The UC Irvine Institute for Clinical and Translational Science NRSA trainee program is funded by the National Institutes of Health National Center for Advancing Translational Science (NCATS) program designed to integrate training in clinical research and translational science into the core curriculum of pre- and postdoctoral trainees in medicine, public health, nursing, pharmaceutical sciences, biomedical and social sciences, physical sciences, engineering, and informatics.
Project Title: Investigating postnatal oxytocin as a treatment for cognitive and synaptic dysfunction in Fragile X model mic
Mentor: Christine Gall PhD
The neuropeptide oxytocin is in clinical trials for treatment of Fragile X syndrome (FXS) and other autism spectrum disorders. These intellectual disorders are characterized by multiple cognitive and social deficits, however almost all of research (human and animal) has only examined the effects of oxytocin on social deficits. The oxytocin system is important for the normal development of functional networks involved in social behaviors, but also for non-social cognition mediated by the hippocampal formation. Fragile X model mice (Fmr1-KO) have fewer oxytocin cells than wildtype (WT) and have lower oxytocin levels. Interestingly, oxytocin KO mice are considered an autism model because they exhibit behavioral deficits similar to Fmr1-KOs and other autism models. Recent studies demonstrated that postnatal oxytocin treatment restructured neural circuitry and restored social deficits measured in later adulthood in two different mouse models of autism. Fmr1-KOs have deficits in spatial and recognition memory tasks and impaired learning-related synaptic plasticity. Although there is some evidence that oxytocin enhances non-social cognition and plasticity in WT animals, exceptionally few studies have tested effects of oxytocin treatment on these deficits in FXS and/or other autism models. Proposed research will test if postnatal oxytocin treatment causes enduring changes that rescue cognitivefunction (Aim 1) and synaptic plasticity (Aim 2) of Fmr1-KOs in adulthood. As oxytocin is being tested in clinical trials with children, it is important to examine how oxytocin treatment during development effects non-social learning and memory as well as hippocampal function
Project Title: Functional Immunoimaging of Regulatory T cells in the GI Tract and the Roles of Biotin and Calcium
Mentor: Michael Calahan, PhD
There are limited effective treatments for autoimmune diseases like Inflammatory Bowel Disease in the GI tract because mucosal immunity remains incompletely understood. However, a growing body of evidence supports a role for regulatory T cells (Tregs) in suppressing these deleterious immune responses, but the cellular dynamics have not been explored. Furthermore, Ca2+ signaling and biotin metabolism appear to affect Tregs in ways that have not been studied. Building on the strong foundation built by Drs. Said and Cahalan in the fields of two-photon microscopy (2-P), Ca2+ signaling, and biotin (vitamin B7) physiology, our goal is to define cellular immunoregulatory events that explain how Tregs function to suppress intestinal inflammation. Utilizing two mouse models for enteritis (induced by adoptive transfer and spontaneous from biotin deficiency) in conjunction with reporter strains that have fluorescently labeled immune cells, we will visualize the cellular dynamics as Tregs interact with naïve and reactive effector T cells (Teffs) and with antigen-presenting cells (APCs) during initiation, progression and suppression of intestinal inflammation. The proposed studies will help to identify the mechanism of action of Tregs in the mesenteric lymph node and intestinal wall. We will use this information to evaluate the cellular basis of several promising therapeutics.
Aim 1. Treg suppression of T cell priming and trafficking in mesenteric lymph nodes. The initiation of intestinal inflammation occurs in mesenteric lymph nodes; we will use 2-P in an adoptive transfer mouse model for enteritis to elucidate the mechanism by which Tregs limit these inflammatory processes and examine the action of a therapeutic Sphingosine-1-phosphate receptor (S1P1) agonist.
1.1 How do Tregs control the choreography of T cell activation in the lymph node? We hypothesize that Tregs use a cellular competition-based mechanism to displace T cells from DCs to limit T cell priming. Furthermore, we propose that antigen-specific Tregs will be more effective than polyclonal Tregs in displacing reactive T cells. Using 2-P in mesenteric lymph nodes, we will be the first to visualize and define the real-time choreography of Tregs as they control initiation, progression and resolution of enteritis.
