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Schwann cells (SCs) and olfactory ensheathing glia (OEG) have shown promise for spinal cord injury repair. We sought their in vivo identification following transplantation into the contused adult rat spinal cord at 1 week post-injury by: (i) DNA in situ hybridization (ISH) with a Y-chromosome specific probe to identify male transplants in female rats and (ii) lentiviral vector-mediated expression of EGFP. Survival, migration, and axon-glia association were quantified from 3 days to 9 weeks post-transplantation. At 3 weeks after transplantation into the lesion, a 60-90% loss of grafted cells was observed. OEG-only grafts survived very poorly within the lesion (

Transgenic mice carrying the bacterial lacZ reporter gene under the control of the regulatory elements of the human myoD gene have been produced. The developmental expression of the myoD reporter transgene in somites, limb buds, visceral arches, and cephalocervical regions was studied in transgenic embryos by β-gal staining. In somites, the spatiotemporal pattern of transgene expression was different from other muscle-specific regulatory and structural genes and revealed that myoD-expressing cells arise in distinct patterns in somites that are dependent on position along the anterior-posterior (AP) body axis (occipital and cervical vs thoracic and more posterior myotomes). Transgene expression did not follow a strict anterior to posterior sequence of activation and therefore was not strictly correlated with somite developmental age. Moreover, the pattern of transgene expression along the dorsal-ventral myotomal axis was dependent on somite position along the anterior-posterior axis. While myoD expression is first detected after the myotome is well-formed, transgene expression in the dorsal and ventral medial lips of the dermatome suggests a function for myoD in the expansion of the myotome. Whole-mount in situ hybridization confirmed that these unique patterns of transgene expression in somites, as well as expression in limb buds, visceral arches, and other myogenic centers, are concordant with the distribution of endogenous myoD transcripts. These results shed new light on the developmental differences between myotomes at different positions along the AP and DV axis and demonstrate a unique axial pattern of somitic myoD expression, suggesting a specific role of myoD in myotome lineage determination and differentiation.

We produced transgenic mice carrying the native sheep beta-lactoglobulin (BLG) or fusion genes composed of the BLG promoter and human serum albumin (HSA) minigenes. BLG was expressed exclusively in the mammary glands of the virgin and lactating transgenic mice evaluated. In contrast, transgenic females carrying the BLG/HSA fusion constructs also expressed the HSA RNA ectopically in skeletal muscle, kidney, brain, spleen, salivary gland and skin. Ectopic expression of HSA RNA was detected only in strains that express the transgene in the mammary gland. There was no obvious correlation between the level of the HSA RNA expressed in the mammary gland and that found ectopically. In three transgenic strains analysed, the expression of HSA RNA in kidney and skeletal muscle increased during pregnancy and lactation, whereas in the brain HSA expression decreased during lactation in one of the strains. HSA protein was synthesized in skeletal muscle and skin of strain #23 and its level was higher in lactating mice compared with virgin mice. Expression of HSA was also analysed in males and was found to be more stringently controlled than in females of the same strains. In situ hybridization analyses localized the expressed transgene in the skin, kidney, brain and salivary glands of various transgenic strains. Distinct strain-specific and cell-type specific HSA expression patterns were observed in the skin. This is in contrast to the exclusive expression of the HSA transgene in epithelial cells surrounding the alveoli of the mammary gland. Taken together, these results suggest that the absence of sufficient mammary-specific regulatory elements in the BLG promoter sequences and/or the juxtaposition of the BLG promoter with the HSA coding sequences leads to novel tissue- and cell-specific expression in ectopic tissues of transgenic mice.

