GENES INVOLVED IN AXONAL REGENERATION IN OLD POST-STROKE RATS Authors: Z ă v ă leanu Alexandra Daniela Greşiţ ă Andrei Scientific Coordinator: Bug ă Ana Maria

INTRODUCTION Brain injuries and their complications are definitely more common among old people. Our study wants to identify all the cellular and molecular mechanisms that fail in regenerating injured axons and neurons after a brain lesion, but also those genes which have a huge importance when it comes to these processes.

MATERIAL AND METHODS There were used thirty young male Sprague-Dawley rats that were divided into 3-day and 14-day post stroke survival groups and nineteen postnatal Sprague-Dawley rats. In addition, nineteen young male Sprague-Dawley rats were used like control group (naive). Cerebral infarction was induced by transcranial interruption of blood flow by transiently lifting the middle cerebral artery with a tungsten hook as previously described. (1) Subsequent to survival times of 3 or 14 days, rats were deeply anesthetized and the blood removed by perfusion with neutral buffered saline. Brains were cut into 2mm slices and the periinfarcted area was microdissected under a microscope and stored at - 70°C until use.

THE GROUPS OF RATS Stroke group (30) Young (naive) rats (19) Juvenile rats (19) 3-days post stroke 14 days post stroke

The middle cerebral artery in rats The infarct area

IMMUNOHISTOCHEMISTRY The kinesin superfamiliy (KIF) genes encode a class of microtubule-based motor proteins that are specialized in intracellular transport of membranous organelles. During cellular division, KIF4 is essential for the organization of central spindles. 5-Bromo-2′-deoxyuridine (5-BrdU) is a thymidine analogue which is incorporated into DNA. 5-BrdU is routinely and extensively used to measure DNA synthesis and to label dividing cells. Consequently 5-BrdU is used to study cell signaling and other processes that induce cell proliferation. The antibody reacts specifically with GFAP in immunoblotting assays and labels astrocytes, Bergmann glia cells and chondrocytes of elastic cartilage in immunohistochemical staining. The antibody reacts with glial specific antigen in frozen or alcohol-fixed tissue sections.

RESULTS A relative high number of genes that are expressed in developmental period was up-regulated after stroke in adult brain. We founded Kif4 gene expression highly up-regulated after lesion in adult brain.

HEAT MAP THAT SHOWS THE NUMBER OF GENES EXPRESSED AFTER THE STROKE AND POSTNATAL

KIF 4 GENE EXPRESSION

Axons and neurons that can be normally observed in young rats brain after exposing to mouse anti-KIF 4. Astrocytes visible near a stroke area after exposing to mouse anti-KIF 4. New cells and blood vessels near the stroke area after exposing to mouse anti-KIF 4

Kif4 and BRDU expressed in the rat’s infarct core (14 days post stroke) Kif4 and GFAP expressed in the rat’s infarct core (14 days post stroke)

CONCLUSION We have found developmental Kif 4 gene an important player in recovery process after stroke in adult brain after stroke. This Kif4 transcription factor can be an important therapeutically target to increase the axonal regeneration and to improve the recovery after stroke in aged people.