Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS) is caused by mutations in SACS gene encoding sacsin, a huge protein highly expressed in cerebellar Purkinje cells (PCs). ARSACS patients, as well as mouse models, display early degeneration of PCs, but the underlying mechanisms remain unexplored, with no available treatments. In this work, we demonstrated aberrant calcium (Ca2+) homeostasis and its impact on PC degeneration in ARSACS. Mechanistically, we found pathological elevation in Ca2+-evoked responses in Sacs-/- PCs, as the result of defective mitochondria and ER trafficking to distal dendrites and strong downregulation of key Ca2+ buffer-proteins. Alteration of cytoskeletal linkers, that we identified as specific sacsin interactors, likely account for faulty organellar trafficking in Sacs-/- cerebellum. Based on this pathogenetic cascade, we treated Sacs-/- mice with Ceftriaxone, a repurposed drug which exerts neuroprotection by limiting neuronal glutamatergic stimulation, and thus Ca2+ fluxes into PCs. Ceftriaxone treatment significantly improved motor performances of Sacs-/- mice, at both pre- and post-symptomatic stages. We correlated this effect to restored Ca2+ homeostasis, which arrests PC degeneration and attenuates secondary neuroinflammation. These findings disclose new key steps in ARSACS pathogenesis and support further optimization of Ceftriaxone in pre-clinical and clinical settings for the treatment of ARSACS patients.
Andrea Del Bondio, Fabiana Longo, Daniele De Ritis, Erica Spirito, Paola Podini, Bernard Brais, Angela Bachi, Angelo Quattrini, Francesca Maltecca
Cephalic tetanus (CT) is a severe form of tetanus that follows head wounds and the intoxication of cranial nerves by tetanus neurotoxin (TeNT). Hallmarks of CT are cerebral palsy, which anticipates the typical spastic paralysis of tetanus, and rapid evolution of cardiorespiratory deficit even without generalized tetanus. How TeNT causes this unexpected flaccid paralysis, and how the canonical spasticity then rapidly evolves into cardiorespiratory defects remain unresolved aspects of CT pathophysiology. Using electrophysiology and immunohistochemistry, we demonstrate that TeNT cleaves its substrate VAMP within facial neuromuscular junctions and causes a botulism-like paralysis overshadowing its canonical spasticity. Meanwhile, TeNT spreads among brainstem neuronal nuclei and, as shown by an assay to monitor the ventilation ability of CT mice, it harms essential functions like respiration. A partial axotomy of the facial nerve revealed a still-unknown ability of TeNT to undergo intra-brainstem diffusion, which allows the toxin to spread onto brainstem nuclei devoid of direct peripheral efferents. Other showing a mechanism possibly involved in the transition from local to generalized tetanus, these findings suggest that patients with idiopathic facial nerve palsy should be immediately considered for CT and treated with antisera to block the potential progression of a life-threatening form of tetanus.
Federico Fabris, Stefano Varani, Marika Tonellato, Ivica Matak, Petra Šoštarić, Patrik Meglić, Matteo Caleo, Aram Megighian, Ornella Rossetto, Cesare Montecucco, Marco Pirazzini
Respiratory syncytial virus (RSV) infection causes significant morbidity and mortality in infants, immunocompromised, and older individuals. There is an urgent need for effective antivirals and vaccines for high risk individuals. We used two complementary in vivo models to analyze RSV-associated human lung pathology and human immune correlates of protection. RSV infection resulted in widespread human lung epithelial damage, a pro-inflammatory innate immune response, and elicited a natural adaptive human immune response that conferred protective immunity. We demonstrated a key role for human T cells in controlling RSV infection. Specifically, primed human CD8+ T cells or CD4+ T cells effectively and independently control RSV replication in human lung tissue in the absence of an RSV-specific antibody response. These preclinical data support the development of RSV vaccines which also elicit effective T cell responses to improve RSV vaccine efficacy.
