University of MIAMI
Department of Medicine,
Katz Family Division of Nephrology and Hypertension

George W. Burke, M.D. - Principal Investigator
Alla Mitrofanova, Ph.D. - Co-Principal Investigator
Alessia Fornoni, M.D.
Antonio Fontanella, B.S.
Miami, Florida
Grant Award - 2 years
“SMPDL3b Modulates Podocyte’s Innate Immunity in Chronic Kidney Disease”

Chronic kidney disease (CKD) is a major health problem worldwide, where glomerular disease
is the most common primary cause leading to end stage renal disease in the US. Among
multiple pathogenic mechanisms responsible for CKD, inflammation plays a significant role.
Podocytes, specialized epithelial cells in glomeruli involved in blood filtration, are one of the
main target cells of injury in CKD. Podocytes exhibit many features of adaptive and innate
immune cells and are involved in inflammatory responses. Our recent studies showed that
cholesterol and sphingolipids are major determinants of podocyte function and survival, and that
sphingomyelin phosphodiesterase acid like 3b (SMPDL3b), a lipid raft associated protein, plays
an important role in podocyte function and regulates the availability of bioactive sphingolipids.
Expression of SMPDL3b is significantly upregulated in patients and mouse models of
glomerular diseases, irrespective of its metabolic or non-metabolic origin. Additionally, we
demonstrated an activation of glomerular stimulator of interferon genes (STING), a cytosolic
DNA sensing pathway, in CKD in association with increased inflammation and levels of
apoptotic podocyte death. Importantly, the recent studies have also demonstrated the
importance of lipids in regulation of STING-mediated immune response and the feedback loop,
where STING expression has a role in the regulation of lipid metabolism. However, whether
SMPDL3b may affect STING activation and contribute to podocyte injury remains unknown.
Release of mitochondrial DNA (mtDNA) into the cytoplasm seems to be one of the central
causes to STING activation. Our preliminary data demonstrated impaired PTEN-induced

putative kinase 1 (PINK1)-mediated mitophagy, a form of selective elimination of damaged or
excessive mitochondria, in both, clinical and experimental CKD. Importantly, these findings are
accompanied by increased mtDNA levels in the cytosolic fraction of kidney cortices in vivo.
However, the mechanism of mtDNA escape in podocytes remains unclear. In this project, we
propose to elucidate the complex mechanism involved in mtDNA leakage into cytosol and
SMPDL3b-dependent STING activation in podocytes leading to CKD development and
progression, which could unveil novel targets to treat CKD.

University of Pennsylvania
Department of Neuroscience

John A. Dani, Ph.D.- Principal Investigator
Philadelphia, Pennsylvania
Grant Award - 5 years
“Potential Common Therapeutic Target to Decrease the Vulnerability to Addictions”

The proposal arises from our studies showing that stress, nicotine, and cocaine all converge onto a common molecular target, causing downregulation of KCC2, which defines the chloride gradient and chloride reversal potential for GABAergic neurons of the midbrain. This drug-induced downregulation of KCC2 function alters GABAergic inhibition and alters midbrain GABAergic circuitry, such that addictive drug use acts to increase its own self-administration.

In new preliminary results, we have found that correcting KCC2 downregulation by boosting its activity, for example with CLP290, corrects midbrain circuitry and decreases drug self-administration.

Therefore, the specific aims of the study are:

1. To determine whether diverse addictive drugs downregulate KCC2 and characterize the time course of functional KCC2 downregulation.

2. To determine whether up or downregulation of KCC2, per se, can affect addictive drug self-administration and examine potential therapeutic avenues to decrease drug use by increasing KCC2 activity.

This application has the objective of enhancing and accelerating research on a potentially common drug target to decrease the vulnerability for abusive addictive drug use.

Albert Einstein Healthcare Network
Moss Rehabilitation Research Institute

Dylan J. Edwards, PhD - Principal Investigator. Moss Rehabilitation Research Institute
Shailesh Kantak, PhD - Co-Principal Investigator. Moss Rehabilitation Research Institute
Aapo Nummenmaa, PhD - Co-Principal Investigator. Massachusetts General Hospital Martinos Center
Philadelphia, Pennsylvania and Boston, Massachusetts
Grant Award - 3 years
“Precision Targeting for Transcranial Magnetic Stimulation Treatment in Stroke”

Transcranial magnetic stimulation (TMS) is approved in the United States in psychiatric and neurosurgical applications. In recent years, there has been advancement in the sophistication of stimulation devices using frameless stereotaxic neuronavigation and brain imaging (MRI), yet treatment targeting and dosing remains rudimentary. There is presently a critical need to enhance the precision of targeting and dosing that might underlie individual variability in treatment response. With recent advances from our ongoing collaborator (Dr. Nummenmaa) at the renowned Martinos Center for Biomedical Imaging at Massachusetts General Hospital, we have the ability to verify in the human TMS laboratories at MRRI (Dr. Edwards, Dr. Kantak), anatomically precise TMS electric fields in the human brain, which has thus far been elusive. Drs. Edwards and Kantak have a track record of clinical TMS mapping and Dr Nummenmaa of advanced electromagnetic modeling for TMS including real-time targeting and dosimetry.

