Methods and compositions for treatment of traumatic brain injury and for modulation of migration of neurogenic cells
A brain-injured, traumatic technique used in nervous system cells, biochemical devices and methods, drug combinations, etc.
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Embodiment 1
[0086] Embodiment 1: Preparation of MSCs and SB623 cells
[0087] Bone marrow aspirate from an adult donor was obtained from Lonza Walkersville, Inc. (Walkersville, MD) and plated on a medium supplemented with 10% fetal bovine serum (Hyclone, Logan, UT), 2 mM L-glutamine (Invitrogen, Carlsbad, UT). CA) and penicillin / streptomycin (Invitrogen) in α-MEM (Mediatech, Herndon, VA). Incubate the cells at 37 °C and 5% CO 2 The cells were cultured under the same conditions for 3 days to obtain a monolayer of adherent cells, and after removal of non-adherent cells, the culture was continued for 2 weeks under the same conditions. During this time, cells were passaged twice by using 0.25% trypsin / EDTA. A portion of cells from the second passage was frozen as MSCs.
[0088] The remaining cells from the second passage were plated and, by using Fugene6 (Roche Diagnostics, Indianapolis, IN), enriched with a plasmid (pCMV -hNICD1-SV40-Neo R ) for transfection. This plasmid also contains...
Embodiment 2
[0090] Example 2: Induction of TBI in a rat model
[0091] A total of 40 animals were identified at baseline (before TBI surgery) as exhibiting normal behavior (50-60% biased swing activity on EBST; 60 second dwell time on Rotorod; and mean Bederson score of at most 0-0.5 ) animals underwent TBI surgery as described below.
[0092] All surgical procedures were performed under sterile conditions. Adult male Sprague-Dawley rats were anesthetized with 1.5% isoflurane and examined for pain reflex. Under deep anesthesia, animals underwent a moderate TBI model as follows. Each animal was placed in a stereotaxic frame where anesthesia was maintained with 1-2% isoflurane administered via a gas mask. After exposure of the skull, a 4-mm craniotomy was performed on the left frontoparietal cortex centered at -2.0 mm AP and +2.0 mm ML relative to bregma. Use a pneumatic metal impactor with a diameter of 3mm to impact the brain at a speed of 6.0m / s, reach a depth of 1.0mm under the dura...
Embodiment 3
[0094] Example 3: Transplantation of SB623 cells
[0095] Among animals that experienced a TBI, only those with the following degrees of behavioral impairment at day 7 post-TBI were selected for transplantation studies: at least 75% biased swing activity on the EBST; 30 seconds or more on the Rotorod A short dwell time; and an average Bederson score of at least 2.5. Those selected animals were randomly assigned to groups receiving SB623 grafts (n=20) or vehicle infusions (n=20). The target area for grafting was the medial cortex, which corresponds to the cortical area surrounding the lesion, based on previously established target sites for similar stereotaxic implants.
[0096] All surgical procedures were performed under sterile conditions. Animals were anesthetized with 1.5% isoflurane and examined for pain reflexes. Once deep anesthesia is achieved (determined by loss of pain reflex), hair is shaved around the surgical incision area (skull area) with sufficient margins t...
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