Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder caused by mutations in the gene encoding fibrillin-1 (FBN1), the main component of extracellular microfibrils. In the aortic wall, these microfibrils maintain structural integrity and sustain hemodynamic load. Pathogenic FBN1 variants are thought to structurally and functionally impair fibrillin-1 microfibrils, leading to progressive aortic aneurysm and dissection, the major causes of morbidity and mortality in MFS. However, the molecular mechanisms whereby these genetic variants translate into structural and mechanical defects are far from being understood.Here we explored the morphology and the nanomechanical characteristics of individual aortic fibrillin-1 microfibrils from MFS patients and non-MFS controls by atomic force microscopy. The topographical assessment revealed a preserved overall pattern and periodicity of the microfibrils, but with morphological irregularities in MFS microfibril beads and interbead segments, consistent with presumed structural fragility. Force spectroscopy revealed a reduction of transverse elastic modulus in patients harboring haploinsufficient FBN1 variants. Nanoindentation analysis was indicative of localized deformation, occurring at markedly lower forces in MFS microfibril beads, suggesting diminished load-bearing capacity.These data provide direct nanoscale evidence of structural and mechanical consequences of FBN1 mutations on human aortic tissue. Altered fibrillin-1 microfibril morphology and reduced stiffness in MFS support a pathogenetic mechanism in which compromised microfibrillar integrity weakens the aortic wall, predisposing it to progressive dilation.
Background: Glaucoma is a leading cause of blindness characterized by progressive degeneration of the optic nerve. Elevated intraocular pressure, typically associated with glaucoma, exerts biomechanical strain on the optic nerve head, where mechanosensitive astrocytes contribute to profibrotic extracellular matrix remodeling. Cross-linking enzymes such as transglutaminase 2 play a role in this process. A unique function of transglutaminase 2 is its ability to catalyze the post-translational modification of proteins through the transamidation of serotonin onto glutamine residues ("serotonylation"). The serotonylation of extracellular matrix proteins, such as fibronectin, could plausibly be involved in extracellular matrix remodeling. Here, we investigated whether optic nerve head astrocytes could serotonylate extracellular proteins in response to a biomechanical insult. Results: Primary human optic nerve head astrocytes subjected to 0-12% cyclic stretch for 24 h exhibited increased extracellular levels of transglutaminase 2 and serotonin. Correspondingly, serotonylation of immunoprecipitated fibronectin samples, detected by immunoblotting, was 2.5-fold higher in stretched cells. Serotonylation of specific glutaminyl residues of fibronectin due to the catalytic effect of transglutaminase 2 was also proven though an in vitro experiment using nanoflow liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS). Conclusions: These findings demonstrate that fibronectin can be serotonylated by optic nerve head astrocytes and that this modification is enhanced by biomechanical stress. While transglutaminase 2 is classically associated with extracellular matrix cross-linking, these results suggest that serotonylation may represent an additional and unexplored mechanism by which transglutaminase 2 contributes to extracellular matrix remodeling in the glaucomatous optic nerve head.
Hutchinson–Gilford progeria (HGPS) is a rare genetic disorder characterized by clinical features that mimic accelerated aging. The classical form of progeria is caused by a heterozygous pathogenic variant in in the LMNA gene resulting in the truncated lamin A protein progerin that accumulates in the nuclear envelope and exerts toxic effects on connective tissue and bone growth. Although connective tissue abnormalities are a hallmark of the disease, the extracellular matrix (ECM) produced by HGPS fibroblasts has not been systematically characterized. Here, we combined analysis of four RNA-seq datasets with functional and secretome analyses of primary dermal fibroblasts from an infant with HGPS. Transcriptomic analyses identified enrichment of basement membrane–related pathways and consistent dysregulation of the matrisome gene COL4A1. Patient fibroblasts recapitulated canonical HGPS phenotypes, including progerin accumulation, altered nuclear morphology, reduced proliferation, early senescence, and changes in cell mechanics. Secretome proteomics revealed selective downregulation of core basement membrane components, including collagen IV chains with corresponding transcriptional reductions and aberrant collagen IV deposition in vitro. In 3D skin equivalents, HGPS fibroblasts failed to support proper dermo-epidermal basement membrane assembly, a defect mirrored by reduced collagen IV staining in patient skin tissue. Together, these findings identify impaired basement membrane composition and organization as a previously underappreciated feature of HGPS and suggest that ECM remodeling may contribute to cutaneous pathology and potentially broader disease manifestations.
Tendons are essential connective tissues that transmit mechanical forces from muscles to bones, enabling locomotion and maintaining joint stability. Proper mechanosensation is critical for preserving their extracellular matrix (ECM) integrity. PIEZO1, a mechanosensitive ion channel, has been implicated in regulating tendon architecture under increased mechanical loads, but its role under baseline physiological conditions remains unclear. Here, we generated tendon-targeted Piezo1 conditional knockout (Piezo1 p-t-ko ) mice using a tamoxifen-inducible Scx-CreERT2 system. All in vivo experiments were performed in female mice. PIEZO1 deficiency resulted in significantly reduced Achilles tendon thickness and smaller collagen fibril diameters by transmission electron microscopy. RNA sequencing and qPCR analyses demonstrated downregulation of key ECM-related genes, including Col1a1, Dcn and Fmod, as well as the tendon transcription factors Mkx. Immunostaining further showed reduced signals of MKX, COL1A1 and DCN. Adjacent muscle morphology and transcriptomes were unaltered, supporting the tendon-selective nature of the observed phenotype. Together, these findings suggest that PIEZO1 contributes to collagen fibril architecture and tendon-associated transcriptional programs in female mouse Achilles tendons under baseline physiological conditions.
Pancreatic ductal adenocarcinoma (PDAC) is defined by a dense, collagen-rich stroma that limits therapeutic efficacy and drives metastasis. While collagen reorganization has been studied in resected PDAC tumors, its structure and organization in metastatic PDAC and response to chemotherapy remains poorly characterized. We evaluated collagen structure by Second Harmonic Generation (SHG) imaging of formalin fixed paraffin embedded tissue from pancreatic tumors and liver metastases of 20 PDAC patients that were untreated or treated with FOLFOX or FOLFIRINOX (FOL). Collagen fiber width, length, alignment, and density were assessed at tumor cores, tumor boundaries, and in the adjacent tissue. We document for the first time fundamental differences in these collagen structure features between normal pancreas and normal liver. Surprisingly, the structure of collagen in primary pancreatic cancer was similar to normal pancreas. Liver metastases showed alterations in collagen structure, revealing a transition from a liver-like collagen phenotype (thinner, shorter less dense collagen fibers) at the tumor border to a tumor-core phenotype with thicker, longer, less aligned, and denser collagen fibers. FOL treatment increased length, width, and density at both primary tumors and liver metastases. Overall, these findings reveal organ specific collagen remodeling in PDAC and suggest that chemotherapy with FOL modulates extracellular matrix (ECM) remodeling. These findings expand the scope of PDAC collagen structural characterization and highlight the complexity of matrix targeting strategies in metastatic disease.
