The steady increase in the incidence of obesity among adults has been paralleled with higher levels of obesity-associated breast cancer. While recent studies have suggested that adipose stromal/stem cells (ASCs) isolated from obese women enhance tumorigenicity, the mechanism(s) by which this occurs remains undefined. Evidence suggests that increased adiposity results in increased leptin secretion from adipose tissue, which has been shown to increased cancer cell proliferation. Previously, our group demonstrated that ASCs isolated from obese women (obASCs) also express higher levels of leptin relative to ASCs isolated from lean women (lnASCs) and that this obASC-derived leptin may account for enhanced breast cancer cell growth. The current study investigates the impact of inhibiting leptin expression in lnASCs and obASCs on breast cancer cell (BCC) growth and progression. Estrogen receptor positive (ER+) BCCs were co-cultured with leptin shRNA lnASCs or leptin shRNA obASCs and changes in the proliferation, migration, invasion, and gene expression of BCCs were investigated. To assess the direct impact of leptin inhibition in obASCs on BCC proliferation, MCF7 cells were injected alone or mixed with control shRNA obASCs or leptin shRNA obASCs into SCID/beige mice. ER+ BCCs were responsive to obASCs during direct co-culture, whereas lnASCs were unable to increase ER+ BCC growth. shRNA silencing of leptin in obASCs negated the enhanced proliferative effects of obASC on BCCs following direct co-culture. BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction. Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver. These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed with MCF7 cells mixed with leptin shRNA obASCs, when compared to tumors formed with MCF7 cells mixed with control shRNA obASCs. This study provides mechanistic insight as to how obesity enhances the proliferation and metastasis of breast cancer cells; specifically, obASC-derived leptin contributes to the aggressiveness of breast cancer in obese women.

Introduction: Obesity has been associated with increased incidence and mortality of breast cancer. While the precise correlation between obesity and breast cancer remains to be determined, recent studies suggest that adipose tissue and adipose stem cells (ASCs) influence breast cancer tumorigenesis and tumor progression. Methods: Breast cancer cells lines were co-cultured with ASCs (n = 24), categorized based on tissue site of origin and body mass index (BMI), and assessed for enhanced proliferation, alterations in gene expression profile with PCR arrays, and enhanced tumorigenesis in immunocompromised mice. The gene expression profile of ASCs was assess with PCR arrays and qRT-PCR and confirmed with Western blot analysis. Inhibitory studies were conducted by delivering estrogen antagonist ICI182,780, leptin neutralizing antibody, or aromatase inhibitor letrozole and assessing breast cancer cell proliferation. To assess the role of leptin in human breast cancers, Oncomine and Kaplan Meier plot analyses were conducted. Results: ASCs derived from the abdominal subcutaneous adipose tissue of obese subjects (BMI > 30) enhanced breast cancer cell proliferation in vitro and tumorigenicity in vivo. These findings were correlated with changes in the gene expression profile of breast cancer cells after co-culturing with ASCs, particularly in estrogen receptor-alpha (ESR1) and progesterone receptor (PGR) expression. Analysis of the gene expression profile of the four groups of ASCs revealed obesity induced alterations in several key genes, including leptin (LEP). Blocking estrogen signaling with ICI182,780, leptin neutralizing antibody, or letrozole diminished the impact of ASCs derived from obese subjects. Women diagnosed with estrogen receptor/progesterone receptor positive (ER⁺/PR⁺) breast cancers that also expressed high levels of leptin had poorer prognosis than women with low leptin expression. Conclusion: ASCs isolated from the abdomen of obese subjects demonstrated increased expression of leptin, through estrogen stimulation, which increased breast cancer cell proliferation. The results from this study demonstrate that abdominal obesity induces significant changes in the biological properties of ASCs and that these alterations enhance ER⁺/PR⁺ breast cancer tumorigenesis through estrogen dependent pathways.