Chandrav De, Raymond J. Pickles, Wenbo Yao, Baolin Liao, Allison E. Boone, Mingyu Choi, Diana M. Battaglia, Frederic B. Askin, Jason K. Whitmire, Guido Silvestri, J. Victor Garcia, Angela Wahl
Respiration can positively impact cerebrospinal fluid (CSF) flow in the brain, yet its effects on central nervous system (CNS) fluid homeostasis including waste clearance function via the glymphatic and meningeal lymphatic systems remain unclear. Here, we investigated the effect of supporting respiratory function via continuous positive airway pressure (CPAP) on glymphatic-lymphatic function in spontaneously breathing anesthetized rodents. To do this, we used a systems approach combining engineering, magnetic resonance imaging, computational fluid dynamics analysis, and physiological testing. We first designed a nasal CPAP device for use in the rat and demonstrated that it functioned similar to clinical devices as evidenced by its ability to open the upper airway, augment end-expiratory lung volume, and improve arterial oxygenation. We further showed that CPAP increased CSF flow speed at the skull base and augmented glymphatic transport regionally. The CPAP-induced augmented CSF flow speed was associated with an increase in intracranial pressure (ICP), including the ICP waveform pulse amplitude. We suggest that the augmented pulse amplitude with CPAP underlies the increase in CSF bulk flow and glymphatic transport. Our results provide new insights into the functional crosstalk at the pulmonary-CSF interface and suggest that CPAP might have therapeutic benefit for sustaining glymphatic-lymphatic function.
Burhan Ozturk, Sunil Koundal, Ehab Al Bizri, Xinan Chen, Zachary H. Gursky, Feng Dai, Andrew S. Lim, Paul Heerdt, Jonathan Kipnis, Allen Tannenbaum, Hedok Lee, Helene Benveniste
Laryngotracheal stenosis (LTS) is pathologic fibrotic narrowing of the larynx and trachea characterized by hypermetabolic fibroblasts and CD4-mediated inflammation. However, the role of CD4 T-cells in promoting LTS fibrosis is unknown. The mechanistic target of rapamycin (mTOR) signaling pathways have been shown to regulate T-cell phenotype. Here we sought to investigate the influence of mTOR signaling in CD4 T-cells on LTS pathogenesis. In this study, human LTS specimens revealed an increased population of CD4-cells expressing the activated isoform of mTOR. In a murine LTS model, targeting mTOR with systemic sirolimus and a sirolimus-eluting airway stent reduced fibrosis and TH17-cells. Selective deletion of mTOR in CD4+-cells reduced TH17-cells and attenuated fibrosis, demonstrating CD4-cells’ pathologic role in LTS. Multispectral immunofluorescence of human LTS revealed increased TH17-cells. In-vitro, TH17-cells increased collagen-1 production by LTS fibroblasts, which was prevented with sirolimus pretreatment of TH17-cells. Collectively, mTOR signaling drives pathologic CD4 T-cell phenotypes in LTS, and targeting mTOR with sirolimus is effective at treating LTS through inhibition of pro-fibrotic TH17-cells. Finally, sirolimus may be delivered locally with a drug-eluting stent, transforming clinical therapy for LTS.
Kevin M. Motz, Ioan A. Lina, Idris Samad, Michael K. Murphy, Madhavi Duvvuri, Ruth J. Davis, Alexander Gelbard, Liam Chung, Yee Chan-Li, Samuel Collins, Jonathan D. Powell, Jennifer H. Elisseeff, Maureen R. Horton, Alexander T. Hillel
Immune responses in people with multiple sclerosis (pwMS) on disease-modifying therapies (DMTs) have been of significant interest throughout the COVID-19 pandemic. Lymphocyte-targeting immunotherapies including anti-CD20 treatments and sphingosine-1-phosphate receptor (S1PR) modulators attenuate antibody responses after vaccination. Evaluation of cellular responses after vaccination is therefore of particular importance in these populations. In this study, we analysed CD4 and CD8 T cell functional responses to SARS-CoV-2 spike peptides in healthy controls and pwMS on five different DMTs by flow cytometry. Although pwMS on rituximab and fingolimod therapies had low antibody responses after both two and three vaccine doses, T cell responses in pwMS on rituximab were preserved after a third vaccination, even when an additional dose of rituximab was administered between vaccine doses two and three. PwMS taking fingolimod had low detectable T cell responses in peripheral blood. CD4 and CD8 T cell responses to SARS-CoV-2 variants of concern Delta and Omicron were lower than to the ancestral Wuhan-Hu-1 variant. Our results indicate the importance of assessing both cellular and humoral responses after vaccination and suggest that even in the absence of robust antibody responses vaccination can generate immune responses in pwMS.
