The Quality and Quantity of Nanoparticles Extracted from Human Adipose Tissue Derived-Mesenchymal Stem Cells

CRISPR/Cas9-Mediated Knockin Application in Cell Therapy: A Non-viral Procedure for Bystander Treatment of Glioma in Mice.

Adipose Tissue–derived Mesenchymal Stem Cells Expressing Prodrug-converting Enzyme Inhibit Human Prostate Tumor Growth

Objective To investigate the effect and related mechanism of apolipoprotein A5 (ApoA5) on adipogenic differentiation of human adipose-derived mesenchymal stem cells (AMSC). Methods Subcutaneous adipose tissue was obtained from 40 patients undergoing abdominal surgery at our hospital from February to July 2015. After induction of human AMSC by collagenase digestion, the adipose tissue was induced to differentiate into mature adipocytes and treated with ApoA5 at 600 and 1 200 ng/ml, respectively (ApoA5 intervention groups). Cells treated without ApoA5 protein were used as control group. The cells were harvested on the 7th and 14th day of differentiation, and the following assays were performed: (1) the effect of ApoA5 on TG content was measured by a TG assay kit; (2) RT-qPCR assay was used to detect the effect of ApoA5 on aP2 and FAS mRNA expression; (3) the effect of ApoA5 on the expression of CIDEC mRNA and protein was detected by RT-qPCR and Western blot; (4) the effect of ApoA5 on the expression of C/EBPβ mRNA and protein was detected by RT-qPCR and Western blot; (5) using lentiviral transfection technique, we overexpressed the gene of CIDEC in AMSC and cells were divided into lentiviral negative control group, lentiviral over-expressed CIDEC group and lentiviral over-expressed CIDEC+ApoA5 intervention group (the ApoA5 intervention concentration was 1 200 ng/ml). Thereby, we examined the effect of ApoA5 on the above indicators in adipogenic differentiation of AMSC in the case of CIDEC overexpression. Results (1) Effect of ApoA5 on TG content in AMSC: on the 7th and 14th day after the intervention, the TG levels were lower in ApoA5 600 and 1 200 ng/ml group AMSC than those in the control group (all P 0.05). (6) The effect of ApoA5 on the expression of aP2 and FAS mRNA in AMSC after CIDEC overexpression: on the 7th day after intervention, the expression levels of aP2 and FAS mRNA in AMSC were higher in the lentivirus over-expressed CIDEC group than in the lentivirus negative control group (both P 0.05). (7) The effect of ApoA5 on the expression of C/EBPβ mRNA and protein in AMSC after CIDEC overexpression: on the 7th day after intervention, the mRNA and relative protein expressions of C/EBPβ in AMSC were higher in lentivirus-overexpressed CIDEC group than in lentivirus negative control group (both P 0.05). Conclusions ApoA5 can inhibit the adipogenic differentiation of AMSC,and this effect may be mediated by down-regulating the expression of CIDEC. Furthermore, our results indicate that CIDEC could be considered as a key factor in adipogenic differentiation. Key words: Apolipoproteins A; Adipogenesis; Adipose-derived mesenchymal stem cells

The aim of this study was to explore the effects of human adipose-derived mesenchymal stem cells (ASCs) on the growth of gastric cancer cells in vivo and vitro and its mechanism. ASCs were isolated from abandoned adipose tissues, and the surface markers were identified by flow cytometry. In vitro experiments, HGC-27 cells cultured in ASCs-conditioned medium (CM) were assigned as the experimental group, while HGC-27 cells cultured in normal medium were as the control group. MTT and colony formation assays were performed to detect cell viability and colony formatting ability, respectively. Annexin-V/PI assay, Western blot, and caspase-3 enzyme activity assay were performed to detect cells apoptosis. The isolated ASCs could be differentiated into adipocytes and osteoblasts in vitro. Flow cytometry showed that CD73 and CD105 were positively expressed in HGC-27 cells. Compared with the mice injected HGC-27 cells only, the tumor formation in mice injected both ASCs and HGC-27 cells was significantly smaller (P < 0.05). The colony formation ability in experimental group was 40.09% smaller than control group (P < 0.05) and the cell apoptosis rate in experimental group was higher than the control group (P < 0.05). Furthermore, the expressions of cleaved PARP, cleaved caspase-3 proteins, and caspase-3 enzyme viability in experimental group were significantly higher than those of control group (P < 0.05). In conclusion, ASCs can effectively inhibit the growth of HGC-27 cells by inducing apoptosis.

