scholarly journals A Novel Bioengineered Functional Motor Unit Platform to Study Neuromuscular Interaction

2020 ◽  
Vol 9 (10) ◽  
pp. 3238
Author(s):  
Jasdeep Saini ◽  
Alessandro Faroni ◽  
Adam J. Reid ◽  
Kamel Mamchaoui ◽  
Vincent Mouly ◽  
...  
Keyword(s):  
Motor Unit ◽  
Motor Neurons ◽  
Acetylcholine Receptors ◽  
In Vivo Studies ◽  
Cholinergic Antagonists ◽  
Myoblast Cell ◽  
Nerve Muscle ◽  
And Function

Background: In many neurodegenerative and muscular disorders, and loss of innervation in sarcopenia, improper reinnervation of muscle and dysfunction of the motor unit (MU) are key pathogenic features. In vivo studies of MUs are constrained due to difficulties isolating and extracting functional MUs, so there is a need for a simplified and reproducible system of engineered in vitro MUs. Objective: to develop and characterise a functional MU model in vitro, permitting the analysis of MU development and function. Methods: an immortalised human myoblast cell line was co-cultured with rat embryo spinal cord explants in a serum-free/growth fact media. MUs developed and the morphology of their components (neuromuscular junction (NMJ), myotubes and motor neurons) were characterised using immunocytochemistry, phase contrast and confocal microscopy. The function of the MU was evaluated through live observations and videography of spontaneous myotube contractions after challenge with cholinergic antagonists and glutamatergic agonists. Results: blocking acetylcholine receptors with α-bungarotoxin resulted in complete, cessation of myotube contractions, which was reversible with tubocurarine. Furthermore, myotube activity was significantly higher with the application of L-glutamic acid. All these observations indicate the formed MU are functional. Conclusion: a functional nerve-muscle co-culture model was established that has potential for drug screening and pathophysiological studies of neuromuscular interactions.

scholarly journals Alcohol-and-HIV-Induced Lysosomal Dysfunction Regulates Extracellular Vesicles Secretion in Vitro and in Liver-Humanized Mice

Biology ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 29
Author(s):  
Raghubendra Singh Dagur ◽  
Moses New-Aaron ◽  
Murali Ganesan ◽  
Weimin Wang ◽  
Svetlana Romanova ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Extracellular Vesicles ◽  
Treated Group ◽  
Human Hepatocytes ◽  
In Vivo Studies ◽  
People Living With Hiv ◽  
Humanized Mouse Model ◽  
And Function

Background: Alcohol abuse is common in people living with HIV-1 and dramaticallyenhances the severity of HIV-induced liver damage by inducing oxidative stress and lysosomaldysfunction in the liver cells. We hypothesize that the increased release of extracellular vesicles(EVs) in hepatocytes and liver humanized mouse model is linked to lysosome dysfunction. Methods:The study was performed on primary human hepatocytes and human hepatoma RLWXP-GFP (Huh7.5 cells stably transfected with CYP2E1 and XPack-GFP) cells and validated on ethanol-fed liverhumanizedfumarylacetoacetate hydrolase (Fah)-/-, Rag2-/-, common cytokine receptor gamma chainknockout (FRG-KO) mice. Cells and mice were infected with HIV-1ADA virus. Results: We observedan increase in the secretion of EVs associated with a decrease in lysosomal activity and expressionof lysosomal-associated membrane protein 1. Next-generation RNA sequencing of primary humanhepatocytes revealed 63 differentially expressed genes, with 13 downregulated and 50 upregulatedgenes in the alcohol–HIV-treated group. Upstream regulator analysis of differentially expressedgenes through Ingenuity Pathway Analysis identified transcriptional regulators affecting downstreamgenes associated with increased oxidative stress, lysosomal associated disease, and function andEVs biogenesis. Our in vitro findings were corroborated by in vivo studies on human hepatocytetransplantedhumanized mice, indicating that intensive EVs’ generation by human hepatocytes andtheir secretion to serum was associated with increased oxidative stress and reduction in lysosomalactivities triggered by HIV infection and ethanol diet. Conclusion: HIV-and-ethanol-metabolisminducedEVs release is tightly controlled by lysosome status in hepatocytes and participates in thedevelopment of double-insult-induced liver injury.

scholarly journals Neuronal over-expression of Oxr1 is protective against ALS-associated mutant TDP-43 mislocalisation in motor neurons and neuromuscular defects in vivo

