Adipose-Derived Mesenchymal Stem Cells Enhance Ovarian Cancer Growth and Metastasis by Increasing Thymosin Beta 4X-Linked Expression.
ABSTRACT: As shown in our previous studies, growth and metastasis of ovarian cancer can be regulated by adipose-derived mesenchymal stem cells (ADSCs). However, the underlying mechanism has not yet been revealed. In this study, a proteomics analysis was performed to compare protein expression treated with and without ADSCs in ovarian cancer cells. Protein levels were altered in ovarian cancer cells due to the treatment of ADSCs. Thymosin beta 4 X-linked (TMSB4X) levels changed dramatically, and this protein was identified as one of the most important candidate molecules contributing to the tumour-promoting effects of ADSCs. Compared with the cells that are cultured in the normal growth medium, the TMSB4X levels cultured in ADSC-conditioned medium increased significantly in ovarian cancer cells. Furthermore, the growth and invasion of cancer cells were decreased, even in the ADSC-conditioned medium treatment group (P < 0.05), by the inhibition of TMSB4X. As shown in the bioluminescence images captured in vivo, increased ovarian cancer's growth and metastasis, along with elevated TMSB4X expression, were observed in the group of ADSC-conditioned medium, and the tumour-promoting effect of ADSCs was attenuated by the inhibition of TMSB4X. Based on our findings, increased TMSB4X expression may play a role in accelerating the ADSC-mediated proliferation, invasion, and migration of ovarian cancers.
Project description:Adipose-derived stem cells (ADSCs) primed with paclitaxel (PTX) are now hypothesized to represent a potential Trojan horse to vehicle and deliver PTX into tumors. We analyzed the anticancer activity of PTX released by ADSCs primed with PTX (PTX-ADSCs) (~20 ng/mL) in a panel of ovarian cancer (OvCa) cells sensitive or resistant to PTX. We used two (2D) and three dimensional (3D) in vitro models (multicellular tumor spheroids, MCTSs, and heterospheroids) to mimic tumor growth in ascites. The coculture of OvCa cells with PTX-ADSCs inhibited cell viability in 2D models and in 3D heterospheroids (SKOV3-MCTSs plus PTX-ADSCs) and counteracted PTX-resistance in Kuramochi cells. The cytotoxic effects of free PTX and of equivalent amounts of PTX secreted in PTX-ADSC-conditioned medium (CM) were compared. PTX-ADSC-CM decreased OvCa cell proliferation, was more active than free PTX and counteracted PTX-resistance in Kuramochi cells (6.0-fold decrease in the IC50 values). Cells cultivated as 3D aggregated MCTSs were more resistant to PTX than 2D cultivation. PTX-ADSC-CM (equivalent-PTX) was more active than PTX in MCTSs and counteracted PTX-resistance in all cell lines. PTX-ADSC-CM also inhibited OvCa-MCTS dissemination on collagen-coated wells. In conclusion, PTX-ADSCs and PTX-MSCs-CM may represent a new option with which to overcome PTX-resistance in OvCa.
Project description:Transplantation of stem cells into damaged hearts has had modest success as a treatment for ischemic heart disease. One of the limitations is the poor stem cell survival in the diseased microenvironment. Prolyl hydroxylase domain protein 2 (PHD2) is a cellular oxygen sensor that regulates 2 key transcription factors involved in cell survival and inflammation: hypoxia-inducible factor and nuclear factor-?B.We studied whether and how PHD2 silencing in human adipose-derived stem cells (ADSCs) enhances their cardioprotective effects after transplantation into infarcted hearts.ADSCs were transduced with lentiviral short hairpin RNA against prolyl hydroxylase domain protein 2 (shPHD2) to silence PHD2. ADSCs, with or without shPHD2, were transplanted after myocardial infarction in mice. ADSCs reduced cardiomyocyte apoptosis, fibrosis, and infarct size and improved cardiac function. shPHD2-ADSCs exerted significantly more protection. PHD2 silencing induced greater ADSC survival, which was abolished by short hairpin RNA against hypoxia-inducible factor-1?. Conditioned medium from shPHD2-ADSCs decreased cardiomyocyte apoptosis. Insulin-like growth factor-1 (IGF-1) levels were significantly higher in the conditioned medium of shPHD2-ADSCs versus ADSCs, and depletion of IGF-1 attenuated the cardioprotective effects of shPHD2-ADSC-conditioned medium. Nuclear factor-?B activation was induced by shPHD2 to induce IGF-1 secretion via binding to IGF-1 gene promoter.PHD2 silencing promotes ADSCs survival in infarcted hearts and enhances their paracrine function to protect cardiomyocytes. The prosurvival effect of shPHD2 on ADSCs is hypoxia-inducible factor-1? dependent, and the enhanced paracrine function of shPHD2-ADSCs is associated with nuclear factor-?B-mediated IGF-1 upregulation. PHD2 silencing in stem cells may be a novel strategy for enhancing the effectiveness of stem cell therapy after myocardial infarction.