Preconditioning of human adipose tissue-derived mesenchymal stem cells with HEK293-coditioned media can influence on the expression of BMP2, BMP6 and BMP11: Potential application in the treatment of renal lesions

Effects of human adipose-derived mesenchymal stem cells and platelet-rich plasma on healing of wounds with full-thickness skin defects in mice

Adipose-derived mesenchymal stromal cells primed by macrophages decrease inflammation and multiple organ lesions improving mice survival after cecal ligation and puncure-induced sepsis

Objective To evaluate the feasibility of differentiation of human adipose mesenchymal stem cells (hADSCs) into endothelial-like cells and the safety of induced cells intraocular application. Methods HADSCs were induced to endothelial-like cells in vitro.The experimental group was added with streptomycin, vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), and the control group was added with the same amount of phosphate buffered saline (PBS), the expression of von Willebrand factor (vWF), a marker of endothelial cells, was observed in the two groups.Six C57BL/6J mice were randomly divided into experimental group and control group by using the random number method, 3 mice for each group, the induced cells were injected intravitreally into C57BL/6J mice as experimental group, and the same amount of PBS buffer was injected intravitreally into C57BL/6J mice as control group.The changes of retinal cells were observed by electron microscopy.The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission. Results After induction of hADSCs in vitro, the endothelial cell marker was expressed.VWF immunofluorescence of the experimental group showed strong green fluorescence, and the color rendering rate was 100%.The control group showed no coloration of vWF immunofluorescence, and the color positive rate was 0.After 1 month of intravitreal injection, the retinal ganglion cells and rod cells were not degraded and necrotic. Conclusions HADSCs can differentiate into endothelial-like cells in vitro, and there is no retinal toxicity in a short-term. Key words: Adipose derived mesenchymal stem cells; Endothelium-like cells; Induced differentiation; Toxicity

The current work investigated the immunological features of insulin-producing cells (IPCs) generated from rat adipose-derived mesenchymal stem cells (ADSCs) both in vitro and in vivo. The research was carried out at Ahvaz Jundishapur University of Medical Sciences in 2023. ADSCs were derived from rat adipose tissues and differentiated into IPCs. The control group included undifferentiated ADSCs. The amount of secreted insulin was measured using ELISA. The expression of major histocompatibility complex-I (MHC-I) and MHC-II, cluster of differentiation 40 (CD40), and CD80 by IPCs in vitro was assessed using Western Blot analysis. The in vivo study was performed on 10 male diabetic rats. The experimental group received 107 IPCs in the peritoneal cavity. The control group received 107 undifferentiated ADSCs. After 4 hours, the expression of CD3a and CD45 by immune cells collected from the peritoneal cavity was measured using flow cytometry. All parameters were statistically analyzed using a t test. The differentiated cells secreted much higher amounts of insulin than the control group (P=0.04). IPCs exhibited higher expression of MHC-I and MHC-II, CD40, and CD80 (P=0.02, P=0.008, P=0.07, and P=0.02, respectively). The experimental group showed higher levels of CD3a and CD45 expression than the control group (P=0.07, P=0.04, respectively). Functional IPCs generated by ADSCs differentiation exhibited immunogenic activity both in vitro and in vivo. Immune-modulating strategies are required for the effective transplantation of the differentiated IPCs generated in our study.

