Neuroscience
Abstract
Pancreatic cancer is a highly innervated gastrointestinal disease in which sympathetic nerves play a critical role in modulating tumor growth and the tumor microenvironment (TME). While recent studies suggest that sympathetic nerves influence various TME components, including lymphoid and myeloid immune cells, their interactions with cancer-associated fibroblasts (CAFs) remain poorly understood. CAFs are a hallmark of pancreatic tumors and are known to upregulate axon guidance and neuroactive cues, suggesting a potential feedback loop with tumor-innervating nerves. Here, we investigated the bidirectional crosstalk between sympathetic nerves and CAFs in human and mouse pancreatic tumors. Using a chemo-genetic ablation model, we selectively eliminated pancreatic sympathetic nerves and found that denervation significantly reduced tumor size in female mice. To further dissect this interaction, we established co-culture systems with immortalized pancreatic fibroblasts and primary sympathetic neuron explants, identifying key transcriptional changes driven by CAF-sympathetic nerve signaling. Our findings demonstrated that sympathetic signaling enhanced CAF activation and extracellular matrix remodeling, while activated CAFs, in turn, induced transcriptional programs in sympathetic neurons associated with nerve injury response. These results establish CAFs as central mediators of the tumor-supportive role of sympathetic nerves, offering further insights into the neural regulation of pancreatic cancer progression.
Authors
Ariana L. Sattler, Parham Diba, Kevin Hawthorne, Carl Pelz, Joe Grieco, Tetiana Korzun, Bryan Chong, M.J. Kuykendall, Rosalie C. Sears, Daniel L. Marks, Mara H. Sherman, Teresa A Zimmers, S. Ece Eksi
Abstract
Oxidative signaling is a central mechanism in alcohol-induced injury and has a strong implication in blood-brain barrier (BBB) dysregulation and neuroinflammation. Here, by targeting oxidative signaling, we hypothesized an innovative approach to develop a clinically relevant therapeutic strategy for alleviating alcohol-mediated neurovascular damage. To accomplish this, we enhanced the endogenous activity of Nrf2 (nuclear factor E2-related factor 2) by treating with an Nrf2 activator III TAT peptide (Nrf2 peptide, NP) and investigated the neuroprotective role of Nrf2 in promoting antioxidant defense properties and reducing BBB damage and transmigration of leukocytes to the brain following alcohol ingestion. We administered the NP subcutaneously to alcohol-ingested mice and evaluated its therapeutic potential in alleviating alcohol-associated neurovascular impairments. We compared the results with control peptide (random sequence with TAT)-treated animals. The studies showed that the NP treatment preserved the oxidant-antioxidant balance, downregulated ICAM-1 and its receptors, and mitigated BBB damage and leukocyte infiltration into the brain. We validated the effect of the NP in Nrf2 knock-out (KO) mice (Nrf2−/−). Thus, this study demonstrates NP’s neurovascular protective effects by regulating the oxidant-antioxidant balance, reducing oxidative stress-induced BBB disruption, and limiting transmigration of immune cells to the brain in a mouse model of alcohol ingestion.
Authors
Bibhuti B. Saikia, Saleena Alikunju, Yemin A. Poovanthodi, Zayan Kassim, P. M. Abdul Muneer
Abstract
Hirschsprung disease (HSCR) is a congenital intestinal disorder characterized by the absence of ganglia in the distal intestine. Despite surgical resection of the aganglionic intestine and pull-through surgery, HSCR patients still experience bowel dysfunction, indicating that latent abnormalities may also exist in the proximal ganglionic intestine. To elucidate possible causes of postoperative bowel dysfunction in HSCR, we investigated differences in the proximal ganglionic intestine using an animal model of HSCR (Ednrb-null mice) and validated our findings in tissue from human HSCR patients. We found that the proximal ganglionic colon of HSCR mice exhibited greater stiffness and fibrosis than their wild-type littermates. Similarly, submucosal fibrosis was significantly greater in the proximal ganglionic intestine of HSCR patients than in intestinal tissue from age and site-matched controls. Furthermore, we observed dysregulated expression of extracellular matrix (ECM)-related genes in the proximal ganglionic intestine of HSCR mice compared to controls. We conclude that increased fibrosis, stiffness, and alterations in ECM composition may contribute to persistent dysfunction of the ganglionic intestine in HSCR. These findings add to the growing body of literature that describe abnormalities in the proximal ganglionic intestine of HSCR and suggest that HSCR is not limited to the aganglionic intestine alone.