Project description:Development of primary follicles in vitro benefits from a three-dimensional matrix that is enriched with paracrine factors secreted from feeder cells and mimics the in vivo environment. In this study, we investigated the role of paracrine signaling from adipose-derived stem cells (ADSCs) in supporting primary follicle development in a biomimetic poly(ethylene glycol) (PEG)-based matrix. Follicles co-cultured with ADSCs and follicles cultured in conditioned medium from ADSCs encapsulated in gels (3D CM) exhibited significantly (p < 0.01 and p = 0.09, respectively) improved survival compared to follicles cultured in conditioned medium collected from ADSCs cultured in flasks (2D CM) and follicles cultured without paracrine support. The gene expression of ADSCs suggested that the stem cells maintained their multipotency in the 3D PEG environment over the culture period, regardless of the presence of the follicles, while under 2D conditions the multipotency markers were downregulated. The differences in cytokine signatures of follicles exposed to 3D and 2D ADSC paracrine factors suggest that early cytokine interactions are key for follicle survival. Taken together, the biomimetic PEG scaffold provides a three-dimensional, in vivo-like environment to induce ADSCs to secrete factors which promote early stage ovarian follicle development and survival.
Project description:Efforts to develop peripheral blood-derived nature killer (NK) cells into therapeutic products have been hampered by these cells' low abundance and histoincompatibility. On the other hand, derivation of NK-like cells from more abundant cell sources such as embryonic stem cells (ESCs) and umbilical cord blood (UCB) requires the selection of rare CD34+ cells. Thus, we sought to convert adipose-derived stem cells (ADSCs), which are abundant and natively CD34+, into NK-like cells. When grown in hematopoietic induction medium, ADSCs formed sphere clusters and expressed hematopoietic markers CD34, CD45, and KDR. Further induction in NK cell-specific medium resulted in a population of cells that expressed NK cell marker CD56, and thus termed ADSC-NK. Alternatively, the hematopoietically induced ADSCs were transduced with NK cell-specific transcription factor E4BP4 prior to induction in NK cell-specific medium. This latter population of cells, termed ADSC-NKE, expressed CD56 and additional NK cell markers such as CD16, CD94, CD158, CD314, FasL, and NKp46. ADSC-NKE was as potent as NK leukemia cell NKL in killing breast cancer cell MCF7 and prostate cancer cells DU145, PC3, LnCap, DuPro, C4-2 and CWR22, but exhibited no killing activity toward normal endothelial and smooth muscle cells. In nude mice test ADSC-NKE was able to significantly delay the progression of tumors formed by MCF7 and PC3. When injected into immunocompetent rats, ADSC-NKE was detectable in bone marrow and spleen for at least 5 weeks. Together, these results suggest that ADSCs can be converted into NK-like cells with anti-tumor activities.
Project description:Tissue regeneration is impaired in aged individuals. Adipose-derived mesenchymal stromal cells (ADSCs), a promising source for cell therapy, were shown to secrete various angiogenic factors and improve vascularization of ischemic tissues. We analyzed how patient age affected the angiogenic properties of ADSCs. ADSCs were isolated from subcutaneous fat tissue of patients with coronary artery disease (CAD; n = 64, 43-77 years old) and without CAD (n = 31, 2-82 years old). ADSC phenotype characterized by flow cytometry was CD90(+)/CD73(+)/CD105(+)/CD45(-)/CD31(-) for all samples, and these cells were capable of adipogenic and osteogenic differentiation. ADSCs from aged patients had shorter telomeres (quantitative reverse transcription polymerase chain reaction) and a tendency to attenuated telomerase activity. ADSC-conditioned media (ADSC-CM) stimulated capillary-like tube formation by endothelial cells (EA.hy926), and this effect significantly decreased with the age of patients both with and without CAD. Angiogenic factors (vascular endothelial growth factor, placental growth factor, hepatocyte growth factor, angiopoetin-1, and angiogenin) in ADSC-CM measured by enzyme-linked immunosorbent assay significantly decreased with patient age, whereas levels of antiangiogenic factors thrombospondin-1 and endostatin did not. Expression of angiogenic factors in ADSCs did not change with patient age (real-time polymerase chain reaction); however, gene expression of factors related to extracellular proteolysis (urokinase and its receptor, plasminogen activator inhibitor-1) and urokinase-type plasminogen activator receptor surface expression increased in ADSCs from aged patients with CAD. ADSCs from aged patients both with and without CAD acquire aging characteristics, and their angiogenic potential declines because of decreasing proangiogenic factor secretion. This could restrict the effectiveness of autologous cell therapy with ADSCs in aged patients.
