OBJECTIVE: To investigate the effectiveness and preliminary mechanisms of icariin (ICA) in enhancing the reparative effects of adipose-derived stem cells (ADSCs) on skin radiation damagies in rats. METHODS: Twelve SPF-grade Sprague Dawley rats [body weight (220±10) g] were subjected to a single dose of 10 Gy X-ray irradiation on a 1.5 cm×1.5 cm area of their dorsal skin, with a dose rate of 200 cGy/min to make skin radiation damage model. After successful modelling, the rats were randomly divided into 4 groups ( n=3), and on day 2, the corresponding cells were injected subcutaneously into the irradiated wounds: group A received 0.1 mL of rat ADSCs (1×10 ⁷cells/mL), group B received 0.1 mL of rat ADSCs (1×10 ⁷cells/mL)+1 μmol/L ICA (0.1 mL), group C received 0.1 mL of rat ADSCs (1×10 ⁷cells/mL) pretreated with a hypoxia-inducible factor 2α (HIF-2α) inhibitor+1 μmol/L ICA (0.1 mL), and group D received 0.1 mL of rat ADSCs (1×10 ⁷cells/mL) pretreated with a Notch1 inhibitor+1 μmol/L ICA (0.1 mL). All treatments were administered as single doses. The skin injury in the irradiated areas of the rats was observed continuously from day 1 to day 7 after modelling. On day 28, the rats were sacrificed, and skin tissues from the irradiated areas were harvested for histological examination (HE staining and Masson staining) to assess the repair status and for quantitative collagen content detection. Immunohistochemical staining was performed to detect CD31 expression, while Western blot and real-time fluorescence quantitative PCR (qRT-PCR) were used to measure the protein and mRNA relative expression levels of vascular endothelial growth factor (VEGF), platelet-derived growth factor BB (PDGF-BB), fibroblast growth factor 2 (FGF-2), interleukin 10 (IL-10), transforming growth factor β (TGF-β), HIF-2α, and Notch1, 2, and 3. RESULTS: All groups exhibited skin ulcers and redness after irradiation. On day 3, exudation of tissue fluid was observed in all groups. On day 7, group B showed significantly smaller skin injury areas compared to the other 3 groups. On day 28, histological examination revealed that the epidermis was thickened and the dermal fibers were slightly disordered with occasional inflammatory cell aggregation in group A. In group B, the epidermis appeared more normal, the dermal fibers were more orderly, and there was an increase in new blood vessels without significant inflammatory cell aggregation. In contrast, groups C and D showed significantly increased epidermal thickness, disordered and disrupted dermal fibers. Group B had higher collagen fiber content than the other 3 groups, and group D had lower content than group A, with significant differences ( P<0.05). Immunohistochemical staining showed that group B had significantly higher CD31 expression than the other 3 groups, while groups C and D had lower expression than group A, with significant differences ( P<0.05). Western blot and qRT-PCR results indicated that group B had significantly higher relative expression levels of VEGF, PDGF-BB, FGF-2, IL-10, TGF-β, HIF-2α, and Notch1, 2, and 3 proteins and mRNAs compared to the other 3 groups ( P<0.05). CONCLUSION ICA may enhance the reparative effects of ADSCs on rat skin radiation damage by promoting angiogenesis and reducing inflammatory responses through the HIF-2α-VEGF-Notch signaling pathway.
Animals ; Rats, Sprague-Dawley ; Skin/pathology* ; Rats ; Vascular Endothelial Growth Factor A/genetics* ; Basic Helix-Loop-Helix Transcription Factors/genetics* ; Signal Transduction ; Flavonoids/pharmacology* ; Adipose Tissue/cytology* ; Stem Cells/cytology* ; Receptors, Notch/metabolism* ; Radiation Injuries, Experimental/metabolism* ; Wound Healing/drug effects* ; Male

