Browsing by Subject "AMPK"

Now showing 1 - 7 of 7
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    Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance
    (Federation of American Societies for Experimental Biology, 2021-03-17) Gampala, Silpa; Zhang, GuangJun; Chang, Chun Ju; Yang, Jer-Yen; Pediatrics, School of Medicine
    Vismodegib, a Smoothened antagonist, is clinically approved for treatment of human basal cell carcinoma (BCC), in the clinical trials of medulloblastoma (MB) and other cancers. However, a significant proportion of these tumors fail to respond to Vismodegib after a period of treatment. Here, we find that AMPK agonists, A769662, and Metformin, can inhibit GLI1 activity and synergize with Vismodegib to suppress MB cell growth in vitro and in vivo. Furthermore, combination of AMPK agonists with Vismodegib is effective in overcoming Vismodegib‐resistant MB. This is the first report demonstrating that combining AMPK agonist (Metformin) and SHH pathway inhibitor (Vismodegib) confers synergy for MB treatment and provides an effective chemotherapeutic regimen that can be used to overcome resistance to Vismodegib in SHH‐driven cancers.
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    Fluid flow-induced activation of subcellular AMPK and its interaction with FAK and Src
    (Elsevier, 2020-01) Guo, Yunxia; Steele, Hannah E.; Li, Bai-Yan; Na, Sungsoo; Biomedical Engineering, School of Engineering and Technology
    AMP-activated protein kinase (AMPK) is a metabolic energy sensor that plays a critical role in cancer cell survival and growth. While the physical microenvironment is believed to influence tumor growth and progression, its role in AMPK regulation remains largely unknown. In the present study, we evaluated AMPK response to mechanical forces and its interaction with other mechano-responsive signaling proteins, FAK and Src. Using genetically encoded biosensors that can detect AMPK activities at different subcellular locations (cytosol, plasma membrane, nucleus, mitochondria, and Golgi apparatus), we observed that AMPK responds to shear stress in a subcellular location-dependent manner in breast cancer cells (MDA-MB-231). While normal epithelial cells (MCF-10A) also similarly responded to shear stress, they are less sensitive to shear stress compared to MDA-MB-231 cells. Inhibition of FAK and Src significantly decreased the basal activity level of AMPK at all five subcellular locations in MDA-MB-231 cells and selectively blocked shear stress-induced AMPK activation. Moreover, testing with cytoskeletal drugs revealed that myosin II might be the critical mediator of shear stress-induced AMPK activation in MDA-MB-231 cells. These findings suggest that breast cancer cells and normal epithelial cells may have different mechanosensitivity in AMPK signaling and that FAK and Src as well as the myosin II-dependent signaling pathway are involved in subcellular AMPK mechanotransduction in breast cancer cells.
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    Mechanotransduction of mitochondrial AMPK and its distinct role in flow-induced breast cancer cell migration
    (Elsevier, 2019-06) Steele, Hannah E.; Guo, Yunxia; Li, Bai-Yan; Na, Sungsoo; Biomedical Engineering, School of Engineering and Technology
    The biophysical microenvironment of the tumor site has significant impact on breast cancer progression and metastasis. The importance of altered mechanotransduction in cancerous tissue has been documented, yet its role in the regulation of cellular metabolism and the potential link between cellular energy and cell migration remain poorly understood. In this study, we investigated the role of mechanotransduction in AMP-activated protein kinase (AMPK) activation in breast cancer cells in response to interstitial fluid flow (IFF). Additionally, we explored the involvement of AMPK in breast cancer cell migration. IFF was applied to the 3D cell-matrix construct. The subcellular signaling activity of Src, FAK, and AMPK was visualized in real-time using fluorescent resonance energy transfer (FRET). We observed that breast cancer cells (MDA-MB-231) are more sensitive to IFF than normal epithelial cells (MCF-10A). AMPK was activated at the mitochondria of MDA-MB-231 cells by IFF, but not in other subcellular compartments (i.e., cytosol, plasma membrane, and nucleus). The inhibition of FAK or Src abolished flow-induced AMPK activation in the mitochondria of MDA-MB-231 cells. We also observed that global AMPK activation reduced MDA-MB-231 cell migration. Interestingly, specific AMPK inhibition in the mitochondria reduced cell migration and blocked flow-induced cell migration. Our results suggest the linkage of FAK/Src and mitochondria-specific AMPK in mechanotransduction and the differential role of AMPK in breast cancer cell migration depending on its subcellular compartment-specific activation.