Project description:Angiogenesis is a complicated process in which perivascular cells play important roles. Multipotent mesenchymal stem/stromal cells (MSCs) from distinct tissues have been proved to be proangiogenic and share functional properties and gene expression profiles with perivascular cells. However, different tissues derived MSCs may exhibit different potential for clinical applications. Accordingly, comparative studies on different MSCs are essential. Here, we characterized MSCs from adipose (ADSCs), umbilical cord (UCMSCs), and endometrium (EMSCs) in terms of the surface antigen expression, differentiation ability, and the ability of angiogenesis promotion on endothelial colony-forming cells (ECFCs) both in vitro and in vivo. No significant differences in immunophenotype and differentiation were observed. In addition, three types of MSCs all located around tubular-like structures formed by ECFCs in coculture system on matrigel. But ECFCs seeded on ADSCs monolayer formed more organized capillary-like network than that on UCMSCs or EMSCs. When suspended with ECFCs in matrigel and implanted into nude mice, ADSCs promoted more functional vessel formation after 7 days. Moreover, in murine hindlimb ischemia model, cotransplantation of ECFCs with ADSCs was significantly superior to UCMSCs and EMSCs in promoting perfusion recovery and limb salvage. Furthermore, ADSC-conditioned medium (CM) contained more proangiogenic factors (such as vascular endothelial growth factor-A, platelet-derived growth factor BB, and basic fibroblast growth factor) and less inhibitory factor (such as thrombospondin-1), when compared with UCMSC-CM and EMSC-CM. And ADSC-CM more durably stabilized the vascular-like structures formed by ECFCs on matrigel and promoted ECFCs migration more efficiently. In summary, MSCs from adipose show significantly efficient promotion on angiogenesis both in vitro and in vivo than UCMSCs and EMSCs. Hence, ADSCs may be recommended as a more suitable source for treating hindlimb ischemia.
Project description:INTRODUCTION: Young patients receiving chemotherapy occasionally face infertility and premature ovarian failure (POF). Numerous investigations reported that adipose-derived stem cells (ADSCs) transplantation could ameliorate the structure and function of injured tissues. The aim of this study was to explore the therapeutic efficacy of ADSC transplantation for chemotherapy-induced ovarian damage. METHODS: Female mice were injected intraperitoneally with 50 mg/kg cyclophosphamide (CTX). After 15 consecutive days of injection, ADSCs were transplanted either directly into bilateral ovaries or via intravenous injection, and the ovaries were excised after either 1 week or 1 month of treatment. The follicles were counted and categorized, and ovarian histologic sections were stained for TUNEL. Ovarian function was evaluated by monitoring ovulation. ADSC tracking, microarray analyses, and real-time polymerase chain reaction (PCR) were used to assess the inner mechanism of injury and repair. RESULTS: The ovarian function of mice exposed to CTX injection improved after ADSC transplantation. The population of follicles at different stages and ovulation significantly increased after the treatment. Immunofluorescence revealed reduced TUNEL staining. The tracking of ADSCs revealed that these cells did not directly differentiate into the follicle component. Microarray analyses indicated that changes in different groups of genes might affect follicle formation or ovulation. CONCLUSIONS: ADSC transplantation improved ovarian function. Our results suggest a potential mechanism for ADSC therapy.
Project description:Stem cells hold great promise for treating cartilage degenerative diseases such as osteoarthritis (OA). The efficacy of stem cell-based therapy for cartilage repair is highly dependent on their interactions with local cells in the joint. This study aims at evaluating the interactions between osteoarthritic chondrocytes (OACs) and adipose-derived stem cells (ADSCs) using three dimensional (3D) biomimetic hydrogels. To examine the effects of cell distribution on such interactions, ADSCs and OACs were co-cultured in 3D using three co-culture models: conditioned medium (CM), bi-layered, and mixed co-culture with varying cell ratios. Furthermore, the effect of transforming growth factor (TGF)-?3 supplementation on ADSC-OAC interactions and the resulting cartilage formation was examined. Outcomes were analyzed using quantitative gene expression, cell proliferation, cartilage matrix production, and histology. TGF-?3 supplementation led to a substantial increase in cartilage matrix depositions in all groups, but had differential effects on OAC-ADSC interactions in different co-culture models. In the absence of TGF-?3, CM or bi-layered co-culture had negligible effects on gene expression or cartilage formation. With TGF-?3 supplementation, CM and bi-layered co-culture inhibited cartilage formation by both ADSCs and OACs. In contrast, a mixed co-culture with moderate OAC ratios (25% and 50%) resulted in synergistic interactions with enhanced cartilage matrix deposition and reduced catabolic marker expression. Our results suggested that the interaction between OACs and ADSCs is highly dependent on cell distribution in 3D and soluble factors, which should be taken into consideration when designing stem cell-based therapy for treating OA patients.
