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BioRxiv : the Preprint Server For... May 2024Duchenne muscular dystrophy (DMD) is a lethal muscle disease caused by the absence of the protein dystrophin. Dystrophin is hypothesized to work as a molecular shock...
Duchenne muscular dystrophy (DMD) is a lethal muscle disease caused by the absence of the protein dystrophin. Dystrophin is hypothesized to work as a molecular shock absorber that limits myofiber membrane damage when undergoing reversible unfolding upon muscle stretching and contraction. Utrophin is a dystrophin homologue that is under investigation as a protein replacement therapy for DMD. However, it remains uncertain whether utrophin can mechanically substitute for dystrophin. Here, we compared the mechanical properties of homologous utrophin and dystrophin fragments encoding the N terminus through spectrin repeat 3 (UtrN-R3, DysN-R3) using two operational modes of atomic force microscopy (AFM), constant speed and constant force. Our comprehensive data, including the statistics of force magnitude at which the folded domains unfold in constant speed mode and the time of unfolding statistics in constant force mode, show consistent results. We recover parameters of the energy landscape of the domains and conducted Monte Carlo simulations which corroborate the conclusions drawn from experimental data. Our results confirm that UtrN-R3 expressed in bacteria exhibits significantly lower mechanical stiffness compared to insect UtrN-R3, while the mechanical stiffness of the homologous region of dystrophin (DysN-R3) is intermediate between bacterial and insect UtrN-R3, showing greater similarity to bacterial UtrN-R3.
PubMed: 38826288
DOI: 10.1101/2024.05.18.593686 -
International Heart Journal 2024When stimulated, vascular smooth muscle cells (VSMCs) change from a differentiated to a dedifferentiated phenotype. Dedifferentiated VSMCs have a key activity in...
When stimulated, vascular smooth muscle cells (VSMCs) change from a differentiated to a dedifferentiated phenotype. Dedifferentiated VSMCs have a key activity in cardiovascular diseases such as in-stent restenosis. MicroRNAs (miRNAs) have crucial functions in conversion of differentiated VSMCs to a dedifferentiated phenotype. We investigated the activity of miR-411-5p in the proliferation, migration, and phenotype switch of rat VSMCs.Based on a microRNA array assay, miR-411-5p expression was found to be significantly increased in cultured VSMCs stimulated by platelet-derived growth factor-BB (PDGF-BB). A CCK-8 assay, transwell assay, and scratch test were performed to measure the effect of miR-411-5p on the proliferation and migration of PDGF-BB-treated VSMCs. MiR-411-5p promoted expression of dedifferentiated phenotype markers such as osteopontin and tropomyosin 4 in PDGF-BB-treated VSMCs. Using mimics and inhibitors, we identified the target of miR-411-5p in PDGF-BB-treated VSMCs and found that calmodulin-regulated spectrin-associated protein-1 (CAMSAP1) was involved in the phenotypic switch mediated by PDGF-BB.By inhibiting expression of CAMSAP1, miR-411-5p enhanced the proliferation, migration, and phenotype switch of VSMCs.Blockade of miR-411-5p interaction with CAMSAP1 is a promising approach to treat in-stent restenosis.
Topics: Animals; MicroRNAs; Muscle, Smooth, Vascular; Rats; Cell Proliferation; Becaplermin; Phenotype; Cell Movement; Cells, Cultured; Myocytes, Smooth Muscle; Rats, Sprague-Dawley; Male; Osteopontin
PubMed: 38825498
DOI: 10.1536/ihj.23-590 -
High Altitude Medicine & Biology May 2024Wang L, Fu G, Han R, Fan P, Yang J, Gong K, Zhao Z, Zhang C, Sun K, Shao GMALAT1 and NEAT1 Are Neuroprotective during Hypoxic Preconditioning in the Mouse Hippocampus...
