking, which is a crucial event for the secretion of cytokines by ADSCs. Our previous study indicated that Rab37 promotes the adiopogenic differentiation of ADSCs. In this study, we explored the role of Rab37 in ADSC-mediated diabetic wound healing. An in vivo study in db/db diabetic mice showed that Rab37-expressing ADSCs shortened the wound closure time, improved re-epithelialization and collagen deposition, and promoted angiogenesis during wound healing. An in vitro study showed that Rab37 promoted the proliferation, migration and endothelial differentiation of ADSCs. LC-MS/MS analysis identified Hsp90α and TIMP1 as up-regulated cytokines in conditioned media of Rab37-ADSCs. The up-regulation of Rab37 enhanced the secretion of Hsp90α and TIMP1 during endothelial differentiation and under high-glucose exposure. Interestingly, Rab37 promoted the expression of TIMP1, but not Hsp90α, during endothelial differentiation. PLA showed that Rab37 can directly bind to Hsp90α orTIMP1 in ADSCs. Moreover, Hsp90α and TIMP1 knockdown compromised the promoting effects of Rab37 on the proliferation, migration and endothelial differentiation of ADSCs. In conclusion, Rab37 promotes the proliferation, migration and endothelial differentiation of ADSCs and accelerates ADSC-mediated diabetic wound healing through regulating the secretion of Hsp90α and TIMP1.

The Application of Adipose Tissue-Derived Mesenchymal Stem Cells (ADMSCs) and a Twin-Herb Formula to the Rodent Wound Healing Model: Use Alone or Together?

Our previous study reported that mesenchymal stem cells (MSCs) accelerated the wound healing process through anti-inflammatory, anti-apoptotic, and pro-angiogenetic effects in a rodent skin excision model. NF3 is a twin-herb formula, which presents similar effects in promoting wound healing. Research focusing on the interaction of MSCs and Chinese medicine is limited. In this study, we applied MSCs and the twin-herb formula to the wound healing model and investigated their interactions. Wound healing was improved in all treatment groups (MSCs only, NF3 only, and MSCs + NF3). The combined therapy further enhanced the effect: more GFP-labelled ADMSCs, collagen I and collagen III expression, Sox9 positive cells, and CD31 positive cells, along with less ED-1 positive cells, were detected; the expressions of proinflammatory cytokine IL-6 and TNF-α were downregulated; and the expression of anti-inflammatory cytokine IL-10 was upregulated. In vitro, NF3 promoted the cell viability and proliferation ability of MSCs, and a higher concentration of protein was detected in the NF3-treated supernatant. A proteomic analysis showed there were 15 and 22 proteins in the supernatants of normal ADMSCs and NF3-treated ADMSCs, respectively. After PCR validation, the expressions of 11 related genes were upregulated. The results of a western blot suggested that the TGFβ/Smad and Wnt pathways were related to the therapeutic effects of the combined treatment. Our study suggests for the first time that NF3 enhanced the therapeutic effect of MSCs in the wound healing model and the TGFβ/Smad and Wnt pathways were related to the procedure.

The effect of PRP and hyperosmolarity simultaneous use on expression profile alteration of miRNAs associated with cartilage differentiation in human adipose tissue-derived mesenchymal stem cells

BackgroundMesenchymal stem cells (MSC) are a type of multipotent stem cell whose differentiation into cartilage cells has been considered in recent years. Platelet-rich plasma (PRP) may impair cartilage differentiation due to its richness in growth factors and hyperosmolarity due to its proximity to the required cartilage environment. ObjectivesThe main purpose of this study was to treat human adipose tissue-derived MSCs concurrently with PRP and hyperosmolarity to investigate the expression profile of micro-RNA (miRNA) involved in the cartilage process differentiation. We examined the effect of PRP and the increase in osmolarity on the expression of miR-27, miR-101, miR-140, miR-145, miR-146, and miR-199. MethodsMesenchymal stem cells were extracted from human adipose tissue and differentiated into chondrocytes and the effect of baseline cultures (diff), PRP (prp), hyperosmolarity (os), base plus hyperosmolarity (diff + os), PRP plus hyperosmolarity (prp + os) next to the control group were studied in cartilage differentiation using specific stains such as Alcian blue, hematoxylin and eosin, and collagen type 2 and 10 immunohistochemistry. In addition, the expression of miR-27, miR-140, miR-199, miR-146, miR-101, and miR-145 was evaluated using real-time PCR. ConclusionHuman adipose tissue-derived MSCs with the ability to differentiate into adipocytes and osteocytes showed the properties of chondrocytes in all differentiation groups. Alkaline phosphatase (ALP) enzyme activity and calcium deposition were lower in the diff + os group than in other groups. Therefore, the diff + os group may be a more suitable environment for cartilage differentiation. Furthermore, 5% PRP concentration and hyperosmolarity showed a positive effect on miR-140, miR-199, miR-27, and, miR-146 and a negative effect on miR-101 and miR-145 on cartilage differentiation.