1.2 How does an S1P1 agonist modulate Treg function to prevent egress of Teffs from mesenteric lymph nodes? We propose a second mechanism by which Tregs limit inflammation at the level of the lymph node by preventing Teffs from entering the circulation. We hypothesize that Tregs function as “guardians of egress” by competing with Teffs for transendothelial migration in lymph node sinuses. We will use the pre-clinical S1P1 receptor agonist, CYM-5442 (a close analog to one being used in a stage 3 clinical trials for Ulcerative Colitis) to examine how it works synergistically with Tregs to sequester Teffs.
Aim 2. Biotin’s effect on Treg activity in the intestine. Biotin is known to play a role in immune function which is underscored by the severe enteritis seen in the recently developed biotin deficiency mouse model. Using our novel preparation of the intestine for 2-P, we will probe the cellular mechanisms of Treg function in the bowel wall including the impact of biotin on this process.
2.1 How do Tregs suppress inflammation in the intestine? We hypothesize that Tregs limit intestinal damage by displacing Teffs from APCs and altering their ability to present costimulatory molecules to Teffs, but Tregs from biotin deficient mice will be less effective. By visualizing the motility characteristics of Tregs, Teffs and APCs, we will determine how Tregs limit intestinal inflammation.
2.2 How does Biotin modulate Treg function in the intestine? We have seen severe colitis and reduced Tregs in a biotin deficient state and the complete reversal with supplementation. Using 2-P, we will visualize the cellular basis of this recovery guided by our hypothesis that biotin affects Treg trafficking and activity in the intestine.
Aim 3. Treg control of Ca2+ signaling to modulate immune responses. Ca2+ signaling in T cells is mediated by Orai1 Ca2+ channels and sustained Ca2+ signaling is necessary for immune responses. We will determine how Tregs suppress Ca2+ signaling in Teffs and evaluate therapeutic Ca2+ channel blockers.
3.1 How do Tregs control Ca2+ signaling in Teffs? Using both the adoptive transfer and biotin deficiency models, we will monitor the real-time frequency and duration of Ca2+ signals in inflammatory cells and how it changes in the presence of Tregs. We hypothesize that Tregs utilize contact-dependent suppression of Ca2+ signaling in Teffs to induce their release from dendritic cells.
3.2 How does Orai1 channel blockade effect enteritis? Orai1 channel blockade has been effective in suppressing Teffs without altering Tregs and we hypothesize that blocking Orai1 will inhibit immune synapse formation between Teffs and APCs giving a competitive advantage to Tregs in the intestine.
Project Title: Clinically translatable multimodal optical imaging platform for quantitative assessment of
cerebral hemodynamic response to cardiac arrest and resuscitation
Mentor: Bruce Tromberg, PhD
Nearly 80% of people who survive cardiac arrest (CA) enter a state of coma, with low likelihood of complete recovery of brain function (Madi and Holzer 2004). The extent of vascular recovery following cardiopulmonary resuscitation (CPR) is closely related to the level of brain damage suffered by the patient (Bisschops 2012). Therefore, there is great need to quantitatively characterize the perfusion, oxygenation, and metabolism of the brain in response to CA and CPR. Biomedical optics techniques, using light to interrogate tissue, can help address this unmet clinical need by quantitatively monitoring cerebral hemodynamic response to perturbation.
We will employ a non-contact, real-time, label-free, quantitative in vivo optical imaging system to interrogate the brain’s response to CA and CPR in a rat model. This technology will include laser speckle imaging (LSI) and multispectral spatial frequency domain imaging (SFDI) instrumentation to interrogate changes in cerebral blood flow and oxygenation. By using LSI and SFDI simultaneously, we can calculate the relative cerebral metabolic rate of oxygen consumption. This valuable physiological parameter will enable quantitative characterization of cerebral recovery after CPR, directly correlated with electroencephalogram (EEG) and arterial blood gas measurements acquired during the experiment.