OBJECTIVE: To investigate the effect of human adipose-derived mesenchymal stem cells (hASCs) exosomes on osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) extracted from osteoporotic mice, and to evaluate the effect of hASCs exosomes on preventing bone loss induced by estrogen deficiency. METHODS: hASCs exosomes were extracted by ultracentrifugation. The osteoporotic mice were established by bilateral ovariectomy (OVX). BMSCs were isolated from osteo-porotic mice and cultured for further analysis. In the experimental group, these BMSCs were exposed to an osteogenic induction medium supplemented with hASCs exosomes to evaluate their potential effects on osteogenesis. In contrast, the control group was treated with the same osteogenic induction medium, but without the addition of hASCs exosomes, to serve as a baseline comparison for the study. To comprehensively assess the osteogenic differentiation of BMSCs influenced by hASCs exosomes, alkaline phosphatase (ALP) staining, ALP activity quantitative analysis and quantitative reverse transcription polymerase chain reaction (qPCR) were performed. These evaluations provided critical insights into the role of hASCs exosomes in promoting osteoblast differentiation and bone formation in osteoporotic conditions. The fluorescence labeled hASCs exosomes were injected via the tail vein to observe the biodistribution of exosomes. Two weeks after OVX, the mice were divided into three groups: The experimental group consisted of estrogen-deficient mice receiving hASCs exosome injections; the negative control group consisted of estrogen-deficient mice receiving phosphate-buffered saline (PBS) injections; and the positive control group consisted of mice that underwent Sham surgery and received PBS injections.The injections were administered once every 3 days, for a total of 8 injections. Afterward, the femurs were collected from the mice, and micro-computed tomography (micro-CT) was performed to measure bone mineral density and conduct bone morphometric analysis. RESULTS: hASCs exosomes were successfully extracted using ultracentrifugation. After the induction by hASCs exosomes, ALP staining and ALP activity in the BMSCs extracted from osteoporotic mice were significantly enhanced, the expression of osteogenesis related genes in BMSCs were significantly up-regulated. More trabecular bone and higher bone mineral density were observed in estrogen-deficient mice injected with hASCs exosomes compared with estrogen-deficient mice injected with PBS, and there was no significant decrease in bone mineral density compared with the Sham operation group. CONCLUSION hASCs exosomes promoted the osteogenic differentiation of BMSCs extracted from osteoporotic mice. hASCs exosomes prevented bone loss induced by estrogen deficiency.
Animals ; Mesenchymal Stem Cells/cytology* ; Exosomes ; Estrogens/deficiency* ; Humans ; Osteogenesis ; Cell Differentiation ; Female ; Mice ; Osteoporosis/prevention & control* ; Ovariectomy ; Adipose Tissue/cytology* ; Cells, Cultured

Adipose tissue-derived mesenchymal stem cells (ADSCs) are crucially involved in various biological processes because of their self-renewal, multi-differentiation, and immunomodulatory activities. Some ADSC's characteristics have been associated with the basic theory of Chinese medicine (CM), especially the Meridian theory. CM can improve the biological properties of ADSCs to facilitate their use in injury treatment, restore immune homeostasis, and inhibit inflammatory responses. Therefore, the combination of CM and ADSCs may be a new promising research direction in integrative medicine of China. This review summarizes the association between CM and ADSCs to assess the potential application value of their combination against various diseases.
Mesenchymal Stem Cells/cytology* ; Adipose Tissue/cytology* ; Humans ; Medicine, Chinese Traditional ; Integrative Medicine ; Animals

