Himangshu Sonowal

Inhibition of actin polymerization decreases osteogeneic differentiation of mesenchymal stem cells through p38 MAPK pathway

BackgroundMesenchymal Stem Cells (MSC) are important candidates for therapeutic applications due to their ex vivo proliferation and differentiation capacity. MSC differentiation is controlled by both intrinsic and extrinsic factors and actin cytoskeleton plays a major role in the event. In the current study, we tried to understand the initial molecular mechanisms and pathways that regulate the differentiation of MSC into osteocytes or adipocytes.ResultsWe observed that actin modification was important during differentiation and differentially regulated during adipogenesis and osteogenesis. Initial disruption of actin polymerization reduced further differentiation of MSC into osteocytes and osteogenic differentiation was accompanied by increase in ERK1/2 and p38 MAPK phosphorylation. However, only p38 MAPK phosphorylation was down regulated upon inhibition of actin polymerization which as accompanied by decreased CD49E expression.ConclusionTaken together, our results show that actin modification is a pre-requisite for MSC differentiation into osteocytes and adipocytes and osteogenic differentiation is regulated through p38 MAPK phosphorylation. Thus by modifying their cytoskeleton the differentiation potential of MSC could be controlled which might have important implications for tissue repair and regeneration.

Aldose reductase inhibitor increases doxorubicin-sensitivity of colon cancer cells and decreases cardiotoxicity

Anthracycline drugs such as doxorubicin (DOX) and daunorubicin remain some of the most active wide-spectrum and cost-effective drugs in cancer therapy. However, colorectal cancer (CRC) cells are inherently resistant to anthracyclines which at higher doses cause cardiotoxicity. Our recent studies indicate that aldose reductase (AR) inhibitors such as fidarestat inhibit CRC growth in vitro and in vivo. Here, we show that treatment of CRC cells with fidarestat increases the efficacy of DOX-induced death in HT-29 and SW480 cells and in nude mice xenografts. AR inhibition also results in higher intracellular accumulation of DOX and decreases the expression of drug transporter proteins MDR1, MRP1, and ABCG2. Further, fidarestat also inhibits DOX–induced increase in troponin-I and various inflammatory markers in the serum and heart and restores cardiac function in mice. These results suggest that fidarestat could be used as adjuvant therapy to enhance DOX sensitivity of CRC cells and to reduce DOX-associated cardiotoxicity.

Aspalatone Prevents VEGF-Induced Lipid Peroxidation, Migration, Tube Formation, and Dysfunction of Human Aortic Endothelial Cells

Although aspalatone (acetylsalicylic acid maltol ester) is recognized as an antithrombotic agent with antioxidative and antiplatelet potential; its efficacy in preventing endothelial dysfunction is not known. In this study, we examined the antiangiogenic, antioxidative, and anti-inflammatory effect of aspalatone in human aortic endothelial cells (HAECs). Specifically, the effect of aspalatone on VEGF-induced HAECs growth, migration, tube formation, and levels of lipid peroxidation-derived malondialdehyde (MDA) was examined. Our results indicate that the treatment of HAECs with aspalatone decreased VEGF-induced cell migration, tube formation, and levels of MDA. Aspalatone also inhibited VEGF-induced decrease in the expression of eNOS and increase in the expression of iNOS, ICAM-1, and VCAM-1. Aspalatone also prevented the VEGF-induced adhesion of monocytes to endothelial cells. Furthermore, aspalatone also prevented VEGF-induced release of inflammatory markers such as Angiopoietin-2, Leptin, EGF, G-CSF, HB-EGF, and HGF in HAECs. Thus, our results suggest that aspalatone could be used to prevent endothelial dysfunction, an important process in the pathophysiology of cardiovascular diseases.

Aldose reductase mediates NLRP3 inflammasome-initiated innate immune response in hyperglycemia-induced thp1 monocytes and male mice

&NA; Despite recent studies that show oxidative stress‐generated reactive oxygen species (ROS) regulate NOD‐like receptor family pyrin domain containing 3 (NLRP3) inflammasome‐mediated innate immune response in various diabetic complications, the mechanism by which ROS activate innate immune response is not well understood. We have shown previously that aldose reductase (AR), besides reducing glucose, reduces lipid aldehydes and their glutathione conjugates and participates in various oxidative stress‐induced inflammatory pathways. To understand the role of AR in ROS‐induced innate immune response, we have investigated the mechanism(s) by which AR activates hyperglycemia‐induced NLRP3 inflammsome‐initiated innate immune response in Thp1 monocytes and in streptozotocin (STZ)‐induced diabetic mice. In Thp1 monocytes, inhibition or ablation of AR prevented high‐glucose‐induced activation of NLRP3 inflammasome and caspase‐1 and release of the innate immune cytokines interleukin (IL)‐1&bgr; and IL‐18. AR inhibition in Thp1 cells also prevented the high‐glucose‐induced generation of ROS, influx of Ca2+, efflux of K+, and activation of Lyn, Syk, and PI3K. Furthermore, the AR inhibitor fidarestat prevented the expression of NLRP inflammasome components in STZ‐induced diabetic mouse heart and aorta, and also prevented the release of various cytokines in the serum. Collectively, our data suggest that AR regulates hyperglycemia‐induced NLRP3 inflammasome‐mediated innate immune response by altering the ROS/Lyn/Syk/PI3K/Ca2+/K+ signals.

