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Browsing by Subject "Cellular reprogramming"
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Item Chronic cAMP activation induces adipocyte browning through discordant biphasic remodeling of transcriptome and chromatin accessibility(Elsevier, 2022) So, Jisun; Taleb, Solaema; Wann, Jamie; Strobel, Olivia; Kim, Kyungchan; Roh, Hyun Cheol; Biochemistry and Molecular Biology, School of MedicineObjective: Adipose tissue thermogenesis has been suggested as a new therapeutic target to promote energy metabolism for obesity and metabolic disease. Cold-inducible thermogenic adipocytes, called beige adipocytes, have attracted significant attention for their potent anti-obesity activity in adult humans. In this study, we identified the mechanisms underlying beige adipocyte recruitment, so-called adipocyte browning, by different stimuli. Methods: We generated a new adipocyte cell line with enhanced browning potentials and determined its transcriptomic and epigenomic responses following cAMP (forskolin, FSK) versus PPARγ activation (rosiglitazone). We performed time-course RNA-seq and compared the treatments and in vivo adipocyte browning. We also developed an improved protocol for Assay for Transposase Accessible Chromatin-sequencing (ATAC-seq) and defined changes in chromatin accessibility in a time course. The RNA-seq and ATAC-seq data were integrated to determine the kinetics of their coordinated regulation and to identify a transcription factor that drives these processes. We conducted functional studies using pharmacological and genetic approaches with specific inhibitors and shRNA-mediated knockdown, respectively. Results: FSK, not rosiglitazone, resulted in a biphasic transcriptomic response, resembling the kinetics of in vivo cold-induced browning. FSK promoted tissue remodeling first and subsequently shifted energy metabolism, concluding with a transcriptomic profile similar to that induced by rosiglitazone. The thermogenic effects of FSK were abolished by PPARγ antagonists, indicating PPARγ as a converging point. ATAC-seq uncovered that FSK leads to a significant chromatin remodeling that precedes or persists beyond transcriptomic changes, whereas rosiglitazone induces minimal changes. Motif analysis identified nuclear factor, interleukin 3 regulated (NFIL3) as a transcriptional regulator connecting the biphasic response of FSK-induced browning, as indicated by disrupted thermogenesis with NFIL3 knockdown. Conclusions: Our findings elucidated unique dynamics of the transcriptomic and epigenomic remodeling in adipocyte browning, providing new mechanistic insights into adipose thermogenesis and molecular targets for obesity treatment.Item Nanotransfection-based vasculogenic cell reprogramming drives functional recovery in a mouse model of ischemic stroke(American Association for the Advancement of Science, 2021-03-19) Lemmerman, Luke R.; Balch, Maria H.H.; Moore, Jordan T.; Alzate-Correa, Diego; Rincon-Benavides, Maria A.; Salazar-Puerta, Ana; Gnyawali, Surya; Harris, Hallie N.; Lawrence, William; Ortega-Pineda, Lilibeth; Wilch, Lauren; Risser, Ian B.; Maxwell, Aidan J.; Duarte-Sanmiguel, Silvia; Dodd, Daniel; Guio-Vega, Gina P.; McTigue, Dana M.; Arnold, W. David; Nimjee, Shahid M.; Sen, Chandan K.; Khanna, Savita; Rink, Cameron; Higuita-Castro, Natalia; Gallego-Perez, Daniel; Surgery, School of MedicineIschemic stroke causes vascular and neuronal tissue deficiencies that could lead to substantial functional impairment and/or death. Although progenitor-based vasculogenic cell therapies have shown promise as a potential rescue strategy following ischemic stroke, current approaches face major hurdles. Here, we used fibroblasts nanotransfected with Etv2, Foxc2, and Fli1 (EFF) to drive reprogramming-based vasculogenesis, intracranially, as a potential therapy for ischemic stroke. Perfusion analyses suggest that intracranial delivery of EFF-nanotransfected fibroblasts led to a dose-dependent increase in perfusion 14 days after injection. MRI and behavioral tests revealed ~70% infarct resolution and up to ~90% motor recovery for mice treated with EFF-nanotransfected fibroblasts. Immunohistological analysis confirmed increases in vascularity and neuronal cellularity, as well as reduced glial scar formation in response to treatment with EFF-nanotransfected fibroblasts. Together, our results suggest that vasculogenic cell therapies based on nanotransfection-driven (i.e., nonviral) cellular reprogramming represent a promising strategy for the treatment of ischemic stroke.Item Systemic Injection of RPE65-Programmed Bone Marrow-Derived Cells Prevents Progression of Chronic Retinal Degeneration(Elsevier, 2017-04-05) Qi, Xiaoping; Pay, S. Louise; Yan, Yuanqing; Thomas, James, Jr.; Lewin, Alfred S.; Chang, Lung-Ji; Grant, Maria B.; Boulton, Michael E.; Ophthalmology, School of MedicineBone marrow stem and progenitor cells can differentiate into a range of non-hematopoietic cell types, including retinal pigment epithelium (RPE)-like cells. In this study, we programmed bone marrow-derived cells (BMDCs) ex vivo by inserting a stable RPE65 transgene using a lentiviral vector. We tested the efficacy of systemically administered RPE65-programmed BMDCs to prevent visual loss in the superoxide dismutase 2 knockdown (Sod2 KD) mouse model of age-related macular degeneration. Here, we present evidence that these RPE65-programmed BMDCs are recruited to the subretinal space, where they repopulate the RPE layer, preserve the photoreceptor layer, retain the thickness of the neural retina, reduce lipofuscin granule formation, and suppress microgliosis. Importantly, electroretinography and optokinetic response tests confirmed that visual function was significantly improved. Mice treated with non-modified BMDCs or BMDCs pre-programmed with LacZ did not exhibit significant improvement in visual deficit. RPE65-BMDC administration was most effective in early disease, when visual function and retinal morphology returned to near normal, and less effective in late-stage disease. This experimental paradigm offers a minimally invasive cellular therapy that can be given systemically overcoming the need for invasive ocular surgery and offering the potential to arrest progression in early AMD and other RPE-based diseases.