Vitalant

RFA-IP-24-046: NATIONWIDE COHORT OF BLOOD DONORS TO ESTIMATE BURDEN OF RESPIRATORY VIRUSES AND IMMUNOLOGIC RESPONSE

MECHANISMS AND CLINICAL EFFECTS OF MICROCHIMERISM IN TRANSFUSED TRAUMA PATIENTS

HEMATOPOIETIC AND IMMUNE DEVELOPMENT IN THE HUMAN CHORION - PROJECT SUMMARY/ABSTRACT: HEMATOPOIESIS OCCURS AT MULTIPLE SITES DURING HUMAN PRENATAL DEVELOPMENT. IN ADDITION TO THE PRIMARY INTRAEMBRYONIC HEMATOPOIETIC TISSUES, IT IS NOW RECOGNIZED THAT THE EXTRA-EMBRYONIC TISSUES – THE PLACENTA AND CHORION – CONTAIN HEMATOPOIETIC STEM CELLS. THE PRESENCE OF HEMATOPOIETIC STEM CELLS IN THESE TISSUES RAISES THE POSSIBILITY THAT THEY ACTIVELY CONTRIBUTE TO THE POOL OF BLOOD CELLS CONTAINED IN THE EXTRA-EMBRYONIC TISSUES. IN THIS PROPOSAL WE FOCUS ON THE HUMAN FETAL MEMBRANES, SPECIFICALLY THE CHORION. MACROPHAGES ARE PRESENT IN THE CHORION AND PRESUMABLY ACT AS SENTINELS AND FIRST RESPONDERS TO INFECTION AS WELL AS INVASION BY MATERNAL CELLS. THE PRESENCE OF THESE MACROPHAGES SUGGESTS THAT THE CHORION IS NOT JUST AN INERT PHYSICAL BARRIER. WE HYPOTHESIZE THAT THE HUMAN CHORION IS AN IMMUNOLOGICALLY ACTIVE BARRIER CONTAINING MACROPHAGES THAT DEVELOP IN SITU FROM HEMATOPOIETIC STEM CELLS THAT TOGETHER WITH OTHER CELLS IN THE CHORION MAINTAIN THE MATERNAL-FETAL INTERFACE TO PREVENT INFECTION AND IMMUNE REJECTION OF THE FETUS DURING PREGNANCY. IN THE FIRST OF THREE SPECIFIC AIMS, WE WILL INVESTIGATE IF CHORIONIC HEMATOPOIETIC STEM CELLS SERVE AS A LOCAL SOURCE OF PRECURSORS FOR MONOCYTIC CELLS FOUND IN THE CHORION. TISSUES ACROSS A RANGE OF GESTATIONAL AGES WILL BE EXAMINED USING FLOW CYTOMETRY, HEMATOPOIETIC PROGENITOR CULTURES, AND A TRANSPLANT MODEL, USING IMMUNODEFICIENT MICE, TO DETERMINE THE TYPES OF HEMATOPOIETIC PRECURSORS THAT EXIST IN THE CHORION. EXPERIMENTS WILL FOCUS ON DEMONSTRATING ACTIVE MYELOPOIESIS IN THE CHORION. IN THE SECOND AIM, WE WILL EXPLORE THE ROLE OF THE CHORIONIC MICROENVIRONMENT IN THE DEVELOPMENT AND FUNCTION OF MACROPHAGES. MESENCHYMAL STROMAL CELLS AND CYTOTROPHOBLASTS COMPRISE THE HEMATOPOIETIC NICHE IN THE CHORION, WHICH WILL BE STUDIED IN CULTURE TO TEST THEIR EFFECTS ON MONOCYTE DEVELOPMENT AND MACROPHAGE MATURATION. WE HYPOTHESIZE THAT THESE CELL TYPES SUPPORT THE DEVELOPMENT OF M2 MACROPHAGES TO FOSTER AN ANTI-INFLAMMATORY ENVIRONMENT IN THE CHORION. THIS WILL BE MORE SPECIFICALLY TESTED IN THE THIRD AIM, IN WHICH WE WILL INVESTIGATE THE IMMUNOMODULATORY ROLE OF CHORION CELLS IN PREVENTING REJECTION BY MATERNAL T-CELLS. PRELIMINARY DATA SHOW THE PRESENCE OF T CELLS IN THE CHORION OF BOTH MATERNAL AND FETAL ORIGIN. THE ORIGINS AND DISTRIBUTION OF T-CELLS IN THE CHORION WILL BE FURTHER INVESTIGATED AT DIFFERENT GESTATIONS. ADDITIONALLY, WE WILL DETERMINE IF THE T-CELLS ARE ACTIVE AND IF THEY ARE ENRICHED IN ANY SPECIFIC T-CELL SUBTYPES. T-CELLS FROM THE ADJACENT DECIDUA WILL BE ANALYZED FOR COMPARISON AS WELL AS FETAL T-CELLS FROM THE SPLEEN. WE WILL EXPLORE THE ROLES OF CHORIONIC MACROPHAGES, MESENCHYMAL STROMAL CELLS, AND CYTOTROPHOBLASTS IN THE REGULATION OF T-CELL ACTIVATION. TOGETHER THESE AIMS WILL SHOW THAT CHORIONIC MACROPHAGES DEVELOP LOCALLY FROM A POPULATION OF STEM CELLS IN THE CHORION. ALSO, THAT THESE MACROPHAGES, IN CONJUNCTION TO OTHER CELL CELLULAR POPULATIONS CONTRIBUTE TO GENERATE AN ACTIVE IMMUNOLOGICAL BARRIER AT THE EXTENSIVE MATERNAL-FETAL INTERFACE THAT INCLUDES THE FETAL MEMBRANES, WHICH FUNCTION TO WARD OFF INFECTION AND INVASION OF MATERNAL LYMPHOCYTES.