Asia-Sophia Wolf, Anthony Ravussin, Marton König, Mathias H. Øverås, Guri Solum, Ingrid Fadum Kjønstad, Adity Chopra, Trygve Holmøy, Hanne F. Harbo, Silje Watterdal Syversen, Kristin Kaasen Jørgensen, Einar A. Høgestøl, John T. Vaage, Elisabeth G. Celius, Fridtjof Lund-Johansen, Ludvig A. Munthe, Gro Owren Nygaard, Siri Mjaaland
Altered mitochondrial function without a well-defined cause has been documented in the patients with ulcerative colitis (UC). In our efforts to understand UC pathogenesis, we observed reduced expression of clustered mitochondrial homologue, CLUH, only in the active UC tissues compared to the unaffected areas from the same patient and healthy controls. Stimulation with bacterial toll like receptor (TLR) ligands similarly reduced CLUH expression in the human primary macrophages. Further, CLUH negatively regulated secretion of pro-inflammatory cytokines IL6, TNF-α and rendered a pro-inflammatory niche in the TLR stimulated macrophage. CLUH was further found to bind to mitochondrial fission protein DRP-1 and also regulated DRP-1 transcription in the human macrophages. In the TLR ligand stimulated macrophages, absence of CLUH led to enhanced DRP-1 availability for mitochondrial fission and smaller dysfunctional mitochondrial pool was observed. Mechanistically, this fissioned mitochondrial pool in turn enhanced mitochondrial ROS production, reduced mitophagy and lysosomal function in the CLUH knockout macrophages. Remarkably, our studies in the mice model of colitis with CLUH knockdown displayed exacerbated disease pathology. Taken together, this is the first report signifying the role of CLUH in UC pathogenesis, by means of regulating inflammation via maintaining mitochondrial-lysosomal functions in the human macrophages and intestinal mucosa.
Shaziya Khan, Desh Raj, Shikha Sahu, Anam Naseer, Nishakumari C. Singh, Sunaina Kumari, Sharmeen Ishteyaque, Jyotsna Sharma, Promila Lakra, Madhav Nilakanth Mugale, Arun Kumar Trivedi, Mrigank Srivastava, Tulika Chandra, Vivek Bhosale, Manoj Kumar Barthwal, Shashi Kumar Gupta, Kalyan Mitra, Aamir Nazir, Uday C. Ghoshal, Amit Lahiri
Obesity-associated metabolic inflammation drives the development of insulin resistance and type 2 diabetes, notably through modulating innate and adaptive immune cells in metabolic organs. The nutrient sensor liver kinase B1 (LKB1) has recently been shown to control cellular metabolism and T cell priming functions of dendritic cells (DCs). Here, we report that hepatic DCs from high-fat diet (HFD)-fed obese mice display increased LKB1 phosphorylation and that LKB1 deficiency in DCs (CD11cΔLKB1) worsened HFD-driven hepatic steatosis and impaired glucose homeostasis. Loss of LKB1 in DCs was associated with increased expression of T helper 17-polarizing cytokines and accumulation of hepatic IL-17A+ T helper cells in HFD-fed mice. Importantly, IL-17A neutralization rescued metabolic perturbations in HFD-fed CD11cΔLKB1 mice. Mechanistically, deficiency of the canonical LKB1 target AMPK in HFD-fed CD11cΔAMPKα1 mice recapitulated neither the hepatic Th17 phenotype nor the disrupted metabolic homeostasis, suggesting the involvement of other and/or additional LKB1 downstream effectors. We indeed provide evidence that the control of Th17 responses by DCs via LKB1 is actually dependent on both AMPKalpha1 and AMPK-related salt-inducible kinase(s) signaling. Altogether, our data reveal a key role for LKB1 signaling in DCs in protection against obesity-induced metabolic dysfunctions by limiting hepatic Th17 responses.
