Showing papers by "Denise R. Cooper published in 2014"
TL;DR: Human adipose-derived stem cells are promising therapeutic cells for TBI, and lncRNAs in the secretome is an important mechanism of cell therapy, although hADSCs showed reduced efficacy in aged rats, which may in part result from decreased homing of the cells to the spleen.
Abstract: Traumatic brain injury (TBI) survivors exhibit motor and cognitive symptoms from the primary injury that can become aggravated over time because of secondary cell death. In the present in vivo study, we examined the beneficial effects of human adipose-derived stem cells (hADSCs) in a controlled cortical impact model of mild TBI using young (6 months) and aged (20 months) F344 rats. Animals were transplanted intravenously with 4 × 106 hADSCs (Tx), conditioned media (CM), or vehicle (unconditioned media) at 3 h after TBI. Significant amelioration of motor and cognitive functions was revealed in young, but not aged, Tx and CM groups. Fluorescent imaging in vivo and ex vivo revealed 1,1′ dioactadecyl-3-3-3′,3′-tetramethylindotricarbocyanine iodide-labeled hADSCs in peripheral organs and brain after TBI. Spatiotemporal deposition of hADSCs differed between young and aged rats, most notably reduced migration to the aged spleen. Significant reduction in cortical damage and hippocampal cell loss was observed in both Tx and CM groups in young rats, whereas less neuroprotection was detected in the aged rats and mainly in the Tx group but not the CM group. CM harvested from hADSCs with silencing of either NEAT1 (nuclear enriched abundant transcript 1) or MALAT1 (metastasis associated lung adenocarcinoma transcript 1), long noncoding RNAs (lncRNAs) known to play a role in gene expression, lost the efficacy in our model. Altogether, hADSCs are promising therapeutic cells for TBI, and lncRNAs in the secretome is an important mechanism of cell therapy. Furthermore, hADSCs showed reduced efficacy in aged rats, which may in part result from decreased homing of the cells to the spleen.
138 citations
TL;DR: The ability of cells to increase phosphorylated SR proteins for PPARγ2 splicing suggests that fluxes in NEAT1 levels during adipogenesis regulate alternative splicing events.
Abstract: Long non-coding (lnc) RNAs serve a multitude of functions in cells. NEAT1 RNA is a highly abundant 4 kb lncRNA in nuclei, and coincides with paraspeckles, nuclear domains that control sequestration of paraspeckle proteins. We examined NEAT1 RNA levels and its function in 3T3-L1 cells during differentiation to adipocytes. Levels of NEAT1 transcript, measured by RT-PCR, fluctuated in a temporal manner over the course of differentiation that suggested its role in alternative splicing of PPARγ mRNA, the major transcription factor driving adipogenesis. When cells were induced to differentiate by a media cocktail of insulin, dexamethasone, and isobutylmethyxanthine (IBMX) on Day 0, NEAT1 levels dropped on Day 4, when the PPARγ2 variant was spliced and when terminal differentiation occurs The appearance of PPARγ2 coordinates with the PPARγ1 variant to drive differentiation of adipocytes. SiRNA used to deplete NEAT1 resulted in the inability of cells to phosphorylate the serine/arginine-rich splicing protein, SRp40. SiRNA treatment for SRp40 resulted in dysregulation of PPARγ1 and, primarily, PPARγ2 mRNA levels. SRp40 associated with NEAT1, as shown by RNA-IP on days 0 and 8, but decreased on day 4, and concentrations increased over that of IgG control. Overexpression of SRp40 increased PPARγ2, but not γ1. Although lncRNA MALAT1 has been investigated in SR protein function, NEAT1 has not been shown to bind SR proteins for phosphorylation such that alternative splicing results. The ability of cells to increase phosphorylated SR proteins for PPARγ2 splicing suggests that fluxes in NEAT1 levels during adipogenesis regulate alternative splicing events.
96 citations
TL;DR: Compared with reports of ADSCs from a number of human depots, there have been no comparisons of the ability of dedifferentiated DFAT cells from the same donor and depot to differentiate or modulate migration of HDFs.
Abstract: Objective: Adipose tissue is a robust source of adipose-derived stem cells (ADSCs) that may be able to provide secreted factors that promote the ability of wounded tissue to heal. However, adipocytes also have the potential to dedifferentiate in culture to cells with stem cell-like properties that may improve their behavior and functionality for certain applications. Approach: ADSCs are adult mesenchymal stem cells that are cultured from the stromal vascular fraction of adipose tissue. However, adipocytes are capable of dedifferentiating into cells with stem cell properties. In this case study, we compare ADSC and dedifferentiated fat (DFAT) cells from the same patient and fat depot for mesenchymal cell markers, embryonic stem cell markers, ability to differentiate to adipocytes and osteoblasts, senescence and telomerase levels, and ability of conditioned media (CM) to stimulate migration of human dermal fibroblasts (HDFs). Innovation and Conclusions: ADSCs and DFAT cells displayed identical levels of CD9...
33 citations
TL;DR: KCδI is a gate-keeper of adipocyte differentiation and is required for mitotic clonal expansion of preadipocytes in adipogenesis, and the splice factor TRA2B regulates alternative splicing of PKC δI during adipogenesis.
11 citations
TL;DR: The relationship between a survival kinase PKCδVIII and hTERT expression in adipose derived stem cells from a lean and obese subject is determined and a larger epigenetic modification in obese ADSC compared to lean ADSC is proposed.
Abstract: Background: Adipose-derived stem cells (ADSC) were isolated and characterized from lean and obese subjects. We previously reported that distinct differences were observed in differentiating lean and obese preadipocytes. Protein kinase C delta (PKCδ) is alternatively spliced and has important roles in apoptosis. PKCδI promotes apoptosis and PKCδVIII promotes survival. Our previous data indicated an increase in the survival kinase, PKCδVIII in ADSC derived from an obese donor. We also determined that obese adipocytes were resistant to apoptosis. Here, we determine the relationship between a survival kinase PKCδVIII and hTERT expression in adipose derived stem cells from a lean and obese subject.
Methods: We evaluated the telomerase activity and human telomerase reverse transcriptase (hTERT) expression in lean and obese ADSC. The lean and obese ADSC were purchased as cryopreserved cells from ZenBio™ (Research Triangle Park, NC, USA). Analyses were performed using PRISM™ software and analyzed using two-tailed Student’s t -test.
Results: We observed an increase in telomerase in differentiating obese ADSC using western blot analysis. We determined the levels of hTERT splice variants. hTERT α+/β+ splice variant was increased after transfected of PKCδVIII. We next determined whether PKCδVIII over-expression affected the levels of telomerase. The results indicate an increase in telomerase with PKCδVIII over-expression.
Conclusions: Over-expression of PKCδVIII in lean ADSC substantially increased expression of hTERT and telomerase. The decreased senescence seen in obese ADSC may in part be attributed to PKCδVIII. Obese ADSC undergo lower senescence and may have increased growth potential. These results propose a larger epigenetic modification in obese ADSC compared to lean ADSC.
4 citations