Neuroscience
Abstract
Sepsis-associated encephalopathy manifesting as delirium is a common problem in critical care medicine. In this study, patients that had delirium due to sepsis had significant cognitive impairments at 12–18 months after hospital discharge when compared with controls and Cambridge Neuropsychological Automated Test Battery–standardized scores in spatial recognition memory, pattern recognition memory, and delayed-matching-to-sample tests but not other cognitive functions. A mouse model of S. pneumoniae pneumonia-induced sepsis, which modeled numerous aspects of the human sepsis-associated multiorgan dysfunction, including encephalopathy, also revealed similar deficits in spatial memory but not new task learning. Both humans and mice had large increases in chemokines for myeloid cell recruitment. Intravital imaging of the brains of septic mice revealed increased neutrophil and CCR2+ inflammatory monocyte recruitment (the latter being far more robust), accompanied by subtle microglial activation. Prevention of CCR2+ inflammatory monocyte recruitment, but not neutrophil recruitment, reduced microglial activation and other signs of neuroinflammation and prevented all signs of cognitive impairment after infection. Therefore, therapeutically targeting CCR2+ inflammatory monocytes at the time of sepsis may provide a novel neuroprotective clinical intervention to prevent the development of persistent cognitive impairments.
Authors
Graciela Andonegui, Erin L. Zelinski, Courtney L. Schubert, Derrice Knight, Laura A. Craig, Brent W. Winston, Simon C. Spanswick, Björn Petri, Craig N. Jenne, Janice C. Sutherland, Rita Nguyen, Natalie Jayawardena, Margaret M. Kelly, Christopher J. Doig, Robert J. Sutherland, Paul Kubes
Abstract
Androgen excess is a hallmark of polycystic ovary syndrome (PCOS), a prevalent yet poorly understood endocrine disorder. Evidence from women and preclinical animal models suggests that elevated perinatal androgens can elicit PCOS onset in adulthood, implying androgen actions in both PCOS ontogeny and adult pathophysiology. Prenatally androgenized (PNA) mice exhibit a robust increase of progesterone-sensitive GABAergic inputs to gonadotropin-releasing hormone (GnRH) neurons implicated in the pathogenesis of PCOS. It is unclear when altered GABAergic wiring develops in the brain, and whether these central abnormalities are dependent upon adult androgen excess. Using GnRH-GFP–transgenic mice, we determined that increased GABA input to GnRH neurons occurs prior to androgen excess and the manifestation of reproductive impairments in PNA mice. These data suggest that brain circuit abnormalities precede the postpubertal development of PCOS traits. Despite the apparent developmental programming of circuit abnormalities, long-term blockade of androgen receptor signaling from early adulthood rescued normal GABAergic wiring onto GnRH neurons, improved ovarian morphology, and restored reproductive cycles in PNA mice. Therefore, androgen excess maintains changes in female brain wiring linked to PCOS features and the blockade of androgen receptor signaling reverses both the central and peripheral PNA-induced PCOS phenotype.
Authors
Mauro S.B. Silva, Melanie Prescott, Rebecca E. Campbell
Abstract
Ischemic stroke is a leading cause of morbidity and mortality in the US; however, there currently exists only one effective acute pharmacological therapeutic intervention. Purinergic signaling has been shown to regulate vascular function and pathological processes, including inflammation and arterial myogenic reactivity, and plays a role in ischemic stroke outcome. Purinergic signaling requires extracellular purines; however, the mechanism of purine release from cells is unclear. Pannexin1 (Panx1) channels are potentially novel purine release channels expressed throughout the vascular tree that couples regulated purine release with purinergic signaling. Therefore, we examined the role of smooth muscle and endothelial cell Panx1, using conditional cell type–specific transgenic mice, in cerebral ischemia/reperfusion injury outcomes. Deletion of endothelial cell Panx1, but not smooth muscle cell Panx1, significantly reduced cerebral infarct volume after ischemia/reperfusion. Infiltration of leukocytes into brain tissue and development of cerebral myogenic tone were both significantly reduced when mice lacked endothelial Panx1. Panx1 regulation of myogenic tone was unique to the cerebral circulation, as mesenteric myogenic reactivity and blood pressure were independent of endothelial Panx1. Overall, deletion of endothelial Panx1 mitigated cerebral ischemic injury by reducing inflammation and myogenic tone development, indicating that endothelial Panx1 is a possible novel target for therapeutic intervention of ischemic stroke.
