Publications by authors named "Dinesh Upadhya"

27 Publications

  • Page 1 of 1

Editorial: Endocrine Modulators of Neurological Processes: Potential Treatment Targets of Pediatric Neurological Diseases.

Front Endocrinol (Lausanne) 2021 18;12:655290. Epub 2021 Feb 18.

Movement Disorders Clinic, Neurology Unit, Department of Neuroscience, Bambino Gesù Children's Hospital (IRCCS), Rome, Italy.

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http://dx.doi.org/10.3389/fendo.2021.655290DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7930474PMC
February 2021

Patient-derived iPSC modeling of rare neurodevelopmental disorders: Molecular pathophysiology and prospective therapies.

Neurosci Biobehav Rev 2021 02 25;121:201-219. Epub 2020 Dec 25.

Centre for Molecular Neurosciences, Kasturba Medical College, Manipal Academy of Higher Education, Manipal, 576104, Karnataka, India. Electronic address:

The pathological alterations that manifest during the early embryonic development due to inherited and acquired factors trigger various neurodevelopmental disorders (NDDs). Besides major NDDs, there are several rare NDDs, exhibiting specific characteristics and varying levels of severity triggered due to genetic and epigenetic anomalies. The rarity of subjects, paucity of neural tissues for detailed analysis, and the unavailability of disease-specific animal models have hampered detailed comprehension of rare NDDs, imposing heightened challenge to the medical and scientific community until a decade ago. The generation of functional neurons and glia through directed differentiation protocols for patient-derived iPSCs, CRISPR/Cas9 technology, and 3D brain organoid models have provided an excellent opportunity and vibrant resource for decoding the etiology of brain development for rare NDDs caused due to monogenic as well as polygenic disorders. The present review identifies cellular and molecular phenotypes demonstrated from patient-derived iPSCs and possible therapeutic opportunities identified for these disorders. New insights to reinforce the existing knowledge of the pathophysiology of these disorders and prospective therapeutic applications are discussed.
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http://dx.doi.org/10.1016/j.neubiorev.2020.12.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7962756PMC
February 2021

Monosodium luminol reinstates redox homeostasis, improves cognition, mood and neurogenesis, and alleviates neuro- and systemic inflammation in a model of Gulf War Illness.

Redox Biol 2020 01 18;28:101389. Epub 2019 Nov 18.

Institute for Regenerative Medicine, Department of Molecular and Cellular Medicine, Texas A&M University College of Medicine, College Station, TX, USA.

Enduring brain dysfunction is amid the highly manifested symptoms in veterans with Gulf War Illness (GWI). Animal studies have established that lasting brain dysfunction in GWI is concomitant with augmented oxidative stress, inflammation, and declined neurogenesis in the brain, and systemic inflammation. We hypothesize that drugs capable of restoring redox homeostasis in GWI will improve cognitive and mood function with modulation of neuroinflammation and neurogenesis. We examined the efficacy of monosodium luminol-GVT (MSL), a drug that promotes redox homeostasis, for improving cognitive and mood function in GWI rats. Young rats were exposed to GWI-related chemicals and moderate restraint stress for four weeks. Four months later, GWI rats received different doses of MSL or vehicle for eight weeks. Behavioral analyses in the last three weeks of treatment revealed that GWI rats receiving higher doses of MSL displayed better cognitive and mood function associated with reinstatement of redox homeostasis. Such restoration was evident from the normalized expression of multiple genes encoding proteins involved in combating oxidative stress in the brain and the return of several oxidative stress markers to control levels in the brain and the circulating blood. Sustained redox homeostasis by MSL also resulted in antiinflammatory and pro-neurogenic effects, which were apparent from reduced densities of hypertrophied astrocytes and activated microglia, and increased neurogenesis with augmented neural stem cell proliferation. Moreover, MSL treatment normalized the concentration of multiple proinflammatory markers in the circulating blood. Thus, MSL treatment reinstated redox homeostasis in an animal model of GWI, which resulted in alleviation of both brain and systemic inflammation, improved neurogenesis, and better cognitive and mood function.
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http://dx.doi.org/10.1016/j.redox.2019.101389DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6888767PMC
January 2020

Promise of extracellular vesicles for diagnosis and treatment of epilepsy.

Epilepsy Behav 2019 Oct 18:106499. Epub 2019 Oct 18.

Institute for Regenerative Medicine, Department of Molecular and Cellular Medicine, Texas A&M University College of Medicine, College Station, TX, USA. Electronic address:

Extracellular vesicles (EVs) released from cells play vital roles in intercellular communication. Moreover, EVs released from stem cells have therapeutic properties. This review confers the potential of brain-derived EVs in the cerebrospinal fluid (CSF) and the serum as sources of epilepsy-related biomarkers, and the promise of mesenchymal stem cell (MSC)-derived EVs for easing status epilepticus (SE)-induced adverse changes in the brain. Extracellular vesicles shed from neurons and glia in the brain can also be found in the circulating blood as EVs cross the blood-brain barrier (BBB). Evaluation of neuron and/or glia-derived EVs in the blood of patients who have epilepsy could help in identifying specific biomarkers for distinct types of epilepsies. Such a liquid biopsy approach is also amenable for repeated analysis in clinical trials for comprehending treatment efficacy, disease progression, and mechanisms of therapeutic interventions. Extracellular vesicle biomarker studies in animal prototypes of epilepsy, in addition, could help in identifying specific micro ribonucleic acid (miRNAs) contributing to epileptogenesis, seizures, or cognitive dysfunction in different types of epilepsy. Furthermore, intranasal (IN) administration of MSC-derived EVs after SE has shown efficacy for restraining SE-induced neuroinflammation, aberrant neurogenesis, and cognitive dysfunction in an animal prototype. Clinical translation of EV therapy as an adjunct to antiepileptic drugs appears attractive to counteract the progression of SE-induced epileptogenic changes, as the risk for thrombosis or tumor is minimal with nanosized EVs. Also, EVs can be engineered to deliver specific miRNAs, proteins, or antiepileptic drugs to the brain since they incorporate into neurons and glia throughout the brain after IN administration. This article is part of the Special Issue "NEWroscience 2018".
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http://dx.doi.org/10.1016/j.yebeh.2019.106499DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7165061PMC
October 2019

Extracellular Vesicles as Therapeutics for Brain Injury and Disease.