NAMPT encapsulated by extracellular vesicles from young adipose-derived mesenchymal stem cells treated tendinopathy in a “One-Stone-Two-Birds” manner

BackgroundTendinopathy is the leading sports-related injury and will cause severe weakness and tenderness. Effective therapy for tendinopathy remains limited, and extracellular vesicles (EVs) derived from adipose tissue-derived mesenchymal stem cells (ADMSCs) have demonstrated great potential in tendinopathy treatment; however, the influence of aging status on EV treatment has not been previously described.ResultsIn this study, it was found that ADMSCs derived from old mice (ADMSCold) demonstrated remarkable cellular senescence and impaired NAD+ metabolism compared with ADMSCs derived from young mice (ADMSCyoung). Lower NAMPT contents were detected in both ADMSCold and its secreted EVs (ADMSCold-EVs). Advanced animal experiments demonstrated that ADMSCyoung-EVs, but not ADMSCold-EVs, alleviated the pathological structural, functional and biomechanical properties in tendinopathy mice. Mechanistic analyses demonstrated that ADMSCyoung-EVs improved cell viability and relieved cellular senescence of tenocytes through the NAMPT/SIRT1/PPARγ/PGC-1α pathway. ADMSCyoung-EVs, but not ADMSCold-EVs, promoted phagocytosis and M2 polarization in macrophages through the NAMPT/SIRT1/Nf-κb p65/NLRP3 pathway. The macrophage/tenocyte crosstalk in tendinopathy was influenced by ADMSCyoung-EV treatment and thus it demonstrated "One-Stone-Two-Birds" effects in tendinopathy treatment.ConclusionsThis study demonstrates an effective novel therapy for tendinopathy and uncovers the influence of donor age on curative effects by clarifying the detailed biological mechanism.Graphical

Лечение остеоартроза коленных суставов на основе применения клеточных технологий

Osteoarthritis (OA) is the most common chronic disease of joint and a major cause of joint pain and disability. Traditional therapies for knee osteoarthritis show little clinical benefit, and the terminal treatment option is joint replacement. Currently, cell therapy is prospective for the treatment of knee osteoarthritis and has shown significant results in animal studies and human clinical trials. The aim: to investigate the treatment results in patients with knee osteoarthritis using adipose tissue stromal-vascular fraction and synovial mesenchymal stem cells.Methods. 60 patients with knee osteoarthritis were included in the study. The stage of knee osteoarthritis was assessed using the Kellgren-Lawrence classification, and patients with stage II-III of osteoarthritis were included in this study. After arthroscopic debridement, patients were divided into 2 groups. The first group was injected with the stromal-vascular fraction (SVF) of synovial membrane, and the second group was injected with adipose tissue mesenchymal stem cells (SVF). The minimum clinical and radiologic follow-up period was 12 months. Results. The assessment of the joints condition after 12 months using different scales showed that in both experimental groups there was a significant improvement of the functional status of the knee of the patients after treatment with MSC and SVF. Functional indicators of knee joints on the WOMAC scale improved by 14.7 and 13.4, on the KOOS scale by 23.6 and 26.4 in the SVF and MSC groups, respectively. Pain syndrome regressed by 1.8 and 2.1 on the VAS scale in the SVF and MSC groups, respectively.Conclusion. The results of both study groups showed that there is a significant difference between WOMAC, VAS and KOOS values before and after surgery (8, 12 weeks and 12 months) in the compared groups towards improvement

Integrated transcriptome-proteome analyses of human stem cells reveal source-dependent differences in their regenerative signature.

