Profiling the mouse brain endothelial transcriptome in health and disease models reveals a core blood-brain barrier dysfunction module.

Blood vessels in the CNS form a specialized and critical structure, the blood-brain barrier (BBB). We present a resource to understand the molecular mechanisms that regulate BBB function in health and dysfunction during disease. Using endothelial cell enrichment and RNA sequencing, we analyzed the gene expression of endothelial cells in mice, comparing brain endothelial cells with peripheral endothelial cells. We also assessed the regulation of CNS endothelial gene expression in models of stroke, multiple sclerosis, traumatic brain injury and seizure, each having profound BBB disruption. We found that although each is caused by a distinct trigger, they exhibit strikingly similar endothelial gene expression changes during BBB disruption, comprising a core BBB dysfunction module that shifts the CNS endothelial cells into a peripheral endothelial cell-like state. The identification of a common pathway for BBB dysfunction suggests that targeting therapeutic agents to limit it may be effective across multiple neurological disorders

Characterization and functionality of proliferative human sertoli cells

It has long been thought that mammalian Sertoli cells are terminally differentiated and nondividing postpuberty. For most previous in vitro studies immature rodent testes have been the source of Sertoli cells and these have shown little proliferative ability when cultured. We have isolated and characterized Sertoli cells from human cadaveric testes from seven donors ranging from 12 to 36 years of age. The cells proliferated readily in vitro under the optimized conditions used with a doubling time of approximately 4 days. Nuclear 5-ethynyl-2´-deoxyuridine (EdU) incorporation confirmed that dividing cells represented the majority of the population. Classical Sertoli cell ultrastructural features, lipid droplet accumulation, and immunoexpression of GATA-4, Sox9, and the FSH receptor (FSHr) were observed by electron and fluorescence microscopy, respectively. Flow cytometry revealed the expression of GATA-4 and Sox9 by more than 99% of the cells, and abundant expression of a number of markers indicative of multipotent mesenchymal cells. Low detection of endogenous alkaline phosphatase activity after passaging showed that few peritubular myoid cells were present. GATA-4 and SOX9 expression were confirmed by reverse transcription polymerase chain reaction (RT-PCR), along with expression of stem cell factor (SCF), glial cell line-derived neurotrophic factor (GDNF), and bone morphogenic protein 4 (BMP4). Tight junctions were formed by Sertoli cells plated on transwell inserts coated with fibronectin as revealed by increased transepithelial electrical resistance (TER) and polarized secretion of the immunoregulatory protein, galectin-1. These primary Sertoli cell populations could be expanded dramatically in vitro and could be cryopreserved. The results show that functional human Sertoli cells can be propagated in vitro from testicular cells isolated from adult testis. The proliferative human Sertoli cells should have important applications in studying infertility, reproductive toxicology, testicular cancer, and spermatogenesis, and due to their unique biological properties potentially could be useful in cell therapy.Fil: Chui, Kitty. MandalMed Inc. ; Estados UnidosFil: Trivedi, Alpa. MandalMed Inc.; Estados Unidos. University of California; Estados UnidosFil: Cheng, C. Yan. Lonza Walkersville; Estados UnidosFil: Cherbavaz, Diana B.. MandalMed Inc.; Estados UnidosFil: Dazin, Paul F.. MandalMed; Estados UnidosFil: Huynh, Ai Lam Thu. MandalMed Inc.; Estados UnidosFil: Mitchel, James B.. Lonza Walkersville; Estados UnidosFil: Rabinovich, Gabriel Adrian. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); ArgentinaFil: Noble Haeusslein, Linda J.. University of California; Estados UnidosFil: John, Constance M.. MandalMed Inc.; Estados Unido

Models of Traumatic Cerebellar Injury

Traumatic brain injury (TBI) is a major cause of morbidity and mortality worldwide. Studies of human TBI demonstrate that the cerebellum is sometimes affected even when the initial mechanical insult is directed to the cerebral cortex. Some of the components of TBI, including ataxia, postural instability, tremor, impairments in balance and fine motor skills, and even cognitive deficits, may be attributed in part to cerebellar damage. Animal models of TBI have begun to explore the vulnerability of the cerebellum. In this paper, we review the clinical presentation, pathogenesis, and putative mechanisms underlying cerebellar damage with an emphasis on experimental models that have been used to further elucidate this poorly understood but important aspect of TBI. Animal models of indirect (supratentorial) trauma to the cerebellum, including fluid percussion, controlled cortical impact, weight drop impact acceleration, and rotational acceleration injuries, are considered. In addition, we describe models that produce direct trauma to the cerebellum as well as those that reproduce specific components of TBI including axotomy, stab injury, in vitro stretch injury, and excitotoxicity. Overall, these models reveal robust characteristics of cerebellar damage including regionally specific Purkinje cell injury or loss, activation of glia in a distinct spatial pattern, and traumatic axonal injury. Further research is needed to better understand the mechanisms underlying the pathogenesis of cerebellar trauma, and the experimental models discussed here offer an important first step toward achieving that objective

Role of Matrix Metalloproteinases and Therapeutic Benefits of Their Inhibition in Spinal Cord Injury