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    Mechanotransduction of subcellular AMPK and its role in breast cancer cell migration
    (2018-04) Steele, Hannah E.; Na, Sungsoo
    The biophysical microenvironment of the tumor site has significant impact on breast cancer progression and metastasis. The importance of altered mechanotransduction in cancerous tissue through the integrin-mediated signaling axis has been documented, yet its role in the regulation of cellular metabolism and the potential link between cellular energy and cell migration remain poorly understood. In this study, we investigated the role of mechanotransduction (via Src and FAK) in AMP-activated protein kinase (AMPK) activation in breast cancer cells in response to interstitial fluid flow. Additionally, we explored the involvement of AMPK in breast cancer cell migration. An in-vitro three-dimensional (3D) cell culture model utilizing collagen-Matrigel matrices was used. Interstitial fluid flow was applied to the 3D cell-matrix construct inside a flow chamber. The sub-cellular signaling activity of Src, FAK, and AMPK was visualized in real-time using fluorescent resonance energy transfer (FRET). We observed that breast cancer cells (MDA-MB-231) are more sensitive to interstitial fluid flow than normal epithelial cells (MCF-10A) in the regulation of FAK and Src. AMPK was activated in the mitochondria of MDA-MB-231 cells by interstitial fluid flow, but not in other subcellular domains (i.e., cytosol, plasma membrane, and nucleus). Subcellular AMPK in MCF-10A cells did not respond to interstitial fluid flow. The inhibition of FAK or Src abolished flow-induced AMPK activation in the mitochondria of MDA-MB-231 cells. We also observed that global AMPK activation reduced MDA-MB-231 cell migration. Interestingly, specific AMPK inhibition in the mitochondria reduced cell migration and blocked interstitial fluid flow-induced cell migration. Our results suggest the linkage of FAK/Src and mitochondria-specific AMPK in mechanotransduction and the dual role of AMPK in breast cancer cell migration depending on its subcellular activation. Therefore, subcellular AMPK activation may play an important and distinct role in cancer invasion and progression.
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    Metformin attenuates an increase of calcium-dependent and ubiquitin-proteasome markers in unloaded muscle
    (American Physiological Society, 2022) Belova, Svetlana P.; Zaripova, Ksenia; Sharlo, Kristina; Kostrominova, Tatiana Y.; Shenkman, Boris S.; Nemirovskaya, Tatiana L.; Anatomy, Cell Biology and Physiology, School of Medicine
    Current study tested a hypothesis that during skeletal muscle unloading, calcium-dependent signaling pathways, markers of protein synthesis, and expression of E3 ubiquitin ligases can be regulated by metformin. Thirty-two male Wistar rats were randomly assigned into one of four groups: nontreated control (3C), control rats treated with metformin (3CM), 3 days of unloading/hindlimb suspension with placebo (3HS), and 3 days of unloading treated with metformin (3HSM). In soleus muscle of HS group level of phospho-AMP-activated protein kinase (p-AMPK) was decreased by 46% while ATP content was increased by 49% when compared with the control group. There was an increase of the level of phospho-CaMK II (483%) and an upregulation of Calcineurin (CaN), SERCA2a, and Calpain-1 mRNA expression (87%, 41%, and 62%, respectively, P < 0.05) in the HS group relative to the control. HS group also had increased mRNA expression of MuRF1, MAFbx, and ubiquitin (167%, 146%, and 191%, respectively, P < 0.05) when compared with the control soleus muscle. Metformin treatment impeded unloading-induced changes in soleus muscle. In conclusion, metformin treatment during 3 days of soleus muscle unloading: 1) prevented the decrease of p-AMPK and increase of ATP content; 2) affected regulation of calcium-dependent signaling pathways via level of CaMK II phosphorylation or CaMK II, CaN, SERCA2a, and Calpain-1 mRNA expression; 3) attenuated an increase in the expression of critical markers of ubiquitin-proteasome pathways MuRF1, MAFbx, and ubiquitin while not affecting the unloading-induced increase of ULK-1 marker of autophagic/lysosomal pathway. NEW & NOTEWORTHY: Current study for the first time tested the hypothesis that during 3 days of soleus muscle unloading, calcium-dependent signaling pathways, markers of protein synthesis, and the expression of E3 ubiquitin ligases can be regulated by metformin. Treatment with metformin during unloading: prevented the decrease of p-AMPK and increase of ATP content, affected regulation of calcium-dependent signaling pathways, and attenuated an increase of critical markers of ubiquitin-proteasome pathways. Nevertheless, metformin treatment has not prevented soleus muscle atrophy.