2019 ◽  
Vol 28 (21) ◽  
pp. 3584-3599 ◽  
Author(s):  
Matthew G Williamson ◽  
Mattéa J Finelli ◽  
James N Sleigh ◽  
Amy Reddington ◽  
David Gordon ◽  
...  
Keyword(s):  
Motor Neurons ◽  
Neurodegenerative Disorder ◽  
Late Onset ◽  
Gene Encoding ◽  
Over Expression ◽  
Neuromuscular Abnormalities ◽  
And Function ◽  
Lateral Sclerosis

Abstract A common pathological hallmark of amyotrophic lateral sclerosis (ALS) and the related neurodegenerative disorder frontotemporal dementia, is the cellular mislocalization of transactive response DNA-binding protein 43 kDa (TDP-43). Additionally, multiple mutations in the TARDBP gene (encoding TDP-43) are associated with familial forms of ALS. While the exact role for TDP-43 in the onset and progression of ALS remains unclear, the identification of factors that can prevent aberrant TDP-43 localization and function could be clinically beneficial. Previously, we discovered that the oxidation resistance 1 (Oxr1) protein could alleviate cellular mislocalization phenotypes associated with TDP-43 mutations, and that over-expression of Oxr1 was able to delay neuromuscular abnormalities in the hSOD1G93A ALS mouse model. Here, to determine whether Oxr1 can protect against TDP-43-associated phenotypes in vitro and in vivo, we used the same genetic approach in a newly described transgenic mouse expressing the human TDP-43 locus harbouring an ALS disease mutation (TDP-43M337V). We show in primary motor neurons from TDP-43M337V mice that genetically-driven Oxr1 over-expression significantly alleviates cytoplasmic mislocalization of mutant TDP-43. We also further quantified newly-identified, late-onset neuromuscular phenotypes of this mutant line, and demonstrate that neuronal Oxr1 over-expression causes a significant reduction in muscle denervation and neuromuscular junction degeneration in homozygous mutants in parallel with improved motor function and a reduction in neuroinflammation. Together these data support the application of Oxr1 as a viable and safe modifier of TDP-43-associated ALS phenotypes.

scholarly journals Structure-function relationships in mitochondrial transcriptional condensates

2022 ◽  
Author(s):  
Marina Feric ◽  
Azadeh Sarfallah ◽  
Furqan Dar ◽  
Dmitry Temiakov ◽  
Rohit V Pappu ◽  
...  
Keyword(s):  
Structure Function ◽  
Computational Modelling ◽  
In Vivo Studies ◽  
Bulk Solution ◽  
The Core ◽  
Functional Consequences ◽  
Cellular Machinery ◽  
And Function

Phase separation organizes many membraneless structures in cells. The functional consequences of concentrating cellular machinery into biomolecular condensates, however, is largely unclear. Here, we use in vitro reconstitutions, in vivo studies, and computational modelling to uncover structure-function relationships of mitochondrial (mt-) transcriptional condensates. In vitro, we find that the core mt-transcription machinery — consisting of POLRMT, TFAM, TFB2M, and DNA — forms viscoelastic, multi-phasic condensates. Strikingly, the rates of condensate-mediated transcription are considerably lower than equivalent reactions in bulk solution. Dampened rates are associated with reduced diffusivities of components that become kinetically arrested in non-equilibrium, vesicular condensates. Perturbation of mt-components in vivo and computational simulations recapitulate the transcription-dependent reorganizations observed in vitro. Our findings demonstrate close, bidirectional interdependence between structure and function of transcriptional condensates.

Structure and function of anhydride-modified forms of human insulin: In silico, in vitro and in vivo studies

2017 ◽  
Vol 96 ◽  
pp. 342-350 ◽  
Author(s):  
Maryam Chinisaz ◽  
Azadeh Ebrahim-Habibi ◽  
Ahmad-Reza Dehpour ◽  
Parichehreh Yaghmaei ◽  
Kazem Parivar ◽  
...  
Keyword(s):  
Human Insulin ◽  
In Silico ◽  
Structure And Function ◽  
In Vivo Studies ◽  
And Function ◽  
Modified Forms

scholarly journals Neurotransmitter regulation of neural development: acetylcholine and nicotinic receptors

2002 ◽  
Vol 74 (3) ◽  
pp. 453-461 ◽  
Author(s):  
ANDRÉA S. TORRÃO ◽  
LUIZ R.G. BRITTO
Keyword(s):  
Nicotinic Acetylcholine Receptors ◽  
Neural Development ◽  
Acetylcholine Receptors ◽  
Nicotinic Receptors ◽  
In Vivo Studies ◽  
Developmental Processes ◽  
Additional Function ◽  
Vertebrate Brain