Authors
Prisca C. Obidike, Britney A. Hsu, Chioma Moneme, Oluyinka O. Olutoye II, Walker D. Short, Mary Hui Li, Swathi Balaji, Yuwen Zhang, Sundeep G. Keswani, Lily S. Cheng
Abstract
Mutation of KRAS in endothelial cells (KRAS-EC) leads to intracerebral hemorrhage (ICH) in brain arteriovenous malformations (bAVM), resulting in severe disabilities or even death. However, it is unclear what causes this hemorrhagic conversion of bAVM. Here, using a locally established, clinically-relevant sporadic bAVM mouse model, created by overexpressing mutant KRAS (KRASG12V) in the brain EC, we demonstrate that KRAS-EC act as trigger for microglia (MG) activation and infiltration of macrophages (Mϕ). Using three-dimensional immunostaining approach with cleared human and mouse bAVM tissues, we demonstrate an abundance of MG/Mϕ around the bAVM nidus. The presence of MG/Mϕ are correlated to the blood-brain barrier leakage in bAVM area. Time-lapsed intravital imaging in Cx3cr1-gfp;Ccr2-rfp reporter mice demonstrate the dynamic activation of MG and infiltration of Mϕ toward mutant KRAS-modified dysplastic vessels. Importantly, a time course analysis showed that these activated/infiltrated MG/Mϕ are present around the bAVMs prior to hemorrhagic conversion, and controlled depletion of MG/Mϕ reduced ICH incidence in bAVM. Inhibition of MG/Mϕ with long-term minocycline treatment attenuated the incidence of ICHs around bAVMs. Our study indicates that MG/Mϕ are involved in destabilization of KRAS-induced bAVM, leading to hemorrhagic conversion/ICH. Thus, modulation of MG/Mϕ may reduce ICH risk in bAVM patients.
Authors
Hyejin Park, Jung-Eun Park, Bridger H. Freeman, Bosco Seong Kyu Yang, Shun-Ming Ting, Alexander K. Suh, Jude P.J. Savarraj, Shuning Huang, Jakob Körbelin, Huimahn Alex Choi, Sean P. Marrelli, Jaroslaw Aronowski, Peng Roc Chen, Eunhee Kim, Eun S. Park
Abstract
Angelman syndrome (AS) is a neurodevelopmental disorder caused by loss of the maternal UBE3A allele, the sole source of UBE3A in mature neurons due to epigenetic silencing of the paternal allele. Although emerging therapies are being developed to restore UBE3A expression by activating the dormant paternal UBE3A allele, existing mouse models for such preclinical studies have limited throughput and utility, creating bottlenecks for both in vitro therapeutic screening and in vivo characterization. To address this, we developed the Ube3a-INSG dual-reporter knock-in mouse, in which an IRES-Nanoluciferase-T2A-Sun1-sfGFP (INSG) cassette was inserted downstream of the endogenous Ube3a stop codon. The INSG model preserves UBE3A protein levels and function while enabling two complementary allele-specific readouts: Sun1-sfGFP and Nanoluciferase. We show that Sun1-sfGFP, a nuclear envelope-localized reporter, enables single-cell fluorescence analysis, whole-brain light-sheet imaging, and nuclear quantification by flow cytometry. Further, Nanoluciferase supports high-throughput luminescence assays for sensitive pharmacological profiling in cultured neurons and non-invasive in vivo bioluminescence imaging for pharmacodynamic assessment. By combining scalable screening, cellular analysis, and real-time in vivo monitoring in a single model, the Ube3a-INSG dual-reporter mouse provides a powerful platform to accelerate therapeutic development centered on UBE3A.
Authors
Hanna Vihma, Lucas M. James, Hannah C. Nourie, Audrey L. Smith, Siyuan Liang, Carlee A. Friar, Tasmai Vulli, Lei Xing, Dale O. Cowley, Alain C. Burette, Benjamin D. Philpot
Abstract
Authors
Rimas V. Lukas, Ruochen Du, Harrshavasan Congivaram, Kathleen McCortney, Karan Dixit, Craig Horbinski, Margaret Schwartz, Raymond Lezon, Lauren Singer, Ditte Primdahl, Jigisha Thakkar, Amy B. Heimberger, Roger Stupp, Priya Kumthekar
Potentiation of fentanyl-induced respiratory depression by alcohol is not fully reversed by naloxone
Abstract
The high frequency of opioid overdose deaths often involves co-use of alcohol, which is reported in approximately 30% of fentanyl fatalities. Both substances depress respiratory function, and their combined effects can be lethal. The present study investigated physiological parameters of respiratory-depressant effects of fentanyl when co-administered with alcohol and its sensitivity to naloxone reversal using whole-body plethysmography in male and female Long-Evans rats. Administration of a high, sedative-like dose of alcohol alone or fentanyl alone resulted in no mortality, but fentanyl+alcohol led to mortality rates of 42% and 33% in females and males, respectively. The fentanyl+alcohol combination reduced minute ventilation and increased apneic pauses compared with either drug alone. Lower, binge-like alcohol doses, when combined with fentanyl, also amplified respiratory depression. Pretreatment with naloxone did not fully restore normal respiration. Naloxone administered after fentanyl+alcohol transiently reversed the decrease in minute ventilation but did not reverse apneic pauses. Fentanyl-dependent rats were partially tolerant to fentanyl- and fentanyl+alcohol-induced respiratory depression, but alcohol-dependent rats exhibited sensitization to alcohol- and fentanyl+alcohol-induced apnea. These findings highlight physiological parameters of severe respiratory risks with fentanyl+alcohol co-use, which are inadequately reversed by naloxone, underscoring the need for targeted strategies to manage opioid+alcohol overdoses.