THE IMPACT OF ILLNESS AND MEDICAL TREATMENTS ON THE ALLOANTIBODY RESPONSE TO PLATELET TRANSFUSION

TRANSFUSION-RELATED IMMUNOMODULATION INFLUENCES INFECTIOUS DISEASE OUTCOMES

METAGENOMIC DETECTION OF EMERGING VIRUSES IN THE BLOOD SUPPLY

MODULATION OF GALECTIN-9 TO TARGET HIV PERSISTENCE - PROJECT SUMMARY WHILE THE ADVENT OF ANTIRETROVIRAL THERAPY (ART) HAS DRAMATICALLY REDUCED THE MORBIDITY AND MORTALITY ASSOCIATED WITH HIV INFECTION, VIRAL ERADICATION IS NOT ACHIEVABLE DUE TO THE PERSISTENCE OF LATENTLY-INFECTED CELLS DURING TREATMENT. ACCUMULATING DATA SUGGEST THAT HIV-INFECTED INDIVIDUALS OFTEN EXPERIENCE PERSISTENT IMMUNE DYSREGULATION, CHRONIC INFLAMMATION, AND ACCELERATED AGING EVEN IN THE SETTING OF ART-MEDIATED VIRAL SUPPRESSION. THESE REALITIES HAVE CREATED A PRONOUNCED INTEREST IN DEVELOPING STRATEGIES TO ERADICATE OR FUNCTIONALLY CURE HIV INFECTION. RECENT WORK FROM OUR GROUP AND OTHERS HAS DEMONSTRATED A HIGHLY IMPORTANT AND PLEIOTROPIC ROLE OF HUMAN GALECTIN-9 (GAL-9), AN IMMUNOREGULATORY Β-GALACTOSIDE-BINDING LECTIN, IN HIV PERSISTENCE. WE HAVE DEMONSTRATED THAT ADMINISTRATION OF GAL-9 POTENTLY REACTIVATES LATENT HIV IN CD4+ T CELLS, BY SIGNALING THROUGH SPECIFIC GLYCANS ON THE TARGET CELL SURFACE. WE HAVE ADDITIONALLY REVEALED THAT GAL-9 EXPRESSION IN TISSUES (E.G. CNS) SPATIALLY OVERLAPS WITH HIV INFECTION, AND HIGH LEVELS OF ENDOGENOUS, CIRCULATING GAL-9 IN PLASMA ARE STRONGLY ASSOCIATED WITH LONG-TERM MAINTENANCE OF INTACT HIV PROVIRAL DNA IN THE CD4+ T CELL COMPARTMENT DURING ART. MOREOVER, OUR RECENT STUDIES IN HIV-INFECTED HUMANIZED MICE HAVE REVEALED THAT EXOGENOUSLY ADMINISTERED GAL-9 SIGNIFICANTLY INCREASES TISSUE-ASSOCIATED HIV DNA AND RNA LEVELS. IN THIS STUDY, WE WILL COMPREHENSIVELY DETERMINE IF THERAPEUTIC INHIBITION OF GAL-9 SIGNALING AT THE TIME OF ART INITIATION CAN ACCELERATE CLEARANCE OF THE LATENT HIV RESERVOIR, RESOLVE HIV-ASSOCIATED INFLAMMATION, AND PREVENT VIRAL REBOUND FOLLOWING ART CESSATION. THESE GOALS ARE WELL ALIGNED WITH THE STATED OBJECTIVE OF PAR-23-297. IN AIM 1 OF OUR STUDY, WE WILL USE ESTABLISHED EX VIVO LATENCY MODELS TO ELUCIDATE THE MOLECULAR AND IMMUNOLOGIC MECHANISMS UNDERLYING GAL-9-MEDIATED EXPANSION OF THE HIV RESERVOIR. IN AIM 2, WE WILL DETERMINE HOW MANIPULATION (SUPPLEMENTATION OR BLOCKADE) OF GAL-9 SIGNALING IMPACTS THE HIV RESERVOIR AND VIRAL REBOUND KINETICS POST-ART CESSATION IN VIVO. OUR MULTIDISCIPLINARY INVESTIGATIVE TEAM AT VITALANT RESEARCH INSTITUTE / UCSF, WEILL CORNELL MEDICINE, AND YALE UNIVERSITY BRINGS A DIVERSE SYSTEMS BIOLOGY APPROACH TO INVESTIGATE THE IMPACT OF GAL-9 ON HIV PERSISTENCE, INCLUDING TOOLS DEVELOPED BY OUR GROUP TO MANIPULATE GAL-9 SIGNALING IN VITRO AND IN VIVO (RECOMBINANT, STABLE FORM EXOGENOUS GAL-9, AND A PANEL OF ANTI-GAL-9 MONOCLONAL ANTIBODIES THAT INTERFERE WITH ITS VIROLOGIC EFFECTS). OUR TRANSLATIONAL DESIGN WILL LEVERAGE CAREFULLY DESIGNED EX VIVO MODELS OF HIV PERSISTENCE, CLINICAL SAMPLES FROM PEOPLE LIVING WITH HIV (PLWH) ON SUPPRESSIVE ART, AND TWO COMPLEMENTARY MURINE MODELS OF HIV LATENCY. OUR FINDINGS WILL GUIDE THE DEVELOPMENT OF NOVEL HIV CURE STRATEGIES THAT LEVERAGE THE IMMUNOMODULATORY ACTIVITIES OF HOST GALECTINS.