Wang L, Fu G, Han R, Fan P, Yang J, Gong K, Zhao Z, Zhang C, Sun K, Shao GMALAT1 and NEAT1 Are Neuroprotective during Hypoxic Preconditioning in the Mouse Hippocampus Possibly by Regulation of NR2B 00:000-000, 2024. The regulation of noncoding ribonucleic acid (ncRNA) has been shown to be involved in cellular and molecular responses to hypoxic preconditioning (HPC), a situation created by the induction of sublethal hypoxia in the brain. The ncRNAs metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) and nuclear paraspeckle assembly transcript 1 (NEAT1) are abundantly expressed in the brain, where they regulate the expression of various genes in nerve cells. However, the exact roles of MALAT1 and NEAT1 in HPC are not fully understood. A mouse model of acute repeated hypoxia was used as a model of HPC, and MALAT1 and NEAT1 levels in the hippocampus were measured using real-time polymerase chain reaction (PCR). The mRNA and protein levels of -methyl-d-aspartate receptor subunit 2 B (NR2B) in the mouse hippocampus were measured using real-time PCR and western blotting, respectively. HT22 cells knocked-down for MALAT1 and NEAT1 were used for testing. Expression of NR2B, which is involved in nerve cell injury under ischemic and hypoxic conditions, was also evaluated. The levels of spectrin and cleaved caspase-3 in MALAT1 and NEAT1 knockdown HT22 cells under oxygen glucose deprivation/reperfusion (OGD/R) were determined by western blotting. HPC increased the expression of MALAT1 and NEAT1 and decreased the expression of NR2B mRNA in the mouse hippocampus ( < 0.05). Knockdown of MALAT1 and NEAT1 increased both NR2B mRNA and protein levels nearly twofold and caused damage under OGD/R conditions in HT22 cells ( < 0.05). MALAT1 and NEAT1 exert neuroprotective effects by influencing the expression of NR2B.
PubMed: 38808452
DOI: 10.1089/ham.2023.0135 -
Molecular Genetics and Genomics : MGG May 2024Hereditary spherocytosis (HS) is one of the most common causes of hereditary hemolytic anemia. The current diagnostic guidelines for HS are mainly based on a combination...
Hereditary spherocytosis (HS) is one of the most common causes of hereditary hemolytic anemia. The current diagnostic guidelines for HS are mainly based on a combination of physical examination and laboratory investigation. However, some patients present with complicated clinical manifestations that cannot be explained by routine diagnostic protocols. Here, we report a rare HS case of mild anemia with extremely high indirect bilirubin levels and high expression of fetal hemoglobin. Using whole exome sequencing analysis, this patient was identified as a heterozygous carrier of a de novo SPTB nonsense mutation (c.605G > A; p.W202*) and a compound heterozygous carrier of known UGT1A1 and KLF1 mutations. This genetic analysis based on the interpretation of the patient's genomic data not only achieved precise diagnosis by an excellent explanation of the complicated phenotype but also provided valuable suggestions for subsequent appropriate approaches for treatment, surveillance and prophylaxis.
Topics: Humans; Spherocytosis, Hereditary; Phenotype; Kruppel-Like Transcription Factors; Spectrin; Glucuronosyltransferase; Exome Sequencing; Codon, Nonsense; Male; Heterozygote; Female
PubMed: 38787432
DOI: 10.1007/s00438-024-02150-5 -
Angiogenesis May 2024The presence of atherosclerotic plaque vessels is a critical factor in plaque destabilization. This may be attributable to the leaky phenotype of these microvessels,...
The presence of atherosclerotic plaque vessels is a critical factor in plaque destabilization. This may be attributable to the leaky phenotype of these microvessels, although direct proof for this notion is lacking. In this study, we investigated molecular and cellular patterns of stable and hemorrhaged human plaque to identify novel drivers of intraplaque vessel dysfunction. From transcriptome data of a human atherosclerotic lesion cohort, we reconstructed a co-expression network, identifying a gene module strongly and selectively correlated with both plaque microvascular density and inflammation. Spectrin Beta Non-Erythrocytic 1 (sptbn1) was identified as one of the central hubs of this module (along with zeb1 and dock1) and was selected for further study based on its predominant endothelial expression. Silencing of sptbn1 enhanced leukocyte transmigration and vascular permeability in vitro, characterized by an increased number of focal adhesions and reduced junctional VE-cadherin. In vivo, sptbn1 knockdown in zebrafish impaired the development of the caudal vein plexus. Mechanistically, increased substrate stiffness was associated with sptbn1 downregulation in endothelial cells in vitro and in human vessels. Plaque SPTBN1 mRNA and protein expression were found to correlate with an enhanced presence of intraplaque hemorrhage and future cardiovascular disease (CVD) events during follow-up. In conclusion, we identify SPTBN1 as a central hub gene in a gene program correlating with plaque vascularisation. SPTBN1 was regulated by substrate stiffness in vitro while silencing blocked vascular development in vivo, and compromised barrier function in vitro. Together, SPTBN1 is identified as a new potential regulator of the leaky phenotype of atherosclerotic plaque microvessels.