Here we propose the use of adeno-associated virus (AAV) vectors as a non-invasive vehicle for the nervous system to deliver genes to spinal motoneurons, based on their retrograde transport from muscle. Long-term protein expression in lower cervical motoneurons was achieved after injections of AAV into the triceps, independently of serotypes 1, 2, or 5. Muscle injections of AAV5-neurotrophin 3 (NT3) resulted in a significant increase in the levels of NT3 in the cervical enlargement, compared to those obtained after injections of AAV5-GFP. Following a dorsal lesion at C4/C5, animals injected with AAV5-NT3 made fewer errors (footslips) in the horizontal ladder test compared to those injected with AAV5-GFP. In parallel, the number and length of corticospinal tract (CST) fibers circumventing the injury site were significantly increased in rats injected with AAV5-NT3. Compared to controls, we observed less astrogliosis and less CST axonal retraction and/or enhanced sprouting in animals injected with AAV5-NT3. In sum, we demonstrate here that the delivery of nt3 via retrograde transport of AAV from triceps to cervical motoneurons leads to reduced functional loss and anatomical reorganization of the CST following injury, without introducing additional injury to the spinal cord.

Due to an ever-growing population of individuals with chronic spinal cord injury, there is a need for experimental models to translate efficacious regenerative and reparative acute therapies to chronic injury application. The present study assessed the ability of fluid grafts of either Schwann cells (SCs) or olfactory ensheathing glia (OEG) to facilitate the growth of supraspinal and afferent axons and promote restitution of hind limb function after transplantation into a 2-month-old, moderate, thoracic (T8) contusion in the rat. The use of cultured glial cells, transduced with lentiviral vectors encoding enhanced green fluorescent protein (EGFP), permitted long-term tracking of the cells following spinal cord transplantation to examine their survival, migration, and axonal association. At 3 months following grafting of 2 million SCs or OEG in 6 μl of DMEM/F12 medium into the injury site, stereological quantification of the three-dimensional reconstructed spinal cords revealed that an average of 17.1 ± 6.8% of the SCs and 2.3 ± 1.4% of the OEG survived from the number transplanted. In the OEG grafted spinal cord, a limited number of glia were unable to prevent central cavitation and were found in patches around the cavity rim. The transplanted SCs, however, formed a substantive graft within the injury site capable of supporting the ingrowth of numerous, densely packed neurofilament-positive axons. The SC grafts were able to support growth of both ascending calcitonin gene-related peptide (CGRP)-positive and supraspinal serotonergic axons and, although no biotinylated dextran amine (BDA)-traced corticospinal axons were present within the center of the grafts, the SC transplants significantly increased corticospinal axon numbers immediately rostral to the injury–graft site compared with injury-only controls. Moreover, SC grafted animals demonstrated modest, though significant, improvements in open field locomotion and exhibited less foot position errors (base of support and foot rotation). Whereas these results demonstrate that SC grafts survive, support axon growth, and can improve functional outcome after chronic contusive spinal cord injury, further development of OEG grafting procedures in this model and putative combination strategies with SC grafts need to be further explored to produce substantial improvements in axon growth and function.

Limited information on the production and secretion of the individual milk proteins at the cellular level in the mammary gland is available. It is generally assumed that all active epithelial cells within the lactating mammary gland of mice share a comparable ability to produce and secrete all the components of milk. This is based on ultrastructural studies and observations on cell membrane turnover related to fat globule membrane loss and vesicular protein exocytosis.

MDP is the minimum structural unit responsible for the immunoadjuvancy of bacterial cell wall peptidoglycans. Many properties of MDP manifest potential therapeutic efficacy for SCI. These include stimulation of phagocytosis of immunocompetent cells, thereby facilitating myelin removal with minimal beneficial proinflammatory cytokine alterations,reduced glutamate release,increased intra- and extra-cellular Mg2+, thereby blocking NMDA receptors, andsuppressor cytokine expression, such as IL-10. The current experiment delivered MDP by injection, 6 hours post T8 moderate contusive spinal cord injury in adult Fischer rats. At 2 wk after injury, histological examination revealed reductions in lesion volume associated with deposits of chondroitin sulfate proteoglycan and more myelinated axons. The activated (ED1+) macrophage/microglia count was higher in the more limited injury site of MDP animals. Concurrently, most were Mac1+, indicating active phagocytosis. Furthermore, MDP treated animals displayed fewer ED1+ cells rostral and caudal to the lesion compared with controls. Behaviorally, MDP animals displayed frequent co-ordination with weight-supported steps (BBB, 13.7 ± 1.6) compared with the occasional stepping in controls (BBB, 10 ± 0.6; n=6, p Supported by CRF, Hollfelder Foundation, and Miami Project