PROPERTIES OF STORED RBCS: MINIMIZATION OF IMMUNE AND VASCULAR REACTIVITY

VIRAL/IMMUNE PARAMETERS OF DENGUE AND WNV IN DONORS: BLOOD SAFETY IMPLICATIONS

MODELING VIRAL ENTRY AND ITS INHIBITION USING SARS-COV

RBC STORAGE EFFECT ON COAGULATION, MICROPARTICLES AND MICROCHIMERISM IN CRITICALLY ILL PATIENTS

EFFECTS OF HUMAN GALECTIN-9 ON THE CNS HIV RESERVOIR

VIRAL ETIOLOGY OF IDIOPATHIC CHRONIC DIARRHEA IN RHESUS MACAQUES

PROTECTIVE B-CELL RESPONSES IN CHIKUNGUNYA VIRUS INFECTION

EFFECT OF BLOOD DONOR SEX AND TESTOSTERONE ON PREDISPOSITION TO HEMOLYSIS IN STORED RED BLOOD CELLS

INNATE SENSING OF CELL-FREE DNA AND THE INTERFERON-MEDIATED CONTROL OF HIV IN VIVO - PROJECT SUMMARY WHILE THE ADVENT OF ANTIRETROVIRAL THERAPY (ART) HAS DRAMATICALLY REDUCED THE MORBIDITY AND MORTALITY ASSOCIATED WITH HIV INFECTION, A CURE IS NOT ACHIEVED DUE TO THE PERSISTENCE OF LATENTLY-INFECTED CELLS DURING TREATMENT. ACCUMULATING DATA SUGGEST THAT HIV-INFECTED INDIVIDUALS OFTEN EXPERIENCE PERSISTENT IMMUNE DYSREGULATION, CHRONIC INFLAMMATION, AND ACCELERATED AGING EVEN IN THE SETTING OF ART-MEDIATED VIRAL SUPPRESSION. THESE REALITIES HAVE CREATED A PRONOUNCED INTEREST IN DEVELOPING STRATEGIES TO CURE HIV INFECTION. IDENTIFYING THE HOST MOLECULAR DETERMINANTS OF HIV PERSISTENCE AND REBOUND KINETICS FOLLOWING CESSATION OF ANTIRETROVIRAL THERAPY (ART) WILL BE CRITICAL IN DEVELOPING EFFECTIVE STRATEGIES TO CURE HIV INFECTION. WE RECENTLY APPLIED A COMPREHENSIVE SYSTEMS PROFILING APPROACH TO PLASMA SAMPLES FROM HIV-INFECTED INDIVIDUALS WHO UNDERWENT ANALYTICAL TREATMENT INTERRUPTION (ATI), TO IDENTIFY CIRCULATING HOST FACTORS THAT ENABLE PREDICTION OF HIV REBOUND KINETICS FOLLOWING ART INTERRUPTION, AND SERVE AS POTENTIAL PHARMACOLOGICAL TARGETS TO PROMOTE DURABLE VIROLOGIC REMISSION IN THE ABSENCE OF ART. THE HOST FACTOR EXHIBITING THE STRONGEST STATISTICAL ASSOCIATION WITH HIV REBOUND TIMING FOLLOWING ART CESSATION WAS CIRCULATING CELL-FREE DNA (CFDNA) IN PLASMA. SPECIFICALLY, INCREASED CFDNA ABUNDANCE IN PLASMA WAS ASSOCIATED WITH DELAYED TIME-TO-REBOUND. CFDNA, RELEASED INTO CIRCULATION DURING PROGRAMMED CELL DEATH, HAS BEEN EXPLOITED EXTENSIVELY IN THE REALM OF CLINICAL ONCOLOGY (OFTEN TERMED “LIQUID BIOPSY”). HOWEVER, CFDNA REMAINS LARGELY UNEXPLORED IN THE SETTING OF HIV INFECTION. IN THIS R01 PROPOSAL, WE RIGOROUSLY EXPLORE CFDNA AS A BIOMARKER OF HIV DISEASE STATES, AND WE INVESTIGATE THE MOLECULAR AND IMMUNOLOGIC MECHANISMS LINKING CFDNA TO VIRAL EXPRESSION AND REBOUND. OUR CENTRAL HYPOTHESES ARE THAT 1) CFDNA REFLECTS THE DEATH RATE OF HIV-INFECTED CELLS IN VIVO; AND 2) CFDNA PROMOTES AN ANTIVIRAL STATE IN THE HOST (E.G. INDUCTION OF TYPE I INTERFERON RESPONSE) WHICH PREVENTS VIRAL REBOUND. OUR PROJECT FEATURES AN INVESTIGATIVE TEAM WITH BASIC, TRANSLATIONAL, AND CLINICAL EXPERTISE, AND LEVERAGES LARGE COLLECTIONS OF CLINICAL SAMPLES FROM WELL-CHARACTERIZED HIV-INFECTED INDIVIDUALS. IN AIM 1, WE WILL APPLY NEXT-GENERATION SEQUENCING (NGS) APPROACHES TO PLASMA SAMPLES FROM HIV-INFECTED INDIVIDUALS TO EXTEND AND ENHANCE THE PROGNOSTIC SIGNIFICANCE OF CFDNA AS A BIOMARKER OF NATURAL HIV CONTROL IN VIVO. IN AIM 2, WE WILL EVALUATE HOW THE UPTAKE AND SENSING OF CFDNA BY HIV TARGET CELLS IMPACTS CELL FATE AND THE REACTIVATION AND REPLICATION OF HIV. IN AIM 3, WE WILL DETERMINE HOW HIV-ASSOCIATED MITOCHONDRIAL DYSFUNCTION LEADS TO CFDNA EXTRUSION AND ULTIMATELY INDUCES TYPE I INTERFERON RESPONSES THAT PREVENT HIV REBOUND. OUR PROJECT WILL ADVANCE THE DEVELOPMENT AND EVALUATION OF HIV CURE STRATEGIES.

