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Neurologic and Histologic Tests Used to Measure Neuroprotective Effectiveness of Virus-Derived Immune-Modulating Proteins

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Abstract

Severe inflammatory disease initiated by neurotrauma and stroke is of primary concern in these intractable pathologies as noted in recent studies and understanding of the pathogenesis of spinal cord injury (SCI) in the rat model. Successful anti-inflammatory treatments should result in neuroprotection and limit the loss of neurological function to injury caused by the initial damage. Continuous subdural infusion offers direct access to the cavity of injury (COI) that forms after balloon crush SCI deep in the spinal cord. Some anti-inflammatory compounds are not likely capable of crossing the blood-spinal cord barrier. Subdural infusion of myxoma virus-derived Serp-1, an anti-thrombotic/anti-thrombolytic, and also of M-T7, a chemokine inhibitor, improved the locomotor scores and pain sensation scores as well as reduced the numbers of macrophages in the COI by 50 and 80%, respectively, while intraperitoneal infusion of either protein had little effect. Injection of a chitosan hydrogel loaded with Serp-1 into the dorsal spinal column crush also resulted in improved neurological deficits and in reduction of the size of the crush lesion 4 weeks after injury. While neurological scores in a simplified hind-end (HE) locomotor test together with a toe-pinch withdrawal test demonstrated improvement in all balloon crush injury and dorsal spinal crush injury rats, a severe inflammation is induced by the injury indicating additional damage to the spinal cord. Thus neurological function testing can be contradictory, rather than corresponding, to the pathogenesis of SCI. The count of macrophages in the COI offers a precise, reliable method of measuring the effectiveness of a neuroprotective treatment of SCI in preclinical studies.

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... In the experiments using the subdural infusion of dexamethasone, M-T7 and Serp-1, the Basso Beattie Bresnahan (BBB) locomotor test used to evaluate SCI rats [63] by others was substituted by a simpler hind end locomotor test and supplemented by a hind end pinch withdrawal test developed by the author [22,23,64] to allow for a more detailed analysis of the neurologic function of SCI rats every day after the SCI. While both tests detected neurological improvements in rats treated with anti-inflammatory infusions in a 7 days long study [22], this was not the case in the follow-up study involving 56 days of treatment with Serp-1 despite the lowering effect of this treatment on the numbers of phagocytic macrophages in the COI [23]. ...
... While both tests detected neurological improvements in rats treated with anti-inflammatory infusions in a 7 days long study [22], this was not the case in the follow-up study involving 56 days of treatment with Serp-1 despite the lowering effect of this treatment on the numbers of phagocytic macrophages in the COI [23]. This observation suggests that scores in both neurological tests and perhaps the BBB test do not adequately represent the pathogenesis of the SCI where severe inflammation likely contributes to further destruction of the spinal cord [1] while the scores improve in untreated rats during the first 4 weeks post-SCI [23,64,65]. In another contradiction, while the phagocytic macrophage-rich inflammation persists beyond 4 weeks in untreated rats and in Serp-1 infused rats [1,23], the scores of both neurological tests stabilized during the week 4 post-SCI [23], an observation also made with the BBB test in treatment experiments lasting beyond 4 weeks [66][67][68]. ...
... In another contradiction, while the phagocytic macrophage-rich inflammation persists beyond 4 weeks in untreated rats and in Serp-1 infused rats [1,23], the scores of both neurological tests stabilized during the week 4 post-SCI [23], an observation also made with the BBB test in treatment experiments lasting beyond 4 weeks [66][67][68]. Thus the usefulness of the neurological test in pre-clinical studies on candidate neuroprotective compounds appears quite limited as it may not correspond to the pathogenesis of the SCI and cannot be used beyond week 4 while the effective treatments are expected to inhibit inflammation by lowering numbers of macrophages and the duration of treatment needs to last well beyond 4 weeks [23,64]. ...
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Chapter
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