Search

Potent beneficial immunomodulatory and anti-inflammatory effects of whole-molecule erythropoietin have been demonstrated in a variety of animal disease models including experimental autoimmune encephalomyelitis (EAE); however, excessive hematopoiesis limits its use in clinical applications. Our group previously generated an Epo-derived small peptide JM4 that is side-effect free and has strong neuroprotective activity without hematologic effects. Here, we investigated the long-term clinical effects of brief treatment with JM4 in chronic relapsing EAE using bioluminescence imaging (BLI) in transgenic mice containing the luciferase gene driven by the murine GFAP promoter. EAE mice treated with JM4 exhibited marked improvement in clinical scores and showed fewer disease flareups than control animals. JM4 therapy concomitantly led to markedly decreased GFAP bioluminescence in the brain and spinal cord in both acute and chronic relapsing EAE mouse models. We found a marker for toxic A1 astrocytes, complement component C3, that is upregulated in the brain and cord of EAE mice and sharply reduced in JM4-treated animals. In addition, an abnormally leaky neurovascular unit permeability was rapidly normalized within 5 days by JM4 therapy. The prolonged therapeutic benefit seen following brief JM4 treatment in EAE mice closely resemble that recently described in humans receiving pulsed immune reconstitution therapy with the disease-modifying compounds, alemtuzumab and cladribine. Our study suggests that JM4 therapy may have widespread clinical applicability for long-term treatment of inflammatory demyelinating diseases and that BLI is a useful noninvasive means of monitoring murine disease activity of the central nervous system. Electronic supplementary material The online version of this article (10.1007/s13311-020-00923-5) contains supplementary material, which is available to authorized users.

There is currently no effective medical treatment for traumatic brain injury (TBI). Beyond the immediate physical damage caused by the initial impact, additional damage evolves due to the inflammatory response that follows brain injury. Here we show that therapy with JM4, a low molecular weight 19-amino acid nonhematopoietic erythropoietin (EPO) peptidyl fragment, containing amino acids 28-46 derived from the first loop of EPO, markedly reduces acute brain injury. Mice underwent controlled cortical injury and received either whole molecule EPO, JM4, or sham-treatment with phosphate-buffered saline. Animals treated with JM4 peptide exhibited a large decrease in number of dead neural cells and a marked reduction in lesion size at both 3 and 8 days postinjury. Therapy with JM4 also led to improved functional recovery and we observed a treatment window for JM4 peptide that remained open for at least 9 h postinjury. The full-length EPO molecule was divided into a series of 6 contiguous peptide segments; the JM4-containing segment and the adjoining downstream region contained the bulk of the death attenuating effects seen with intact EPO molecule following TBI. These findings indicate that the JM4 molecule substantially blocks cell death and brain injury following acute brain trauma and, as such, presents an excellent opportunity to explore the therapeutic potential of a small-peptide EPO derivative in the medical treatment of TBI.

Under inflammatory conditions, leukocytes migrate into the tissue in a process regulated by adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1). We have reported that leukocyte ligation to VCAM-1 initiates an endothelial cell signaling cascade that is necessary for VCAM-1-dependent leukocyte transendothelial migration. It is unknown whether the cytoplasmic domain of VCAM-1 is involved in initiating the VCAM-1 signaling cascade to enable leukocyte transendothelial migration. Notably, the 19 amino acid VCAM-1 cytoplasmic domain is identical in most mammalian species, including human, mouse and common shrew. This interspecies conservation suggests that the VCAM-1 cytoplasmic domain is important for VCAM-1 signaling function. To test this hypothesis, we generated stably transfected endothelial cells with VCAM-1 constructs containing mutations in the three serines and/or the one tyrosine of the cytoplasmic domain. Using this model, we examined the ability of these mutants to signal via activation of the downstream mediator, Rac 1. We also examined the mutants’ ability to support VCAM-1-dependent leukocyte transendothelial migration under conditions of physiological laminar flow. Mutating the four putative phosphorylation sites resulted in a loss of signaling and a loss of leukocyte transendothelial migration as compared to wildtype VCAM-1. This data suggests that the cytoplasmic domain of VCAM-1 is critical to VCAM-1 signaling and function.