Curr Pharm Des 2019 ;25(33):3500-3505

Institute for Regenerative Medicine, Department of Molecular and Cellular Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX, United States.

Extracellular vesicles (EVs) are gaining tremendous importance in comprehending central nervous system (CNS) function and treating neurological disorders because of their role in intercellular communication and reparative processes, and suitability as drug delivery vehicles. Since EVs have lipid membranes, they cross the blood-brain barrier easily and communicate with target neurons and glia even deep inside the brain. EVs from various sources have been isolated, characterized, and tailored for promoting beneficial effects in conditions, such as brain injury and disease. Particularly, EVs isolated from mesenchymal stem cells and neural stem cells have shown promise for alleviating brain dysfunction after injury or disease. Such properties of stem cell-derived EVs have great importance for clinical applications, as EV therapy can avoid several concerns typically associated with cell therapy. This minireview confers the competence of EVs for improving brain function by modulating CNS injury and disease.
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http://dx.doi.org/10.2174/1381612825666191014164950DOI Listing
June 2020

A Model of Chronic Temporal Lobe Epilepsy Presenting Constantly Rhythmic and Robust Spontaneous Seizures, Co-morbidities and Hippocampal Neuropathology.

Aging Dis 2019 Oct 1;10(5):915-936. Epub 2019 Oct 1.

Institute for Regenerative Medicine, Department of Molecular and Cellular Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX, USA.

Many animal prototypes illustrating the various attributes of human temporal lobe epilepsy (TLE) are available. These models have been invaluable for comprehending multiple epileptogenic processes, modifications in electrophysiological properties, neuronal hyperexcitability, neurodegeneration, neural plasticity, and chronic neuroinflammation in TLE. Some models have also uncovered the efficacy of new antiepileptic drugs or biologics for alleviating epileptogenesis, cognitive impairments, or spontaneous recurrent seizures (SRS). Nonetheless, the suitability of these models for testing candidate therapeutics in conditions such as chronic TLE is debatable because of a lower frequency of SRS and an inconsistent pattern of SRS activity over days, weeks or months. An ideal prototype of chronic TLE for investigating novel therapeutics would need to display a large number of SRS with a dependable frequency and severity and related co-morbidities. This study presents a new kainic acid (KA) model of chronic TLE generated through induction of status epilepticus (SE) in 6-8 weeks old male F344 rats. A rigorous characterization in the chronic epilepsy period validated that the animal prototype mimicked the most salient features of robust chronic TLE. Animals displayed a constant frequency and intensity of SRS across weeks and months in the 5th and 6th month after SE, as well as cognitive and mood impairments. Moreover, SRS frequency displayed a rhythmic pattern with 24-hour periodicity and a consistently higher number of SRS in the daylight period. Besides, the model showed many neuropathological features of chronic TLE, which include a partial loss of inhibitory interneurons, reduced neurogenesis with persistent aberrant migration of newly born neurons, chronic neuroinflammation typified by hypertrophied astrocytes and rod-shaped microglia, and a significant aberrant mossy fiber sprouting in the hippocampus. This consistent chronic seizure model is ideal for investigating the efficacy of various antiepileptic drugs and biologics as well as understanding multiple pathophysiological mechanisms underlying chronic epilepsy.
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http://dx.doi.org/10.14336/AD.2019.0720DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6764729PMC
October 2019

Human induced pluripotent stem cell-derived MGE cell grafting after status epilepticus attenuates chronic epilepsy and comorbidities via synaptic integration.

Proc Natl Acad Sci U S A 2019 01 17;116(1):287-296. Epub 2018 Dec 17.

Institute for Regenerative Medicine, Texas A&M Health Science Center College of Medicine, Temple, TX 76502;

Medial ganglionic eminence (MGE)-like interneuron precursors derived from human induced pluripotent stem cells (hiPSCs) are ideal for developing patient-specific cell therapy in temporal lobe epilepsy (TLE). However, their efficacy for alleviating spontaneous recurrent seizures (SRS) or cognitive, memory, and mood impairments has never been tested in models of TLE. Through comprehensive video- electroencephalographic recordings and a battery of behavioral tests in a rat model, we demonstrate that grafting of hiPSC-derived MGE-like interneuron precursors into the hippocampus after status epilepticus (SE) greatly restrained SRS and alleviated cognitive, memory, and mood dysfunction in the chronic phase of TLE. Graft-derived cells survived well, extensively migrated into different subfields of the hippocampus, and differentiated into distinct subclasses of inhibitory interneurons expressing various calcium-binding proteins and neuropeptides. Moreover, grafting of hiPSC-MGE cells after SE mediated several neuroprotective and antiepileptogenic effects in the host hippocampus, as evidenced by reductions in host interneuron loss, abnormal neurogenesis, and aberrant mossy fiber sprouting in the dentate gyrus (DG). Furthermore, axons from graft-derived interneurons made synapses on the dendrites of host excitatory neurons in the DG and the CA1 subfield of the hippocampus, implying an excellent graft-host synaptic integration. Remarkably, seizure-suppressing effects of grafts were significantly reduced when the activity of graft-derived interneurons was silenced by a designer drug while using donor hiPSC-MGE cells expressing designer receptors exclusively activated by designer drugs (DREADDs). These results implied the direct involvement of graft-derived interneurons in seizure control likely through enhanced inhibitory synaptic transmission. Collectively, the results support a patient-specific MGE cell grafting approach for treating TLE.
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http://dx.doi.org/10.1073/pnas.1814185115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6320542PMC
January 2019

Inhibition of ERK1/2 or AKT Activity Equally Enhances Radiation Sensitization in B16F10 Cells.