Plant polysaccharides are effective components that widely present in traditional Chinese medicine(TCM), exhibiting rich biological activities. However, as most plant polysaccharides cannot be directly absorbed and utilized by the human digestive system, it is now believed that their mode of action mainly involves interaction with intestinal microbiota, leading to the production of functional small molecules. The efficacy of Astragalus polysaccharide(APS) is extensive, including weight loss, improvement of fatty liver, reduction of blood lipids, and enhancement of insulin sensitivity, which may also be related to the regulation of intestinal microbiota. Adipose tissue senescence is an important characteristic of the physiological aging process in the body, often occurring prior to the aging of other important organs. Its main features include the accumulation of senescent cells and exacerbation of inflammation within the tissue. Therefore, to explore the potential protective effects of APS on aging, the improvement of adipose tissue aging phenotype in naturally aging mice was observed using APS, and combined with metagenomic metabolomics, corresponding microbial metabolic functional molecules were identified. Furthermore, functional tests in cell aging models were conducted. The results showed that APS significantly improved the adipocyte aging characteristics of naturally aging mice: specifically reducing aging-induced adipocyte hypertrophy; decreasing the protein expression of aging markers cyclin-dependent kinase inhibitor p21(P21) and multiple tumor suppressor 1(P16); lowering the tissue inflammation reaction. Metagenomic metabolomic analysis of serum from mice in each group revealed that APS significantly increased the content of indole-3-lactic acid(ILA) in naturally aging mice. Further in vitro studies showed that ILA could improve the aging of 3T3-L1 mouse embryonic fibroblasts induced by bleomycin, reduce the protein expression of the aging marker P21, alleviate inflammation, and enhance the ability of preadipocytes to mature. Therefore, APS had the efficacy of protecting naturally aging mice, and its action may be related to the increase in the intestinal microbiota metabolite ILA. This study suggested that TCM may serve as an important entry point for explaining the mechanism of action of TCM by regulating intestinal microbiota and their functional metabolites.
Animals ; Mice ; Aging/drug effects* ; Adipose Tissue/metabolism* ; Polysaccharides/pharmacology* ; Indoles/pharmacology* ; Male ; Astragalus Plant/chemistry* ; 3T3-L1 Cells ; Humans ; Adipocytes/cytology* ; Mice, Inbred C57BL ; Cellular Senescence/drug effects* ; Drugs, Chinese Herbal/administration & dosage* ; Gastrointestinal Microbiome/drug effects*

Objective To investigate the therapeutic effect of SPK1 gene transfected adipose derived mesenchymal stem cells(ADMSC)on experimental autoimmune encephalomyelitis mice and the effect on T helper cell 17(Th17)/regulatory T(Treg) cells balance. Methods EAE was induced by myelin oligodendrocyte glycoprotein 35-55 in mice.Totally 44 mice were randomly divided into four groups:normal control group(NC group),model group(EAE group),ADMSC group,and ADMSC-SPK1 group.Forty days after injection,the pathological changes of brain and spinal cord,Th17/Treg-related inflammatory markers in brain tissue,expressions of interleukin-17A(IL-17A)and forkhead box protein p3(Foxp3)in brain and spinal cord tissue,and flow cytometric results of spleen immune cells were detected. Results Forty days after the injection,serious inflammatory cell infiltration and demyelination occurred in the brain and spinal cord of EAE group,whereas demyelination and axonal injury were improved in ADMSC group and ADMSC-SPK1 group.Compared with EAE group,the ADMSC group and ADMSC-SPK1 group had significantly improved levels of IL-17A(
Adipose Tissue/cytology* ; Animals ; Cytokines ; Encephalomyelitis, Autoimmune, Experimental/therapy* ; Interleukin-17 ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stem Cells/cytology* ; Mice ; Mice, Inbred C57BL ; Phosphotransferases (Alcohol Group Acceptor)/genetics* ; T-Lymphocytes, Regulatory/cytology* ; Th17 Cells/cytology* ; Transfection