To investigate the effect of daphnetin (DAP) combined with insulin-like growth factor 1 (IGF-1) gene transfection on chondrogenic differentiation of adipose-derived mesenchymal stem cells (ADSCs) in rats. Rat ADSCs were isolated and amplified by enzymatic digestion. The third generation ADSCs were treated with IGF-1 gene transfection as experimental group and normal ADSCs as control group. The cells of the two groups were treated with different concentrations of DAP (0, 30, 60, 90 μg/mL), respectively. Cell proliferation was detected by cell counting kit 8 (CCK-8) after cultured for 72 hours. After 14 days, real-time fluorescence quantitative PCR and Western blot were used to detect the mRNA and protein expressions of chondrocyte markers (collagen type Ⅱ and Aggrecan) in each group; and toluidine blue staining and collagen type Ⅱ immunohistochemical staining were performed. CCK-8 assay showed that with the increase of DAP concentration, the cell absorbance ( A) value of the control group and the experimental group increased gradually ( P<0.05). At the same DAP concentration, the cell A value of the experimental group was significantly higher than that of the control group ( P<0.05). Real-time fluorescence quantitative PCR and Western blot showed that with the increase of DAP concentration, the relative mRNA and protein expressions of collagen type Ⅱ and Aggrecan in the control group did not change significantly, and there was no significant difference among the different concentration groups ( P>0.05). But the mRNA and protein expressions of collagen type Ⅱ and Aggrecan in the experimental group increased gradually, and the 60 and 90 μg/mL DAP concentration groups were significantly higher than 0 μg/mL DAP concentration group ( P<0.05). At the same DAP concentration, the relative mRNA and protein expressions of collagen type Ⅱ and Aggrecan were significantly higher in the experimental group than in the control group ( P<0.05). Toluidine blue staining showed that with the increase of DAP concentration, there was no significant difference in cell staining between the control group and the experimental group. At the same DAP concentration, the cells in the experimental group were slightly darker than those in the control group. Immunohistochemical staining of collagen type Ⅱ showed that with the increase of DAP concentration, there was no significant difference in the cytoplasmic brown-yellow coloring of the cells in the control group. The cytoplasmic brown-yellow coloring of the cells in the experimental group gradually deepened, with 60 and 90 μg/mL DAP concentration groups significantly deeper than 0 μg/mL DAP concentration group. At the same DAP concentration, the color of the cells in the experimental group was significantly deeper than that in the control group. DAP can promote the proliferation of ADSCs in rats. The differentiation of ADSCs into chondrocytes induced by DAP alone was slightly, but DAP combined with IGF-1 gene transfection has obvious synergistic effect to promote chondrogenic differentiation of ADSCs.

To investigate the regulation of adipose-derived mesenchymal stem cells (ADSC) on helper T cells and regulatory T cells in allergic rhinitis(AR) mouse model and the underlying mechanisms. Using random number table, 60 Balb/c mice were divided into 6 groups (represented by: sensitized/challenged/treated ), they were the experimental group 1(OVA/OVA/high dose ADSC), the experimental group 2(OVA/OVA/low dose ADSC), the experimental group 3(OVA/OVA/PBS), the experimental group 4(OVA/OVA/0), the control group 1(PBS/PBS/0) and the control group 2(0/0/0). The mouse ADSC were isolated and cultured through conventional method, and AR mouse model was built with OVA and aluminum. The mice were injected with high (3×10(6)), low (1×10(6)) ADSC respectively labeled by CM-Dil for 3 consecutive days via tail-vein injection and sacrificed 48 hours later. Finally, levels of IL-4, IL-6, IL-10 and IFN -γ in serum were examined by ELISA; expressions of the four cytokines in spleen were examined by q RT-PCR; migration of ADSC to mouse model nasal mucosa were observed through fluorescence microscope; eosinophil infiltration were observed by the nasal HE staining. Mouse ADSC was isolated, cultured and identified successfully. There was significant difference in symptom scores of AR models (compared with 0/0/0 group, P<0.01). The IL-4 and IL-6 levels of OVA/OVA/high ADSC group were significantly lower than OVA/OVA/0 group (group 1: (17.95±7.78), (27.51±5.93) pg/ml; group 4: (56.82±9.12), (70.03±7.22) pg/ml), the IFN-γ and IL-10 levels increased significantly (group 1: (367.74±13.79), (417.10±72.40) pg/ml; group 4: (199.46±11.25), (122.50±15.57) pg/ml) in serum. These differences were statistically significant(P<0.01). Compared with OVA/OVA/low ADSC group, the IL-4 and IL-6 levels decreased significantly (group 1: (17.95±7.78), (27.51±5.93) pg/ml; group 2: (41.57±12.27), (56.21±9.23)pg/ml) of OVA / OVA / high ADSC group, and the IFN-γ and IL-10 increased significantly (group 1: (367.74±13.79), (417.10±72.40)pg/ml; group 2: (281.77±30.41), (203.45±87.10) pg/ml). These differences were statistically significant(P<0.01). At the same time, the corresponding changes observed at the levels of the cytokines' mRNA. ADSC labeled by CM-Dil could migrate to the mouse nasal mucosa. OVA/OVA/high ADSC group showed the more red fluorescence than the OVA/OVA/low ADSC group. The eosinophils in nasal mucosa of the two groups reduced compared with the normal control. ADSC injected via tail-vein can migrate to nasal mucosa and play non-specific immune effects, that may to effect the releases of some cytokines then to regulate the Th1/Th2 imbalance and the function of Treg cell, finally that be dose-related in a certain extent.

