scholarly journals Knockin mouse models demonstrate differential contributions of synaptotagmin-1 and -2 as receptors for botulinum neurotoxins

2021 ◽  
Vol 17 (10) ◽  
pp. e1009994
Author(s):  
Hatim Thaker ◽  
Jie Zhang ◽  
Shin-Ichiro Miyashita ◽  
Vivian Cristofaro ◽  
SunHyun Park ◽  
...  
Keyword(s):  
Mouse Models ◽  
Neuromuscular Junctions ◽  
Critical Role ◽  
Mutant Mice ◽  
Autonomic Nerves ◽  
Vesicle Membrane ◽  
Wide Range ◽  
Synaptotagmin 1 ◽  
Botulinum Neurotoxins

Botulinum neurotoxins (BoNTs) are the most potent toxins known and are also utilized to treat a wide range of disorders including muscle spasm, overactive bladder, and pain. BoNTs’ ability to target neurons determines their specificity, potency, and therapeutic efficacy. Homologous synaptic vesicle membrane proteins synaptotagmin-1 (Syt1) and synaptotagmin-2 (Syt2) have been identified as receptors for BoNT family members including BoNT/B, DC, and G, but their contributions at physiologically relevant toxin concentrations in vivo have yet to be validated and established. Here we generated two knockin mutant mouse models containing three designed point-mutations that specifically disrupt BoNT binding in endogenous Syt1 or Syt2, respectively. Utilizing digit abduction score assay by injecting toxins into the leg muscle, we found that Syt1 mutant mice showed similar sensitivity as the wild type mice, whereas Syt2 mutant mice showed reduced sensitivity to BoNT/B, DC, and G, demonstrating that Syt2 is the dominant receptor at skeletal neuromuscular junctions. We further developed an in vivo bladder injection assay for analyzing BoNT action on bladder tissues and demonstrated that Syt1 is the dominant toxin receptor in autonomic nerves controlling bladder tissues. These findings establish the critical role of protein receptors for the potency and specificity of BoNTs in vivo and demonstrate the differential contributions of Syt1 and Syt2 in two sets of clinically relevant target tissues.

scholarly journals A Critical Role of the p75 Tumor Necrosis Factor Receptor (p75TNF-R) in Organ Inflammation Independent of  TNF, Lymphotoxin α, or the p55TNF-R

1998 ◽  
Vol 188 (7) ◽  
pp. 1343-1352 ◽  
Author(s):  
Eleni Douni ◽  
George Kollias
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Critical Role ◽  
Tumor Necrosis Factor Receptor ◽  
Peripheral Blood Mononuclear ◽  
Enhanced Production ◽  
Wide Range ◽  
Lymphotoxin Α ◽  
Necrosis Factor

Despite overwhelming evidence that enhanced production of the p75 tumor necrosis factor receptor (p75TNF-R) accompanies development of specific human inflammatory pathologies such as multi-organ failure during sepsis, inflammatory liver disease, pancreatitis, respiratory distress syndrome, or AIDS, the function of this receptor remains poorly defined in vivo. We show here that at levels relevant to human disease, production of the human p75TNF-R in transgenic mice results in a severe inflammatory syndrome involving mainly the pancreas, liver, kidney, and lung, and characterized by constitutively increased NF-κB activity in the peripheral blood mononuclear cell compartment. This process is shown to evolve independently of the presence of TNF, lymphotoxin α, or the p55TNF-R, although coexpression of a human TNF transgene accelerated pathology. These results establish an independent role for enhanced p75TNF-R production in the pathogenesis of inflammatory disease and implicate the direct involvement of this receptor in a wide range of human inflammatory pathologies.

scholarly journals The Diverse Roles of the Global Transcriptional Regulator PhoP in the Lifecycle of Yersinia pestis

