This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Magdalena Kucia, PI Stem cells provide a novel potential source to replace dead neurons and supporting tissue in a brain damaged by chronic ischemia as seen for example in sleep apnea (SA). Some investigations of animals and short-term human bone marrow (BM) transplants have demonstrated that bone marrow cells provide a source of neurons and can repair brain damage (e.g., during stroke). The mechanisms of this functional improvement are currently the focus of intense research, creating a need for new study methodologies to assess the effectiveness of such strategies. Elucidation of stem cell-related mechanisms of regeneration is crucial to developing effective stem cell-based therapies that could extend the lifespan of patients with diseases that are treatable by tissue regeneration. One such disorder is chronic brain damage due to hypoxia resulting from sleep apnea. Based on preliminary data, a novel hypothesis is presented that the pool of CXCR4+ epiblast-derived VSELs is deposited in the BM during early development. These cells subsequently serve as a reserve mobile pool of stem cells that may be mobilized into peripheral blood and play an important role in brain regeneration - where they are chemoattracted by an SDF-1 gradient. Furthermore, it is hypothesized that an age-related decrease in the marrow pool of these circulating VSELs may contribute to aging of the central nervous system (CNS), resulting in less effective repair. To investigate these issues, four specific aims were proposed.
Specific Aim 1. Neural differentiation of bone marrow-derived VSELs. We have presented evidence that BM contains a population of VSELs and that these epiblast-derived cells are deposited there early in development during rapid body growth/expansion. We will first optimize their isolation from bone marrow and neural differentiation (ability to grow neurospheres). Next we will evaluate the age-related presence of VSELs in bone marrow tissue. Once it is determined whether VSELs circulate under normal steady-state conditions in the peripheral blood (PB) at very low but detectable levels, we will study their mobilization in a murine model of sleep apnea.
Specific Aim 2. Optimize mobilization of VSELs into peripheral blood. Since mobilized peripheral blood (mPB) may be a source of VSELs for potential neural regeneration, we will optimize their mobilization into PB. We will test the effect of various mobilizing agents involving selected growth factors (G-CSF, Flt3-ligand, VEGF, HGF) and small-molecule inhibitors (CXCR4-antagonist T140, C3aR antagonist) on the efficacy of their mobilization. We also will investigate mobilization of these cells in response to hypoxia damage and the role of the SDF-1?CXCR4 axis in this process. It is hypothesized that CXCR4+ VSELs are mobilized and subsequently chemo-attracted into a damaged brain in an SDF-1-dependent manner.
Specific Aim 3. Develop an approach to expand VSELs. Since the number of VSELs that can be isolated from the BM and mPB of older individuals is relatively low, an efficient ex vivo expansion system may be needed to obtain a sufficient number of these cells for neural regeneration. It also is possible that ex vivo culture-derived VSELs will better engraft and regenerate brain cells. We will employ selected strategies to expand these cells ex vivo involving cocktails of selected growth factors, BM stroma support, and our new strategy based on the expansion of stem cells in the presence of membrane-derived microvesicles isolated from embryonic stem cells (ESMV). We observed that purified VSEL cells are able to form spheres in co-cultures with C2C12 myoblastic cell line feeder layers that resemble embryoid bodies. Cells from these spheres may again (up to 5-7 passages) grow new secondary spheres, or if plated into cultures promoting tissue differentiation, expand into cells from all three germ-cell layers. We will employ this system to expand neural cells from VSEL-derived spheres.
Specific Aim 4. Determine the efficacy of VSELs in brain regeneration in vivo in a murine model of sleep apnea (SA). The contribution of VSELs to functional regeneration of damaged tissues will be tested in an in vivo mouse model of SA. We will compare the regeneration potential of syngeneic VSELs isolated from enhanced green immunofluorescence protein (EGFP+) transgenic mice to rescue brain cells damaged by hypoxia and the role of the SDF-1?CXCR4 axis in this process. We will employ freshly isolated VSELs from BM or mPB as well as VSEL expanded in ex vivo cultures. As a readout of brain regeneration, we will use selected behavioral tests.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Exploratory Grants (P20)
Project #
5P20RR018733-09
Application #
8360670
Study Section
National Center for Research Resources Initial Review Group (RIRG)
Project Start
2011-07-01
Project End
2012-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
9
Fiscal Year
2011
Total Cost
$239,359
Indirect Cost
Name
University of Louisville
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
057588857
City
Louisville
State
KY
Country
United States
Zip Code
40292
Garbett, Nichola C; Brock, Guy N; Chaires, Jonathan B et al. (2017) Characterization and classification of lupus patients based on plasma thermograms. PLoS One 12:e0186398
Kendrick, Sarah K; Zheng, Qi; Garbett, Nichola C et al. (2017) Application and interpretation of functional data analysis techniques to differential scanning calorimetry data from lupus patients. PLoS One 12:e0186232
Garbett, Nichola C; Brock, Guy N (2016) Differential scanning calorimetry as a complementary diagnostic tool for the evaluation of biological samples. Biochim Biophys Acta 1860:981-989
Zhao, Guoping; Neely, Aaron M; Schwarzer, Christian et al. (2016) N-(3-oxo-acyl) homoserine lactone inhibits tumor growth independent of Bcl-2 proteins. Oncotarget 7:5924-42
Donninger, Howard; Schmidt, M Lee; Mezzanotte, Jessica et al. (2016) Ras signaling through RASSF proteins. Semin Cell Dev Biol 58:86-95
Lanceta, Lilibeth; Mattingly, Jacob M; Li, Chi et al. (2015) How Heme Oxygenase-1 Prevents Heme-Induced Cell Death. PLoS One 10:e0134144
Schwarzer, Christian; Fu, Zhu; Morita, Takeshi et al. (2015) Paraoxonase 2 serves a proapopotic function in mouse and human cells in response to the Pseudomonas aeruginosa quorum-sensing molecule N-(3-Oxododecanoyl)-homoserine lactone. J Biol Chem 290:7247-58
Donninger, Howard; Calvisi, Diego F; Barnoud, Thibaut et al. (2015) NORE1A is a Ras senescence effector that controls the apoptotic/senescent balance of p53 via HIPK2. J Cell Biol 208:777-89
Garbett, Nichola C; Mekmaysy, Chongkham S; DeLeeuw, Lynn et al. (2015) Clinical application of plasma thermograms. Utility, practical approaches and considerations. Methods 76:41-50
Zhao, Guoping; Lu, Huayi; Li, Chi (2015) Proapoptotic activities of protein disulfide isomerase (PDI) and PDIA3 protein, a role of the Bcl-2 protein Bak. J Biol Chem 290:8949-63

Showing the most recent 10 out of 146 publications