Objective To induce adipose-derived stem cells (ADSCs) to differentiate into intermediate mesoderm (IM)-like cells ,with IM-like cells for recellularizing kidney scaffolds,and then to obtain a tissue-engineering kidney with renal structures and functions through co-culture.Methods After inguinal fat pads of Wistar rats were surgically harvested,the primary ADSCs were isolated,induced,and cultured for stem cell identification. ADSCs were inducted to differentiate into IM-like cells by adding glycogen synthase kinase-3 inhibitor (CHIR99021) and fibroblast growth factor 9 (FGF9) at different stages. Seven days later,the IM-like cells were identified. The induced IM-like cells and well-prepared kidney decellularized scaffolds were co-cultured for 10 days to obtain recellularized tissue-engineered kidneys and their differentiation was identified.Results The ADSCs harvested had osteogenic and adipogenic abilities and could express the stem cell surface markers. After 7 days of induction,the positive expressions of odd-skipped related 1 and paired-box 2 were observed in IM-like cells by immunofluorescence technique. After 10 days of co-culture with kidney decellularized scaffolds,the positive expressions of Wilms'tumor 1,GATA-binding protein-3,and E-cadherin were observed by immunofluorescence technique.Conclusion ADSCs can be induced into IM-like cells,and renal cell differentiation can be observed through combining the induced IM-like cells with kidney decellularized scaffolds.
Adipose Tissue ; Animals ; Cell Differentiation ; Cells, Cultured ; Kidney ; growth & development ; Mesoderm ; cytology ; Rats ; Rats, Wistar ; Regeneration ; Stem Cells ; cytology ; Tissue Engineering ; Tissue Scaffolds

OBJECTIVE: How to increase the long-term retention rate of autologous fat grafting has been widely discussed. This study aimed to evaluate long-term fat graft retention rates for the most widely used fat processing methods in the area of facial esthetic surgery, including centrifugation, filtration, and sedimentation, using three-dimensional (3D) imaging. DATA SOURCES: PubMed, Embase, Wiley/Cochrane Library, and Web of Science databases were comprehensively searched from inception to July 2018 according to the guidelines of the American Society of Plastic Surgeons Fat Graft Task Force Assessment Methodology. STUDY SELECTION: Articles were screened using predetermined inclusion and exclusion criteria. Data collected included patient characteristics, follow-up devices, fat grafting techniques, and clinical outcomes. Patient cohorts were pooled, and fat graft retention rates were calculated. Complications were summarized according to different clinical characteristics. RESULTS: Of 77 articles, 10 clinical studies met the inclusion criteria and reported quantified measurement outcomes with 3D imaging which provide precise volumetric data with approximately 2% standard deviation compared to real volumes. Data of 515 patients were included. Fat grafting retention varied from 21% to 82%. We found filtration and centrifugation techniques could result in better retention outcomes. However, retention varied within each processing technique, with no significant difference among the 3 techniques. Twenty-two complications were reported among 515 patients, including donor-site hematoma (1 case), mild post-operative erythema (2 cases), mild volumetric asymmetries (2 cases), chronic edema (2 cases), overcorrection (2 cases), skin irregularity (6 cases), and headache or dysesthesia (7 cases). CONCLUSIONS Filtration and centrifugation techniques may result in better fat grafting retention outcomes than gravity sedimentation; however, more accurate statistical evidence is needed. Controversies continue to exist with respect to the performance of the different fat-processing techniques in fat graft retention.
Adipocytes ; cytology ; Adipose Tissue ; cytology ; Centrifugation ; methods ; Filtration ; methods ; Humans ; Imaging, Three-Dimensional ; methods