EFFECTS OF CELL-INTRINSIC IMMUNITY ON ESTABLISHMENT AND REVERSAL OF HIV LATENCY

METAGENOMIC ANALYSIS OF THE HUMAN VIROME

MECHANISMS REGULATING ALLOIMMUNIZATION AND TOLERANCE WITH PATHOGEN REDUCTION AND TRANSFUSION OF ALLOGENEIC PLATELETS

EXTRACELLULAR VESICLES AS BIOMARKERS AND THERAPEUTIC TARGETS FOR ATHEROSCLEROTIC CARDIOVASCULAR DISEASE IN PERSONS LIVING WITH HIV - PROJECT SUMMARY: DESPITE ANTIRETROVIRAL THERAPY (ART), PEOPLE LIVING WITH HIV (PLWH) HAVE A TWO-FOLD INCREASED RISK OF CARDIOVASCULAR DISEASE (CVD) COMPARED WITH THE GENERAL POPULATION. WHILE PERSISTENT SYSTEMIC INFLAMMATION AND IMMUNE ACTIVATION APPEAR TO PREDICT THIS RISK, IT HAS BEEN CHALLENGING TO IDENTIFY THE PATHOPHYSIOLOGIC PATHWAYS THAT CAN BE TARGETED TO REDUCE INFLAMMATION AND CARDIOVASCULAR RISK IN TREATED HIV INFECTION. EXTRACELLULAR VESICLES (EVS) PLAY A SIGNIFICANT ROLE IN REGULATING PATHOPHYSIOLOGIC PATHWAYS LIKE INFLAMMATION AND ANGIOGENESIS, AND THEY ARE IMPLICATED IN KEY ASPECTS OF HEART DISEASE. TO DATE, LIMITED DATA EXIST ON THE MOLECULAR MECHANISMS UNDERLYING THE EFFECT OF EVS ON ATHEROSCLEROSIS PROGRESSION, LEAVING OPEN THE QUESTION OF HOW EVS MIGHT IMPACT THE RESOLUTION OF INFLAMMATION TO INFLUENCE CVD RISK IN HIV. THIS PROPOSAL IS INNOVATIVE IN THAT IT WILL EMPLOY HIGH-THROUGHPUT TESTING PLATFORMS FOR CHARACTERIZING EVS AND THEIR CARGO ON LONGITUDINAL SAMPLES FROM WELL-CHARACTERIZED COHORTS OF PERSONS WITH HIV AND CVD AND WILL UTILIZE A NOVEL MOUSE MODEL OF HIV ATHEROSCLEROSIS AND CUTTING-EDGE IN VITRO MODELS TO IDENTIFY AND VALIDATE NOVEL INFLAMMATORY MEDIATORS CARRIED BY EVS AND PROVIDE POTENTIAL NOVEL TARGETS FOR INTERVENTION. IN AIM 1, WE WILL TEST THE HYPOTHESIS THAT 1) IMMUNOLOGICALLY ACTIVE EVS DERIVED FROM HIV+ ADULTS ON ART WILL CARRY PRO- INFLAMMATORY MEDIATORS THAT WILL BE ASSOCIATED WITH CAROTID ARTERY INTIMA-MEDIA THICKNESS PROGRESSION; AND 2) EVS FROM HIV+ ADULTS WITH HIGHER CVD RISK WILL CARRY A UNIQUE MIRNA SIGNATURE AND WILL BE ASSOCIATED WITH A SPECIFIC INFLAMMATORY PATHWAY (E.G. NLRP3 INFLAMMASOME SIGNALING/CASPASE-1 ACTIVATION). IN AIM 2, WE WILL DEVELOP A NOVEL ACCELERATED ATHEROSCLEROSIS MOUSE MODEL ASSOCIATED WITH HIV INFECTION TO TEST THE HYPOTHESIS THAT 1) SYSTEMIC ADMINISTRATION OF EVS DERIVED FROM HIV-1 TG26/APOE−/− MICE WILL MORE STRONGLY INDUCE PRODUCTION OF PROINFLAMMATORY MEDIATORS WHEN INFUSED INTO RECIPIENT MICE AND WILL TRANSFER MOLECULES THAT ALTER EXOSOMAL MIRNA EXPRESSION IN RECIPIENT CELLS; 2) DNA-CONTAINING EVS FROM HIV-1 TG26/APOE−/−MICE WILL INDUCE EPIGENETIC CHANGES IN RECIPIENT CELLS; AND 3) THE ADMINISTRATION OF EVS DERIVED FROM VX-765 OR MCC950-TREATED ATHEROSCLEROTIC MICE WILL PRODUCE CIRCULATING EVS THAT LACK THE ABILITY TO AFFECT THE EXOSOMAL MIRNA EXPRESSION IN RECIPIENT CELLS. IN AIM 3, WE WILL TEST THE HYPOTHESIS THAT 1) EVS FROM THOSE WITH HIGH CVD RISK WILL DIRECTLY CAUSE CARDIOMYOCYTE AND ENDOTHELIAL CELL DYSFUNCTION AND PROMOTE INCREASED CELL DEATH COMPARED TO CONTROLS; 2) CASPASE INHIBITORS (CASPASE-1/INFLAMMASOME INHIBITION) WILL MITIGATE APOPTOSIS IN CARDIOMYOCYTES AND ENDOTHELIAL CELLS MEDIATED BY EVS DERIVED FROM THOSE WITH HIGH CVD RISK; AND 3) EV SIGNALING WILL BE DEPENDENT ON EV RNA CONTENT FOR DELIVERY OF A PRO-INFLAMMATORY SIGNAL. WE ALSO WILL FOCUS ON THE DEVELOPMENT OF GENOME-WIDE CRISPR SCREENS TO IDENTIFY THE CAUSAL VARIANTS, GENES, AND MOLECULAR MECHANISMS ON CARDIAC CELL FUNCTION. WE AIM TO CREATE A NEW PARADIGM FOR DISCOVERY OF NOVEL EV-BASED INTERACTIONS THAT CAN BE TARGETED TO PREVENT IMMUNE ACTIVATION AND INFLAMMATION THAT PERSISTS IN PLWH.