Authors
Miranda E Good, Stephanie A. Eucker, Jun Li, Hannah M. Bacon, Susan M. Lang, Joshua T. Butcher, Tyler J. Johnson, Ronald P. Gaykema, Manoj K. Patel, Zhiyi Zuo, Brant E. Isakson
Abstract
Pain is the predominant symptom of osteoarthritis, but the connection between joint damage and the genesis of pain is not well understood. Loss of articular cartilage is a hallmark of osteoarthritis, and it occurs through enzymatic degradation of aggrecan by cleavage mediated by a disintegrin and metalloproteinase with thrombospondin motif 4 (ADAMTS-4) or ADAMTS-5 in the interglobular domain (E373–374A). Further cleavage by MMPs (N341–342F) releases a 32-amino-acid aggrecan fragment (32-mer). We investigated the role of this 32-mer in driving joint pain. We found that the 32-mer excites dorsal root ganglion nociceptive neurons, both in culture and in intact explants. Treatment of cultured sensory neurons with the 32-mer induced expression of the proalgesic chemokine CCL2. These effects were mediated through TLR2, which we demonstrated was expressed by nociceptive neurons. In addition, intra-articular injection of the 32-mer fragment provoked knee hyperalgesia in WT but not Tlr2-null mice. Blocking the production or action of the 32-mer in transgenic mice prevented the development of knee hyperalgesia in a murine model of osteoarthritis. These findings suggest that the aggrecan 32-mer fragment directly activates TLR2 on joint nociceptors and is an important mediator of the development of osteoarthritis-associated joint pain.
Authors
Rachel E. Miller, Shingo Ishihara, Phuong B. Tran, Suzanne B. Golub, Karena Last, Richard J. Miller, Amanda J. Fosang, Anne-Marie Malfait
Abstract
Fabry disease, the most common lysosomal storage disease, affects multiple organs and results in a shortened life span. This disease is caused by a deficiency of the lysosomal enzyme α-galactosidase A, which leads to glycosphingolipid accumulation in many cell types. Neuropathic pain is an early and severely debilitating symptom in patients with Fabry disease, but the cellular and molecular mechanisms that cause the pain are unknown. We generated a rat model of Fabry disease, the first nonmouse model to our knowledge. Fabry rats had substantial serum and tissue accumulation of α-galactosyl glycosphingolipids and had pronounced mechanical pain behavior. Additionally, Fabry rat dorsal root ganglia displayed global N-glycan alterations, sensory neurons were laden with inclusions, and sensory neuron somata exhibited prominent sensitization to mechanical force. We found that the cation channel transient receptor potential ankyrin 1 (TRPA1) is sensitized in Fabry rat sensory neurons and that TRPA1 antagonism reversed the behavioral mechanical sensitization. This study points toward TRPA1 as a potentially novel target to treat the pain experienced by patients with Fabry disease.
Authors
James J. Miller, Kazuhiro Aoki, Francie Moehring, Carly A. Murphy, Crystal L. O’Hara, Michael Tiemeyer, Cheryl L. Stucky, Nancy M. Dahms
Abstract
In multiple sclerosis (MS), a demyelinating inflammatory disease of the CNS, and its animal model (experimental autoimmune encephalomyelitis; EAE), circulating immune cells gain access to the CNS across the blood-brain barrier to cause inflammation, myelin destruction, and neuronal damage. Here, we discovered that calnexin, an ER chaperone, is highly abundant in human brain endothelial cells of MS patients. Conversely, mice lacking calnexin exhibited resistance to EAE induction, no evidence of immune cell infiltration into the CNS, and no induction of inflammation markers within the CNS. Furthermore, calnexin deficiency in mice did not alter the development or function of the immune system. Instead, the loss of calnexin led to a defect in brain endothelial cell function that resulted in reduced T cell trafficking across the blood-brain barrier. These findings identify calnexin in brain endothelial cells as a potentially novel target for developing strategies aimed at managing or preventing the pathogenic cascade that drives neuroinflammation and destruction of the myelin sheath in MS.
Authors
Joanna Jung, Paul Eggleton, Alison Robinson, Jessica Wang, Nick Gutowski, Janet Holley, Jia Newcombe, Elzbieta Dudek, Amber M. Paul, Douglas Zochodne, Allison Kraus, Christopher Power, Luis B. Agellon, Marek Michalak
Abstract
CHD7, an ATP-dependent chromatin remodeler, is disrupted in CHARGE syndrome, an autosomal dominant disorder characterized by variably penetrant abnormalities in craniofacial, cardiac, and nervous system tissues. The inner ear is uniquely sensitive to CHD7 levels and is the most commonly affected organ in individuals with CHARGE. Interestingly, upregulation or downregulation of retinoic acid (RA) signaling during embryogenesis also leads to developmental defects similar to those in CHARGE syndrome, suggesting that CHD7 and RA may have common target genes or signaling pathways. Here, we tested three separate potential mechanisms for CHD7 and RA interaction: (a) direct binding of CHD7 with RA receptors, (b) regulation of CHD7 levels by RA, and (c) CHD7 binding and regulation of RA-related genes. We show that CHD7 directly regulates expression of Aldh1a3, the gene encoding the RA synthetic enzyme ALDH1A3 and that loss of Aldh1a3 partially rescues Chd7 mutant mouse inner ear defects. Together, these studies indicate that ALDH1A3 acts with CHD7 in a common genetic pathway to regulate inner ear development, providing insights into how CHD7 and RA regulate gene expression and morphogenesis in the developing embryo.
