Browsing by Author "Khanna, Savita"
Now showing 1 - 10 of 12
Results Per Page
Sort Options
Item Cutaneous Epithelial to Mesenchymal Transition Activator ZEB1 Regulates Wound Angiogenesis and Closure in a Glycemic Status–Dependent Manner(American Diabetes Association, 2019-11) Singh, Kanhaiya; Sinha, Mithun; Pal, Durba; Tabasum, Saba; Gnyawali, Surya C.; Khona, Dolly; Sarkar, Subendu; Mohanty, Sujit K.; Soto-Gonzalez, Fidel; Khanna, Savita; Roy, Sashwati; Sen, Chandan K.; Surgery, School of MedicineEpithelial to mesenchymal transition (EMT) and wound vascularization are two critical interrelated processes that enable cutaneous wound healing. Zinc finger E-box binding homeobox 1 (ZEB1), primarily studied in the context of tumor biology, is a potent EMT activator. ZEB1 is also known to contribute to endothelial cell survival as well as stimulate tumor angiogenesis. The role of ZEB1 in cutaneous wounds was assessed using Zeb1+/− mice, as Zeb1−/− mice are not viable. Quantitative stable isotope labeling by amino acids in cell culture (SILAC) proteomics was used to elucidate the effect of elevated ZEB1, as noted during hyperglycemia. Under different glycemic conditions, ZEB1 binding to E-cadherin promoter was investigated using chromatin immunoprecipitation. Cutaneous wounding resulted in loss of epithelial marker E-cadherin with concomitant gain of ZEB1. The dominant proteins downregulated after ZEB1 overexpression functionally represented adherens junction pathway. Zeb1+/− mice exhibited compromised wound closure complicated by defective EMT and poor wound angiogenesis. Under hyperglycemic conditions, ZEB1 lost its ability to bind E-cadherin promoter. Keratinocyte E-cadherin, thus upregulated, resisted EMT required for wound healing. Diabetic wound healing was improved in ZEB+/− as well as in db/db mice subjected to ZEB1 knockdown. This work recognizes ZEB1 as a key regulator of cutaneous wound healing that is of particular relevance to diabetic wound complication.Item Electroceutical Fabric Lowers Zeta Potential and Eradicates Coronavirus Infectivity upon Contact(2020-05-15) Sen, Abhishek; Khona, Dolly; Ghatak, Subhadip; Gopalakrishnan, Vinoj; Cornetta, Kenneth; Roy, Sashwati; Khanna, Savita; Sen, Chandan; Indiana Center for Regenerative Medicine and Engineering, IU School of MedicineCoronavirus with intact infectivity attached to PPE surfaces pose significant threat to the spread of COVID-19. We tested the hypothesis that an electroceutical fabric, generating weak potential difference of 0.5V, disrupts the infectivity of coronavirus upon contact by destabilizing the electrokinetic properties of the virion. Respiratory coronavirus particles (105) were placed in direct contact with the fabric for 1 or 5 minutes. Viral particles (2.5-4x104) were recovered from the fabric. Following one minute of contact, zeta potential of the coronavirus was significantly lowered indicating destabilization of its electrokinetic properties. Size-distribution plot showed appearance of aggregation of the virus. Testing of the cytopathic effects of the virus showed eradication of infectivity as quantitatively assessed by PI-calcein and MTT cell viability tests. This work provides the rationale to consider the studied electroceutical fabric, or other materials with comparable property, as material of choice for the development of PPE in the fight against COVID-19.Item Electroceutical fabric lowers zeta potential and eradicates coronavirus infectivity upon contact(Nature, 2021-12) Ghatak, Subhadip; Khona, Dolly K.; Sen, Abhishek; Huang, Kaixiang; Jagdale, Gargi; Singh, Kanhaiya; Gopalakrishnan, Vinoj; Cornetta, Kenneth G.; Roy, Sashwati; Khanna, Savita; Baker, Lane A.; Sen, Chandan K.; Medical and Molecular Genetics, School of MedicineCoronavirus with intact infectivity attached to PPE surfaces pose significant threat to the spread of COVID-19. We tested the hypothesis that an electroceutical fabric, generating