World J Oncol 2018 Feb 8;9(1):21-28. Epub 2018 Mar 8.

Yenepoya Research Centre, Yenepoya University, Mangalore, India.

Background: The aim of the study was to evaluate the radiation sensitizing ability of ERK1/2, PI3K-AKT and JNK inhibitors in highly radiation resistant and metastatic B16F10 cells which carry wild-type and .

Methods: Mouse melanoma cell line B16F10 was exposed to 1.0, 2.0 and 3.0 Gy of electron beam radiation. Phosphorylated ERK1/2, AKT and JNK levels were estimated by ELISA. Cells were exposed to 2.0 and 3.0 Gy of radiation with or without prior pharmacological inhibition of ERK1/2, AKT as well as JNK pathways. Cell death induced by radiation as well as upon inhibition of these pathways was measured by TUNEL assay using flow cytometry.

Results: Exposure of B16F10 cells to 1.0, 2.0 and 3.0 Gy of electron beam irradiation triggered an increase in all the three phosphorylated proteins compared to sham-treated and control groups. B16F10 cells pre-treated with either ERK1/2 or AKT inhibitors equally enhanced radiation-induced cell death at 2.0 as well as 3.0 Gy (P < 0.001), while inhibition of JNK pathway increased radiation-induced cell death to a lesser extent. Interestingly combined inhibition of ERK1/2 or AKT pathways did not show additional cell death compared to individual ERK1/2 or AKT inhibition. This indicates that ERK1/2 or AKT mediates radiation resistance through common downstream molecules in B16F10 cells.

Conclusions: Even without activating mutations in or genes, ERK1/2 and AKT play a critical role in B16F10 cell survival upon radiation exposure and possibly act through common downstream effector/s.
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http://dx.doi.org/10.14740/wjon1088wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5862079PMC
February 2018

Prospects of Cannabidiol for Easing Status Epilepticus-Induced Epileptogenesis and Related Comorbidities.

Mol Neurobiol 2018 Aug 25;55(8):6956-6964. Epub 2018 Jan 25.

Institute for Regenerative Medicine and Department of Molecular and Cellular Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX, 77845, USA.

The hippocampus is one of the most susceptible regions in the brain to be distraught with status epilepticus (SE) induced injury. SE can occur from numerous causes and is more frequent in children and the elderly population. Administration of a combination of antiepileptic drugs can abolish acute seizures in most instances of SE but cannot prevent the morbidity typically seen in survivors of SE such as cognitive and mood impairments and spontaneous recurrent seizures. This is primarily due to the inefficiency of antiepileptic drugs to modify the evolution of SE-induced initial precipitating injury into a series of epileptogenic changes followed by a state of chronic epilepsy. Chronic epilepsy is typified by spontaneous recurrent seizures, cognitive dysfunction, and depression, which are associated with persistent inflammation, significantly waned neurogenesis, and abnormal synaptic reorganization. Thus, alternative approaches that are efficient not only for curtailing SE-induced initial brain injury, neuroinflammation, aberrant neurogenesis, and abnormal synaptic reorganization but also for thwarting or restraining the progression of SE into a chronic epileptic state are needed. In this review, we confer the promise of cannabidiol, an active ingredient of Cannabis sativa, for preventing or easing SE-induced neurodegeneration, neuroinflammation, cognitive and mood impairments, and the spontaneous recurrent seizures.
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http://dx.doi.org/10.1007/s12035-018-0898-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7023903PMC
August 2018

Resveratrol for Easing Status Epilepticus Induced Brain Injury, Inflammation, Epileptogenesis, and Cognitive and Memory Dysfunction-Are We There Yet?

Front Neurol 2017 13;8:603. Epub 2017 Nov 13.

Olin E. Teague Veterans' Medical Center, Central Texas Veterans Health Care System, Temple, Texas, United States.

Status epilepticus (SE) is a medical emergency exemplified by self-sustaining, unceasing seizures or swiftly recurring seizure events with no recovery between seizures. The early phase after SE event is associated with neurodegeneration, neuroinflammation, and abnormal neurogenesis in the hippocampus though the extent of these changes depends on the severity and duration of seizures. In many instances, over a period, the initial precipitating injury caused by SE leads to temporal lobe epilepsy (TLE), typified by spontaneous recurrent seizures, cognitive, memory and mood impairments associated with chronic inflammation, reduced neurogenesis, abnormal synaptic reorganization, and multiple molecular changes in the hippocampus. While antiepileptic drugs are efficacious for terminating or greatly reducing seizures in most cases of SE, they have proved ineffective for easing SE-induced epileptogenesis and TLE. Despite considerable advances in elucidating SE-induced multiple cellular, electrophysiological, and molecular changes in the brain, efficient strategies that prevent SE-induced TLE development are yet to be discovered. This review critically confers the efficacy and promise of resveratrol, a phytoalexin found in the skin of red grapes, for easing SE-induced neurodegeneration, neuroinflammation, aberrant neurogenesis, and for restraining the evolution of SE-induced brain injury into a chronic epileptic state typified by spontaneous recurrent seizures, and learning, memory, and mood impairments.
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http://dx.doi.org/10.3389/fneur.2017.00603DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5694141PMC
November 2017

Chronic Oxidative Stress, Mitochondrial Dysfunction, Nrf2 Activation and Inflammation in the Hippocampus Accompany Heightened Systemic Inflammation and Oxidative Stress in an Animal Model of Gulf War Illness.