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    Metformin Corrects Abnormal Circadian Rhythm and Kir4.1 Channels in Diabetes
    (The Association for Research in Vision and Ophthalmology, 2020-06-22) Alex, Alpha; Luo, Qianyi; Mathew, Deepa; Di, Rong; Bhatwadekar, Ashay D.; Ophthalmology, School of Medicine
    Purpose Diabetic retinopathy (DR) is a leading cause of visual impairment. Müller cells in DR are dysfunctional due to downregulation of the inwardly rectifying potassium channel Kir4.1. Metformin, a commonly used oral antidiabetic drug, is known to elicit its action through 5′ adenosine monophosphate-activated protein kinase (AMPK), a cellular metabolic regulator; however, its effect on Kir4.1 channels is unknown. For this study, we hypothesized that metformin treatment would correct circadian rhythm disruption and Kir4.1 channel dysfunction in db/db mice. Methods Metformin was given orally to db/db mice. Wheel-running activity, retinal levels of Kir4.1, and AMPK phosphorylation were determined at study termination. In parallel, rat retinal Müller cell line (rMC-1) cells were treated using metformin and 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) to assess the effect of AMPK activation on the Kir4.1 channel. Results The wheel-running activity of the db/db mice was improved following the metformin treatment. The Kir4.1 level in Müller cells was corrected after metformin treatment. Metformin treatment led to an upregulation of clock regulatory genes such as melanopsin (Opn4) and aralkylamine N-acetyltransferase (Aanat). In rMC-1 cells, AMPK activation via AICAR and metformin resulted in increased Kir4.1 and intermediate core clock component Bmal-1 protein expression. The silencing of Prkaa1 (gene for AMPKα1) led to decreased Kir4.1 and Bmal-1 protein expression. Conclusions Our findings demonstrate that metformin corrects abnormal circadian rhythm and Kir4.1 channels in db/db mouse a model of type 2 diabetes. Metformin could represent a critical pharmacological agent for preventing Müller cell dysfunction observed in human DR.
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    Osteocyte signaling and its effects on the activities of osteoblasts and breast cancer cells
    (2021-05) Ahandoust, Sina; Na, Sungsoo; Yokota, Hiroki; Li, Jiliang
    Bone is a common location for breast cancer cell metastasis, and progression of tumor in bone can lead to bone loss and affect human health. Osteocytes have important roles in bone homeostasis and osteogenesis, and their interaction with metastasized cancer cells are known to affect progression of metastasized tumor. However, the potential role of metabolic signaling and actin- cytoskeleton-associated moesin in the interaction of osteocytes and tumor cells remain poorly understood. In this study, we first examined the roles of metabolic signaling, specifically global AMPK modulators and mitochondria-specific AMPK inhibitor (Mito-AIP), as well as mechanical force in beta catenin signaling through interaction between osteocytes and precursor osteoblasts as well as osteocytes and breast cancer cells. We also evaluated the role of metabolic signaling in Rho GTPases including RhoA, Rac1 and Cdc42. We observed that AMPK activator (A769662) and Mito-AMPK stimulated beta catenin translocation to the nucleus, indicating the activation of Wnt signaling, while Mito-AIP did not significantly affect beta catenin activation in osteoblasts. We also observed that osteocyte conditioned medium (CM) treated with Mito-AIP substantially increased beta catenin signaling in osteoblasts, while decreasing beta catenin signaling in breast cancer cells. CM of osteocytes treated with fluid flow increased beta catenin signaling in breast cancer cells. A769662 and Mito-AIP also decreased the activities of RhoA, Rac1, and Cdc42 in cancer cells which are known to regulate cancer cell migration. Additionally, we evaluated the roles of intracellular and extracellular moesin (MSN) protein in well-established oncogenic signaling proteins, such as FAK, Src, and RhoA as well beta catenin signaling. Constitutively active MSN (MSN+) significantly increased FAK and Src activities in cancer cells, but decreased the activity of RhoA. Surprisingly, CM of mesenchymal stem cells treated with MSN+ decreased the activities of FAK, Src, and RhoA, suggesting the inhibitory role of extracellular MSN in tumor-promoting signaling. Our results suggest the distinct role of AMPK signaling, specifically at mitochondria of osteocytes, in the activities of beta-catenin signaling in osteoblasts and breast cancer cells and the distinct role of intracellular and extracellular MSN in these two types of cell.