PubMed: 38780883
DOI: 10.1007/s10456-024-09921-z -
BioRxiv : the Preprint Server For... May 2024The plasma membrane and the underlying skeleton form a protective barrier for eukaryotic cells. The molecules forming this complex composite material constantly...
The plasma membrane and the underlying skeleton form a protective barrier for eukaryotic cells. The molecules forming this complex composite material constantly rearrange under mechanical stress to confer this protective capacity. One of those molecules, spectrin, is ubiquitous in the membrane skeleton and primarily located proximal to the inner leaflet of the plasma membrane and engages in protein-lipid interactions via a set of membrane-anchoring domains. Spectrin is linked by short actin filaments and its conformation varies in different types of cells. In this work, we developed a generalized network model for the membrane skeleton integrated with myosin contractility and membrane mechanics to investigate the response of the spectrin meshwork to mechanical loading. We observed that the force generated by membrane bending is important to maintain a smooth skeletal structure. This suggests that the membrane is not just supported by the skeleton, but has an active contribution to the stability of the cell structure. We found that spectrin and myosin turnover are necessary for the transition between stress and rest states in the skeleton. Our model reveals that the actin-spectrin meshwork dynamics are balanced by the membrane forces with area constraint and volume restriction promoting the stability of the membrane skeleton. Furthermore, we showed that cell attachment to the substrate promotes shape stabilization. Thus, our proposed model gives insight into the shared mechanisms of the membrane skeleton associated with myosin and membrane that can be tested in different types of cells.
PubMed: 38746295
DOI: 10.1101/2024.04.29.591779 -
JCI Insight May 2024Childhood-onset essential hypertension (COEH) is an uncommon form of hypertension that manifests in childhood or adolescence and, in the United States,...
Childhood-onset essential hypertension (COEH) is an uncommon form of hypertension that manifests in childhood or adolescence and, in the United States, disproportionately affects children of African ancestry. The etiology of COEH is unknown, but its childhood onset, low prevalence, high heritability, and skewed ancestral demography suggest the potential to identify rare genetic variation segregating in a Mendelian manner among affected individuals and thereby implicate genes important to disease pathogenesis. However, no COEH genes have been reported to date. Here, we identify recessive segregation of rare and putatively damaging missense variation in the spectrin domain of spectrin repeat containing nuclear envelope protein 1 (SYNE1), a cardiovascular candidate gene, in 3 of 16 families with early-onset COEH without an antecedent family history. By leveraging exome sequence data from an additional 48 COEH families, 1,700 in-house trios, and publicly available data sets, we demonstrate that compound heterozygous SYNE1 variation in these COEH individuals occurred more often than expected by chance and that this class of biallelic rare variation was significantly enriched among individuals of African genetic ancestry. Using in vitro shRNA knockdown of SYNE1, we show that reduced SYNE1 expression resulted in a substantial decrease in the elasticity of smooth muscle vascular cells that could be rescued by pharmacological inhibition of the downstream RhoA/Rho-associated protein kinase pathway. These results provide insights into the molecular genetics and underlying pathophysiology of COEH and suggest a role for precision therapeutics in the future.
Topics: Adolescent; Child; Female; Humans; Male; Age of Onset; Cytoskeletal Proteins; Essential Hypertension; Exome; Exome Sequencing; Genetic Predisposition to Disease; Mutation, Missense; Nerve Tissue Proteins; Nuclear Proteins; Pedigree; rhoA GTP-Binding Protein; United States; Infant, Newborn; Infant; Child, Preschool; Young Adult
PubMed: 38716726
DOI: 10.1172/jci.insight.172152 -
Frontiers in Physiology 2024The integrity of the erythrocyte membrane cytoskeletal network controls the morphology, specific surface area, material exchange, and state of erythrocytes in the blood...