Front Mol Neurosci 2017 14;10:182. Epub 2017 Jun 14.

Research Service, Olin E. Teague Veterans' Medical Center, Central Texas Veterans Health Care System, TempleTX, United States.

Memory and mood dysfunction are the key symptoms of Gulf war illness (GWI), a lingering multi-symptom ailment afflicting >200,000 veterans who served in the Persian Gulf War-1. Research probing the source of the disease has demonstrated that concomitant exposures to anti-nerve gas agent pyridostigmine bromide (PB), pesticides, and war-related stress are among the chief causes of GWI. Indeed, exposures to GWI-related chemicals (GWIR-Cs) and mild stress in animal models cause memory and mood impairments alongside reduced neurogenesis and chronic low-level inflammation in the hippocampus. In the current study, we examined whether exposure to GWIR-Cs and stress causes chronic changes in the expression of genes related to increased oxidative stress, mitochondrial dysfunction, and inflammation in the hippocampus. We also investigated whether GWI is linked with chronically increased activation of Nrf2 (a master regulator of antioxidant response) in the hippocampus, and inflammation and enhanced oxidative stress at the systemic level. Adult male rats were exposed daily to low-doses of PB and pesticides (DEET and permethrin), in combination with 5 min of restraint stress for 4 weeks. Analysis of the hippocampus performed 6 months after the exposure revealed increased expression of many genes related to oxidative stress response and/or antioxidant activity (, and ), reactive oxygen species metabolism (, and ) and oxygen transport ( and ). Furthermore, multiple genes relevant to mitochondrial respiration (, and ) and neuroinflammation (, and ) were up-regulated, alongside 73-88% reduction in the expression of anti-inflammatory genes and , and nuclear translocation and increased expression of Nrf2 protein. These hippocampal changes were associated with elevated levels of pro-inflammatory cytokines and chemokines (Tnfa, IL1b, IL1a, Tgfb, and Fgf2) and lipid peroxidation byproduct malondialdehyde in the serum, suggesting the presence of an incessant systemic inflammation and elevated oxidative stress. These results imply that chronic oxidative stress, inflammation, and mitochondrial dysfunction in the hippocampus, and heightened systemic inflammation and oxidative stress likely underlie the persistent memory and mood dysfunction observed in GWI.
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http://dx.doi.org/10.3389/fnmol.2017.00182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5469946PMC
June 2017

Intranasal MSC-derived A1-exosomes ease inflammation, and prevent abnormal neurogenesis and memory dysfunction after status epilepticus.

Proc Natl Acad Sci U S A 2017 04 10;114(17):E3536-E3545. Epub 2017 Apr 10.

Institute for Regenerative Medicine, Texas A&M Health Science Center College of Medicine, Temple, TX 76502;

Status epilepticus (SE), a medical emergency that is typically terminated through antiepileptic drug treatment, leads to hippocampus dysfunction typified by neurodegeneration, inflammation, altered neurogenesis, as well as cognitive and memory deficits. Here, we examined the effects of intranasal (IN) administration of extracellular vesicles (EVs) secreted from human bone marrow-derived mesenchymal stem cells (MSCs) on SE-induced adverse changes. The EVs used in this study are referred to as A1-exosomes because of their robust antiinflammatory properties. We subjected young mice to pilocarpine-induced SE for 2 h and then administered A1-exosomes or vehicle IN twice over 24 h. The A1-exosomes reached the hippocampus within 6 h of administration, and animals receiving them exhibited diminished loss of glutamatergic and GABAergic neurons and greatly reduced inflammation in the hippocampus. Moreover, the neuroprotective and antiinflammatory effects of A1-exosomes were coupled with long-term preservation of normal hippocampal neurogenesis and cognitive and memory function, in contrast to waned and abnormal neurogenesis, persistent inflammation, and functional deficits in animals receiving vehicle. These results provide evidence that IN administration of A1-exosomes is efficient for minimizing the adverse effects of SE in the hippocampus and preventing SE-induced cognitive and memory impairments.
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http://dx.doi.org/10.1073/pnas.1703920114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5410779PMC
April 2017

Chemotherapy Resistance Mechanisms in Advanced Skin Cancer.

Oncol Rev 2017 Mar 24;11(1):326. Epub 2017 Mar 24.

Department of Biochemistry, Yenepoya Medical College , Mangaluru, India.

Melanoma is a most dangerous and deadly type of skin cancer, and considered intrinsically resistant to both radiotherapy and chemotherapy. It has become a major public health concern as the incidence of melanoma has been rising steadily over recent decades with a 5-year survival remaining less than 5%. Detection of the disease in early stage may be curable, but late stage metastatic disease that has spread to other organs has an extremely poor prognosis with a median survival of less than 10 months. Since metastatic melanoma is unresponsive to therapy that is currently available, research is now focused on different treatment strategies such as combinations of surgery, chemotherapy and radiotherapy. The molecular basis of resistance to chemotherapy seen in melanoma is multifactorial; defective drug transport system, altered apoptotic pathway, deregulation of apoptosis and/or changes in enzymatic systems that mediate cellular metabolic machinery. Understanding of alterations in molecular processes involved in drug resistance may help in developing new therapeutic approaches to treatment of malignant melanoma.
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http://dx.doi.org/10.4081/oncol.2017.326DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5379221PMC
March 2017

Neural Stem Cell or Human Induced Pluripotent Stem Cell-Derived GABA-ergic Progenitor Cell Grafting in an Animal Model of Chronic Temporal Lobe Epilepsy.