Pathogens ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 1039
Author(s):  
Hana S. Fukuto ◽  
Gloria I. Viboud ◽  
Viveka Vadyvaloo
Keyword(s):  
Yersinia Pestis ◽  
Current Knowledge ◽  
Response Regulator ◽  
Expression Profiles ◽  
Critical Role ◽  
Gene Expression Profiles ◽  
Acanthamoeba Castellanii ◽  
Wide Range

Yersinia pestis, the causative agent of plague, has a complex infectious cycle that alternates between mammalian hosts (rodents and humans) and insect vectors (fleas). Consequently, it must adapt to a wide range of host environments to achieve successful propagation. Y. pestis PhoP is a response regulator of the PhoP/PhoQ two-component signal transduction system that plays a critical role in the pathogen’s adaptation to hostile conditions. PhoP is activated in response to various host-associated stress signals detected by the sensor kinase PhoQ and mediates changes in global gene expression profiles that lead to cellular responses. Y. pestis PhoP is required for resistance to antimicrobial peptides, as well as growth under low Mg2+ and other stress conditions, and controls a number of metabolic pathways, including an alternate carbon catabolism. Loss of phoP function in Y. pestis causes severe defects in survival inside mammalian macrophages and neutrophils in vitro, and a mild attenuation in murine plague models in vivo, suggesting its role in pathogenesis. A Y. pestisphoP mutant also exhibits reduced ability to form biofilm and to block fleas in vivo, indicating that the gene is also important for establishing a transmissible infection in this vector. Additionally, phoP promotes the survival of Y. pestis inside the soil-dwelling amoeba Acanthamoeba castellanii, a potential reservoir while the pathogen is quiescent. In this review, we summarize our current knowledge on the mechanisms of PhoP-mediated gene regulation in Y. pestis and examine the significance of the roles played by the PhoP regulon at each stage of the Y. pestis life cycle.

scholarly journals Smcr8 deficiency disrupts axonal transport-dependent lysosomal function and promotes axonal swellings and gain of toxicity in C9ALS/FTD mouse models

2019 ◽  
Vol 28 (23) ◽  
pp. 3940-3953 ◽  
Author(s):  
Chen Liang ◽  
Qiang Shao ◽  
Wei Zhang ◽  
Mei Yang ◽  
Qing Chang ◽  
...  
Keyword(s):  
Axonal Transport ◽  
Mouse Models ◽  
Motor Neurons ◽  
Neuromuscular Junctions ◽  
Motor Behavior ◽  
In Vivo Studies ◽  
Lysosomal Function ◽  
Repeat Expansions ◽  
Axonal Swellings

Abstract G4C2 repeat expansions in an intron of C9ORF72 cause the most common familial amyotrophic lateral sclerosis and frontotemporal dementia (collectively, C9ALS/FTD). Mechanisms and mediators of C9ALS/FTD pathogenesis remain poorly understood. C9orf72 and Smcr8 form a protein complex. Here, we show that expression of Smcr8, like C9orf72, is reduced in C9ALS/FTD mouse models and patient tissues. Since Smcr8 is highly conserved between human and mouse, we evaluated the effects of Smcr8 downregulation in mice. Smcr8 knockout (KO) mice exhibited motor behavior deficits, which resemble those of C9ALS/FTD mouse models, and displayed axonal swellings in their spinal cords and neuromuscular junctions. These deficits are caused by impaired autophagy-lysosomal functions due to disrupted axonal transport in mutant motor neurons. Consistent with its interaction with C9orf72 and their downregulation in patient tissues, Smcr8 deficiency exacerbated autophagy-lysosomal impairment in C9orf72 KO mice. The disease relevance of Smcr8 downregulation was reflected by exacerbated axonal swellings and gain of toxicity pathology arising from Smcr8 haploinsufficiency in a mouse model of C9ALS/FTD. Thus, our in vivo studies suggested that Smcr8 deficiency impairs axonal transport dependent autophagy-lysosomal function and exacerbates axonal degeneration and gain of toxicity in C9ALS/FTD mouse models.