The integrity of the erythrocyte membrane cytoskeletal network controls the morphology, specific surface area, material exchange, and state of erythrocytes in the blood circulation. The antioxidant properties of resveratrol have been reported, but studies on the effect of resveratrol on the hypoxia-induced mechanical properties of erythrocytes are rare. In this study, the effects of different concentrations of resveratrol on the protection of red blood cell mor-phology and changes in intracellular redox levels were examined to select an appropriate concentration for further study. The Young's modulus and surface roughness of the red blood cells and blood viscosity were measured via atomic force microsco-py and a blood rheometer, respectively. Flow cytometry, free hemoglobin levels, and membrane lipid peroxidation levels were used to characterize cell membrane damage in the presence and absence of resveratrol after hypoxia. The effects of oxida-tive stress on the erythrocyte membrane proteins band 3 and spectrin were further investigated by immunofluorescent label-ing and Western blotting. Resveratrol changed the surface roughness and Young's modulus of the erythrocyte mem-brane, reduced the rate of eryptosis in erythrocytes after hypoxia, and stabilized the intracellular redox level. Further data showed that resveratrol protected the erythrocyte membrane proteins band 3 and spectrin. Moreover, resistance to band 3 pro-tein tyrosine phosphorylation and sulfhydryl oxidation can protect the stability of the erythrocyte membrane skeleton net-work, thereby protecting erythrocyte deformability under hypoxia. The results of the present study may provide new insights into the roles of resveratrol in the prevention of hypoxia and as an antioxidant.
PubMed: 38706947
DOI: 10.3389/fphys.2024.1399154 -
MicroPublication Biology 2024The dynamics of microtubule-mediated protrusions, termed Interplanar Amida Network (IPAN) in pupal wing, involve cell shape changes. The molecular mechanisms underlying...
The dynamics of microtubule-mediated protrusions, termed Interplanar Amida Network (IPAN) in pupal wing, involve cell shape changes. The molecular mechanisms underlying these processes are yet to be fully understood. This study delineates the stages of cell shape alterations during the disassembly of microtubule protrusions and underscores the pivotal role of α-Spectrin in driving these changes by regulating both the microtubule and actomyosin networks. Our findings also demonstrate that α-Spectrin is required for the apical relaxation of wing epithelia during protrusion disassembly, indicating its substantial contribution to the robustness of 3D tissue morphogenesis.
PubMed: 38690064
DOI: 10.17912/micropub.biology.001169 -
Scientific Reports Apr 2024SPTBN2 is a protein-coding gene that is closely related to the development of malignant tumors. However, its prognostic value and biological function in pan-cancer,...
SPTBN2 is a protein-coding gene that is closely related to the development of malignant tumors. However, its prognostic value and biological function in pan-cancer, especially pancreatic cancer (PAAD), have not been reported. In the present study, a novel exploration of the value and potential mechanism of SPTBN2 in PAAD was conducted using multi-omics in the background of pan-cancer. Via various database analysis, up-regulated expression of SPTBN2 was detected in most of the tumor tissues examined. Overexpression of SPTBN2 in PAAD and kidney renal clear cell cancer patients potentially affected overall survival, disease-specific survival, and progression-free interval. In PAAD, SPTBN2 can be used as an independent factor affecting prognosis. Mutations and amplification of SPTBN2 were detected, with abnormal methylation of SPTBN2 affecting its expression and the survival outcome of PAAD patients. Immunoassay results demonstrate that SPTBN2 was a potential biomarker for predicting therapeutic response in PAAD, and may influence the immunotherapy efficacy of PAAD by regulating levels of CD8 + T cells and neutrophil infiltration. Results from an enrichment analysis indicated that SPTBN2 may regulate the development of PAAD via immune pathways. Thus, SPTBN2 is a potential prognostic biomarker and immunotherapy target based on its crucial role in the development of PAAD.
Topics: Humans; Pancreatic Neoplasms; Biomarkers, Tumor; Prognosis; Gene Expression Regulation, Neoplastic; Spectrin; Male; Female; Mutation; DNA Methylation; Multiomics
PubMed: 38684762
DOI: 10.1038/s41598-024-60780-6