Authors
Emma V. Frye, Lyndsay E. Hastings, Aniah N. Matthews, Adriana Gregory-Flores, Janaina C.M. Vendruscolo, Lindsay A. Kryszak, Shelley N Jackson, Aidan J. Hampson, Nora D. Volkow, Leandro F. Vendruscolo, Renata C.N. Marchette, George F. Koob
Abstract
Acute severe joint pain is a major symptom in gouty arthritis (GA), and its adequate treatment represents an unmet medical need. Mrgprb2, a specific mast cell receptor, has been implicated in the generation of chronic pain by mobilizing mast cell degranulation, yet its significance in GA pain and joint inflammation is still not well defined. Here, we found that Mrgprb2 was expressed in mouse synovial mast cells. In a murine model of GA, acute blockade or genetic deletion of Mrgprb2 significantly attenuated arthritis pain and hyperexcitability of joint nociceptors with significant reductions in innate immune cell recruitment in the synovium. Under naive conditions, activation of synovial Mrgprb2 was sufficient to excite peripheral terminals of joint nociceptors to induce acute joint hypernociception via the mobilization of mast cell degranulation. Additionally, the level of the neuropeptide substance P (SP) was elevated in the synovium of GA model mice. Using humanized MRGPRX2-knockin mice, we revealed that SP contributed to joint pain and inflammation by activating mast cells through Mrgprb2/MRGPRX2. These findings suggest that synovial mast cell–expressed Mrgprb2/MRGPRX2 merits consideration as a key neuroimmune player and a potential therapeutic target for treating GA pain and joint inflammation.
Authors
Lin Yang, Chengxi Liu, Jin Xiao, Yu Song, Huan Chen, Dan Li, Cong Zou, Tao Hong, Yinglan Liu, Dake Qi, Nathachit Limjunyawong, Wenjie Liu, Lintao Qu
Abstract
Amyotrophic lateral sclerosis (ALS) is a progressive motor neuron disease. Emerging evidence suggests manifestations beyond the neuromuscular system. Bone alterations are part of the ALS clinical picture; it remains unclear whether they are secondary to muscle denervation or due to an autonomous process. We investigated skeletal involvement in the SOD1(G93A) mouse model at presymptomatic (P45) and symptomatic (P110) stage through biomechanical and transcriptomic approaches. Three-point bending revealed significant reductions in femoral rigidity and maximum bending force in SOD1 mutants at P45, indicating early structural deficits. Micro-CT analysis demonstrated reduced trabecular bone mineral density and thickness at P45, with progressive trabecular loss and cortical thinning by P110. Histological examination revealed marked osteoblast loss at P45 suggesting impaired bone formation as the primary early mechanism. Transcriptomics of bulk bone and cultured osteoblasts from P45 mice identified dysregulation of bone differentiation, including downregulation of osteoblast differentiation genes and upregulation of negative regulators of ossification and increased cell senescence signatures. Unfolded protein response was upregulated in SOD1 osteoblasts. Immunohistochemistry confirmed the senescence phenotype with increased p16Ink4a level in SOD1 osteoblasts. These findings suggest that bone deterioration precede overt motor symptoms and are linked to osteoblast premature senescence.
Authors
Burak Özkan, Jan-Moritz Ramge, Diana Wiesner, Jelena Scekic-Zahirovic, Stefano Antonucci, Sandra Nungeß, Dorothea Gebauer, Anita Ignatius, Jochen H. Weishaupt, Melanie Haffner-Luntzer, Francesco Roselli
Abstract
Commotio retinae (CR) resulting from retinal trauma can lead to focal photoreceptor degeneration and permanent vision loss. Currently no therapies exist for CR-induced retinal degeneration, in part due to a lacking large animal model that replicates human injury pathology and allows testing of therapeutics. Severe CR is clinically characterized by subretinal fluid and focal photoreceptor outer nuclear layer thinning. To develop a porcine CR model, we developed a laser-guided projectile apparatus and optimized projectile delivery procedure using porcine cadaveric eyes embedded in a 3D-printed porcine skull. Scleral and corneal impacts, resulted in retinal damage consistent with patient injury but corneal impacts also led to cornea damage and opacification, which precluded follow up imaging. In live porcine eyes, scleral impacts of 39.5 m/s induced transient blood retinal barrier breakdown evidenced by subretinal fluid on optical coherence tomography (OCT), leakage observed on fluorescein and indocyanine green angiography, and transient photoreceptor outer segment disruption seen by OCT and multifocal electroretinography. Impacts above 39.5 m/s induced longer-lasting photoreceptor degeneration, but only transient blood retinal barrier breakdown. This porcine model, combined with clinically relevant imaging and diagnostic modalities will be valuable for testing the safety and efficacy of therapies to restore vision after focal photoreceptor degeneration.
Authors
Juan Amaral, Irina Bunea, Arvydas Maminishkis, Maria M. Campos, Francesca Barone, Rohan Gupta, Mitra Farnoodian, Jonathan Newport, M. Joseph Phillips, Ruchi Sharma, David M. Gamm, Kapil Bharti, Richard J. Blanch
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