Mesenchymal stem cells (MSCs) are gaining increasing prominence as an effective regenerative cellular therapy. However, ensuring consistent and reliable effects across clinical populations has proved to be challenging. In part, this can be attributed to heterogeneity in the intrinsic molecular and regenerative signature of MSCs, which is dependent on their source of origin. The present work uses integrated omics-based profiling, at different functional levels, to compare the anti-inflammatory, immunomodulatory, and angiogenic properties between MSCs from neonatal (umbilical cord MSC [UC-MSC]) and adult (adipose tissue MSC [AD-MSC], and bone marrow MSC [BM-MSC]) sources. Using multi-parametric analyses, we identified that UC-MSCs promote a more robust host innate immune response; in contrast, adult-MSCs appear to facilitate remodeling of the extracellular matrix (ECM) with stronger activation of angiogenic cascades. These data should help facilitate the standardization of source-specific MSCs, such that their regenerative signatures can be confidently used to target specific disease processes.

Cytotoxicity and Genotoxicity of Graphene Oxide Scaffold: Human Adipose Tissue Derived Mesenchymal Stromal/stem Cells (AT-MSCs) Culture

Cytotoxicity and Genotoxicity of Graphene Oxide Scaffold: Human Adipose Tissue Derived Mesenchymal Stromal/stem Cells (AT-MSCs) Culture

Effects of resveratrol and its analogues on the cell cycle of equine mesenchymal stem/stromal cells.

Resveratrol (RSV; trans-3,5,4'-trihydroxystilbene) strongly activates sirtuin 1, and it and its analogue V29 enhance the proliferation of mesenchymal stem/stromal cells (MSCs).Although culture medium containing 5-azacytydine and RSV inhibits senescence of adipose tissue-derived MSCs isolated from horses with metabolic syndrome, few studies have reported the effects of RSV on equine bone marrow-derived MSCs (eBMMSCs) isolated from horses without metabolic syndrome. The aim of this study was to investigate the effects of RSV and V29 on the cell cycle of eBMMSCs. Following treatment with 5 µM RSV or 10 µM V29, the cell proliferation capacity of eBMMSCs derived from seven horses was evaluated by EdU (5-ethynyl-2'-deoxyuridine) and Ki-67 antibody assays. Brightfield images of cells and immunofluorescent images of EdU, Ki-67, and DAPI staining were recorded by fluorescence microscopy, and the number of cells positive for each was quantified and compared by Friedman's test at P<0.05. The growth fraction of eBMMSCs was significantly increased by RSV and V29 as measured by the EdU assay (control 28.1% ± 13.8%, V29 31.8% ± 14.6%, RSV 32.0% ± 10.8%; mean ± SD; P<0.05) but not as measured by the Ki-67 antibody assay (control 27.0% ± 11.2%, V29 27.4% ± 10.8%, RSV 27.7% ± 6.8%). RSV and V29 promoted progression of the cell cycle of eBMMSCs into the S phase and may be useful for eBMMSC expansion.

Amelioration of DSS-induced colitis in mice by TNF-α-stimulated mesenchymal stem cells derived from feline adipose tissue via COX-2/PGE2 activation.

Mesenchymal stem cells (MSCs) have been investigated as therapeutic agents for inflammatory bowel disease (IBD). Stimulation of MSCs with pro-inflammatory cytokines is an approach to enhance their immunomodulatory effects. However, further investigation is required to support their application in immune-mediated disorders and companion animals. This study aimed to assess the therapeutic effect of tumor necrosis factor (TNF)-α-stimulated feline adipose tissue-derived MSCs (fAT-MSCs) in a dextran sulfate sodium (DSS)-induced colitis mouse model. Colitis mice was made by drinking water with 3% DSS and fAT-MSCs were injected intraperitoneally. Colons were collected on day 10. The severity of the disease was evaluated and compared. Raw 264.7 cells were cultured with the conditioned medium to determine the mechanism, using quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay. TNF-α-stimulated fAT-MSCs more improved severity of DSS-induced colitis in disease activity, colon length, histologic score, and inflammatory cytokine. In sectionized colon tissues, the group comprising TNF-α-stimulated fAT-MSCs had higher proportion of CD11b+CD206+ macrophages than in the other groups. In vitro, TNF-α-stimulation increased cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) secretion from fAT-MSCs. The conditioned medium from TNF-α-stimulated fAT-MSCs enhanced the expression of interleukin-10 and arginase-1 in LPS-activated Raw 264.7 cells. These results represent that TNF-α-stimulated fat-mscs ameliorate the inflamed colon more effectively. Furthermore, we demonstrated that the effectiveness was interlinked with the COX-2/PGE2 pathway.