BACKGROUND: Several patients experience persistent otorrhea after a flawless surgical procedure because of insufficient epithelial healing. Several efforts, such as autologous tissue allograft and xenograft, have been made to halt otorrhea. However, a stable technology to induce temporal epithelial repair is yet to be established. Therefore, this study aims to investigate whether implantation of seeding adipose-derived mesenchymal stem cell (ADMSC) aggregates on extracellular matrix (ECM; herein, ADMSC aggregate-ECM) into damaged skin wound promotes skin regeneration. METHODS: ADMSC aggregate-ECM was prepared using a previously described procedure that isolated ADMSCs from rabbits and applied to the auricle and auditory meatus wound beds of New Zealand white rabbits. Wound healing was assessed by general observation and hematoxylin and eosin (H&E) staining. Secretion of growth factor of the tissue was evaluated by western blotting. Two other groups, namely, ECM and control, were used. Comparisons of three groups were conducted by one-way analysis of variance analysis. RESULTS: ADMSCs adhered tightly to the ECM and quickly formed cell sheets. At 2 weeks, general observation and H&E staining indicated that the wound healing rates in the ADMSC aggregate-ECM (69.02 ± 6.36%) and ECM (59.32 ± 4.10%) groups were higher than that in the control group (43.74 ± 12.15%; P = 0.005, P < 0.001, respectively) in ear auricle excisional wounds. At 7 weeks, The scar elevation index was evidently reduced in the ADMSC aggregate-ECM (2.08 ± 0.87) and ECM (2.31 ± 0.33) groups compared with the control group (4.06 ± 0.45; P < 0.001, P < 0.001, respectively). In addition, the scar elevation index of the ADMSC aggregate-ECM group reached the lowest rate 4 weeks in advance. In auditory meatus excisional wounds, the ADMSC aggregate-ECM group had the largest range of normal skin-like structure at 4 weeks. The ADMSC aggregate-ECM and ECM groups secreted increased amounts of growth factors that contributed to skin regeneration at weeks 1 and 2, respectively. CONCLUSIONS ADMSC aggregate-ECM and ECM are effective repair materials for wound healing, especially ADMSC aggregate-ECM. This approach will provide a meaningful experimental basis for mastoid epithelium repair in subsequent clinical trials.
Adipose Tissue ; cytology ; Animals ; Cell Differentiation ; physiology ; Cell Proliferation ; physiology ; Cells, Cultured ; Ear Auricle ; cytology ; Extracellular Matrix ; chemistry ; Flow Cytometry ; Mesenchymal Stem Cell Transplantation ; methods ; Mesenchymal Stem Cells ; cytology ; Microscopy, Electron, Scanning ; Osteogenesis ; physiology ; Rabbits ; Real-Time Polymerase Chain Reaction

BackgroundCorneal stromal cells (CSCs) are components of the corneal endothelial microenvironment that can be induced to form a functional tissue-engineered corneal endothelium. Adipose-derived mesenchymal stem cells (ADSCs) have been reported as an important component of regenerative medicine and cell therapy for corneal stromal damage. We have demonstrated that the treatment with ADSCs leads to phenotypic changes in CSCs in vitro. However, the underlying mechanisms of such ADSC-induced changes in CSCs remain unclear.

MethodsADSCs and CSCs were isolated from New Zealand white rabbits and cultured in vitro. An Exosome Isolation Kit, Western blotting, and nanoparticle tracking analysis (NTA) were used to isolate and confirm the exosomes from ADSC culture medium. Meanwhile, the optimal exosome concentration and treatment time were selected. Cell Counting Kit-8 and annexin V-fluorescein isothiocyanate/propidium iodide assays were used to assess the effect of ADSC- derived exosomes on the proliferation and apoptosis of CSCs. To evaluate the effects of ADSC- derived exosomes on CSC invasion activity, Western blotting was used to detect the expression of matrix metalloproteinases (MMPs) and collagens.

Results:ADSCs and CSCs were successfully isolated from New Zealand rabbits. The optimal concentration and treatment time of exosomes for the following study were 100 μg/ml and 96 h, respectively. NTA revealed that the ADSC-derived exosomes appeared as nanoparticles (40-200 nm), and Western blotting confirmed positive expression of CD9, CD81, flotillin-1, and HSP70 versus ADSC cytoplasmic proteins (all P < 0.01). ADSC-derived exosomes (50 μg/ml and 100 μg/ml) significantly promoted proliferation and inhibited apoptosis (mainly early apoptosis) of CSCs versus non-exosome-treated CSCs (all P < 0.05). Interestingly, MMPs were downregulated and extracellular matrix (ECM)-related proteins including collagens and fibronectin were upregulated in the exosome-treated CSCs versus non-exosome-treated CSCs (MMP1: t = 80.103, P < 0.01; MMP2: t = 114.778, P < 0.01; MMP3: t = 56.208, P < 0.01; and MMP9: t = 60.617, P < 0.01; collagen I: t = -82.742, P < 0.01; collagen II: t = -72.818, P < 0.01; collagen III: t = -104.452, P < 0.01; collagen IV: t = -133.426, P < 0.01, and collagen V: t = -294.019, P < 0.01; and fibronectin: t = -92.491, P < 0.01, respectively).