The microtubule modulator RanBP10 plays a critical role in regulation of platelet discoid shape and degranulation

Blood ◽  
2009 ◽  
Vol 114 (27) ◽  
pp. 5532-5540 ◽  
Author(s):  
Stefan Kunert ◽  
Imke Meyer ◽  
Silke Fleischhauer ◽  
Martin Wannack ◽  
Janine Fiedler ◽  
...  
Keyword(s):  
Fetal Liver ◽  
Critical Role ◽  
Mutant Mice ◽  
Axis Ratio ◽  
Prolonged Bleeding Time ◽  
Normal Platelet ◽  
And Function ◽  
Granule Release

Abstract Terminally mature megakaryocytes undergo dramatic cellular reorganization to produce hundreds of virtually identical platelets. A hallmark feature of this process is the generation of an elaborate system of branched protrusions called proplatelets. We recently identified RanBP10 as a tubulin-binding protein that is concentrated along polymerized microtubules in mature megakaryocytes. RanBP10 depletion in vitro caused the disturbance of polymerized filaments. Here we study the function of RanBP10 in vivo by generating deficient mice using a gene-trap approach. Mutant mice show normal platelet counts, and fetal liver-derived megakaryocytes reveal only slightly reduced proplatelet formation. However, ultrastructural analysis unveiled a significantly increased geometric axis ratio for resting platelets, and many platelets exhibited disorders in microtubule filament numbers and localization. Mutant mice showed a markedly prolonged bleeding time. Granule release, a process that depends on internal contraction of the microtubule marginal coil, also was reduced. Flow cytometry analysis revealed reduced expression of CD62P and CD63 after PAR4-peptide stimulation. These data suggest that RanBP10 plays an essential role in hemostasis and in maintaining microtubule dynamics with respect to both platelet shape and function.

Abstract 16515: Differential Functions of HCN1 and HCN4 Pacemaker Channels Within the Sinoatrial Node for the Stable and Precise Beating of the Heart

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Stefanie Fenske ◽  
Vanessa Marks ◽  
Stefanie Koenigsbauer ◽  
Sami I Hassan ◽  
Tilman Ziegler ◽  
...  
Keyword(s):  
Heart Rate ◽  
Mouse Models ◽  
Sinoatrial Node ◽  
Critical Role ◽  
Functional Characterization ◽  
Heart Beat ◽  
Fine Tuning ◽  
Hcn Channels ◽  
Pacemaker Cells

The heart beat is initiated by the generation of spontaneous action potentials in pacemaker cells of the sinoatrial node (SAN) region. The maintenance of a stable heart beat requires mechanisms which protect the SAN pacemaker cells from potential perturbing influences which arise from inside and outside the sinoatrial network. Our previous work suggests that the hyperpolarization-activated cyclic nucleotide gated channel subtype 1 (HCN1) protects against such perturbations and thereby increases the stability, the precision and the safety of the sinoatrial network. Here, we investigate the role of HCN4 channels within this context. Using genetic mouse models deficient for HCN channels as well as mouse models expressing engineered HCN channels, we performed a detailed functional characterization of pacemaker mechanisms in single isolated sinoatrial node cells, explanted beating sinoatrial node preparations, with telemetric in vivo electrocardiography, echocardiography, and in vivo electrophysiology. We provide evidence that HCN4 has a critical role in counteracting and balancing potentially destabilizing effects of the autonomic nervous system on the regulation of the heart rate. Specifically, HCN4 channels smooth the transition of the heart rate to a new equilibrium. Furthermore, we provide evidence that the cAMP- dependent fine tuning of HCN4 channel activity could provide the exact dosage of current to balance and counteract overshooting responses of the heart rate to autonomic regulation. In the absence of such a protecting effect, mice display a brady- tachy syndrome.

scholarly journals Hypoexcitability precedes denervation in the large fast-contracting motor units in two unrelated mouse models of ALS