weak potential difference of 0.5 V, disrupts the infectivity of coronavirus upon contact by destabilizing the electrokinetic properties of the virion. Porcine respiratory coronavirus AR310 particles (105) were placed in direct contact with the fabric for 1 or 5 min. Following one minute of contact, zeta potential of the porcine coronavirus was significantly lowered indicating destabilization of its electrokinetic properties. Size-distribution plot showed appearance of aggregation of the virus. Testing of the cytopathic effects of the virus showed eradication of infectivity as quantitatively assessed by PI-calcein and MTT cell viability tests. This work provides the rationale to consider the studied electroceutical fabric, or other materials with comparable property, as material of choice for the development of PPE in the fight against COVID-19.Item Mitochondria as Target for Tumor Management of Hemangioendothelioma(Liebert, 2020) Gordillo, Gayle M.; Biswas, Ayan; Singh, Kanhaiya; Sen, Abhishek; Guda, Poornachander R.; Miller, Caroline; Pan, Xueliang; Khanna, Savita; Cadenas, Enrique; Sen, Chandan K.; Surgery, School of MedicineAims: Hemangioendothelioma (HE) may be benign or malignant. Mouse hemangioendothelioma endothelial (EOMA) cells are validated to study mechanisms in HE. This work demonstrates that EOMA cells heavily rely on mitochondria to thrive. Thus, a combination therapy, including weak X-ray therapy (XRT, 0.5 Gy) and a standardized natural berry extract (NBE) was tested. This NBE is known to be effective in managing experimental HE and has been awarded with the Food and Drug Administration Investigational New Drug (FDA-IND) number 140318 for clinical studies on infantile hemangioma. Results: NBE treatment alone selectively attenuated basal oxygen consumption rate of EOMA cells. NBE specifically sensitized EOMA, but not murine aortic endothelial cells to XRT-dependent attenuation of mitochondrial respiration and adenosine triphosphate (ATP) production. Combination treatment, selectively and potently, influenced mitochondrial dynamics in EOMA cells such that fission was augmented. This was achieved by lowering of mitochondrial sirtuin 3 (SIRT3) causing increased phosphorylation of AMP-activated protein kinase (AMPK). A key role of SIRT3 in loss of EOMA cell viability caused by the combination therapy was evident when pyrroloquinoline quinone, an inducer of SIRT3, pretreatment rescued these cells. Innovation and Conclusion: Mitochondria-targeting NBE significantly extended survival of HE-affected mice. The beneficial effect of NBE in combination with weak X-ray therapy was, however, far more potent with threefold increase in murine survival. The observation that safe natural products may target tumor cell mitochondria and sharply lower radiation dosage required for tumor management warrants clinical testing.Item A Modified Collagen Dressing Induces Transition of Inflammatory to Reparative Phenotype of Wound Macrophages(Nature Research, 2019-10-04) Das, Amitava; Abas, Motaz; Biswas, Nirupam; Banerjee, Pradipta; Ghosh, Nandini; Rawat, Atul; Khanna, Savita; Roy, Sashwati; Sen, Chandan K.; Surgery, School of MedicineCollagen containing wound-care dressings are extensively used. However, the mechanism of action of these dressings remain unclear. Earlier studies utilizing a modified collagen gel (MCG) dressing demonstrated improved vascularization of ischemic wounds and better healing outcomes. Wound macrophages are pivotal in facilitating wound angiogenesis and timely healing. The current study was designed to investigate the effect of MCG on wound macrophage phenotype and function. MCG augmented recruitment of macrophage at the wound-site, attenuated pro-inflammatory and promoted anti-inflammatory macrophage polarization. Additionally, MCG increased anti-inflammatory IL-10, IL-4 and pro-angiogenic VEGF production, indicating a direct role of MCG in resolving wound