The intention of the current study is to validate advanced real-time mapping methods that would be useful in rTMS treatment studies including depression, neurosurgery, and stroke.

University of ROCHESTER MEDICAL CENTER
DEPARTMENTS OF Psychiatry, Ophthalmology, and Radiology

Steven Silverstein, Ph.D. - Co-Principal Investigator
Joy Choi, M.D. - Co-Principal Investigator*
Rochester, New York
Grant Award - 3 years
“Retinal Vasculopathy as a Proxy for Cerebral Vascular Health and a Marker of Cognitive Improvement in Advanced Heart Failure Patients after LVAD Implantation”

The purpose of this project is to evaluate changes in retinal microvasculature as a biomarker of changes in cerebral blood flow and cognition in heart failure patients from before to after left ventricular assist device (LVAD) implantation. The retinal vasculature measure we will use is optical coherence tomography angiography (OCTA).

We will compare three groups of people: heart failure patients who receive LVAD, heart failure patients who do not receive LVAD, and a group of people with non-proliferative diabetic retinopathy. Groups will be compared at three timepoints corresponding to pre-LVAD implant for the LVAD group, and then three and six months later.

The specific aims of the study are as follows:

1. To determine whether cognition and cerebral blood flow (CBF) improve after LVAD implantation, and whether the extent of post-LVAD-implant CBF change predicts change in cognition scores over time;

2. To determine the degree to which measures of retinal vasculature and neural layer thickness predict baseline data on CBF and cognition, and whether changes in retinal vasculature and neural layer thickness from before to after LVAD implant predict the extent of change on CBF and cognition during this time period;

3. To determine whether retinal biomarkers of brain health predict changes in patient quality of life and other aspects of cardiac disease-relevant psychological functioning (e.g., levels of depression symptoms) from pre- to post-LVAD implant; and

4. To determine the degree to which:

   1. measures of neural functioning in the eye (considered to be a proxy of brain neuronal health) are affected by LVAD treatment, and

   2. changes in retinal function improve the prediction of improvements in cognitive and overall functioning (from Aim 3) in LVAD patients, over and above prediction achieved by retinal vascular measures.

We predict a significant group x time interaction effect, wherein improvements in cognition, retinal vasculature and neural thickness, and visual perception improve significantly over time for the LVAD group, but do not change over time for the other two groups. We also expect to find that retinal microvasculature measures predict both baseline functioning and degree of change in the other outcome measures, which would further establish OCTA as a useful biomarker in cardiology and neuropsychiatry.

University of MIAMI
Department of Medicine,
Katz Family Division of Nephrology and Hypertension

George W. Burke, M.D. - Principal Investigator
Alla Mitrofanova, Ph.D. - Co-Principal Investigator*
Miami, Florida
Grant Award - 2 years
“Role of the cGAS-STING Pathway in Glomerular Diseases”

*As a result of this work, Dr. Mitrofanova was awarded the 2021 Carl W. Gottschalk Research Scholar Grant. You can read the interview with her here.

Chronic kidney disease is a major health problem worldwide with a significant impact on the cardiovascular disease and no cure options. A glomerulus, which is tuft of the smallest blood vessels constantly filtering blood, suffers first in a case of kidney disease. Podocytes are highly specialized cells of the glomerular filtration barrier that play a pivotal role in maintaining glomerular filtration function and are target cells of injury in chronic kidney disease, including focal segmental glomerulosclerosis (FSGS) and diabetic kidney disease (DKD). Chronic inflammation and oxidative stress considerably promote the progression of DKD and FSGS.

Our recent studies showed that cholesterol and sphingolipids are major determinants of podocyte function and survival, and that sphingomyelin phosphodiesterase acid like 3b (SMPDL3b), a lipid raft associated protein that regulates cell plasma membrane fluidity, plays an important role in podocyte function. We demonstrated that SMPDL3b expression is suppressed in glomeruli of patients with recurrence of FSGS and upregulated in patients with DKD. This results in altered sphingolipid metabolism and disbalance in bioactive sphingolipids such as ceramide-1-phosphate and, possibly, sphingosine-1-phosphate in podocytes. Both bioactive sphingolipids are known to regulate many important cell functions, such as inflammation, senescence, programmed cell death, metabolic reactions, or energy production. We furthermore demonstrated that in DKD, podocyte mitochondrial dysfunction occurs, which is associated with reduced oxygen consumption capabilities and accumulation of cardiolipin, the most important phospholipid of the inner mitochondrial membrane. Similarly, abnormalities in mitochondrial function were found to cause severe early-onset FSGS with the innate immune activation. More recently, the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway, a component of a cell innate immune system, has been shown to regulate inflammation and energy homeostasis under obesity conditions, in acute kidney injury and chronic kidney disease.