Curr Protoc Stem Cell Biol 2016 08 17;38:2D.7.1-2D.7.47. Epub 2016 Aug 17.

Institute for Regenerative Medicine, Texas A&M University Health Science Center College of Medicine, Temple, Texas.

Grafting of neural stem cells (NSCs) or GABA-ergic progenitor cells (GPCs) into the hippocampus could offer an alternative therapy to hippocampal resection in patients with drug-resistant chronic epilepsy, which afflicts >30% of temporal lobe epilepsy (TLE) cases. Multipotent, self-renewing NSCs could be expanded from multiple regions of the developing and adult brain, human embryonic stem cells (hESCs), and human induced pluripotent stem cells (hiPSCs). On the other hand, GPCs could be generated from the medial and lateral ganglionic eminences of the embryonic brain and from hESCs and hiPSCs. To provide comprehensive methodologies involved in testing the efficacy of transplantation of NSCs and GPCs in a rat model of chronic TLE, NSCs derived from the rat medial ganglionic eminence (MGE) and MGE-like GPCs derived from hiPSCs are taken as examples in this unit. The topics comprise description of the required materials, reagents and equipment, methods for obtaining rat MGE-NSCs and hiPSC-derived MGE-like GPCs in culture, generation of chronically epileptic rats, intrahippocampal grafting procedure, post-grafting evaluation of the effects of grafts on spontaneous recurrent seizures and cognitive and mood impairments, analyses of the yield and the fate of graft-derived cells, and the effects of grafts on the host hippocampus. © 2016 by John Wiley & Sons, Inc.
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http://dx.doi.org/10.1002/cpsc.9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5313261PMC
August 2016

GABA-ergic cell therapy for epilepsy: Advances, limitations and challenges.

Neurosci Biobehav Rev 2016 Mar 31;62:35-47. Epub 2015 Dec 31.

Institute for Regenerative Medicine, Texas A&M Health Science Center College of Medicine, Temple, TX, United States; Research Service, Olin E. Teague Veterans' Medical Center, Central Texas Veterans Health Care System, Temple, TX, United States; Department of Molecular and Cellular Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX, United States.

Diminution in the number of gamma-amino butyric acid positive (GABA-ergic) interneurons and their axon terminals, and/or alterations in functional inhibition are conspicuous brain alterations believed to contribute to the persistence of seizures in acquired epilepsies such as temporal lobe epilepsy. This has steered a perception that replacement of lost GABA-ergic interneurons would improve inhibitory synaptic neurotransmission in the epileptic brain region and thereby reduce the occurrence of seizures. Indeed, studies using animal prototypes have reported that grafting of GABA-ergic progenitors derived from multiple sources into epileptic regions can reduce seizures. This review deliberates recent advances, limitations and challenges concerning the development of GABA-ergic cell therapy for epilepsy. The efficacy and limitations of grafts of primary GABA-ergic progenitors from the embryonic lateral ganglionic eminence and medial ganglionic eminence (MGE), neural stem/progenitor cells expanded from MGE, and MGE-like progenitors generated from human pluripotent stem cells for alleviating seizures and co-morbidities of epilepsy are conferred. Additional studies required for possible clinical application of GABA-ergic cell therapy for epilepsy are also summarized.
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http://dx.doi.org/10.1016/j.neubiorev.2015.12.014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4869704PMC
March 2016

Fibroblast growth factor receptor 2 (FGFR2) is required for corneal epithelial cell proliferation and differentiation during embryonic development.

PLoS One 2015 23;10(1):e0117089. Epub 2015 Jan 23.

Dept. of Ophthalmology, Mason Eye Institute, University of Missouri, Columbia, Missouri, United States of America.

Fibroblast growth factors (FGFs) play important roles in many aspects of embryonic development. During eye development, the lens and corneal epithelium are derived from the same surface ectodermal tissue. FGF receptor (FGFR)-signaling is essential for lens cell differentiation and survival, but its role in corneal development has not been fully investigated. In this study, we examined the corneal defects in Fgfr2 conditional knockout mice in which Cre expression is activated at lens induction stage by Pax6 P0 promoter. The cornea in LeCre, Fgfr2(loxP/loxP) mice (referred as Fgfr2(CKO)) was analyzed to assess changes in cell proliferation, differentiation and survival. We found that Fgfr2(CKO) cornea was much thinner in epithelial and stromal layer when compared to WT cornea. At embryonic day 12.5-13.5 (E12.5-13.5) shortly after the lens vesicle detaches from the overlying surface ectoderm, cell proliferation (judged by labeling indices of Ki-67, BrdU and phospho-histone H3) was significantly reduced in corneal epithelium in Fgfr2(CKO) mice. At later stage, cell differentiation markers for corneal epithelium and underlying stromal mesenchyme, keratin-12 and keratocan respectively, were not expressed in Fgfr2(CKO) cornea. Furthermore, Pax6, a transcription factor essential for eye development, was not present in the Fgfr2(CKO) mutant corneal epithelial at E16.5 but was expressed normally at E12.5, suggesting that FGFR2-signaling is required for maintaining Pax6 expression in this tissue. Interestingly, the role of FGFR2 in corneal epithelial development is independent of ERK1/2-signaling. In contrast to the lens, FGFR2 is not required for cell survival in cornea. This study demonstrates for the first time that FGFR2 plays an essential role in controlling cell proliferation and differentiation, and maintaining Pax6 levels in corneal epithelium via ERK-independent pathways during embryonic development.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0117089PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4304804PMC
January 2016

Nuclear DNA fragmentation negatively affects zona binding competence of Y bearing mouse spermatozoa.

J Assist Reprod Genet 2013 Dec 19;30(12):1611-5. Epub 2013 Oct 19.