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
María de Lourdes Martínez-Silva ◽  
Rebecca D Imhoff-Manuel ◽  
Aarti Sharma ◽  
CJ Heckman ◽  
Neil A Shneider ◽  
...  
Keyword(s):  
Motor Unit ◽  
Mouse Models ◽  
Neuromuscular Junctions ◽  
Motor Units ◽  
Loss Of Function ◽  
Disease Markers ◽  
Physiological Type ◽  
Lateral Sclerosis ◽  
Motoneuron Degeneration

Hyperexcitability has been suggested to contribute to motoneuron degeneration in amyotrophic lateral sclerosis (ALS). If this is so, and given that the physiological type of a motor unit determines the relative susceptibility of its motoneuron in ALS, then one would expect the most vulnerable motoneurons to display the strongest hyperexcitability prior to their degeneration, whereas the less vulnerable should display a moderate hyperexcitability, if any. We tested this hypothesis in vivo in two unrelated ALS mouse models by correlating the electrical properties of motoneurons with their physiological types, identified based on their motor unit contractile properties. We found that, far from being hyperexcitable, the most vulnerable motoneurons become unable to fire repetitively despite the fact that their neuromuscular junctions were still functional. Disease markers confirm that this loss of function is an early sign of degeneration. Our results indicate that intrinsic hyperexcitability is unlikely to be the cause of motoneuron degeneration.

Gene-targeting and transgenic approaches to IGF and IGF binding protein function

1995 ◽  
Vol 269 (4) ◽  
pp. E613-E622 ◽  
Author(s):  
T. L. Wood
Keyword(s):  
Mouse Models ◽  
Protein Function ◽  
Binding Proteins ◽  
Germ Line ◽  
Critical Role ◽  
Somatic Growth ◽  
Igf Binding Proteins ◽  
Igf Binding ◽  
Mouse Lines

The ability to manipulate genetic information in the germ line of mice has provided powerful approaches to study gene function in vivo. These approaches have included the establishment of mouse lines in which a specified gene or genes are overexpressed, ectopically expressed, or deleted. Transgenic and gene-targeted mouse lines have been used extensively to study the function of the insulin-like growth factors (IGF), IGF-I and IGF-II, and their receptors and binding proteins. In the IGF system, these technologies have elucidated the roles of the IGFs in fetal and somatic growth and have demonstrated a critical role for this system in transformation and tumorigenesis. Analysis of combinatorial crosses of gene-targeted mouse lines also has suggested the existence of an as yet unidentified IGF receptor that regulates fetal growth. Similar approaches using transgenic and gene-targeted mouse models have been initiated to study the in vivo functions of the IGF binding proteins. These mouse models provide important tools to test specific functional questions in vivo as well as to study the long-term physiological consequences of chronic gene alterations.

scholarly journals Genetically Engineered Mouse Models of Liver Tumorigenesis Reveal a Wide Histological Spectrum of Neoplastic and Non-Neoplastic Liver Lesions

Cancers ◽  
2020 ◽  
Vol 12 (8) ◽  
pp. 2265
Author(s):  
Katja Steiger ◽  
Nina Gross ◽  
Sebastian A. Widholz ◽  
Roland Rad ◽  
Wilko Weichert ◽  
...  
Keyword(s):  
Mouse Models ◽  
Liver Tumors ◽  
Genetically Engineered ◽  
Liver Lesions ◽  
Genetically Engineered Mouse Models ◽  
Chemically Induced ◽  
Wide Range ◽  
Correct Application ◽  
Huge Variety