This optical imaging platform can provide valuable insight into cerebral response to CA and CPR under different clinical interventions. The technology has the potential to (1) provide metrics for assessing cerebral response to CA, CPR, and arousal from coma in a clinical setting, and (2) motivate potential improvements to intensive care for patients with CA-induced coma.
Project Title: Noninvasive, point-of-care monitoring of neonatal intestinal blood flow
Mentor: Bernard Choi PhD
Project Summary: Intestinal failure (IF) in neonates is a serious condition characterized by a reduction in the functional intestinal mass, leading to undernourishment. IF is caused by several developmental issues associated with poor blood flow to the intestines in preterm infants. We intend to design and build two optical imaging devices, based on the principles of Laser Speckle Imaging (LSI), to non-invasively measure blood flow. The first device will be used in the neonatal intensive care unit (NICU) at CHOC Children’s Hospital (CHOC) to monitor intestinal blood flow, and the second device will be used in the CHOC operating room (OR) during surgery to map intestinal blood flow. The first LSI device will allow clinicians to objectively monitor and assess the health of the neonatal intestine. In doing so, we expect to assist clinicians in diagnosing neonates with IF sooner, which would allow surgical intervention to have the greatest impact. The surgical interventions required to treat IF are challenging, as there is currently no tool used to reliably identify ischemic regions within the intestine during surgery. The second LSI device will provide wide field, real-time blood flow maps of the intestines that would allow surgeons to objectively identify ischemic portions of the intestines that need to be resected. After the ischemic tissue is removed, the LSI device can also be used to immediately assess whether successful anastomosis of the remaining healthy intestine has been achieved.
Project Title: Targeting the Rho/ROCK pathway for treatment of VHL-deficient Clear Cell Renal Cell Carcinoma
Mentor: Olga Razorenova, PhD
The Von Hippel-Lindau (VHL) tumor-suppressor gene is mutated/lost in about 90% of Clear Cell Renal Cell Carcinomas (CC-RCCs). Because VHL-deficient CC-RCCs resist current therapies and frequently metastasize, identifying effective new therapies will be crucial for treatment of CC-RCC patients. By screening the Library of Pharmacologically Active Compounds, we identified the Rho-associated protein kinase (ROCK) inhibitor Y- 27632, which selectively targets VHL-deficient cells. Our follow-up in vitro experiments have shown that knockdown of Rho GTPase C (RhoC) and ROCK1 leads to VHL-deficient CC-RCC death and proliferation defect. In this proposal, we hypothesize that VHL loss leads to over-activation of the Rho/ROCK pathway due to the loss of RhoC/VHL interaction. We further hypothesize that targeting the Rho/ROCK pathway in vivo will lead to suppression of VHL-deficient tumors with minimal toxicity to normal tissues. We will test these hypotheses through the following specific aims: (1) To determine the mechanism of the synthetic lethal interaction between VHL deficiency and ROCK inhibition in CC-RCC; (2) To assess the therapeutic potential of Rho/ROCK inhibitors in vivo. Under Aim 1 we will assess the impact of inhibition of upstream ROCK regulators on proliferation/survival of VHL-deficient CC-RCC. We will then further investigate the effect of VHL deficiency on Rho expression and activity. Under Aim 2 we will test the effect of Rho/ROCK inhibitors on primary tumor growth and metastasis in vivo using an orthotopic mouse model of CC-RCC. We expect this project to yield important results and identify a new class of therapeutics for CC-RCC.