More than 2.5 million patients in the U.S. require treatment for pressure ulcers annually, and the elderly are at particularly high risk for pressure ulcer development. Current therapy for pressure ulcers consists of conservative medical management for shallow lesions and aggressive debridement and surgery for deeper lesions. The current study uses a murine model to address the hypothesis that adipose-derived stromal/stem cell (ASC) treatment would accelerate and enhance pressure ulcer repair. The dorsal skin of both young (2 months old [mo]) and old (20 mo) C57BL/6J female mice was sandwiched between external magnets for 12 hours over 2 consecutive days to initiate a pressure ulcer. One day following the induction, mice were injected with ASCs isolated from congenic mice transgenic for the green fluorescent protein under a ubiquitous promoter. Relative to phosphate-buffered saline-treated controls, ASC-treated mice displayed a cell concentration-dependent acceleration of wound closure, improved epidermal/dermal architecture, increased adipogenesis, and reduced inflammatory cell infiltration. The ASC-induced improvements occurred in both young and elderly recipients, although the expression profile of angiogenic, immunomodulatory, and reparative mRNAs differed as a function of age. The results are consistent with clinical reports that fat grafting improved skin architecture in thermal injuries; the authors of this published study have invoked ASC-based mechanisms to account for their clinical outcomes. Thus, the current proof-of-principle study sets the stage for clinical translation of autologous and/or allogeneic ASC treatment of pressure ulcers. Significance: Adipose-derived stromal/stem cells (ASCs) promote the healing of pressure ulcer wounds in both young and old mice. ASCs enhance wound healing rates through adipogenic differentiation and regeneration of the underlying architecture of the skin.

Introduction: While administration of ex vivo culture-expanded stem cells has been used to study immunosuppressive mechanisms in multiple models of autoimmune diseases, less is known about the uncultured, nonexpanded stromal vascular fraction (SVF)-based therapy. The SVF is composed of a heterogeneous population of cells and has been used clinically to treat acute and chronic diseases, alleviating symptoms in a range of tissues and organs. Methods: In this study, the ability of human SVF cells was compared with culture-expanded adipose stem cells (ASCs) and bone-derived marrow stromal cells (BMSCs) as a treatment of myelin oligodendrocyte glycoprotein (35–55)-induced experimental autoimmune encephalitis in C57Bl/6J mice, a well-studied multiple sclerosis model (MS). A total of 1 × 10⁶ BMSCs, ASCs, or SVF cells were administered intraperitoneally concomitantly with the induction of disease. Mice were monitored daily for clinical signs of disease by three independent, blinded investigators and rated on a scale of 0 to 5. Spinal cords were obtained after euthanasia at day 30 and processed for histological staining using luxol fast blue, toluidine blue, and hematoxylin and eosin to measure myelin and infiltrating immune cells. Blood was collected from mice at day 30 and analyzed by enzyme-linked immunosorbent assay to measure serum levels of inflammatory cytokines. Results: The data indicate that intraperitoneal administration of all cell types significantly ameliorates the severity of disease. Furthermore, the data also demonstrate, for the first time, that the SVF was as effective as the more commonly cultured BMSCs and ASCs in an MS model. All cell therapies also demonstrated a similar reduction in tissue damage, inflammatory infiltrates, and sera levels of IFNγ and IL-12. While IFNγ levels were reduced to comparable levels between treatment groups, levels of IL-12 were significantly lower in SVF-treated than BMSC-treated or ASC-treated mice. Conclusions: Based on these data, it is evident that SVF cells have relevant therapeutic potential in an animal model of chronic MS and might represent a valuable tool for stem cell-based therapy in chronic inflammatory disease of the central nervous system. SVF offers advantages of direct and rapid isolation procedure in a xenobiotic-free environment.

Background: Obesity is associated with a higher risk of developing cancer and co-morbidities that are part of the metabolic syndrome. Adipose tissue is recognized as an endocrine organ, as it affects a number of physiological functions, and contains adipose tissue-derived stem cells (ASCs). ASCs can differentiate into cells of multiple lineages, and as such are applicable to tissue engineering and regenerative medicine. Yet the question of whether ASC functionality is affected by the donor’s body mass index (BMI) still exists.Results: ASCs were isolated from patients having different BMIs (BMI-ASCs), within the ranges of 18.5-32.8. It was hypothesized that overweight BMI-ASCs would be more compromised in early adipogenic and osteogenic potential, and ability to form coloniesin vitro. BMI was inversely correlated with ASC proliferation and colony forming potential as assessed by CyQUANT proliferation assay (fluorescence- based measurement of cellular DNA content), and colony forming assays. BMI was positively correlated with early time point (day 7) but not later time point (day 15) intracytoplasmic lipid accumulation as assessed by Oil-Red-O staining. Alizarin red staining and RT-PCR for alkaline phosphatase demonstrated that elevated BMI resulted in compromised ASC mineralization of extracellular matrix and decreased alkaline phosphatase mRNA expression.Conclusions: These data demonstrate that elevated BMI resulted in reduced ASC proliferation, and potentially compromised osteogenic capacityin vitro; thus BMI is an important criterion to consider in selecting ASC donors for clinical applications.