Division of Clinical Embryology, Department of Obstetrics & Gynecology, Kasturba Medical College, Manipal University, Manipal, 576 104, India.

Purpose: To investigate the influence of sperm DNA integrity on the zona binding ability of mouse spermatozoa in relation to their sex chromosomal constitution.

Method(s): In this prospective experimental study, the sperm DNA fragmentation was induced by exposing testicular area of Swiss Albino mice (Mus musculus) to different doses of γ-radiation (0, 2.5, 5.0 and 10.0 Gy). Sperm DNA fragmentation was quantified by single cell gel electrophoresis (comet assay). In vitro sperm zona binding assay was performed and the numbers of zona bound X and Y bearing spermatozoa were determined using fluorescence in situ hybridization (FISH).

Result(s): The assessment of zona pellucida bound X and Y-bearing spermatozoa using fluorescence in situ hybridization has revealed a unique binding pattern. The number of zona bound Y-spermatozoa declined significantly (P < 0.01 to 0.0001) with increase in the DNA damage. The skewed binding pattern of X and Y-bearing sperm was strongly correlated with the extent of sperm DNA damage.

Conclusion(s): The zona pellucida may have a role in preventing DNA damaged mouse sperm binding especially towards Y-bearing sperm. However, the exact mechanism behind this observation needs to be elucidated further.
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http://dx.doi.org/10.1007/s10815-013-0123-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3843170PMC
December 2013

MAPK1 is required for establishing the pattern of cell proliferation and for cell survival during lens development.

Development 2013 Apr;140(7):1573-82

Department of Ophthalmology, University of Missouri, Columbia, MO 65212, USA.

The mitogen-activated protein kinases (MAPKs; also known as ERKs) are key intracellular signaling molecules that are ubiquitously expressed in tissues and were assumed to be functionally equivalent. Here, we use the mouse lens as a model system to investigate whether MAPK1 plays a specific role during development. MAPK3 is known to be dispensable for lens development. We demonstrate that, although MAPK1 is uniformly expressed in the lens epithelium, its deletion significantly reduces cell proliferation in the peripheral region, an area referred to as the lens germinative zone in which most active cell division occurs during normal lens development. By contrast, cell proliferation in the central region is minimally affected by MAPK1 deletion. Cell cycle regulators, including cyclin D1 and survivin, are downregulated in the germinative zone of the MAPK1-deficient lens. Interestingly, loss of MAPK1 subsequently induces upregulation of phosphorylated MAPK3 (pMAPK3) levels in the lens epithelium; however, this increase in pMAPK3 is not sufficient to restore cell proliferation in the germinative zone. Additionally, MAPK1 plays an essential role in epithelial cell survival but is dispensable for fiber cell differentiation during lens development. Our data indicate that MAPK1/3 control cell proliferation in the lens epithelium in a spatially defined manner; MAPK1 plays a unique role in establishing the highly mitotic zone in the peripheral region, whereas the two MAPKs share a redundant role in controlling cell proliferation in the central region of the lens epithelium.
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http://dx.doi.org/10.1242/dev.081042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3596996PMC
April 2013

The extent of paternal sperm DNA damage influences early post-natal survival of first generation mouse offspring.

Eur J Obstet Gynecol Reprod Biol 2013 Feb 12;166(2):164-7. Epub 2012 Oct 12.

Clinical Embryology, Department of Obstetrics & Gynecology, Kasturba Medical College, Manipal University, Manipal 576104, India.

Objectives: To study the post-natal characteristics and the survival of offspring derived from DNA damaged sperm.

Study Design: This experimental prospective study was conducted on Swiss Albino mice (Mus musculus). Sperm DNA damage was induced by different doses of γ-irradiation in male mice who were then mated with healthy female mice. The post-natal characteristics including the survival of first generation offspring were studied and then correlated with the amount of paternal sperm DNA damage.

Results: A significant reduction of survival in the early post-natal period was observed in the first generation offspring derived from the DNA damaged sperm, and a strong association was observed between the extent of sperm DNA damage and the survival of the offspring.

Conclusion: The DNA damage load in sperm at the time of fertilization influences early post-natal survival of the mouse offspring.
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http://dx.doi.org/10.1016/j.ejogrb.2012.09.021DOI Listing
February 2013

Sperm processing by swim-up and density gradient is effective in elimination of sperm with DNA damage.

J Assist Reprod Genet 2012 Jun 13;29(6):557-63. Epub 2012 Mar 13.

Division of Reproductive Medicine, Clinical Embryology Laboratory, Kasturba Medical College Manipal University, Manipal 576 104, India.

Purpose: DNA damage may occur during sperm processing, thereby negatively influencing fertilizing ability of the sperm. The present study was designed to compare the effectiveness of gradient and swim-up, either alone or in combination, to eliminate sperm with DNA damage.

Methods: A total of 51 subjects visiting the University infertility clinic with normozoospermic parameters, oligozoospermia and teratozoospermia were included. Semen characteristics were analysed by standard criteria; Terminal deoxy nucelotidyl transferase mediated dUTP nick end labeling assay was employed for DNA damage assessment.

Results: The percentage of TUNEL positive sperm after sperm processing was significantly lower in normozoospermic (P < 0.05), oligozoospermic (P < 0.001) and teratozoospermic samples (P < 0.01). No difference was observed in the incidence of TUNEL positive sperm between the various techniques, suggesting that they are comparable.

Conclusions: Sperm preparation has been found to result in enrichment of sperm with intact chromatin, which is likely to improve the chances of achieving a viable pregnancy.
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http://dx.doi.org/10.1007/s10815-012-9742-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3370043PMC
June 2012

Combination of swim-up and density gradient separation methods effectively eliminate DNA damaged sperm.