Conclusion:The findings indicate that ADSCs might play an important role in CSC viability regulation and ECM remodeling, partially through the secretion of exosomes.

Adipose Tissue ; cytology ; Animals ; Cell Proliferation ; physiology ; Cell Survival ; physiology ; Cells, Cultured ; Exosomes ; metabolism ; Extracellular Matrix ; metabolism ; Fibroblasts ; cytology ; metabolism ; Matrix Metalloproteinases ; metabolism ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Rabbits

Objective To investigate the effect of adipose mesenchymal stem cells(AMSCs) on the peripheral blood lymphocyte(PBL) in psoriasis vulgaris(PV) patients and the expression and secretion profiles of related inflammatory cytokines in the PBL.Methods AMSCs from three PV patients were co-cultured with PBL. Peripheral blood regulatory cells(Treg) and T helper cell 17(Th17)ratio was measured by flow cytometry. The anti- and pro-inflammatory cytokines expressed and secreted by PBL were detected by quantitative real-time polymerase chain reaction(qRT-PCR) and enzyme-linked immunosorbent assay(ELISA).Results The Treg/total lymphocyte ratio was significantly higher in the healthy people AMSCs+PBL co-culture group[(3.2±0.5)%;P=0.001],but AMSCs in patients had a tendency to promote the proliferation of Treg cells [(1.3±0.2)%],with no significant difference(P=0.485) when compared with the PBL culture alone group[(1.0±0.1)%]. qRT-PCR showed that the ability of PBL in expressing Treg transcription factor forkhead box p3 and transforming growth factor(TGF)-Β mRNA was significantly lower in psoriasis AMSCs+PBL co-culture group than in the healthy people AMSCs+PBL co-culture group(P=0.00,P=0.03),AMSCs had a tendency to promote the expression of interlukin(IL)-10 in peripheral blood lymphocytes,but there was no significant difference(P=0.09).ELISA showed the PBL in healthy people AMSCs+PBL co-culture group secreted the anti-inflammatory cytokine IL-10[(156.9±41.8) ng/Μl] and TGF-Β[(2774.1 ± 526.4) ng/Μl];in contrast,the abilities of PBL in PV patient AMSCs+PBL co-culture group in secreting the anti-inflammatory cytokines has a downward trend:IL-10[(90.4±28.8) ng/Μl] and TGF-Β[(1597.9±55.7) ng/Μl],although the differences were not statistically significant. After the co-culture,the proportion of Th17 cells in the psoriasis AMSCs+PBL co-culture group[(0.8±0.3)%] showed a decreasing trend when compared with the PBL culture alone group[(1.1±0.1)%],although the results were not statistically significant. Also,the proportion of Th17 cells showed no significant difference between PV patient AMSCs+PBL co-culture group and healthy people AMSCs+PBL co-culture group. Finally,both the psoriasis AMSCs+PBL co-culture group and the healthy people AMSCs+PBL co-culture group showed no obvious inhibitory effect on the expression and secretion of Th17 transcription factor retinoid-related orphan nuclear receptor Γt and pro-inflammatory cytokines IL-17 and IL-23 in PBL,and there was no significant difference between these two groups.Conclusions AMSCs in PV patients have decreased ability in regulating the anti-inflammatory function of peripheral blood Treg lymphocytes. However,they have no effect on the proinflammatory effect of peripheral blood Th17 lymphocytes.
Adipose Tissue ; cytology ; Cytokines ; immunology ; Forkhead Transcription Factors ; immunology ; Humans ; Inflammation ; immunology ; Mesenchymal Stem Cells ; cytology ; Psoriasis ; immunology ; T-Lymphocytes, Regulatory ; immunology ; Th17 Cells ; immunology