This review will focus on matrix metalloproteinases (MMPs) and their inhibitors in the context of spinal cord injury (SCI). MMPs have a specific cellular and temporal pattern of expression in the injured spinal cord. Here we consider their diverse functions in the acutely injured cord and during wound healing. Excessive activity of MMPs, and in particular gelatinase B (MMP-9), in the acutely injured cord contributes to disruption of the blood-spinal cord barrier, and the influx of leukocytes into the injured cord, as well as apoptosis. MMP-9 and MMP-2 regulate inflammation and neuropathic pain after peripheral nerve injury and may contribute to SCI-induced pain. Early pharmacologic inhibition of MMPs or the gelatinases (MMP-2 and MMP-9) results in an improvement in long-term neurological recovery and is associated with reduced glial scarring and neuropathic pain. During wound healing, gelatinase A (MMP-2) plays a critical role in limiting the formation of an inhibitory glial scar, and mice that are genetically deficient in this protease showed impaired recovery. Together, these findings illustrate complex, temporally distinct roles of MMPs in SCIs. As early gelatinase activity is detrimental, there is an emerging interest in developing gelatinase-targeted therapeutics that would be specifically tailored to the acute injured spinal cord. Thus, we focus this review on the development of selective gelatinase inhibitors

Models of traumatic cerebellar injury.

Traumatic brain injury (TBI) is a major cause of morbidity and mortality worldwide. Studies of human TBI demonstrate that the cerebellum is sometimes affected even when the initial mechanical insult is directed to the cerebral cortex. Some of the components of TBI, including ataxia, postural instability, tremor, impairments in balance and fine motor skills, and even cognitive deficits, may be attributed in part to cerebellar damage. Animal models of TBI have begun to explore the vulnerability of the cerebellum. In this paper, we review the clinical presentation, pathogenesis, and putative mechanisms underlying cerebellar damage with an emphasis on experimental models that have been used to further elucidate this poorly understood but important aspect of TBI. Animal models of indirect (supratentorial) trauma to the cerebellum, including fluid percussion, controlled cortical impact, weight drop impact acceleration, and rotational acceleration injuries, are considered. In addition, we describe models that produce direct trauma to the cerebellum as well as those that reproduce specific components of TBI including axotomy, stab injury, in vitro stretch injury, and excitotoxicity. Overall, these models reveal robust characteristics of cerebellar damage including regionally specific Purkinje cell injury or loss, activation of glia in a distinct spatial pattern, and traumatic axonal injury. Further research is needed to better understand the mechanisms underlying the pathogenesis of cerebellar trauma, and the experimental models discussed here offer an important first step toward achieving that objective

RIGOR Guidelines: Escalating STAIR and STEPS for Effective Translational Research

Stroke continues to be a serious and significant health problem in the USA and worldwide. This article will emphasize the need for good laboratory practices, transparent scientific reporting, and the use of translational research models representative of the disease state to develop effective treatments. This will allow for the testing and development of new innovative strategies so that efficacious therapies can be developed to treat ischemic and hemorrhagic stroke. This article recommends guidelines for effective translational research, most importantly, the need for study blinding, study group randomization, power analysis, accurate statistical analysis, and a conflict of interest statement. Additional guidelines to ensure reproducibility of results and confirmation of efficacy in multiple species are discussed

Traumatic Injury to the Immature Frontal Lobe: A New Murine Model of Long-Term Motor Impairment in the Absence of Psychosocial or Cognitive Deficits

Traumatic brain injury in children commonly involves the frontal lobes, and is associated with distinct structural and behavioral changes. Despite the clinical significance of injuries localized to this region during brain development, the mechanisms underlying secondary damage and long-term recovery are poorly understood. Here we have characterized the first model of unilateral focal traumatic injury to the developing frontal lobe. Male C57Bl/6J mice at postnatal day (p) 21, an age approximating a toddler-aged child, received a controlled cortical impact or sham surgery to the left frontal lobe and were euthanized 1 and 7 d later. A necrotic cavity and local inflammatory response were largely confined to the unilateral frontal lobe, dorsal corpus callosum and striatum anterior to Bregma. While cell death and accumulated beta-amyloid precursor protein were characteristic features of the peri-contusional motor cortex, corpus callosum, cingulum and dorsal striatum, underlying structures including the hippocampus showed no overt pathology. To determine the long-term functional consequences of injury at p21, two additional cohorts were subjected to a battery of behavioral tests in adolescence (p35-45) or adulthood (p70-80). In both cohorts, brain-injured mice showed normal levels of anxiety, sociability, spatial learning and memory. The signature phenotypic features were deficits in motor function and motor learning, coincident with a reduction in ipsilateral cortical brain volumes. Together, these findings demonstrate classic morphological features of a focal traumatic injury, including early cell death and axonal injury, and long-term volumetric loss of cortical volumes. The presence of deficits in sensorimotor function and coordination in the absence of abnormal findings related to anxiety, sociability and memory, likely reflect several variables including the unique location of the injury and the emergence of favorable compensatory mechanisms during subsequent brain development