Project description:BACKGROUND:Cartilage tissue engineering is a promising technique for repairing cartilage defect. Due to the limitation of cell number and proliferation, mesenchymal stem cells (MSCs) have been developed as a substitute to chondrocytes as a cartilage cell-source. This study aimed to develop cartilage tissue from human adipose-derived stem cells (ADSCs) cultured on a Bombyx mori silk fibroin scaffold and supplemented with 10% platelet-rich plasma (PRP). METHODS:Human ADSCs and PRP were characterized. A silk fibroin scaffold with 500??m pore size was fabricated through salt leaching. ADSCs were then cultured on the scaffold (ADSC-SS) and supplemented with 10% PRP for 21?days to examine cell proliferation, chondrogenesis, osteogenesis, and surface marker expression. The messenger ribonucleic acid (mRNA) expression of type 2 collagen, aggrecan, and type 1 collagen was analysed. The presence of type 2 collagen confirming chondrogenesis was validated using immunocytochemistry. The negative and positive controls were ADSC-SS supplemented with 10% foetal bovine serum (FBS) and ADSC-SS supplemented with commercial chondrogenesis medium, respectively. RESULTS:Cells isolated from adipose tissue were characterized as ADSCs. Proliferation of the ADSC-SS PRP was significantly increased (p?<?0.05) compared to that of controls. Chondrogenesis was observed in ADSC-SS PRP and was confirmed through the increase in glycosaminoglycans (GAG) and transforming growth factor-?1 (TGF-?1) secretion, the absence of mineral deposition, and increased surface marker proteins on chondrogenic progenitors. The mRNA expression of type 2 collagen in ADSC-SS PRP was significantly increased (p?<?0.05) compared to that in the negative control on days 7 and 21; however, aggrecan was significantly increased on day 14 compared to the controls. ADSC-SS PRP showed stable mRNA expression of type 1 collagen up to 14?days and it was significantly decreased on day 21. Confocal analysis showed the presence of type 2 collagen in the ADSC-SS PRP and positive control groups, with high distribution outside the cells forming the extracellular matrix (ECM) on day 21. CONCLUSION:Our study showed that ADSC-SS with supplemented 10% PRP medium can effectively support chondrogenesis of ADSCs in vitro and promising for further development as an alternative for cartilage tissue engineering in vivo.
Project description:Mesenteric adipose tissue hyperplasia is a hallmark of Crohn's disease (CD). Recently, we showed that mesenteric adipose-derived stromal cells (ADSCs) from CD, ulcerative colitis, and control patients synthesize and release adipokines in a disease-dependent manner. Here we examined the expression profiles of CD and control patient-derived mesenteric ADSCs and studied the effects of their extracellular mediators on colonocyte signaling in vitro and experimental colitis in vivo. ADSCs were isolated from mesenteric fat of control and CD patients. Microarray profiling and network analysis were performed in ADSCs and human colonocytes treated with conditioned media from cultured ADSCs. Mice with acute colitis received daily injections of conditioned media from patient-derived ADSCs, vehicle, or apolactoferrin. Proliferative responses were evaluated in conditioned media-treated colonocytes and mouse colonic epithelium. Total protein was isolated from cultured colonocytes after treatment with apolactoferrin for Western blot analysis of phosphorylated intracellular signaling kinases. Microarray profiling revealed differential mRNA expression in CD patient-derived ADSCs compared with controls, including lactoferrin. Administration of CD patient-derived medium or apolactoferrin increased colonocyte proliferation compared with controls. Conditioned media from CD patient-derived ADSCs or apolactoferrin attenuated colitis severity in mice and enhanced colonocyte proliferation in vivo. ADSCs from control and CD patients show disease-dependent inflammatory responses and alter colonic epithelial cell signaling in vitro and in vivo. Furthermore, we demonstrate lactoferrin production by adipose tissue, specifically mesenteric ADSCs. We suggest that mesenteric ADSC-derived lactoferrin may mediate protective effects and participate in the pathophysiology of CD by promoting colonocyte proliferation and the resolution of inflammation.