Hendrik J.P. van der Zande, Eline C. Brombacher, Joost M. Lambooij, Leonard R. Pelgrom, Anna Zawistowska-Deniziak, Thiago A. Patente, Graham A. Heieis, Frank Otto, Arifa Ozir-Fazalalikhan, Maria Yazdanbakhsh, Bart Everts, Bruno Guigas
Elevation of glucagon levels and increase in alpha cell proliferation is associated with states of hyperglycemia in diabetes. A better understanding of the molecular mechanisms governing glucagon secretion could have major implications in understanding abnormal responses to hypoglycemia in diabetes patients and provide novel avenues for diabetes management. Our previous studies have highlighted the role of nutrient signaling via mTOR complex 1 (mTORC1) regulation that controls glucagon secretion and alpha cell mass and that hyperglucagonemia can improve glucose homeostasis by diminishing glucagon action in the liver. However, it is unclear if short-term effects of mTORC1 activation are sufficient to induce glucagon secretion without changes in alpha cell mass and whether short-term hyperglucagonemia reduces liver glucagon action in a reversible manner. Using mice with inducible induction of the regulator of the mTORC1 complex (Rheb) in alpha cells (αRhebTg), we showed that short-term activation of mTORC1 signaling is sufficient to induce hyperglucagonemia as a result of increased glucagon secretion. Hyperglucagonemia in the αRhebTg was also associated with an increase in alpha cell size and mass expansion. This model allowed us to identify the effects of chronic and short-term hyperglucagonemia on glucose homeostasis by regulating glucagon signaling in the liver. Short-term hyperglucagonemia impaired glucose tolerance, which was reversible over time. Decrease in liver glucagon effects in αRhebTg mice was associated with reduced expression of the glucagon receptor (GCGR) and genes involved in gluconeogenesis, amino acid metabolism, and urea production. However, only genes regulating gluconeogenesis returned to baseline upon improvement of glycemia. Overall, these studies demonstrate that hyperglucagonemia exerts a biphasic response on glucose metabolism: short-term hyperglucagonemia leads to glucose intolerance, whereas chronic exposure to glucagon generates decrease on hepatic glucagon action along with improved glucose tolerance.
Camila Lubaczeuski, Nadejda Bozadjieva-Kramer, Ruy A. Louzada, George K. Gittes, Gil Leibowitz, Ernesto Bernal-Mizrachi
Elevated blood glucose levels, or hyperglycemia, can increase brain excitability and amyloid-beta (Aβ) release offering a mechanistic link between type-2-diabetes and Alzheimer’s disease (AD). Since the cellular mechanisms governing this relationship are poorly understood, we explored whether ATP-sensitive potassium (KATP) channels, which couple changes in energy availability with cellular excitability, play a role in AD pathogenesis. First, we demonstrate that KATP channel subunits, Kir6.2/KCNJ11 and SUR1/ABCC8 are expressed on excitatory and inhibitory neurons in the human brain and cortical expression of KCNJ11 and ABCC8 changes with Alzheimer’s pathology in humans and mice. Next, we explored whether eliminating neuronal KATP channel activity uncoupled the relationship between metabolism, excitability, and Aβ pathology in a novel mouse model of cerebral amyloidosis and neuronal KATP channel ablation (e.g. APP/PS1, Kir6.2-/- mouse). Using both acute and chronic paradigms, we demonstrate that Kir6.2-KATP channels are metabolic sensors that regulate hyperglycemic-dependent increases in interstitial fluid levels of Aβ, amyloidogenic processing of APP, and amyloid plaque formation, which may be dependent on lactate release. These studies identify a new role for Kir6.2-KATP channels in Alzheimer’s disease and suggest that pharmacological manipulation of Kir6.2-KATP channels holds therapeutic promise in reducing Aβ pathology in diabetic or prediabetic patients.
John Grizzanti, William R. Moritz, Morgan C. Pait, Molly Stanley, Sarah D. Kaye, Caitlin M. Carroll, Nicholas J. Constantino, Lily J. Deitelzweig, James A. Snipes, Derek Kellar, Emily E. Caesar, Ryan J. Pettit-Mee, Stephen M. Day, Jonathon P. Sens, Noelle I. Nicol, Jasmeen Dhillon, Maria S. Remedi, Drew D. Kiraly, Celeste M. Karch, Colin G. Nichols, David M. Holtzman, Shannon L. Macauley
No posts were found with this tag.