Project Title: Neuronal activity patterns predict brain viability in a rodent model of ischemic stroke
Mentor: Robert Frostig, PhD
Improved stroke treatments are necessary as current therapeutic strategies are only effective for a subset of stroke patients and only reduce damage or its functional consequences. We have previously demonstrated a novel, noninvasive ischemic stroke treatment in rodents in which intermittent sensory (whisker) stimulation delivered early after ischemic onset induces retrograde reperfusion via collateral blood vessels and completely protects the ischemic cortex from impending infarct. However, the same intermittent sensory stimulation results in exacerbated damage if delivered hours after ischemic onset. Interestingly, the area of activity evoked normally by whisker stimulation is comparable in size to the late treated animals’ infarct size and approximately twice as large as that of occlusion controls, indicating that single whisker stimulation evokes neuronal activity capable of accounting for either the respective damage or protection of the region. These findings highlight the relevance of neuronal activity in the outcome of ischemic stroke. The relationship between neuronal activity and enhanced post-ischemic collateral blood flow remains unclear and should be studied to determine how neuronal activity contributes to such surprising protection. My main hypothesis is that neuronal activity after ischemic onset critically contributes to the fate of the ischemic cortex. Using multisite recordings across horizontal locations and all cortical depths, I propose to investigate the following aims in a rodent permanent Middle Cerebral Artery occlusion (pMCAO) model: Specific Aim 1: Evoked activity and activation spread after pMCAO. Specific Aim 2: Evoked activity and the effects of sensory stimulation on stroke outcome.
Project Title: Neuronal activity patterns predict brain viability in a rodent model of ischemic stroke
Mentor: Bruce Tromberg, PhD
I propose to accelerate the translation of diffuse optical spectroscopic imaging (DOSI) as a new method for assessing an array of cardiovascular disease (CVD) risk factors. DOSI is a quantitative, model-based technique that utilizes a broadband wavelength range, corrects for light scattering in tissue, and probes several centimeters into tissue. DOSI is particularly relevant for cardiovascular disease research because it measures absolute concentrations of the primary light absorber in blood, hemoglobin, as well as the oxygenation state of hemoglobin, lipid content, and water content. These features of DOSI allow for hemodynamic measurements, compositional analysis, and reliable comparisons across operators, instruments, and even separate institutions. In particular, I propose testing and validating the ability of DOSI to assess endothelial function, an important feature of healthy vasculature. I hypothesize that DOSI can provide a non-invasive means for assessing endothelial function, is able to sense the presence of CVD, and can be used to monitor the effectiveness of interventions to improve health.
We will study a population of patients with coronary artery disease (CAD) and compare with healthy control subjects. In particular, we will compare the DOSI assessment of endothelial function to the EndoPAT assessment, a well-characterized, non-invasive means for endothelial function assessment. We will also characterize the range of orthogonal information content available from DOSI, including metabolic, biochemical composition, and vascular reactivity factors. In addition, we will use DOSI to monitor CAD patients while they undergo interventions to improve cardiovascular health, testing the ability of DOSI to sense restoration of endothelial function.
Project Title: Bladder Cancer Dipstick Test: An Innovative Approach for Simultaneous Biomarker Discovery and Assay Development
Mentor: Andrej Luptak, PhD
Biomarkers harness the potential to improve the standard care of the current medical practices. Conventional methods for biomarker discovery rely on analytical techniques to identify biomarkers based on clinical or experimental observations, and once identified, validation and assay development follows. Unfortunately, discovery to translation of biomarkers for clinical practices require labor intensive, time consuming, and expensive procedures; thus, this process remains an enormous challenge. Due to this pitfall, not many reliable biomarkers have been identified and implemented for clinical practices. Therefore, new approaches are in urgent need to improve the effectiveness of biomarker discovery and translation. In this proposal, I aim to tackle this obstacle by utilizing in vitro selection technique with patient urine samples to evolve and isolate molecular binders that will recognize a panel of biomarkers capable of distinguishing cancerous from non-cancerous urine. Recent research has demonstrated that in vitro selection technique has the capabilities to generate molecular binders highly specific to a microorganism from a complex mixture sample without prior target separation and identification steps. Therefore, I hypothesize this method is also feasible for diseases. Applying in vitro selection technique with functional DNA aptamers encoded with signal generating capabilities forms a synergetic combination suited for the concurrency of biomarker discovery and assay development. Achieving this will be extraordinary and paradigm shifting for the future of medicine.