LEVERAGING STORED PLATELET BIOENERGETICS TO IMPROVE HEMOSTATIC FUNCTION UPON TRANSFUSION - PROJECT SUMMARY / ABSTRACT THE OVERARCHING AIM OF THIS PROPOSAL IS TO ADVANCE THE FIELD OF PERSONALIZED TRANSFUSION MEDICINE BY IDENTIFYING OPTIMAL SUPPLEMENTATION AND STORAGE PROCEDURES TO YIELD PLATELET PRODUCTS WITH MAXIMUM HEMOSTATIC CAPACITY TO IMPROVE OUTCOMES IN ACTIVELY BLEEDING PATIENTS. THERE HAS BEEN RENEWED INTEREST IN USING COLD-STORED PLATELETS (CS-PLT) FOR BLEEDING PATIENTS DUE TO THEIR INCREASED HEMOSTATIC FUNCTION AS COMPARED TO CONVENTIONAL ROOM TEMPERATURE-STORED PLATELETS (RT-PLT). WE HAVE IDENTIFIED THAT THERE IS A METABOLIC SIGNATURE ASSOCIATED WITH CS-PLT HEMOSTATIC FUNCTION UNDER PHYSIOLOGICALLY RELEVANT FLOW CONDITIONS. THIS SIGNATURE SUGGESTS THAT CS-PLT USE THIOL REDOX SWITCHES, AN UNDERAPPRECIATED AREA OF REDOX BIOLOGY, TO PROTECT FROM OXIDATIVE STRESS DURING STORAGE. YET THESE SWITCHES MAY ALSO CONFER ENHANCED HEMOSTATIC ACTIVITY. THE MECHANISMS BY WHICH THIOL REDOX SWITCHES MODULATE PLATELET ACTIVATION, ADHESION, AGGREGATION, AND PROCOAGULANT CAPACITY ARE NOT WELL DEFINED. IN ADDITION, THIS POINTS TO THE OPPORTUNITY TO ALTER THE REDOX POTENTIAL OF PLATELET STORAGE SOLUTIONS THROUGH METABOLIC SUPPLEMENTATION TO CREATE A PLATELET PRODUCT WITH IMPROVED HEMOSTATIC FUNCTION. LASTLY, THERE IS A PAUCITY OF DATA CONCERNING THE IN VIVO HEMOSTATIC EFFICACY OF LONG-TERM STORED CS-PLT WITH OR WITHOUT METABOLIC SUPPLEMENTATION. THUS, OUR CENTRAL HYPOTHESIS IS THAT COLD STORAGE INDUCES A REDOX ENVIRONMENT THAT IS ASSOCIATED WITH INCREASED HEMOSTATIC FUNCTION, AND THAT BY ADJUSTING THE REDOX POTENTIAL OF RT-PLT TO MIMIC THAT INCURRED BY COLD STORAGE WILL CONFER IMPROVED HEMOSTATIC CAPACITY. TO TEST THIS, WE HAVE PROPOSED THREE AIMS: (I) TO IDENTIFY THE THIOL REDOX STATUS OF CS-PLT COMPARED TO RT-PLT AND IDENTIFY THOSE SWITCHES THAT ARE ASSOCIATED WITH PLATELET HEMOSTATIC FUNCTION; (II) TO DETERMINE WHETHER METABOLIC SUPPLEMENTATION OF STORED PLATELETS TO INCREASE THE REDOX POTENTIAL POTENTIATES HEMOSTATIC FUNCTION; (III) AND TO DEMONSTRATE IN VIVO HEMOSTATIC ACTIVITY OF ALTERNATIVELY STORED PLATELET PRODUCTS IN MURINE MODELS. OUR COLLABORATIVE TEAM AND PREMIER TRANSFUSION MEDICINE RESEARCH ENVIRONMENT ARE IDEALLY SUITED TO PERFORM THESE STUDIES, TO INCLUDE THE USE OF MICROFLUIDIC MODELS OF HEMOSTASIS AND THROMBOSIS AND MURINE MODELS WHICH ASSESS PLATELET PRODUCT SAFETY AND EFFICACY. MOREOVER, THIS PROPOSAL ADDRESSES TWO SIGNIFICANT UNMET NEEDS. FIRST, THE NEED TO EXTEND AVAILABILITY AND/OR SHELF LIFE OF PRODUCTS DUE TO CRITICALLY LOW SUPPLIES. THIS IS ADDRESSED THROUGH OUR ASSESSMENT OF CS-PLT STORED UP TO 21 DAYS AND IDENTIFICATION OF SUPPLEMENTATION STRATEGIES TO IMPROVE THE METABOLIC “AGE” OF STORED PLATELETS. SECOND, THE NEED TO TRANSFUSE PATIENTS WITH PRODUCTS THAT WILL BE EFFECTIVE IN STOPPING BLEEDING. WE WILL PROVIDE MUCH NEEDED DATA ON IN VIVO HEMOSTATIC EFFICACY OF CS-PLT AND DETERMINE IF METABOLIC SUPPLEMENTATION OF PLATELETS CAN ENHANCE HEMOSTATIC FUNCTION. FINDINGS GENERATED BY THIS PROPOSAL WILL PROVIDE DETAILED MECHANISTIC DATA ON CS-PLT WITH THE AIM OF BOTH IMPROVING PLATELET INVENTORY MANAGEMENT AND MOST IMPORTANTLY, OUTCOMES IN ACTIVELY BLEEDING PATIENTS VIA PERSONALIZED TRANSFUSION MEDICINE.