J Turk Ger Gynecol Assoc 2011 1;12(3):148-52. Epub 2011 Sep 1.

Clinical Embryology, Division of Reproductive Medicine, Kasturba Medical College, Manipal University, Manipal, India.

Objective: The aim of this experimental prospective study was to investigate the efficacy of single and combination sperm wash methods for their ability to isolate DNA intact spermatozoa.

Material And Methods: Sperm DNA damage was introduced by local testicular irradiation in male mice and the extent of damage was quantified by comet assay. The spermatozoa were subjected to single (swim up or density gradient method) and also a combination of sperm wash techniques. The DNA integrity in various sub-fractions of wash techniques was evaluated.

Results: The amount of DNA damaged sperm did not differ between individual fractions when single wash technique was applied. However, a combination of density gradient and swim-up techniques significantly reduced (p<0.01) the number of DNA damaged sperm in the final population.

Conclusion: The combination of density gradient separation and swim-up method is effective in eliminating DNA damaged spermatozoa.
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http://dx.doi.org/10.5152/jtgga.2011.35DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3939271PMC
March 2014

Predictive and prognostic significance of glutathione levels and DNA damage in cervix cancer patients undergoing radiotherapy.

Int J Radiat Oncol Biol Phys 2010 Oct 3;78(2):343-9. Epub 2010 Feb 3.

Department of Radiotherapy, Shirdi Saibaba Cancer Hospital, Kasturba Medical College, Manipal, Karnataka, India.

Purpose: To assess the predictive significance of serum glutathione (GSH) and tumor tissue DNA damage in the treatment of cervical cancer patients undergoing chemoradiotherapy.

Methods And Materials: This study included subjects undergoing hysterectomy (for normal cervix tissue) and cervical cancer patients who underwent conventional concurrent chemoradiotherapy (cisplatin once per week for 5 weeks with concurrent external radiotherapy of 2 Gy per fraction for 5 weeks, followed by two applications of intracavitary brachytherapy once per week after 2 weeks' rest). Blood was collected after two fractions, whereas both blood and tissues were collected after five fractions of radiotherapy in separate groups of subjects. Serum for total GSH content and tissues were processed for single-cell gel electrophoresis (SCGE) assay for DNA damage analysis. Clinical tumor radioresponse was assessed 2 months after the completion of treatment as complete responders (CR) (100% shrinkage), partial responders (PR) (>50%), and nonresponders (NR) (<50%).

Results: Serum GSH content depleted significantly after a total dose of 4 Gy and 10 Gy of radiotherapy with a single dose of cisplatin, which was significantly lesser in NR than of CR patients. Similarly, Olive Tail Moment, the index of DNA damage, indicated significantly higher values in the fifth fraction of radiotherapy (5-RT) than in pretreatment. The DNA damage after 5-RT in the NR subgroup was significantly lower than that of CR.

Conclusions: Serum GSH analysis and tumor tissue SCGE assay found to be useful parameters for predicting chemoradioresponse prior to and also at an early stage of treatment of cervical cancers.
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http://dx.doi.org/10.1016/j.ijrobp.2009.08.014DOI Listing
October 2010

Association between the extent of DNA damage in the spermatozoa, fertilization and developmental competence in preimplantation stage embryos.

J Turk Ger Gynecol Assoc 2010 1;11(4):182-6. Epub 2010 Dec 1.

Clinical Embryology, Division of Reproductive Medicine, Department of Obstetrics and Gynecology, Kasturba Medical College, Manipal University, Manipal, India.

Objective: To examine the fertilizing ability and DNA damage response of preimplantation stage embryos derived from the γ-irradiated mouse sperm carrying varying amounts of DNA strand-breaks.

Material And Methods: The DNA damage in the sperm was induced by exposing the testicular area to different doses of γ-radiation. After mating with healthy female mice, sperm zona binding, fertilizing ability of DNA damaged sperm and developmental competence of embryos derived from the DNA damaged sperm were assessed.

Results: The in vivo zona binding ability and fertilizing ability of DNA damaged sperm was significantly affected in the 5.0 and 10.0 Gy sperm irradiation groups. Although the development of the embryos derived from the DNA damaged sperm was not significantly affected until day 2.5 post-coitus, further development was significantly altered, as evidenced by the total cell number in the embryos.

Conclusion: The sperm carrying DNA strand breaks still has the ability to fertilize the oocyte normally. However, the events like zona-binding and successful fertilization depend on the extent of sperm DNA fragmentation. The study has also showed a great heterogeneity in embryonic development at peri-implantation period with respect to the degree of sperm DNA damage.
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http://dx.doi.org/10.5152/jtgga.2010.34DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3939148PMC
March 2014

Transgenerational changes in somatic and germ line genetic integrity of first-generation offspring derived from the DNA damaged sperm.

Fertil Steril 2010 May 9;93(8):2486-90. Epub 2009 Jul 9.

Division of Reproductive Medicine, Kasturba Medical College, Manipal University, Manipal, India.

Objective: To report a more quantitative approach to study the influence of varying levels of sperm DNA damage on transgenerational changes in genomic instability in a mouse model.

Design: Experimental prospective study.

Setting: Embryology research laboratory.

Animal(s): Swiss albino mice.

Intervention(s): The sperm DNA damage was induced by different doses of gamma-irradiation to male mice followed by mating with healthy female mice.

Main Outcome Measure(s): Genomic integrity in embryos, fetus, and spermatozoa of F1 mice derived from the DNA-damaged sperm.

Result(s): The transgenerational changes in genetic integrity were attributed by a dose-dependent increase in the frequency of micronuclei in preimplantation embryos and a concomitant increase in genomic instability in fetal liver cells and sperm chromatin modifications in F1 males. A strong positive correlation was observed between the extent of sperm DNA damage and somatic and germ-line genomic instability.

