Browsing by Author "Surma, Michelle"

Now showing 1 - 7 of 7
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    Arp2/3 mediated dynamic lamellipodia of the hPSC colony edges promote liposome-based DNA delivery
    (Oxford University Press, 2024) Surma, Michelle; Anbarasu, Kavitha; Das, Arupratan; Ophthalmology, School of Medicine
    Cationic liposome-mediated delivery of drugs, DNA, or RNA plays a pivotal role in small molecule therapy, gene editing, and immunization. However, our current knowledge regarding the cellular structures that facilitate this process remains limited. Here, we used human pluripotent stem cells (hPSCs), which form compact colonies consisting of dynamically active cells at the periphery and epithelial-like cells at the core. We discovered that cells at the colony edges selectively got transfected by cationic liposomes through actin-related protein 2/3 (Arp2/3) dependent dynamic lamellipodia, which is augmented by myosin II inhibition. Conversely, cells at the core establish tight junctions at their apical surfaces, impeding liposomal access to the basal lamellipodia and thereby inhibiting transfection. In contrast, liposomes incorporating mannosylated lipids are internalized throughout the entire colony via receptor-mediated endocytosis. These findings contribute a novel mechanistic insight into enhancing therapeutic delivery via liposomes, particularly in cell types characterized by dynamic lamellipodia, such as immune cells or those comprising the epithelial layer.
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    Disruption of both ROCK1 and ROCK2 genes in cardiomyocytes promotes autophagy and reduces cardiac fibrosis during aging
    (Wiley, 2019-06) Shi, Jianjian; Surma, Michelle; Yang, Yang; Wei, Lei; Pediatrics, School of Medicine
    In this study, we investigated the pathophysiological impact of Rho-associated coiled-coil–containing protein kinase (ROCK)1 and ROCK2 double deletion vs. single deletion on cardiac remodeling. Utilizing a cardiomyocyte-specific and tamoxifen-inducible MerCreMer recombinase (MCM), 3 mouse lines (MCM/ROCK1fl/fl/ROCK2fl/fl, MCM/ROCK1fl/fl, and MCM/ROCK2fl/fl) were generated. As early as 5 d after inducible deletion, the double ROCK knockout hearts exhibited reduced phosphorylation of myosin light chain (MLC) and focal adhesion kinase (FAK), supporting a role for ROCK activity in regulating the nonsarcomeric cytoskeleton. Moreover, the autophagy marker microtubule-associated proteins 1A-1B light chain 3B was increased in the double ROCK knockout, and these early molecular features persisted throughout aging. Mechanistically, the double ROCK knockout promoted age-associated or starvation-induced autophagy concomitant with reduced protein kinase B (AKT), mammalian target of rapamycin (mTOR), Unc-51–like kinase signaling, and cardiac fibrosis. In contrast, ROCK2 knockout hearts showed increased phosphorylated (p)-MLC and p-FAK levels, which were mostly attributable to a compensatory ROCK1 overactivation. Autophagy was inhibited at the baseline accompanying increased mTOR activity, leading to increased cardiac fibrosis in the ROCK2 knockout hearts. Finally, the loss of ROCK1 had no significant effect on p-MLC and p-FAK levels, mTOR signaling, or autophagy at baseline. In summary, deletions of ROCK isoforms in cardiomyocytes have different, even opposite, effects on endogenous ROCK activity and the MLC/FAK/AKT/mTOR signaling pathway, which is involved in autophagy and fibrosis of the heart.
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    Dissecting the roles of ROCK isoforms in stress-induced cell detachment
    (Taylor & Francis, 2013) Shi, Jianjian; Surma, Michelle; Zhang, Lumin; Wei, Lei; Pediatrics, School of Medicine
    The homologous Rho kinases, ROCK1 and ROCK2, are involved in stress fiber assembly and cell adhesion and are assumed to be functionally redundant. Using mouse embryonic fibroblasts (MEFs) derived from ROCK1(-/-) and ROCK2(-/-) mice, we have recently reported that they play different roles in regulating doxorubicin-induced stress fiber disassembly and cell detachment: ROCK1 is involved in destabilizing the actin cytoskeleton and cell detachment, whereas ROCK2 is required for stabilizing the actin cytoskeleton and cell adhesion. Here, we present additional insights into the roles of ROCK1 and ROCK2 in regulating stress-induced impairment of cell-matrix and cell-cell adhesion. In response to doxorubicin, ROCK1(-/-) MEFs showed significant preservation of both focal adhesions and adherens junctions, while ROCK2(-/-) MEFs exhibited impaired focal adhesions but preserved adherens junctions compared with the wild-type MEFs. Additionally, inhibition of focal adhesion or adherens junction formations by chemical inhibitors abolished the anti-detachment effects of ROCK1 deletion. Finally, ROCK1(-/-) MEFs, but not ROCK2(-/-) MEFs, also exhibited preserved central stress fibers and reduced cell detachment in response to serum starvation. These results add new insights into a novel mechanism underlying the anti-detachment effects of ROCK1 deletion mediated by reduced peripheral actomyosin contraction and increased actin stabilization to promote cell-cell and cell-matrix adhesion. Our studies further support the differential roles of ROCK isoforms in regulating stress-induced loss of central stress fibers and focal adhesions as well as cell detachment.