EFFECT OF BLOOD DONOR SEX AND INTER-INDIVIDUAL VARIABILITY IN PLASMA TESTOSTERONE ON THE TRANSFUSION EFFECTIVENESS AND HEMOSTATIC POTENTIAL OF RED BLOOD CELLS AND PLATELETS - PROJECT SUMMARY / ABSTRACT THE LONG-TERM GOAL OF THIS PROPOSAL IS TO ADVANCE TRANSFUSION EFFECTIVENESS AND SAFETY BY ADDRESSING THE GAPS IN KNOWLEDGE OF TESTOSTERONE-MEDIATED CELLULAR CHANGES THAT IMPACT RED BLOOD CELL (RBC) AND PLATELET FUNCTION IN STORAGE AND AFTER TRANSFUSION. WE HYPOTHESIZE THAT DISRUPTION OF PHYSIOLOGICAL TESTOSTERONE SIGNALING INDUCED BY GENETIC, IDIOPATHIC OR THERAPEUTIC INTERVENTION (TESTOSTERONE REPLACEMENT THERAPY, TRT) CONTRIBUTES TO INTER-DONOR HETEROGENEITY IN THE QUALITY OF BLOOD COMPONENTS, AND TO ALTERED TRANSFUSION EFFECTIVENESS. THE SCIENTIFIC PREMISE IS BASED ON OUR FINDINGS THAT SUPRAPHYSIOLOGICAL CONCERTATION (>890 NG/DL; COMMON IN TRT) OF FREE (BIOAVAILABLE) TESTOSTERONE IN PLASMA IS CORRELATED WITH INCREASED OXIDATIVE HEMOLYSIS AND DECREASED MEMBRANE DEFORMABILITY. RELEVANT TO TRANSFUSION EFFECTIVENESS, GAMMA-IRRADIATED RBC UNITS FROM TRT DONORS HAD LOWER SURVIVAL SHORTLY AFTER INFUSION INTO IMMUNODEFICIENT MICE COMPARED WITH NON-TRT CONTROLS, AND PATIENTS’ HEMOGLOBIN INCREMENTS FOLLOWING A SINGLE RBC UNIT TRANSFUSION WERE REDUCED FOR RBC UNITS FROM TRT DONORS COMPARED TO THOSE FROM NON-TRT MALE DONORS (75% OF NON-TRT). OUR STUDIES IN PLATELETS SUGGESTED THAT EXOGENOUS TESTOSTERONE HAS A PRIMING EFFECT ON PLATELETS EVIDENCED BY INCREASED AGGREGATION AND MITOCHONDRIAL BIOENERGETICS, AND THAT TRT WAS ASSOCIATED WITH INCREASED EXPRESSION OF PLATELET ACTIVATION MARKERS IN STORAGE. IN AIM 1, WE WILL QUANTIFY THE CONCENTRATIONS OF FREE AND TOTAL TESTOSTERONE IN PLASMA FROM MALE DONORS FROM NHLBI’S REDS-III RBC-OMICS STUDY WITH COMPLETE GENOTYPED INFORMATION AND LINKED DATA OF RECIPIENT OUTCOMES AFTER RBC TRANSFUSION EVENTS. BY MEASURING PLASMA TESTOSTERONE CONCENTRATION, WE WILL BE ABLE TO DETERMINE THE PREVALENCE OF SUB- PHYSIOLOGICAL (HYPOGONADISM) AND SUPRAPHYSIOLOGICAL TESTOSTERONE AMONG MALE DONORS; CONDUCT GENOME-WIDE ASSOCIATION (GWA) STUDIES TO IDENTIFY SINGLE NUCLEOTIDE POLYMORPHISM (SNP) ASSOCIATED WITH DONOR PLASMA TESTOSTERONE CONCENTRATION; AND DEFINE THE IMPACT OF TESTOSTERONE CONCENTRATION AND IDENTIFIED SNPS ON TRANSFUSION EFFECTIVENESS (MEASURED BY CHANGES IN PATIENT’S HEMOGLOBIN AND BILIRUBIN INCREMENTS) USING THE REDS-III VEIN-TO-VEIN DATABASE. IN AIM 2, WE EVALUATE THE IMPACT OF HYPOGONADISM AND TRT ON PLATELET PHENOTYPE AND HEMOSTATIC FUNCTION IN THERAPY, IN STORAGE, AND IN A MOUSE MODEL OF TRANSFUSION. THE GOAL IS TO EVALUATE THE FEASIBILITY OF EXPANDING THE UTILIZATION OF BLOOD COMPONENTS DERIVED FROM INDIVIDUALS ON TRT. WE ANTICIPATE THAT THIS PROJECT’S OUTCOMES WILL ALLOW US TO: IDENTIFY GENETIC DETERMINANTS OF PLASMA TESTOSTERONE THAT IMPACT THE EFFECTIVENESS OF TESTOSTERONE THERAPY IN PATIENTS, AND RBC/PLATELET SURVIVAL IN STORAGE AND AFTER TRANSFUSION; BENEFIT THE EMERGING FIELD OF PRECISION TRANSFUSION MEDICINE BY ESTABLISHING BLOOD DONOR TESTOSTERONE THRESHOLD VALUES FOR TRANSFUSION EFFECTIVENESS AND SAFETY IN VULNERABLE PATIENT POPULATIONS (NEONATES/ANDROGEN SENSITIVE); IMPROVE TRT DONOR SCREENING TO MINIMIZE RECIPIENT EXPOSURE TO SUPRAPHYSIOLOGICAL FREE TESTOSTERONE; REEXAMINE CURRENT POLICIES THAT LIMIT OR BAN PLATELET DONATION BY TRT DONORS; AND EXPLORE DIFFERENTIAL UTILIZATION BASED ON TRT PLATELET CHARACTERISTICS IN CLINICAL SCENARIOS OF ACUTE BLEEDING LIKE TRAUMA AND SURGERY.

IMPROVING BLOOD SAFETY AND HIV TESTING IN BRAZIL: A RANDOMIZED CONTROLLED TRIAL

DOES RBC STORAGE AGE EFFECT INFLAMMATION, IMMUNE FUNCTION AND SUSCEPTIBILITY TO TRANSFUSION ASSOCIATED MICROCHIMERISM IN CRITICALLY III PATIENTS? ADV