Several neurotransmitter systems have been related to developmental processes during the past decade. In this review, we discuss the evidence that the nicotinic acetylcholine receptors could have an additional function during development that may be unrelated to their role in cholinergic neurotransmission in the vertebrate brain. Both temporal expression data and in vitro and in vivo studies with nicotinic agonists and antagonists have provided direct support for a role of nicotinic receptors in neural developmental processes such as neurite outgrowth and differentiation. A similar picture has emerged for other neurotransmitter and receptor systems as well, which generates a new view of neural processes during both development and mature life.

scholarly journals Experimental Renovascular Disease Induces Endothelial Cell Mitochondrial Damage and Impairs Endothelium-Dependent Relaxation of Renal Artery Segments

2020 ◽  
Vol 33 (8) ◽  
pp. 765-774
Author(s):  
Arash Aghajani Nargesi ◽  
Xiang-Yang Zhu ◽  
Ishran M Saadiq ◽  
Kyra L Jordan ◽  
Amir Lerman ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Renal Artery ◽  
Tube Formation ◽  
Mitochondrial Damage ◽  
In Vivo Studies ◽  
Mitochondrial Morphology ◽  
Renovascular Disease ◽  
And Function

Abstract BACKGROUND Mitochondria modulate endothelial cell (EC) function, but may be damaged during renal disease. We hypothesized that the ischemic and metabolic constituents of swine renovascular disease (RVD) induce mitochondrial damage and impair the function of renal artery ECs. METHODS Pigs were studied after 16 weeks of metabolic syndrome (MetS), renal artery stenosis (RAS), or MetS + RAS, and Lean pigs served as control (n = 6 each). Mitochondrial morphology, homeostasis, and function were measured in isolated primary stenotic-kidney artery ECs. EC functions were assessed in vitro, whereas vasoreactivity of renal artery segments was characterized in organ baths. RESULTS Lean + RAS and MetS + RAS ECs showed increased mitochondrial area and decreased matrix density. Mitochondrial biogenesis was impaired in MetS and MetS + RAS compared with their respective controls. Mitochondrial membrane potential similarly decreased in MetS, Lean + RAS, and MetS + RAS groups, whereas production of reactive oxygen species increased in MetS vs. Lean, but further increased in both RAS groups. EC tube formation was impaired in MetS, RAS, and MetS + RAS vs. Lean, but EC proliferation and endothelial-dependent relaxation of renal artery segments were blunted in MetS vs. Lean, but further attenuated in Lean + RAS and MetS + RAS. CONCLUSIONS MetS and RAS damage mitochondria in pig renal artery ECs, which may impair EC function. Coexisting MetS and RAS did not aggravate EC mitochondrial damage in the short time of our in vivo studies, suggesting that mitochondrial injury is associated with impaired renal artery EC function.

scholarly journals ZFX Promotes Proliferation and Metastasis of Pancreatic Cancer Cells via the MAPK Pathway

2018 ◽  
Vol 48 (1) ◽  
pp. 274-284 ◽  
Author(s):  
Xiaoling Song ◽  
Minghui Zhu ◽  
Fahong Zhang ◽  
Fei Zhang ◽  
Yijian Zhang ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Mapk Pathway ◽  
In Vivo Studies ◽  
Human Pancreatic Cancer ◽  
Cell Functions ◽  
Pancreatic Cancer Cells ◽  
Pathway Activation ◽  
And Function

Background/Aims: The role of ZFX in tumourigenesis is unclear. We aimed to study ZFX expression, regulation, and function and the clinical implications of this protein in human pancreatic cancer (PCa). Methods: One hundred and twenty patients with histologically confirmed PCa who underwent surgery were recruited for this study. Tumour samples and PCa cell lines were used to examine ZFX. Various cell functions related to tumourigenesis were assessed. In vivo mouse tumour xenografts were used to confirm the in vitro results. Results: Patients with ZFX-positive tumours had worse overall survival than patients with ZFX-negative tumours. The depletion of ZFX using lentiviral shRNAs significantly inhibited cell proliferation by inducing cell cycle arrest in G0/G1 phase and resulted in increased cell apoptosis and invasive repression. In vivo studies confirmed that ZFX promoted tumour growth. Mechanistically, MAPK pathway activation was involved in the oncogenic functions of ZFX. Conclusions: ZFX acts as a putative oncogene in PCa and could be a novel therapeutic target for this disease.

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