Based on our previous studies, we will examine if changes in expression of SMPDL3b causes sphingolipid-mediated mitochondrial dysfunction leading to podocyte injury through chronic activation of the cGAS-STING signaling pathway.

2019 CMRF Grantees

University of Colorado
Department of Medicine

Lori A. Walker, Ph.D.- Principal Investigator
Denver, Colorado
Grant Award - 3 years
“Elucidating the Mechanisms Underlying Cannabinoid-mediated Changes in Smooth Muscle Contractility”

Currently, it is estimated that in the United States, over 35 million people regularly (2X week or more) use cannabis with 8.1 million people using cannabis daily. However, with the increase in cannabis use, there is a growing number of observational case reports linking cannabis use to adverse cardiovascular outcomes including myocardial infarction and stroke, attributed in large part to aberrant contraction of blood vessels.

Since its discovery in the early 1990s, the endocannabinoid system (ECS) has been increasingly implicated in key signaling pathways in both health and disease and is emerging as a key regulator of cellular homeostasis. While the ECS is thought to be involved in homeostasis, there is no consensus regarding the effects of cannabinoids on the cardiovascular system, as increases in the ECS have been linked to both increases and decreases in blood pressure.

Accordingly, we will conduct a mechanistic study using a rodent model for hypertension that examines the direct effects of cannabinoid receptor stimulation on vascular (blood vessel) smooth muscle at the level of isolated muscle and in the whole animal. We have compelling preliminary data demonstrating differential effects of cannabinoid receptor stimulation on different types of smooth muscle; visceral (gut) smooth muscle has reduced contractility with cannabinoid receptor stimulation whereas vascular smooth muscle contractility is increased by cannabinoid receptor stimulation, representing a potential novel mechanism underlying systemic hypertension.

It is possible that the vascular cannabinoid receptors will represent novel targets for the design of new drugs for the treatment of hypertension.

University of Colorado
Department of Medicine

Mary Weiser-Evans, Ph.D.-Principal Investigator
Denver, Colorado
Grant Award - 1 year
“Epigenetic Control of Pathological Vascular Remodeling: Role of Smooth Muscle PTEN Regulation”

Cardiovascular diseases remain the leading cause of death in the United States and globally, accompanied by a huge financial burden. In the United States, coronary artery disease is the leading cause of death, estimated to underlie one third of all deaths in those over 35 years of age.

Atherosclerosis and restenosis are chronic and acute inflammatory vascular diseases, respectively, characterized by significant vascular remodeling. Activation of resident vascular smooth muscle cells (SMCs) plays a critical role in remodeling and is a key event promoting disease progression. This is marked by a shift from a differentiated, contractile phenotype to a dedifferentiated, inflammatory phenotype. While the concept of SMC phenotypic modulation is well-accepted, the mechanisms regulating these SMC transitions are complex. Importantly, there are no therapeutics that target this shift to prevent both the loss of the SMC contractile phenotype and the increased inflammatory environment.

Our previous work demonstrated that regulation of the tumor suppressor, PTEN, is critical in pathological vascular remodeling. We showed that PTEN inactivation promotes SMC phenotypic transition to an inflammatory phenotype and established that loss of PTEN directly correlated with SMC dedifferentiation and human atherosclerosis progression and complexity. Further, our studies indicate that systemic upregulation of PTEN is sufficient to maintain the mature differentiated SMC phenotype and prevent vascular disease progression, an entirely novel concept for the treatment of cardiovascular disease.

Guided by our preliminary results, we will examine the effects of PTEN upregulation as a highly significant therapeutic target through direct effects on SMC phenotype and crosstalk with recruited immune cells.

University of Pennsylvania
Department of Neuroscience

John A. Dani, Ph.D.- Principal Investigator
Philadelphia, Pennsylvania
Grant Award - 3 years
“Early Life Experiences Change Midbrain Circuits & Alter Neuropsychiatric Risk”

A person’s risk for mental illness is defined mainly by the individual’s genetic predisposition modified by the individual’s lifetime experiences. Adolescence is a particularly vulnerable time for modifying an individual’s risk profile. Our recent research has focused on two common adolescent exposures that alter the risk for neuropsychiatric disease: stress and nicotine. Nearly 1/3 of high school students say they are overwhelmed by stress, and nicotine vaping with electronic cigarettes (e-cigs) is increasing rapidly among adolescents. Our preliminary results that underlie this application have shown that adolescent stress or nicotine exposure produces long-lasting changes in midbrain, mesocorticolimbic circuitry. This circuitry is particularly important for shaping behavior, and its dysfunction contributes to mental illness. Adolescent exposure to nicotine is particularly important right now because nicotine vaping with e-cigs recently increased nine-fold among US high school students and six-fold among middle school students. In addition, more than twice as many students reported using e-cigs compared to regular cigarettes. Guided by our preliminary results, we will examine the mechanisms underlying the changes in midbrain circuitry induced by adolescent nicotine exposure or by stress. Acting at those mechanistic targets, we also will attempt to reverse the circuitry therapeutically back toward the original condition.