Browsing by Author "Anbarasu, Kavitha"

Now showing 1 - 2 of 2
  • Thumbnail Image
    Item
    Enhanced mitochondrial biogenesis promotes neuroprotection in human pluripotent stem cell derived retinal ganglion cells
    (Springer Nature, 2023-02-24) Surma, Michelle; Anbarasu, Kavitha; Dutta, Sayanta; Olivera Perez, Leonardo J.; Huang, Kang-Chieh; Meyer, Jason S.; Das, Arupratan; Ophthalmology, School of Medicine
    Mitochondrial dysfunctions are widely afflicted in central nervous system (CNS) disorders with minimal understanding on how to improve mitochondrial homeostasis to promote neuroprotection. Here we have used human stem cell differentiated retinal ganglion cells (hRGCs) of the CNS, which are highly sensitive towards mitochondrial dysfunctions due to their unique structure and function, to identify mechanisms for improving mitochondrial quality control (MQC). We show that hRGCs are efficient in maintaining mitochondrial homeostasis through rapid degradation and biogenesis of mitochondria under acute damage. Using a glaucomatous Optineurin mutant (E50K) stem cell line, we show that at basal level mutant hRGCs possess less mitochondrial mass and suffer mitochondrial swelling due to excess ATP production load. Activation of mitochondrial biogenesis through pharmacological inhibition of the Tank binding kinase 1 (TBK1) restores energy homeostasis, mitigates mitochondrial swelling with neuroprotection against acute mitochondrial damage for glaucomatous E50K hRGCs, revealing a novel neuroprotection mechanism.
  • Thumbnail Image
    Item
    Human Stem Cell Differentiated Retinal Ganglion Cells for Developing Glaucoma Neuroprotection and Cell Replacement Strategies
    (2024-07) Anbarasu, Kavitha; Das, Arupratan; Corson, Timothy; Meyer, Jason; Graham, Brett; Janga, Sarath
    Progressive loss of retinal ganglion cells (RGCs) leads to glaucoma. Early diagnosis offers an opportunity to protect existing RGCs. In advanced glaucoma, most RGCs are lost causing blindness and cell replacement therapy the only option. We used a human stem cell-based RGC differentiation model to develop neuroprotection by restoring mitochondrial homeostasis and enhancing RGC differentiation efficiency to increase the success of cell replacement therapy. Unmyelinated axons in RGCs require high levels of ATP, making disrupted mitochondria a risk factor in glaucoma. Our goal was to restore mitochondrial homeostasis through mitophagy (mitochondrial autophagy) and mitobiogenesis (mitochondrial biogenesis). Mutations in the mitophagy protein Optineurin (OPTNE50K) are found in patients with normal tension glaucoma and hence, we also used RGCs with the E50K mutation. We discovered that hRGCE50Ks suffer from mitobiogenesis issues, Parkin/Pink mediated mitophagy defects, and have OPTNE50K-Tank binding kinase-1 (TBK1) aggregates. hRGCE50Ks have lower mitochondrial mass and a higher mitochondrial load. We inhibited TBK1 to induce mitochondrial biogenesis and dissolve OPTNE50K-TBK1 aggregates. Our results show TBK1 inhibition triggered mitobiogenesis, dissolved aggregates, decreased mitochondrial ATP production load, and increased spare respiratory capacity, leading to neuroprotection. With complete RGC loss, enhancing differentiation to progenitor cells with lower cell division capacity can improve the success of cell replacement therapy and reduce teratoma formation and poor tissue integration. We observed that stem cells use proteasomes for mitochondrial degradation, while hRGCs use the lysosomal mitophagy pathway. Our results indicate that proteasomal activity declines during differentiation to hRGCs. Inhibition of proteasomal activity during early differentiation resulted in higher and faster RGC differentiation, with similar effects seen in motor neuron differentiation. We did not observe metabolic reprogramming in differentiating cells upon proteasomal activity inhibition but saw changes in cell cycle distribution, specifically an increase in the number of cells in the G1 phase. Proteomics analysis post-inhibitory treatment showed elevated neuronal differentiation proteins. Our results can be translated to minimize injection cell numbers and other risks of cell replacement therapy. In summary, my research identifies novel mechanisms for restoring mitochondrial homeostasis for neuroprotection in glaucomatous RGCs and develops an enhanced differentiation strategy to aid the success of cell replacement therapy.