Human adipose-derived mesenchymal stem cells (Ad-MSCs) have been proposed as suitable option for cell-based therapies to support bone regeneration. In the bone environment, Ad-MSCs will receive stimuli from resident cells that may favor their osteogenic differentiation. There is recent evidence that this process can be further improved by extremely low frequency pulsed electromagnetic fields (ELF-PEMFs). Thus, the project aimed at (i) investigating whether co-culture conditions of human osteoblasts (OBs) and Ad-MSCs have an impact on their proliferation and osteogenic differentiation; (ii) whether this effect can be further improved by repetitive exposure to two specific ELF-PEMFs (16 and 26 Hz); (iii) and the effect of these ELF-PEMFs on human osteoclasts (OCs). Osteogenic differentiation was improved by co-culturing OBs and Ad-MSCs when compared to the individual mono-cultures. An OB to Ad-MSC ratio of 3:1 had best effects on total protein content, alkaline phosphatase (AP) activity, and matrix mineralization. Osteogenic differentiation was further improved by both ELF-PEMFs investigated. Interestingly, only repetitive exposure to 26 Hz ELF-PEMF increased Trap5B activity in OCs. Considering this result, a treatment with gradually increasing frequency might be of interest, as the lower frequency (16 Hz) could enhance bone formation, while the higher frequency (26 Hz) could enhance bone remodeling.

Effects of exosomes from human adipose-derived mesenchymal stem cells on pulmonary vascular endothelial cells injury in septic mice and its mechanism

Objective: To observe the effect of basic fibroblast growth factor (bFGF) treatment on efficacy of adipose-derived mesenchymal stem cells (ADSCs) in cirrhotic rats. Methods: A rat model of liver cirrhosis was established via intraperitoneal injection of carbon tetrachloride (CCl(4)) for 10 weeks. Thirty SD rats were randomly divided into 3 groups (n = 10): control group served as group A, and 0.5ml of phosphate-buffered saline (PBS) was transfused into the tail vein; ADSCs single-dose transplantation group served as group B, and 1×10(6) ADSCs were transplanted into the tail vein; bFGF-treated ADSCs treatment group served as group C, and 1×10(6) bFGF-treated ADSCs were transplanted through tail vein. Liver function, pathological and cytokine changes, and the in vivo survival transformation condition of the transplanted cells were measured at one week after transplantation. F test and an independent sample t test were used. Results: bFGF treatment had significantly promoted the proliferation, differentiation and overexpression of hepatocyte growth factor (HGF) in ADSCs [ADSCs single: (2 137.16 ± 261.52) pg/ml vs. ADSCs (bFGF): (4 776.23 ± 532.44) pg/ml, P < 0.05]. The bFGF-treated ADSCs treatment group had statistically significant differences in promoting the recovery of liver function [alanine aminotransferase (ALT): ADSCs single (190.8 ± 34.98) vs. ADSCs (bFGF): (117.8 ± 35.81) pg/ml; aspartate aminotransferase (AST): ADSCs single (295.2 ± 33.71) U/L vs. ADSCs (bFGF): (183.8 ± 41.29) U/L, P ​​< 0.05). There was no statistically significant difference in serum albumin levels between the control group, ADSCs single group, and ADSCs (bFGF) group. Compared with the control group, the serum albumin level of ADSCs (bFGF) group was increased significantly (P < 0.05), and the difference between the control group and the ADSCs single transplantation group in improving liver regeneration and reducing liver damage was statistically significant (P < 0.05). Masson trichrome staining showed that the percentage of the liver fibrosis area in the bFGF-treated ADSCs treatment group was 6.78% ± 0.56%, which was significantly higher than that of the control and ADSCs single dose transplantation group (7.96% ± 0.64%) (P < 0.05). Western blot analysis showed that the expression of HGF protein in the bFGF-treated ADSCs treatment group was significantly up-regulated, while the expression of α-smooth muscle actin was significantly down-regulated, and the difference was significant in the ADSCs single transplantation group. A double fluorescent staining method showed that the numbers of stem cells implanted in the liver tissue of the bFGF-treated ADSCs treatment group were higher than that of the ADSCs single transplantation group. In-vitro cell experiments confirmed that bFGF had significantly promoted the overexpression of HGF in ADSCs. Conclusion: bFGF-treated ADSCs transplantation can significantly improve liver function and fibrosis as compared with ADSCs single-dose transplantation in cirrhotic rats.

The Effectiveness of a Human Patient Simulator in the ATLS Shock Skills Station