Conclusion(s): Sperm-mediated transgenerational genomic instability is dependent on the amount of DNA damage present in the sire's sperm at the time of fertilization.
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http://dx.doi.org/10.1016/j.fertnstert.2009.06.015DOI Listing
May 2010

Ability of deoxyribonucleic acid-damaged sperm to withstand freeze-thaw-induced damage during cryopreservation.

Fertil Steril 2009 Sep 30;92(3):959-963. Epub 2008 Sep 30.

Division of Reproductive Medicine, Kasturba Medical College, Manipal University, Manipal, India.

Objective: To understand the association between sperm DNA damage and the ability of DNA-damaged sperm to withstand the freeze-thaw process during cryopreservation.

Design: Experimental prospective study.

Setting(s): Embryology research laboratory.

Animals: Eight- to 12-week-old Swiss strain male albino mice (Mus musculus).

Intervention(s): Sperm carrying a known amount of DNA damage was subjected to cryopreservation and thereafter evaluated for survival and freeze-thaw-induced DNA damage.

Main Outcome Measure(s): Elucidation of association between the amount of initial sperm DNA damage, cryosurvival, and freeze-thaw-induced DNA modification.

Result(s): A strong correlation (R = -0.87) was observed between the amount of initial sperm DNA damage and postthaw survival. However, no significant enhancement in DNA damage was observed by the cryopreservation of spermatozoa with various amounts of DNA damage.

Conclusion(s): Cryopreservation of DNA-damaged sperm does not deteriorate the DNA quality, but sperm survival is compromised.
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http://dx.doi.org/10.1016/j.fertnstert.2008.07.1754DOI Listing
September 2009

Protection of ionizing radiation-induced cytogenetic damage by hydroalcoholic extract of Cynodon dactylon in Chinese hamster lung fibroblast cells and human peripheral blood lymphocytes.

J Environ Pathol Toxicol Oncol 2008 ;27(2):101-12

Department of Radiobiology, Manipal Life Sciences Center, Manipal University, Manipal 576 104, Karnataka, India.

The radiomodulatory potential of hydroalcoholic extract of a medicinal plant Cynodon dactylon (family: Poaceae) against radiation-induced cytogenetic damage was analyzed using Chinese hamster lung fibroblast (V79) cells and human peripheral blood lymphocytes (HPBLs) growing in vitro. Induction of micronuclei was used as an index of cytogenetic damage, evaluated in cytokinesis blocked binucleate cells. The hydroalcoholic Cynodon dactylon extract (CDE) rendered protection against the radiation-induced DNA damage, as evidenced by the significant (p<0.001) reduction in micronucleated binucleate cells (MNBNC%) after various doses of CDE treatment in V79 cells and HPBLs. The optimum dose of CDE (40 and 50 microg/ml in HPBLs and V79 cells, respectively) with the greatest reduction in micronuclei was further used in combination with various doses of gamma radiation (0.5, 1, 2, 3, and 4 Gy) exposed 1 h after CDE treatment. A linear dose-dependent MNBNC% increase in radiation alone group was observed, while 40/50 microg/ml CDE significantly resulted in the reduction of MNBNC%, compared to the respective radiation alone groups. CDE resulted in a dose-dependent increase in free radical scavenging ability against various free radicals, viz., 2, 2-diphenyl-2-picryl-hydrazyl (DPPH); 2, 2-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS); superoxide anion (O2*-); hydroxyl radical (OH*) and nitric oxide radical (NO*) generated in vitro. Also, an excellent (70%) inhibition of lipid peroxidation in vitro was observed at a dose of 300 microg/ml CDE, attaining the saturation point at higher doses. The present findings demonstrated the radioprotective effect of CDE, also rendering protection against radiation-induced genomic instability and DNA damage. The observed radioprotective effect may be partly attributed to the free radical scavenging and antilipid peroxidative potential of CDE.
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http://dx.doi.org/10.1615/jenvironpatholtoxicoloncol.v27.i2.30DOI Listing
July 2008

Effect of cryopreservation on sperm DNA integrity in patients with teratospermia.

Fertil Steril 2008 Jun 22;89(6):1723-7. Epub 2007 Oct 22.

Division of Reproductive Medicine, Department of Obstetrics and Gynecology, Kasturba Medical College, Manipal University, Manipal, India.

Objective: To test whether sperm with abnormal head morphology are more likely to undergo DNA damage and/or chromatin modification during the process of freeze-thawing.

Design: In this prospective study, the semen samples from forty-four men attending the infertility clinic were included. Samples were divided into aliquots to allow direct comparison of fresh and frozen spermatozoa from the same ejaculate. The sperm morphology and the sperm DNA damage were evaluated before and after cryopreservation. The relationship between sperm head abnormalities and freeze-thaw-induced DNA modification was assessed.

Setting(s): University hospital fertility center.

Patient(s): Men attending infertility clinic for semen analysis.

Intervention(s): The normospermic and teratospermic semen samples were evaluated for DNA damage before and after cryopreservation by comet assay and acridine orange bindability test.

Main Outcome Measure(s): Elucidation of association between sperm morphologic defect and cryodamage.

Result(s): A threefold increase in the amount of DNA damage was observed in teratospermic samples compared with their normospermic counterparts, indicating a higher susceptibility of morphologically abnormal sperm to cryodamage.

Conclusion(s): The susceptibility of morphologically abnormal sperm to DNA damage/chromatin modification during the freeze-thaw process is significantly higher than that of sperm with normal morphology.
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http://dx.doi.org/10.1016/j.fertnstert.2007.06.087DOI Listing
June 2008