inflammation and improving angiogenesis. At the wound-site, impairment in clearance of apoptotic cell bioburden enables chronic inflammation. Engulfment of apoptotic cells by macrophages (efferocytosis) resolves inflammation via a miR-21-PDCD4-IL-10 pathway. MCG-treated wound macrophages exhibited a significantly bolstered efferocytosis index. Such favorable outcome significantly induced miR-21 expression. MCG-mediated IL-10 production was dampened under conditions of miR-21 knockdown pointing towards miR-21 as a causative factor. Pharmacological inhibition of JNK attenuated IL-10 production by MCG, implicating miR-21-JNK pathway in MCG-mediated IL-10 production by macrophages. This work provides direct evidence demonstrating that a collagen-based wound-care dressing may influence wound macrophage function and therefore modify wound inflammation outcomes.Item Myo-Inositol in Fermented Sugar Matrix Improves Human Macrophage Function(Wiley, 2022) Ghosh, Nandini; Das, Amitava; Biswas, Nirupam; Mahajan, Sanskruti P.; Madeshiya, Amit K.; Khanna, Savita; Sen, Chandan K.; Roy, Sashwati; Surgery, School of MedicineScope Reactive oxygen species production by innate immune cells plays a central role in host defense against invading pathogens at wound-site. A weakened hos-defense results in persistent infection leading to wound chronicity. Fermented Papaya Preparation (FPP), a complex sugar matrix, bolstered respiratory burst activity and improved wound healing outcomes in chronic wound patients. The objective of the current study was to identify underlying molecular factor/s responsible for augmenting macrophage host defense mechanisms following FPP supplementation. Methods and results In depth LC-MS/MS analysis of cells supplemented with FPP led to identification of myo-inositol as a key determinant of FPP activity towards improving macrophage function. Myo-inositol, in quantities that is present in FPP, significantly improved macrophage respiratory burst and phagocytosis via de novo synthesis pathway of ISYNA1. Additionally, myo-inositol transporters, HMIT and SMIT1, played a significant role in such activity. Blocking these pathways using siRNA attenuated FPP-induced improved macrophage host defense activities. FPP supplementation emerges as a novel approach to increase intracellular myo-inositol levels. Such supplementation also modified wound microenvironment in chronic wound patients to augment myo-inositol levels in wound fluid. Conclusion These observations indicate that myo-inositol in FPP influences multiple aspects of macrophage function critical for host defense against invading pathogens.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 Neurogenic Tissue Nanotransfection in the Management of Cutaneous Diabetic Polyneuropathy(Elsevier, 2020-08) Roy, Sashwati; Sen, Chandan K.; Ghatak, Subhadip; Higuita-Castro, Natalia; Palakurti, Ravichand; Nalluri, Nagajyothi; Clark, Andrew; Stewart, Richard; Gallego-Perez, Daniel; Prater, Daniel N.; Khanna, Savita; Surgery, School of MedicineThis work rests on our recent report on the successful use of tissue nanotransfection (TNT) delivery of Ascl1, Brn2, and Myt1l (TNTABM) to directly convert skin fibroblasts into electrophysiologically active induced neuronal cells (iN) in vivo. Here we report that in addition to successful neurogenic conversion of cells, TNTABM caused neurotrophic enrichment of the skin stroma. Thus, we asked whether such neurotrophic milieu of the skin can be leveraged to rescue pre-existing nerve fibers under chronic diabetic conditions. Topical cutaneous TNTABM caused elevation of endogenous NGF and other co-regulated neurotrophic factors such as Nt3. TNTABM spared loss of cutaneous PGP9.5+ mature nerve fibers in db/db diabetic mice. This is the first study demonstrating that under conditions of in vivo reprogramming, changes in the tissue microenvironment can be leveraged for therapeutic purposes such as the