Changes In Extracellular Matrices Research Articles

PGE receptor EP4 signaling increased hyaluronan‐vesican complex in abdominal aortic aneurysm

Chronic inflammation and dysregulation of extracellular matrices are hallmarks of abdominal aortic aneurysm (AAA) pathophysiology. We have demonstrated that PGE receptor EP4 signaling in vascular smooth muscle cells (VSMCs) has a pivotal role in AAA formation via immune infiltrates and impairment of elastic fiber repair. However, EP4 signaling‐mediated changes in extracellular matrices have not been fully understood, and expressions of the chondroitin sulfate proteoglycan versican are controversial. Versican has multiple binding partners (e.g., hyaluronan and fibrillin‐1) and modulates VSMC phenotypes. We, therefore, aimed to clarify an expression change in versican in AAA.We administrated angiotensin‐II (AngII) to VSMC‐specific EP4 overexpression mice (EP4‐Tg) to induce AAA. The expression level of versican V0/V1 was gradually increased in the tunica media of EP4‐Tg mice 4 days after AngII administration, but not in Non‐Tg mice. This increase in versican V0/V1 became prominent 14 days after AngII administration in EP4‐Tg mice. Protein expression of versican V0/V1 was increased in the tunica media of human AAA tissues. EP4 stimulation in VSMCs derived from EP4‐Tg mice increased levels of versican mRNAs (20.0±1.4‐fold, n=5, p<0.01) and versican V0/V1 proteins (4.1±0.7‐fold, n=5, p<0.01), while administration of EP1/3 or EP2 agonist did not. The EP4‐mediated increase in versican mRNAs was completely suppressed by Actinomycin D. Versican V0/V1 are cleaved by a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS‐1) and ADAMTS‐5. The expression levels of ADAMTS‐1, ADAMTS‐5, and cleaved versican protein expressions were not changed in EP4‐Tg VSMCs. The expression level of Hyaluronan synthase1 (Has1) mRNA was also increased in EP4‐Tg VSMCs (66.9±10.1‐fold, n=5, p<0.01). Western blotting revealed that versican V0/V1 proteins of approximately 200‐400 kDa in size in the supernatants of EP4‐Tg VSMCs stimulated with an EP4 agonist were increased by hyaluronan digestion with hyaluronidase (6.3±1.5‐fold, n=6, p<0.01). Transmission electron microscopic analysis demonstrated that elastin contractile units between elastic laminae and VSMCs were disrupted in EP4‐Tg mice administrated with 10 days of AngII, but not in Non‐Tg mice.These data suggested that EP4 signaling in VSMCs increased hyaluronan‐versican V0/V1 complex. Versican V0/V1 proteins were increased in the early stage of a mouse AAA model. It has been suggested that versican has inhibitory roles in elastic fiber formation. EP4‐mediated hyaluronan‐versican V0/V1 complex may exacerbate AAA via disruption of elastin contractile units.

Multi-level consistent changes of the ECM pathway identified in a typical keratoconus twin\u2019s family by multi-omics analysis

BackgroundKeratoconus (KC) is a common, degenerative disorder of the cornea, and genetic factors play a key role in its development. However, the genetic etiology of KC is still unclear. This study used the family of twins as material, using, for the first time, multi-omics analysis, to systematically display the changes in KC candidate factors in patients at the DNA, RNA, and protein levels. These can evaluate candidate pathogenic factors in depth and lock onto pathogenic targets.ResultsThe twins in this study presented classic phenotypes, clear diagnoses, complete case data, and clinical samples, which are excellent materials for genetically studying KC. Whole-exome sequencing was conducted on both the twins and their parents. Transcriptome sequencing was conducted on proband’s and health individual’s primary human corneal fibroblast cells. Quantitative Real-time PCR and western blot were used to validate the differential gene expressions between the proband and controls. By integrating genomics, transcriptome, and protein level data, multiple consecutive events of KC were systematically analyzed to help better understand the molecular mechanism and genetic basis of KC. The results showed that the accumulation of rare, micro-effect risk variants was the pathogenic factor in this Chinese KC family. Consistent changes in extracellular matrices (ECMs) at the DNA and RNA levels suggested that ECM related changes play a key role in KC pathogenesis. The major gene variants (WNT16, CD248, COL6A2, COL4A3 and ADAMTS3) may affect the expression of related collagens or ECM proteins, thus reducing the amount of ECM in corneas and resulting in KC.ConclusionsThis study, the first to explore the genetic etiology of KC via multi-omics analysis under the polygenetic model, has provided new insights into the genetic mechanisms underlying KC and an effective strategy for studying KC pathogenesis in the future.