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    Distinct roles for ROCK1 and ROCK2 in the regulation of cell detachment
    (Springer Nature, 2013-02-07) Shi, Jianjian; Wu, Xiangbing; Surma, Michelle; Vemula, Sasidhar; Zhang, Lumin; Yang, Yu; Kapur, Reuben; Wei, Lei; Pediatrics, School of Medicine
    This study, using mouse embryonic fibroblast (MEF) cells derived from ROCK1(-/-) and ROCK2(-/-) mice, is designed to dissect roles for ROCK1 and ROCK2 in regulating actin cytoskeleton reorganization induced by doxorubicin, a chemotherapeutic drug. ROCK1(-/-) MEFs exhibited improved actin cytoskeleton stability characterized by attenuated periphery actomyosin ring formation and preserved central stress fibers, associated with decreased myosin light chain 2 (MLC2) phosphorylation but preserved cofilin phosphorylation. These effects resulted in a significant reduction in cell shrinkage, detachment, and predetachment apoptosis. In contrast, ROCK2(-/-) MEFs showed increased periphery membrane folding and impaired cell adhesion, associated with reduced phosphorylation of both MLC2 and cofilin. Treatment with inhibitor of myosin (blebbistatin), inhibitor of actin polymerization (cytochalasin D), and ROCK pan-inhibitor (Y27632) confirmed the contributions of actomyosin contraction and stress fiber instability to stress-induced actin cytoskeleton reorganization. These results support a novel concept that ROCK1 is involved in destabilizing actin cytoskeleton through regulating MLC2 phosphorylation and peripheral actomyosin contraction, whereas ROCK2 is required for stabilizing actin cytoskeleton through regulating cofilin phosphorylation. Consequently, ROCK1 and ROCK2 can be functional different in regulating stress-induced stress fiber disassembly and cell detachment.
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    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.
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    Novel Insights into the Roles of Rho Kinase in Cancer
    (Springer, 2016-08) Wei, Lei; Surma, Michelle; Shi, Stephanie; Lambert-Cheatham, Nathan; Shi, Jianjian; Department of Pediatrics, IU School of Medicine
    Rho-associated coiled-coil kinase (ROCK) is a major downstream effector of the small GTPase RhoA. The ROCK family, consisting of ROCK1 and ROCK2, plays a central role in the organization of the actin cytoskeleton, and is involved in a wide range of fundamental cellular functions such as contraction, adhesion, migration, proliferation, and apoptosis. Since the discovery of effective inhibitors such as fasudil and Y27632, the biological roles of ROCK have been extensively explored in numerous diseases, including cancer. Accumulating evidence supports the concept that ROCK plays important roles in tumor development and progression through regulating many key cellular functions associated with malignancy, including tumorigenicity, tumor growth, metastasis, angiogenesis, tumor cell apoptosis/survival and chemoresistance as well. This review focuses on the new advances of the most recent 5 years from the studies on the roles of ROCK in cancer development and progression; the discussion is mainly focused on the potential value of ROCK inhibitors in cancer therapy.
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    ROCK2 inhibition enhances the thermogenic program in white and brown fat tissue in mice
    (Wiley, 2020-01) Wei, Lei; Surma, Michelle; Yang, Yang; Tersey, Sarah; Shi, Jianjian; Pediatrics, School of Medicine
    The RhoA/ROCK-mediated actin cytoskeleton dynamics have been implicated in adipogenesis. The two ROCK isoforms, ROCK1 and ROCK2, are highly homologous. The contribution of ROCK2 to adipogenesis in vivo has not been elucidated. The present study aimed at the in vivo and in vitro roles of ROCK2 in the regulation of adipogenesis and the development of obesity. We performed molecular, histological and metabolic analyses in ROCK2+/− and ROCK2+/KD mouse models, the latter harboring an allele with a kinase-dead (KD) mutation. Both ROCK2+/− and ROCK2+/KD mouse models showed a lean body mass phenotype during aging, associated with increased amounts of beige cells in subcutaneous white adipose tissue (sWAT) and increased thermogenic gene expression in all fat depots. ROCK2+/− mice on a high-fat diet showed increased energy expenditure accompanying by reduced obesity, and improved insulin sensitivity. In vitro differentiated ROCK2+/− stromal-vascular (SV) cells revealed increased beige adipogenesis associated with increased thermogenic gene expressions. Treatment with a selective ROCK2 inhibitor, KD025, to inhibit ROCK2 activity in differentiated SV cells reproduced the pro-beige phenotype of ROCK2+/− SV cells. In conclusion, ROCK2 activity-mediated actin cytoskeleton dynamics contribute to the inhibition of beige adipogenesis in WAT, and also promotes age-related and diet-induced fat mass gain and insulin resistance.