In vitro effects of the myokine irisin on human adipose tissue-derived mesenchymal stem cells during proliferation and osteogenic differentiation

Purpose: Irisin is a hormone-like molecule secreted from skeletal muscle in response to exercise both in mice and in humans and identified as an important effector in the crosstalk between muscle and bone. Although a number of studies report that irisin increased osteoblast differentiation in vitro and cortical bone mass in vivo, the models used are exclusively rodent ones. Due to the lack of reports on human cell models, the aim of our work was to investigate the in vitro effects of irisin on the proliferation and the osteogenic differentiation processes in human adipose tissue-derived mesenchymal stem cells (hAMSCs). Methods: hAMSCs were obtained by enzymatic digestion and mechanical dispersion, and cultured in growth medium. Cells were exposed to 10 and 100 ng/ml irisin for the entire experimental period and refreshed every two days. The proliferation was performed in growth medium containing 2.5% fetal bovine serum, and measured by cell counting at 24-48-72 hours. Alkaline phosphatase (ALP) activity and Ca2+ depositions were quantified by fluorometric assay during up to 35 days of osteogenic induction. Results: Cell proliferation assay showed that 100 ng/ml irisin significantly increased the proliferation process (p&lt;0.01) vs control, with a decrease of cell doubling time from 88 to 63 hours. Osteodifferentiation with 10 and 100 ng/ml irisin showed significant increases in ALP activity vs control (p&lt;0.01) after 14 days. Moreover, both tested concentrations of irisin were able to accelerate the deposition of mineralized matrix, resulting in significant increments in the production of Ca2+ nodules vs control after 35 days (p&lt;0.01). Conclusions: This work showed the in vitro effects of irisin on a human cell model of AMSCs. The preliminary results show this myokine to be an important effector on cell proliferation and during osteo-differentiation of hAMSCs, supporting the hypothesis that irisin could represent a potent new anabolic treatment to bring about gain of bone mass.

N,N'-Diphenyl-1,4-phenylenediamine Antioxidant's Potential Role in Enhancing the Pancreatic Antioxidant, Immunomodulatory, and Anti-Apoptotic Therapeutic Capabilities of Adipose-Derived Stem Cells in Type I Diabetic Rats.

Mesenchymal stem cells (MSCs) are considered to be a promising therapeutic protocol for diabetes mellitus (DM) management. The latter is attributed to their differentiation potentiality to pancreatic β-cells, angiogenesis, and immune-modulatory capabilities by releasing various paracrine factors. Interestingly, antioxidant co-administration increased the MSCs' hypoglycemic and regenerative activities. Thus, this study aims to evaluate the therapeutic implication of type 1 DM after the co-administration of adipose tissue-derived-MSCs (AD-MSCs) and N,N'-d iphenyl-1,4-phenylenediamine (DPPD), compared to the single injection of either of them alone. In our four week long experiment, six rat groups were used as control, DPPD (250 mg/kg, i.p.), STZ-diabetic (D), D+DPPD, D+AD-MSCs (1 × 106 cell/rat, i.p.), and D+AD-MSCs+DPPD groups. Within this context, a single injection of AD-MSCs or DPPD into diabetic rats showed significant pancreatic anti-inflammatory, immunomodulation, antioxidant, and anti-apoptotic capacities, superior to AD-MSCs injection. However, AD-MSCs and DPPD co-administration into diabetic rats manifested the highest hypoglycemic and pancreatic regenerative activities in managing diabetes compared to the single shot of AD-MSCs or DPPD. These results highlight the synergetic role of DPPD as an antioxidant in enhancing AD-MSCs' therapeutic applications.