Project Grant R01AA031486

MOLECULAR TARGETING OF THE CEREBROVASCULATURE DURING PRENATAL ALCOHOL EXPOSURE - PROJECT SUMMARY FETAL ALCOHOL SPECTRUM DISORDERS (FASD) CONSTITUTE AN ARRAY OF NEUROCOGNITIVE AND BEHAVIORAL DEFICITS EXHIBITED BY INDIVIDUALS WHO EXPERIENCE SOME DISABILITY AS A RESULT OF PRENATAL ALCOHOL EXPOSURE (PAE). IN THE UNITED STATES, THE INCIDENCE OF FASD IS ESTIMATED TO BE 20-50/1,000 CHILDREN. WITH NO CURATIVE TREATMENTS AVAILABLE, IT IS OF PARAMOUNT IMPORTANCE TO CONSIDER FRESH PERSPECTIVES AND IMPROVE OUR UNDERSTANDING OF FACTORS DRIVING FASD-ASSOCIATED DEFICITS. MUCH RESEARCH HAS FOCUSED ON NEURONAL-SPECIFIC DEFECTS IN THE CENTRAL NERVOUS SYSTEM WITH LESS ATTENTION GIVEN TO THE VASCULAR COMPARTMENT AND IN PARTICULAR TO ENDOTHELIAL CELL-SPECIFIC DEFECTS MEDIATED BY PAE. CONSIDERING THE INTIMATE RELATIONSHIP BETWEEN THE DEVELOPING NEURONAL AND VASCULAR NETWORKS DURING EMBRYOGENESIS, IT IS LIKELY THAT ABNORMAL VASCULAR DEVELOPMENT CONTRIBUTES TO PAE-ASSOCIATED BRAIN PATHOLOGY. USING AN ESTABLISHED MOUSE MODEL OF MODERATE PAE, WE FOUND THAT MICRORNA (MIR)-150-5P WAS SIGNIFICANTLY INCREASED IN THE PAE EMBRYONIC CORTICES AND IN BRAIN MICROVASCULAR ENDOTHELIAL CELLS (BMVECS) ISOLATED FROM THOSE CORTICES COMPARED TO CONTROLS. MIR-150-5P IS A NEGATIVE REGULATOR OF ANGIOGENESIS AND THE BLOOD BRAIN BARRIER (BBB), AND ITS TARGETS INCLUDE ENDOTHELIAL-SPECIFIC TRANSCRIPTION FACTORS AND MATRIX METALLOPROTEINASES AS WELL AS ADHERENS AND TIGHT JUNCTION PROTEINS, WHICH WERE SIGNIFICANTLY REDUCED IN THE SAME TISSUES. ADDITIONALLY, WE FOUND THAT THE VASCULATURE OF PAE EMBRYONIC CORTICES DISPLAYED SIGNIFICANT ALTERATIONS INCLUDING REDUCTIONS IN VESSEL AREA AND VOLUME, THE MEAN DIAMETER OF BLOOD VESSELS, AND BBB INTEGRITY. INHIBITION OF MIR-150-5P OR OVEREXPRESSION OF ITS TARGETS RESTORED ANGIOGENESIS AND DECREASED PERMEABILITY OF BMVECS EXPOSED TO ALCOHOL. OUR LONG-TERM GOALS ARE TO DISCOVER NOVEL TARGETS AND DEVELOP STRATEGIES THAT MAY BE USEFUL FOR REVERSING OR ALLEVIATING THE ADVERSE CONSEQUENCES OF PAE. THE OVERALL OBJECTIVE OF THIS PROPOSAL IS TO IDENTIFY MOLECULAR MECHANISMS AND REGULATORS OF MIR-150-5P THAT LEAD TO ENDOTHELIAL CELL AND VASCULAR DYSFUNCTION, WHICH COULD BE HARNESSED AS POTENTIAL THERAPEUTIC TARGETS FOR IMPROVING PAE-MEDIATED EFFECTS. OUR SPECIFIC AIMS ARE TO FIRST INVESTIGATE THE MECHANISMS BY WHICH MIR-150-5P LEVELS ARE ENHANCED IN BMVECS, EXAMINING ITS TRANSCRIPTIONAL AND POST-TRANSCRIPTIONAL REGULATION. WE WILL THEN EVALUATE THE EFFECT OF MIR-150-5P ON THE MOLECULAR AND CELLULAR PROPERTIES OF BMVECS AND THE NEUROVASCULAR UNIT (NVU) FOLLOWING PAE, PRESERVING PHYSIOLOGICAL CONTEXT AND SPATIAL INFORMATION. FINALLY, WE WILL EXAMINE WHETHER THE MIR-150-5P-MEDIATED DEVIATIONS IN VASCULAR MORPHOLOGY AND FUNCTION PERSIST INTO POSTNATAL LIFE FOLLOWING PAE, USING TISSUE CLEARING COMBINED WITH FLUORESCENCE MICROSCOPY OF THE CORTICAL VASCULATURE AS WELL AS FUNCTIONAL STUDIES OF EX VIVO CEREBRAL VESSELS. THIS WORK WILL UNCOVER NOVEL MOLECULAR MECHANISMS THAT CONTRIBUTE TO VASCULAR DEFECTS IN PAE AND WILL PROVIDE INSIGHT INTO RNA-BASED STRATEGIES THAT COULD BE DEVELOPED FOR IMPROVING PAE-MEDIATED EFFECTS.