Aldose reductase inhibitor, fidarestat prevents doxorubicin-induced endothelial cell death and dysfunction

Graphical abstract Figure. No Caption available. Abstract Despite doxorubicin (Dox) being one of the most widely used chemotherapy agents for breast, blood and lung cancers, its use in colon cancer is limited due to increased drug resistance and severe cardiotoxic side effects that increase mortality associated with its use at high doses. Therefore, better adjuvant therapies are warranted to improve the chemotherapeutic efficacy and to decrease cardiotoxicity. We have recently shown that aldose reductase inhibitor, fidarestat, increases the Dox‐induced colon cancer cell death and reduces cardiomyopathy. However, the efficacy of fidarestat in the prevention of Dox‐induced endothelial dysfunction, a pathological event critical to cardiovascular complications, is not known. Here, we have examined the effect of fidarestat on Dox‐induced endothelial cell toxicity and dysfunction in vitro and in vivo. Incubation of human umbilical vein endothelial cells (HUVECs) with Dox significantly increased the endothelial cell death, and pre‐treatment of fidarestat prevented it. Further, fidarestat prevented the Dox‐induced oxidative stress, formation of reactive oxygen species (ROS) and activation of Caspase‐3 in HUVECs. Fidarestat also prevented Dox‐induced monocyte adhesion to HUVECs and expression of ICAM‐1 and VCAM‐1. Fidarestat pre‐treatment to HUVECs restored the Dox‐induced decrease in the Nitric Oxide (NO)‐levels and eNOS expression. Treatment of HUVECs with Dox caused a significant increase in the activation of NF‐&kgr;B and expression of various inflammatory cytokines and chemokines which were prevented by fidarestat pre‐treatment. Most importantly, fidarestat prevented the Dox‐induced mouse cardiac cell hypertrophy and expression of eNOS, iNOS, and 3‐Nitrotyrosine in the aorta tissues. Further, fidarestat blunted the Dox‐induced expression of various inflammatory cytokines and chemokines in vivo. Thus, our results suggest that by preventing Dox‐induced endothelial cytotoxicity and dysfunction, AR inhibitors could avert cardiotoxicity associated with anthracycline chemotherapy.

Vialinin A, an Edible Mushroom-Derived p-Terphenyl Antioxidant, Prevents VEGF-Induced Neovascularization In Vitro and In Vivo

Increased side toxicities and development of drug resistance are the major concern for the cancer chemotherapy using synthetic drugs. Therefore, identification of novel natural antioxidants with potential therapeutic efficacies is important. In the present study, we have examined how the antioxidant and anti-inflammatory activities of vialinin A, a p-terphenyl compound derived from Chinese edible mushroom T. terrestris and T. vialis, prevents human umbilical vascular endothelial cell (HUVEC) neovascularization in vitro and in vivo models. Pretreatment of HUVECs with vialinin A prevents vascular endothelial growth factor- (VEGF) induced HUVEC cell growth in a dose-dependent manner. Further, vialinin A also inhibits VEGF-induced migration as well as tube formation of HUVECs. Treatment of HUVECs prevents VEGF-induced generation of reactive oxygen species (ROS) and malondialdehyde (MDA) and also inhibits VEGF-induced NF-κB nuclear translocation as well as DNA-binding activity. The VEGF-induced release of various angiogenic cytokines and chemokines in HUVECs was also significantly blunted by vialinin A. Most importantly, in a mouse model of Matrigel plug assay, vialinin A prevents the formation of new blood vessels and the expression of CD31 and vWF. Thus, our results indicate a novel role of vialinin A in the prevention of neovascularization and suggest that anticancer effects of vialinin A could be mediated through its potent antioxidant and antiangiogenic properties.

Abstract 1682: Combined treatment of aldose reductase inhibitor, fidarestat, and anti-PD1 antibodies synergistically prevents T cell- and NK cell-mediated colon cancer growth in murine models

