NNS 2015 Program & Abstract Book

tablished before stereotactic injection of control or green fluorescent

protein (GFP) expressing hNSCs into the PBBI penumbra one week later.

Animals were sacrificed at defined time points post-transplantation.

Brains were sectioned and assessed for GFP fluorescence.

Results:

The cells at week 1 post transplantation were undiffer-

entiated and morphologically indistinguishable from undifferentiated

cells

in vitro

. By week 5, a robust transplant could be seen sending out

processes as far as 2–3 millimeters. By week 8 the number of pro-

cesses was greater and they could be observed in the hind brain

(

12 mm from transplant site). The GFP processes emanating from

the transplant appear to follow intact white matter tracts in PBBI

brain. The processes were found to cross internal capsule in ipsilateral

hemisphere and traverse corpus callosum into contralateral hemi-

sphere.

Conclusion:

PBBI is conducive to human fetal neural stem cells

engraftment and survival. The transplanted cells appear to be capable

of sending processes over long distances.

Support:

This work was supported by State of Florida funding.

Keywords: Penetrating traumatic brain injury, Neural stem cell

transplants, FDA, Neurogenesis, Light sheet microscopy

D3-02

TRANSPLANTATION OF HUMAN INDUCIBLE PLUR-

IPOTENT STEM CELL-DERIVED NEURAL STEM CELLS

PROMOTES LOCOMOTOR RECOVERY AFTER SCI

Qi Lin Cao

, Yiyan Zheng, QIn Wang, shenglan li, ying liu, dong kim,

Qi LIn Cao

UT Medical School at Houston, Department of Neurosurgery, Hous-

ton, USA

Transplantation of hiPSC-derived neural stem cells (NSCs) could be

one of the most promised novel reparative strategies to promote

functional recovery after spinal cord injury (SCI). One of the major

challenges to fully realized the full therapeutic potential of hiPSC is

to purify the desired NSCs from differentiating hiPSCs in vitro be-

fore transplantation. In this study, we use the neural stem cell spe-

cific hiPSC reporter line and fluorescence-activated cell sorting

(FACS) to purify NSCs and then test its therapeutic potential for

SCI. The nestin-EGFP reporter hiPSCs, in which EGFP cassette has

been inserted to the nestin locus of hiPSC via homologous recom-

bination, are induced for neural differentiation and GFP expressing

NSCs are purified by FACS. The purified cells express NSC but not

iPSC markers. Importantly, the purified NSCs continue to proliferate

for a long time in vitro and differentiate into neurons, astrocytes and

oligodendrocytes under respective differentiation conditions. NOD-

SCID mice receive moderate contusion at T9 and then grafts of

hiPSC-derived NSCs, human fibroblasts or control medium at 8 days

after SCI. Robust survival of grafted NSCs is observed in all animals

receiving grafts at 2 months after transplantation. Some grafted

NSCs differentiate into NeuN

mature neurons and more into dou-

blecortin

immature neurons. Astrocyte or oligodendrcoyte differ-

entiation is also observed in grafted NSCs. The volumes of spared

white and gray are significantly increased in animals received NSC

graft. Furthermore, recovery of hindlimb locomotor function is

significantly enhanced in animals receiving grafted of hiPSC-derived

NSCs. No teratoma formation is observed in any animals receiving

hiPSC-derived NSCs. Our results show that the multipotential NSCs

can be purified from NSC specific reporter hiPSC line by FACS.

Importantly, purified NSCs can survive and differentiate into both

neurons and glias and promote functional recovery after transplan-

tation following traumatic SCI. These results suggest that hiPSC-

derived NSCs have great therapeutic potential for SCI and other

neurological diseases.

Keywords: spinal cord injury, neural stem cell, hiPSC

D4 Poster Session VII - Group D: Transplantation

D4-01

INTRACEREBROVENTRICULAR TRANSPLANTATION OF

ADULT NEURAL STEM CELLS (NSCS) AFTER TBI: PROOF-

OF-CONCEPT FOR ACTIVATION OF HOST NSCS

Regina Armstrong

, Genevieve Sullivan

USUHS, Anatomy, Physiology & Genetics, Bethesda, USA

Transplantation of neural stem cells (NSCs) may promote brain repair

by replacing lost cells and by interacting with the host tissue to

modulate the immune response and stimulate endogenous regenera-

tive capacity. Determining an effective NSC delivery route in the CNS

is particularly challenging for TBI that involves diffuse rather than

focal lesions. In clinical management of TBI, ventriculostomy is often

performed in patients with acute TBI after failure to control intra-

cranial pressure by other means. Ventriculostomy may provide a route

of access to the lateral ventricle for therapeutic delivery of stem cells.

The current studies provide a proof-of-concept test of in-

tracerebroventricular (ICV) delivery of adult NSCs into the lateral

ventricle in an impact-acceleration TBI model with traumatic axonal

injury in the corpus callosum. We also test the effect of NSC trans-

plantation in stimulating a regenerative response in endogenous NSCs

in the host subventricular zone (SVZ) based on activation of Sonic

hedgehog (Shh) signaling. Shh maintains the NSC niche in adult CNS.

Gli1 transcription indicates active Shh signaling. TBI from impact to

the skull at bregma was produced in adult

Gli1CreERT2

mice crossed

RosaTdTomato

reporter mice. NSCs were isolated from adult

UBI-

GFP

mice, which ubiquitously express green fluorescent protein

(GFP), and microinjected unilaterally into the lateral ventricle of the

Gli1CreERT2;RosaTdTomato

mice 2 weeks after the TBI or sham

procedure.

Gli1CreERT2;RosaTdTomato

host mice were then ad-

ministered tamoxifen to label endogenous cells responding to Shh

signaling after ICV transplantation. Mice were sacrificed for tissue

analysis at 4 weeks post-TBI. Transplanted GFP-NSCs survived,

maintained an immature phenotype, and were localized along the

ependymal lining of the ventricle and adhering to the choroid plexus.

Endogenous NSCs in the adjacent SVZ were fate-labeled with

TdTomato, indicating active Shh signaling, but were not changed

significantly due to the injury or ICV transplantation of GFP-NSCs.

An inflammatory response to transplanted GFP-NSCs was not ob-

served in sham or TBI mice. Funded by the DoD in the Center for

Neuroscience and Regenerative Medicine (CNRM).

Keywords: Transplantation, neural stem cell, Sonic hedgehog

D4-02

DIFFERENTIATION OF FDA-APPROVED HUMAN NEURAL

STEM CELLS WITH FUNCTIONAL IMPROVEMENT AFTER

A PENETRATING TBI

Aminul Ahmed

, Shyam Gajavelli

, Markus Spurlock

, Lai Yee

Leung

, Deborah Shear

, Frank Tortella

, Ross Bullock

University of Miami, Miami Project to Cure Paralysis, Miami,

United States

A-106