Genetically engineered mouse models (GEMM) are an elegant tool to study liver carcinogenesis in vivo. Newly designed mouse models need detailed (histopathological) phenotyping when described for the first time to avoid misinterpretation and misconclusions. Many chemically induced models for hepatocarcinogenesis comprise a huge variety of histologically benign and malignant neoplastic, as well as non-neoplastic, lesions. Such comprehensive categorization data for GEMM are still missing. In this study, 874 microscopically categorized liver lesions from 369 macroscopically detected liver “tumors” from five different GEMM for liver tumorigenesis were included. The histologic spectrum of diagnosis included a wide range of both benign and malignant neoplastic (approx. 82%) and non-neoplastic (approx. 18%) lesions including hyperplasia, reactive bile duct changes or oval cell proliferations with huge variations among the various models and genetic backgrounds. Our study therefore critically demonstrates that models of liver tumorigenesis can harbor a huge variety of histopathologically distinct diagnosis and, depending on the genotype, notable variations are expectable. These findings are extremely important to warrant the correct application of GEMM in liver cancer research and clearly emphasize the role of basic histopathology as still being a crucial tool in modern biomedical research.

scholarly journals Polyphosphate blocks tumour metastasis via anti-angiogenic activity

2007 ◽  
Vol 406 (1) ◽  
pp. 49-55 ◽  
Author(s):  
Kyu Yeon Han ◽  
Bok Sil Hong ◽  
Yae Jin Yoon ◽  
Chang Min Yoon ◽  
Yoon-Keun Kim ◽  
...  
Keyword(s):  
Critical Role ◽  
Wide Distribution ◽  
Mitogen Activated Protein Kinase ◽  
Cell Surface Receptor ◽  
Inorganic Polyphosphate ◽  
Angiogenic Activity ◽  
Tumour Metastasis ◽  
Wide Range

PolyP (inorganic polyphosphate) is a linear polymer of many tens or hundreds of orthophosphate residues found in a wide range of organisms, including bacteria, fungi, insects, plants and vertebrates. Despite its wide distribution in mammalian tissues and plasma, the biological functions of polyP on tumour metastasis and angiogenesis have not been previously examined. In the present study, we have shown that polyP effectively blocked in vivo pulmonary metastasis of B16BL6 cells by suppression of neovascularization, whereas it did not affect proliferation or adhesion to extracellular matrix proteins. PolyP not only inhibited bFGF (basic fibroblast growth factor)-induced proliferation and ERK (extracellular-signal-regulated kinase)/p38 MAPK (mitogen-activated protein kinase) activation of human endothelial cells, but also blocked the binding of bFGF to its cognate cell-surface receptor. Furthermore, polyP inhibited bFGF-induced in vitro and in vivo angiogenesis, suggesting that polyP possesses an anti-angiogenic activity. Since neovascularization is essential for tumour metastasis, our present findings clearly indicate that polyP has an in vivo anti-metastatic activity via its anti-angiogenic activity. Taken together with the fact that angiogenesis occurs under various normal and pathological conditions, our observations suggest that endogenous polyP may play a critical role during embryonic development, wound healing and inflammation, as well as in the progress of pathological diseases such as rheumatoid arthritis and cancer.

A Brief Review of the Modelling of the Time Dependent Mechanical Properties of Tissue Engineering Scaffolds

2010 ◽  
Vol 6 ◽  
pp. 19-33 ◽  
Author(s):  
N.K. Bawolin ◽  
W.J. Zhang ◽  
Xiong Biao Chen
Keyword(s):  
Mechanical Properties ◽  
Tissue Engineering ◽  
Treatment Process ◽  
Effective Properties ◽  
Critical Role ◽  
Time Dependent ◽  
Tissue Engineering Scaffolds ◽  
Wide Range ◽  
Scaffold Degradation

The functionality of tissue scaffolds in vivo plays a critical role in the treatment process. Due to the time dependent nature of the mechanical properties of the constituent phases of the scaffold, a wide range of mechanical property histories may be observed during the treatment process, possibly influencing outcomes. The critical nature of the mechanical properties in load bearing applications indicates a need for the simultaneous modelling of both scaffold degradation and tissue regeneration with time, and the resulting effective properties of the tissue engineering construct. To this end, a review of the literature is conducted to identify the various existing approaches to modelling scaffold degradation, tissue behavior, and the dependency of the two processes on one another.

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