BROAD SPECTRUM ANTIVIRALS TARGETING ENVELOPE PROTEOLYSIS AND VIRAL UNCOATING

A MULTIDISCIPLINARY APPROACH TO EDUCATION IN PEDIATRIC TRANSFUSION MEDICINE

PERSISTENCE OF WNV RNA IN BLOOD AND THE RISK OF TRANSFUSION-TRANSMISSION

A PROSPECTIVE EVALUATION OF CHRONIC B.MICROTI INFECTION IN SEROREACTIVE BLOOD DON

SEROLOGICAL PREVALENCE OF VIRAL HEMORRHAGIC FEVERS IN EQUATORIAL AFRICA

PROTECTIVE B-CELL RESPONSES IN CHIKUNGUNYA VIRUS INFECTION

VALIDATING THE LINK BETWEEN NXPH2 AND ALLOIMMUNIZATION

OPTIMIZATION OF XMRV AND OTHER MLV-RELATED VIRUS DETECTION FOR SCREENING OF BLOOD

NATURAL HISTORY OF ACUTE AND CHRONIC HCV IN BLOOD DONORS

HIGH THROUGHPUT MEASUREMENT OF ENVELOPE GENE DIVERSITY FOR AN HIV INCIDENCE ASSAY

EFFECTS OF CELL-INTRINSIC IMMUNITY ON ESTABLISHMENT AND REVERSAL OF HIV LATENCY

DEFINING DRIVERS OF MTDNA LEVELS IN BLOOD COMPONENTS AND THEIR IMPACT ON TRANSFUSION-RELATED LUNG INJURY - PROJECT SUMMARY BLOOD TRANSFUSION HAS PROVEN TO BE A LIFE-SAVING TREATMENT FOR THOSE WITH ACUTE AND CHRONIC BLOOD LOSS. NONETHELESS, BLOOD TRANSFUSION POSES ITS OWN RISKS, AND IN RECENT YEARS THE DANGERS OF TRANSFUSION ABOVE AND BEYOND THE INFECTIOUS RISKS HAVE GAINED ATTENTION. RELATIVELY LESS ATTENTION HAS BEEN PAID TO THE NON-INFECTIOUS COMPARED TO THE INFECTIOUS CONSEQUENCES OF TRANSFUSION IN THE TIME SINCE HIV APPEARED AS AN INFECTIOUS RISK OF TRANSFUSION. IN THIS PROPOSAL WE WILL EXPLORE A RECENTLY DISCOVERED GENETIC ASSOCIATION UNDERLYING MTDNA LEVELS IN RBC UNITS AND THE POTENTIAL TO INCREASE RISK OF ACUTE LUNG INJURY, WITH IMPLICATIONS FOR TRANSFUSION MEDICINE AND BEYOND. THIS PROPOSAL WILL EXPLORE A NEWLY DETERMINED LINK BETWEEN ANKLE1 AND MTDNA IN BLOOD DONOR PRODUCTS. THE IDEA THAT A NOVEL GENETIC MARKER OF BLOOD DONORS WHO COULD POTENTIALLY INCREASE THE RISK OF ACUTE LUNG INJURY AFTER TRANSFUSION WOULD REPRESENT AN IMPORTANT STEP IN FORWARDING THE CONCEPT OF PRECISION TRANSFUSION MEDICINE AND ENHANCING TRANSFUSION SAFETY. IN ADDITION TO POTENTIALLY ASSOCIATING ANKLE1 GENETIC VARIATION WITH MTDNA LEVELS AND ACUTE LUNG INJURY, THIS PROPOSAL WILL DEFINE WHICH COMPONENTS OF BLOOD HARBOR THE MAJORITY OF MTDNA. FURTHERMORE, LINKING MTDNA LEVEL WITH ANKLE1 PROTEIN EXPRESSION WOULD PROVIDE AN IMPORTANT MECHANISTIC LINK BETWEEN THE GWAS FINDINGS PRESENTED IN OUR PRELIMINARY DATA. IF ANKLE1 PROTEIN LEVEL IS SHOWN TO BE ASSOCIATED WITH MTDNA, IT WOULD PROVIDE A POTENTIAL DRUGGABLE TARGET TO AFFECT MTDNA IN BLOOD PRODUCTS. IN ADDITION TO STUDYING A NOVEL TOPIC, SEVERAL NEW ASSAYS WILL BE OPTIMIZED FOR THIS PROJECT. ANKLE1 WILL BE MEASURED VIA ELISA AND FLOW CYTOMETRY TO QUANTIFY EXPRESSION AT THE BULK AND CELLULAR LEVEL. WE WILL ALSO OPTIMIZE HOW DIFFERENT SAMPLE TYPES (WHOLE BLOOD VS. PLASMA) CAN BE ASSAYED, INCLUDING METHODS TO PREVENT HEMOGLOBIN FROM INTERFERING WITH THE ASSAYS USED. THIS PROPOSAL WILL ACCOMPLISH TWO MAIN GOALS. FIRST, WE WILL DETERMINE IF MTDNA IN BLOOD PRODUCTS PREDICTS SUBSEQUENT ACUTE LUNG INJURY IN TRANSFUSION RECIPIENTS. FINDING SUCH AN ASSOCIATION WOULD OPEN THE DOOR TO DECREASING THE RISK OF BLOOD TRANSFUSION BY EITHER SCREENING BLOOD PRODUCTS OR DEVELOPING METHODS TO REDUCE MTDNA LEVELS IN PRODUCTS AND TEST THESE MODIFIED PRODUCTS IN ANIMAL MODELS. SECOND, WE WILL DETERMINE WHERE MTDNA IS MOST PLENTIFUL IN BLOOD PRODUCTS AND EXPLORE THE HYPOTHESIS THAT ANKLE1 LEVELS WILL INVERSELY CORRELATE WITH MTDNA BURDEN. UNDERSTANDING THE REGULATION OF MTDNA WOULD ALLOW DEVELOPMENT OF POTENTIAL THERAPEUTICS TO MODIFY LEVELS IN BLOOD PRODUCTS.

POSSIBLE TRANSFUSION TRANSMITTED CEREBRAL AMYLOID ANGIOPATHY: EVALUATION OF TRANSMISSION OF AΒ USING THE RADAR REPOSITORY - PROJECT SUMMARY/ABSTRACT CEREBRAL AMYLOID ANGIOPATHY (CAA) IS A COMMON CAUSE OF SPONTANEOUS INTRACEREBRAL HEMORRHAGE (ICH), WITH A CASE FATALITY RATE OF 30-50%. CAA IS CHARACTERIZED BY AGGREGATION OF LIKELY MISFOLDED DEPOSITS OF AMYLOID Β (AΒ) IN THE CEREBRAL VASCULATURE, LEADING TO VASCULAR WEAKENING AND REPEATED, UNPROVOKED ICH. OVER THE PAST DECADE EVIDENCE HAS BEEN ACCUMULATING THAT AΒ EXHIBITS PRION-LIKE PROPERTIES, BEING OVERTLY TRANSMISSIBLE IN EXPERIMENTAL SETTINGS THROUGH A RANGE OF DIFFERENT PARENTERAL ROUTES. MORE PRESSINGLY, STILL, PARENTERAL IATROGENIC AΒ TRANSMISSIBILITY, LEADING TO CAA DEVELOPMENT, HAS ALSO BEEN DEMONSTRATED IN HUMANS, FOLLOWING NEUROSURGERY AND IN PATIENTS TREATED WITH CADAVERIC HUMAN GROWTH HORMONE. GIVEN THAT PRIONS ARE READILY TRANSFUSION TRANSMISSIBLE, THIS RAISES THE DISCONCERTING POSSIBILITY THAT CAA AND ITS RELATED AΒ-ASSOCIATED CONDITIONS, ARE POTENTIALLY TRANSFUSION TRANSMISSIBLE. A LARGE, BI-NATIONAL, RETROSPECTIVE COHORT STUDY USING THE SWEDISH-DANISH SCANDAT TRANSFUSION DATABASE FOUND THAT 0.1% OF TRANSFUSED PATIENTS IN SWEDEN AND DENMARK RECEIVED RED- CELL UNITS FROM BLOOD DONORS WHO LATER SUFFERED REPEATED, UNPROVOKED ICH, AND THAT THESE PATIENTS WERE AT A NEAR-3-FOLD INCREASED RISK OF THEMSELVES SUFFERING AN UNPROVOKED ICH, AS COMPARED TO PATIENTS WHO DID NOT RECEIVE RED-CELL UNITS FROM AFFECTED BLOOD DONORS. UNDER THE HYPOTHESIS THAT THE OBSERVED ASSOCIATION WAS DRIVEN BY AΒ TRANSMISSION AND CAA DEVELOPMENT IN AFFECTED DONORS AND RECIPIENTS, THE CLINICAL IMPLICATIONS COULD BE VERY LARGE, ESPECIALLY CONSIDERING THAT AΒ AGGREGATION IS ALSO STRONGLY ASSOCIATED WITH ALZHEIMER’S DISEASE. HOWEVER, EXTERNAL REPLICATION OF THESE PRELIMINARY FINDINGS, IDEALLY WITH BIOCHEMICAL AND MECHANISTIC CORROBORATION, IS URGENTLY NEEDED. WE THEREFORE PROPOSE TO UTILIZE THE EXISTING NHLBI REDS ALLOGENEIC DONOR AND RECIPIENT (RADAR) REPOSITORY, WITH SAMPLES FROM BOTH BLOOD DONORS AND PRE- AND POST- TRANSFUSION SAMPLES OF TRANSFUSED, TO (A) INVESTIGATE THE PREVALENCE OF AMYLOID Β PATHOLOGY THROUGH MEASURES OF THE ASSOCIATED PTAU217 PROTEIN AMONG BLOOD DONORS, AND (B) INVESTIGATE THE POSSIBILITY THAT AMYLOID Β IS TRANSFUSION- TRANSMISSIBLE FROM AFFECTED DONORS TO THEIR TRANSFUSED PATIENTS.

