In vitro induction and in vivo expression of bcl-2 in the hNT neurons.
Bcl-2 encodes membrane-associated proteins that suppress programmed cell death in cells of various origins. Compelling evidence suggests that bcl-2 is also involved in neuronal differentiation and axonal regeneration. The human Neuro-Teratocarcinoma (hNT) neurons constitute a terminally differentiated human neuronal cell line that is derived from the Ntera-2/clone D1 (NT2) precursors upon retinoic acid (RA) treatment. After transplantation into the central nervous system (CNS), the hNT neurons survive, engraft, maintain their neuronal identity, and extend long neurite outgrowth. We were particularly interested in the intracellular determinants that confer these post-transplant characteristics to the hNT neurons. Thus, we asked whether the hNT neurons express bcl-2 after transplantation into the rat striatum and if RA induction of the neuronal lineage is mediated by bcl-2. The grafted hNT neurons were first identified using three different antibodies that recognize human-specific epitopes, anti-hMit, anti-hNuc, and NuMA. After a 1-month post-transplant survival time, NuMA immunostaining revealed that 12% of the hNT neurons survived the transplantation. These neurons extended long neuritic processes within the striatum, as demonstrated using the human-specific antibody against the midsize neurofilament subunit HO14. Importantly, we found that 85% of the implanted hNT neurons expressed bcl-2 and that the in vitro induction of the neuronal lineage from the NT2 precursors with RA resulted in an upregulation of bcl-2 expression. Together, these data suggest that the differentiation of the hNT neurons to a neuronal lineage could be mediated at least partially by bcl-2.
Daadi
M M
MM
Center for Aging and Brain Repair, University of South Florida College of Medicine, Tampa, Florida, USA. mdaadi@laytonbio.com
Saporta
S
S
Willing
A E
AE
Zigova
T
T
McGrogan
M P
MP
Sanberg
P R
PR
D000818:Animals; D034961:Antigens, Nuclear; D016380:Brain Tissue Transplantation; D018797:Cell Cycle Proteins; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D019070:Cell Lineage / Q000187:drug effects / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology; D051246:Glucose Transport Proteins, Facilitative; D006085:Graft Survival / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D017072:Neostriatum / Q000254:growth & development / Q000378:metabolism / Q000601:surgery; D016501:Neurites / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D016900:Neurofilament Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000378:metabolism / Q000637:transplantation*; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D033581:Stem Cell Transplantation; D013234:Stem Cells / Q000187:drug effects / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism / Q000637:transplantation; D015854:Up-Regulation / Q000187:drug effects / Q000502:physiology
D034961:Antigens, Nuclear; D018797:Cell Cycle Proteins; D051246:Glucose Transport Proteins, Facilitative; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins; C050045:NUMA1 protein, human; D016900:Neurofilament Proteins; D034521:Nuclear Matrix-Associated Proteins; D009687:Nuclear Proteins; C489081:Numa1 protein, rat; D019253:Proto-Oncogene Proteins c-bcl-2; C437315:SLC2A13 protein, human; C486673:Slc2a13 protein, rat; C061195:neurofilament protein M; D014212:Tretinoin