rescue of pre-existing nerve fibers from its predictable path of loss under conditions of diabetes.Item Novel Bacterial Diversity and Fragmented eDNA Identified in Hyperbiofilm-Forming Pseudomonas aeruginosa Rugose Small Colony Variant(Elsevier, 2020-02-21) Deng, Binbin; Ghatak, Subhadip; Sarkar, Subendu; Singh, Kanhaiya; Ghatak, Piya Das; Mathew-Steiner, Shomita S.; Roy, Sashwati; Khanna, Savita; Wozniak, Daniel J.; McComb, David W.; Sen, Chandan K.; Surgery, School of MedicinePseudomonas aeruginosa biofilms represent a major threat to health care. Rugose small colony variants (RSCV) of P. aeruginosa, isolated from chronic infections, display hyperbiofilm phenotype. RSCV biofilms are highly resistant to antibiotics and host defenses. This work shows that RSCV biofilm aggregates consist of two distinct bacterial subpopulations that are uniquely organized displaying contrasting physiological characteristics. Compared with that of PAO1, the extracellular polymeric substance of RSCV PAO1ΔwspF biofilms presented unique ultrastructural characteristics. Unlike PAO1, PAO1ΔwspF released fragmented extracellular DNA (eDNA) from live cells. Fragmented eDNA, thus released, was responsible for resistance of PAO1ΔwspF biofilm to disruption by DNaseI. When added to PAO1, such fragmented eDNA enhanced biofilm formation. Disruption of PAO1ΔwspF biofilm was achieved by aurine tricarboxylic acid, an inhibitor of DNA-protein interaction. This work provides critical novel insights into the contrasting structural and functional characteristics of a hyperbiofilm-forming clinical bacterial variant relative to its own wild-type strain.Item Skin Transcriptome of Middle-Aged Women Supplemented With Natural Herbo-mineral Shilajit Shows Induction of Microvascular and Extracellular Matrix Mechanisms(Taylor & Francis, 2019-06-04) Das, Amitava; Masry, Mohamed S. El; Gnyawali, Surya C.; Ghatak, Subhadip; Singh, Kanhaiya; Stewart, Richard; Lewis, Madeline; Saha, Abhijoy; Gordillo, Gayle; Khanna, Savita; Surgery, School of MedicineObjective: Shilajit is a pale-brown to blackish-brown organic mineral substance available from Himalayan rocks. We demonstrated that in type I obese humans, shilajit supplementation significantly upregulated extracellular matrix (ECM)–related genes in the skeletal muscle. Such an effect was highly synergistic with exercise. The present study (clinicaltrials.gov ) aimed to evaluate the effects of shilajit supplementation on skin gene expression profile and microperfusion in healthy adult females. Methods: The study design comprised six total study visits including a baseline visit (V1) and a final 14-week visit (V6) following oral shilajit supplementation (125 or 250 mg bid). A skin biopsy of the left inner upper arm of each subject was collected at visit 2 and visit 6 for gene expression profiling using Affymetrix Clariom™ D Assay. Skin perfusion was determined by MATLAB processing of dermascopic images. Transcriptome data were normalized and subjected to statistical analysis. The differentially regulated genes were subjected to Ingenuity Pathway Analysis (IPA®). The expression of the differentially regulated genes identified by IPA® were verified using real-time polymerasechain reaction (RT-PCR). Results: Supplementation with shilajit for 14 weeks was not associated with any reported adverse effect within this period. At a higher dose (250 mg bid), shilajit improved skin perfusion when compared to baseline or the placebo. Pathway analysis identified shilajit-inducible genes relevant to endothelial cell migration, growth of blood vessels, and ECM which were validated by quantitative real-time polymerasechain reaction (RT-PCR) analysis. Conclusions: This work provides maiden evidence demonstrating that oral shilajit supplementation in adult healthy women induced genes relevant to endothelial cell migration and growth of blood vessels. Shilajit supplementation improved skin microperfusion.