Authors
Hui Yao, Sophie F. Hill, Jennifer M. Skidmore, Ethan D. Sperry, Donald L. Swiderski, Gilson J. Sanchez, Cynthia F. Bartels, Yehoash Raphael, Peter C. Scacheri, Shigeki Iwase, Donna M. Martin
Abstract
We identify 2 homozygous mutations in the ε-subunit of the muscle acetylcholine receptor (AChR) in 3 patients with severe congenital myasthenia: εR218W in the pre-M1 region in 2 patients and εE184K in the β8-β9 linker in 1 patient. Arg218 is conserved in all eukaryotic members of the Cys-loop receptor superfamily, while Glu184 is conserved in the α-, δ-, and ε-subunits of AChRs from all species. εR218W reduces channel gating efficiency 338-fold and AChR expression on the cell surface 5-fold, whereas εE184K reduces channel gating efficiency 11-fold but does not alter AChR cell surface expression. Determinations of the effective channel gating rate constants, combined with mutant cycle analyses, demonstrate strong energetic coupling between εR218 and εE184, and between εR218 and εE45 from the β1-β2 linker, as also observed for equivalent residues in the principal coupling pathway of the α-subunit. Thus, efficient and rapid gating of the AChR channel is achieved not only by coupling between conserved residues within the principal coupling pathway of the α-subunit, but also between corresponding residues in the ε-subunit.
Authors
Xin-Ming Shen, Joan M. Brengman, Shelley Shen, Hacer Durmus, Veeramani Preethish-Kumar, Nur Yuceyar, Seena Vengalil, Atchayaram Nalini, Feza Deymeer, Steven M. Sine, Andrew G. Engel
Abstract
Tumor microenvironments can promote stem cell maintenance, tumor growth, and therapeutic resistance, findings linked by the tumor-initiating cell hypothesis. Standard of care for glioblastoma (GBM) includes temozolomide chemotherapy, which is not curative, due, in part, to residual therapy-resistant brain tumor-initiating cells (BTICs). Temozolomide efficacy may be increased by targeting carbonic anhydrase 9 (CA9), a hypoxia-responsive gene important for maintaining the altered pH gradient of tumor cells. Using patient-derived GBM xenograft cells, we explored whether CA9 and CA12 inhibitor SLC-0111 could decrease GBM growth in combination with temozolomide or influence percentages of BTICs after chemotherapy. In multiple GBMs, SLC-0111 used concurrently with temozolomide reduced cell growth and induced cell cycle arrest via DNA damage in vitro. In addition, this treatment shifted tumor metabolism to a suppressed bioenergetic state in vivo. SLC-0111 also inhibited the enrichment of BTICs after temozolomide treatment determined via CD133 expression and neurosphere formation capacity. GBM xenografts treated with SLC-0111 in combination with temozolomide regressed significantly, and this effect was greater than that of temozolomide or SLC-0111 alone. We determined that SLC-0111 improves the efficacy of temozolomide to extend survival of GBM-bearing mice and should be explored as a treatment strategy in combination with current standard of care.
Authors
Nathaniel H. Boyd, Kiera Walker, Joshua Fried, James R. Hackney, Paul C. McDonald, Gloria A. Benavides, Raffaella Spina, Alessandra Audia, Sarah E. Scott, Catherine J. Libby, Anh Nhat Tran, Mark O. Bevensee, Corinne Griguer, Susan Nozell, G. Yancey Gillespie, Burt Nabors, Krishna P. Bhat, Eli E. Bar, Victor Darley-Usmar, Bo Xu, Emily Gordon, Sara J. Cooper, Shoukat Dedhar, Anita B. Hjelmeland
Identification of ion-channel modulators that protect against aminoglycoside-induced hair cell death
Abstract
Aminoglycoside antibiotics are used to treat life-threatening bacterial infections but can cause deafness due to hair cell death in the inner ear. Compounds have been described that protect zebrafish lateral line hair cells from aminoglycosides, but few are effective in the cochlea. As the aminoglycosides interact with several ion channels, including the mechanoelectrical transducer (MET) channels by which they can enter hair cells, we screened 160 ion-channel modulators, seeking compounds that protect cochlear outer hair cells (OHCs) from aminoglycoside-induced death in vitro. Using zebrafish, 72 compounds were identified that either reduced loading of the MET-channel blocker FM 1-43FX, decreased Texas red–conjugated neomycin labeling, or reduced neomycin-induced hair cell death. After testing these 72 compounds, and 6 structurally similar compounds that failed in zebrafish, 13 were found that protected against gentamicin-induced death of OHCs in mouse cochlear cultures, 6 of which are permeant blockers of the hair cell MET channel. None of these compounds abrogated aminoglycoside antibacterial efficacy. By selecting those without adverse effects at high concentrations, 5 emerged as leads for developing pharmaceutical otoprotectants to alleviate an increasing clinical problem.
Authors
Emma J. Kenyon, Nerissa K. Kirkwood, Siân R. Kitcher, Molly O’Reilly, Marco Derudas, Daire M. Cantillon, Richard J. Goodyear, Abigail Secker, Sarah Baxendale, James C. Bull, Simon J. Waddell, Tanya T. Whitfield, Simon E. Ward, Corné J. Kros, Guy P. Richardson
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