Immune checkpoint inhibitors such as anti-programmed death 1 (PD1) and anti-PD-L1 monoclonal antibodies though have great potential for cancer immunotherapy, the acquired drug resistance and unwanted immune side effects limit their use. However, the combination therapy using conventional drugs and immune checkpoint inhibitors not only synergistically increase anti-tumor responses but it also reduces immune-associated adverse events. In our earlier studies, we have shown that inhibition of polyol pathway enzyme, aldose reductase (AR) prevents colon cancer growth and metastasis in culture as well as in mouse models. In this study, we investigated whether fidarestat, a potent AR inhibitor that has already gone through Phase-III clinical trial for diabetic neuropathy and found to safe for human use, can synergize with anti PD-1 to increase antitumor activity and decrease immune-associated adverse effects. Mice bearing syngeneic CT26 colon carcinoma tumors were treated with anti PD-1 alone or in combination with fidarestat, and the tumor growth was monitored. Our results indicate that fidarestat in combination with anti-PD1 synergistically increases tumor regression within 28 days of treatment. Further, a combination of fidarestat and anti-PD1 significantly increased the CD8+ T cells and natural killer (NK) cells and decreased the Myeloid-derived suppressor cells (MDSCs) at tumor milieu. Fidarestat plus anti-PD1 also increased CD8+ T cell and NK cell effector functions as determined by IFNg and granzyme B. Systemically, combination therapy also increased the number of CD8+ T cells and NK cells and decreased MDSCs in the mouse spleens. Further, the effector functions of CD8+ T cells and NK cells were increased with the combination treatment in mice spleens. Moreover, the levels of various inflammatory cytokines and chemokines in the serum and liver were significantly less in fidarestat plus anti-PD1 -treated mice as compared to anti-PD1 alone -treated mice indicating that fidarestat rescues immune-mediated adverse events in various organs. Thus, our results suggest that by regulating the number as well as effector function of CD8+ T cells, NK cells and MDSCs, AR inhibitor in combination with anti-PD1 would not only regress the tumor growth but would also provide a long-lasting specific immunity against colon cancer, which would decrease the chances of tumor recurrence. Citation Format: Ashish Saxena, Himangshu Sonowal, Satish K. Srivastava, Kota V. Ramana. Combined treatment of aldose reductase inhibitor, fidarestat, and anti-PD1 antibodies synergistically prevents T cell- and NK cell-mediated colon cancer growth in murine models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1682.

Didymin prevents hyperglycemia-induced human umbilical endothelial cells dysfunction and death

Graphical abstract Figure. No Caption available. ABSTRACT Although didymin, a flavonoid‐O‐glycosides compound naturally found in the citrus fruits, has been reported to be a potent anticancer agent in the prevention of various cancers, its role in the prevention of cardiovascular complications is unclear. Most importantly, its effect in the prevention of endothelial dysfunction, a pathological process involved in the atherogenesis, is unknown. We have examined the efficacy of didymin in preventing the high glucose (HG; 25 mM)‐induced human umbilical vein endothelial cells (HUVECs) dysfunction. Our results indicate that incubation of HUVECs with HG resulted in the loss of cell viability, and pre‐incubation of didymin prevented it. Further, didymin prevented the HG‐induced generation of reactive oxygen species (ROS) as well as lipid peroxidation product, malondialdehyde. Pretreatment of HUVECs with didymin also prevented the HG‐induced decrease in eNOS and increase in iNOS expressions. Further, didymin prevented the HG‐induced monocytes cell adhesion to endothelial cells, expressions of ICAM‐1 and VCAM‐1 and activation of NF‐&kgr;B. Didymin also prevented the release of various inflammatory cytokines and chemokines in HG‐treated HUVECs. In conclusion, our results demonstrate that didymin with its anti‐oxidative and anti‐inflammatory actions prevents hyperglycemia‐induced endothelial dysfunction and death. Thus, it could be developed as a potential natural therapeutic agent for the prevention of cardiovascular complications in diabetes.

Aldose Reductase Inhibitor Protects against Hyperglycemic Stress by Activating Nrf2-Dependent Antioxidant Proteins

We have shown earlier that pretreatment of cultured cells with aldose reductase (AR) inhibitors prevents hyperglycemia-induced mitogenic and proinflammatory responses. However, the effects of AR inhibitors on Nrf2-mediated anti-inflammatory responses have not been elucidated yet. We have investigated how AR inhibitor fidarestat protects high glucose- (HG-) induced cell viability changes by increasing the expression of Nrf2 and its dependent phase II antioxidant enzymes. Fidarestat pretreatment prevents HG (25 mM)-induced Thp1 monocyte viability. Further, treatment of Thp1 monocytes with fidarestat caused a time-dependent increase in the expression as well as the DNA-binding activity of Nrf2. In addition, fidarestat augmented the HG-induced Nrf2 expression and activity and also upregulated the expression of Nrf2-dependent proteins such as hemeoxygenase-1 (HO1) and NQO1 in Thp1 cells. Similarly, treatment with AR inhibitor also induced the expression of Nrf2 and HO1 in STZ-induced diabetic mice heart and kidney tissues. Further, AR inhibition increased the HG-induced expression of antioxidant enzymes such as SOD and catalase and activation of AMPK-α1 in Thp1 cells. Our results thus suggest that pretreatment with AR inhibitor prepares the monocytes against hyperglycemic stress by overexpressing the Nrf2-dependent antioxidative proteins.