BIOMARKERS OF SPONTANEOUS ACUTE HEPATITIS C VIRUS RESOLUTION

SYSTEMIC EFFECTS OF BONE MARROW-DERIVED MSCS ON VASCULAR STABILITY

INNATE SENSING OF CELL-FREE DNA AND THE INTERFERON-MEDIATED CONTROL OF HIV IN VIVO - PROJECT SUMMARY WHILE THE ADVENT OF ANTIRETROVIRAL THERAPY (ART) HAS DRAMATICALLY REDUCED THE MORBIDITY AND MORTALITY ASSOCIATED WITH HIV INFECTION, A CURE IS NOT ACHIEVED DUE TO THE PERSISTENCE OF LATENTLY-INFECTED CELLS DURING TREATMENT. ACCUMULATING DATA SUGGEST THAT HIV-INFECTED INDIVIDUALS OFTEN EXPERIENCE PERSISTENT IMMUNE DYSREGULATION, CHRONIC INFLAMMATION, AND ACCELERATED AGING EVEN IN THE SETTING OF ART-MEDIATED VIRAL SUPPRESSION. THESE REALITIES HAVE CREATED A PRONOUNCED INTEREST IN DEVELOPING STRATEGIES TO CURE HIV INFECTION. IDENTIFYING THE HOST MOLECULAR DETERMINANTS OF HIV PERSISTENCE AND REBOUND KINETICS FOLLOWING CESSATION OF ANTIRETROVIRAL THERAPY (ART) WILL BE CRITICAL IN DEVELOPING EFFECTIVE STRATEGIES TO CURE HIV INFECTION. WE RECENTLY APPLIED A COMPREHENSIVE SYSTEMS PROFILING APPROACH TO PLASMA SAMPLES FROM HIV-INFECTED INDIVIDUALS WHO UNDERWENT ANALYTICAL TREATMENT INTERRUPTION (ATI), TO IDENTIFY CIRCULATING HOST FACTORS THAT ENABLE PREDICTION OF HIV REBOUND KINETICS FOLLOWING ART INTERRUPTION, AND SERVE AS POTENTIAL PHARMACOLOGICAL TARGETS TO PROMOTE DURABLE VIROLOGIC REMISSION IN THE ABSENCE OF ART. THE HOST FACTOR EXHIBITING THE STRONGEST STATISTICAL ASSOCIATION WITH HIV REBOUND TIMING FOLLOWING ART CESSATION WAS CIRCULATING CELL-FREE DNA (CFDNA) IN PLASMA. SPECIFICALLY, INCREASED CFDNA ABUNDANCE IN PLASMA WAS ASSOCIATED WITH DELAYED TIME-TO-REBOUND. CFDNA, RELEASED INTO CIRCULATION DURING PROGRAMMED CELL DEATH, HAS BEEN EXPLOITED EXTENSIVELY IN THE REALM OF CLINICAL ONCOLOGY (OFTEN TERMED “LIQUID BIOPSY”). HOWEVER, CFDNA REMAINS LARGELY UNEXPLORED IN THE SETTING OF HIV INFECTION. IN THIS R01 PROPOSAL, WE RIGOROUSLY EXPLORE CFDNA AS A BIOMARKER OF HIV DISEASE STATES, AND WE INVESTIGATE THE MOLECULAR AND IMMUNOLOGIC MECHANISMS LINKING CFDNA TO VIRAL EXPRESSION AND REBOUND. OUR CENTRAL HYPOTHESES ARE THAT 1) CFDNA REFLECTS THE DEATH RATE OF HIV-INFECTED CELLS IN VIVO; AND 2) CFDNA PROMOTES AN ANTIVIRAL STATE IN THE HOST (E.G. INDUCTION OF TYPE I INTERFERON RESPONSE) WHICH PREVENTS VIRAL REBOUND. OUR PROJECT FEATURES AN INVESTIGATIVE TEAM WITH BASIC, TRANSLATIONAL, AND CLINICAL EXPERTISE, AND LEVERAGES LARGE COLLECTIONS OF CLINICAL SAMPLES FROM WELL-CHARACTERIZED HIV-INFECTED INDIVIDUALS. IN AIM 1, WE WILL APPLY NEXT-GENERATION SEQUENCING (NGS) APPROACHES TO PLASMA SAMPLES FROM HIV-INFECTED INDIVIDUALS TO VALIDATE AND ENHANCE THE PROGNOSTIC SIGNIFICANCE OF CFDNA AS A BIOMARKER OF NATURAL HIV CONTROL IN VIVO. IN AIM 2, WE WILL EVALUATE HOW THE UPTAKE AND SENSING OF CFDNA BY HIV TARGET CELLS IMPACTS CELL FATE AND THE REACTIVATION AND REPLICATION OF HIV. IN AIM 3, WE WILL DETERMINE HOW HIV-ASSOCIATED MITOCHONDRIAL DYSFUNCTION LEADS TO CFDNA EXTRUSION AND ULTIMATELY INDUCES TYPE I INTERFERON RESPONSES THAT PREVENT HIV REBOUND. OUR PROJECT WILL ADVANCE THE DEVELOPMENT AND EVALUATION OF HIV CURE STRATEGIES.

CLINICAL OUTCOMES OF ZIKA VIRUS IN SICKLE CELL DISEASE