Bioimpedance Analysis of Epithelial Monolayers after Exposure to Nanosecond Pulsed Electric Fields.

Exposures to pulsed electric fields (PEFs) are known to affect cell membranes and consequently also cell-cell interactions as well as associated characteristics. Bioimpedance analysis offers direct and non-invasive insights into structural and functional changes of cell membranes and extracellular matrices through a rigorous evaluation of electrical parameters. Accordingly, the multi-frequency impedance of confluent monolayers of rat liver epithelial WB-F344 cells was monitored in situ before and after exposure to nanosecond PEFs (nsPEFs). The results were fitted by two Cole models in series to obtain the Cole parameters for the monolayer. For an interpretation of the results, dielectric parameters, were correlated with changes of the TJ protein zonula occludens (ZO-1) and the paracellular permeability of the monolayer Cole parameters in general change as a function of pulse number and time. The findings demonstrate that impedance analysis is an effective method to monitor changes of TJs cell-cell contacts and paracellular permeability and relate them to exposure parameters.

Periodontal Biological Events Associated with Orthodontic Tooth Movement: The Biomechanics of the Cytoskeleton and the Extracellular Matrix.

The mechanical stimuli generated by orthodontic forces cause deformation of extracellular matrices and cells, vascular changes, inflammation, and the release of active biological agents generating a complex multifactorial sequence of biological events culminating in bone remodelling enabling orthodontic tooth movement. Orthodontic forces on the teeth generate stresses in periodontal tissues according to a number of variables including the type (continuous, interrupted, or intermittent), magnitude, direction, and frequency of the applied load. Whether the strain is compressive or tensile determines whether bone deposition or bone resorption will occur. The mechanically induced strains mediate structural changes in extracellular matrices and in cells, consequently affecting cellular gene expression and function. In the extracellular matrix, mechanosensing molecules integrated into the structure of various proteins can be activated upon load-induced protein unfolding. These specialized molecules have the capacity to sense and then to convert microenvironmental biomechanical stimuli into intracellular biochemical signals that interact to generate a coordinated tissue response. It is also possible that the applied force may directly cause nuclear deformation with configurational changes in chromatin, thus influencing gene expression. In this review article we summarize the current general concepts of mechanotransduction influencing the remodelling of periodontal tissues thus enabling tooth movement in response to applied orthodontic loads.

Inflammatory responses and morphological changes of radiofrequency‐induced rat sciatic nerve fibres

Recently, the pulsed radiofrequency (PRF, pulsed fashion) procedure was introduced for neuropathic pain management. Since PRF-induced lesions do not need high temperature compared with conventional continuous RF (CRF)-induced lesions, there is no significant loss of sensory fibres after application. The progressive changes in the expression of eight representative inflammatory cytokines - glial fibrillary acidic protein, tumour necrosis factor alpha, interleukin-6, cyclooxygenase-II, inducible nitric oxide synthase, nuclear factor-κB, monocyte chemoattractant protein-1 and macrophage inflammatory protein-1α - as well as the morphological changes in extracellular matrices by Masson's trichrome and Verhoeff-van Gieson staining and atomic force microscopy were assessed for normal (control), sham (no RF current), PRF and CRF (positive control) rats. The RF procedure used in this study was similar to the methods used in human clinical trials. All proposed inflammatory proteins showed up-regulation after RF thermal treatments. Specifically, PRF-treated rats showed recovery of up-regulated inflammatory cytokines on day 30 after application, while CRF rats showed significant up-regulation that persisted until day 30. The control and sham groups showed normal unmyelinated axon and collagen structures. PRF-induced lesions showed less fibre destruction than CRF-induced lesions. PRF-induced lesions also showed mild axonal damage and little swelling of the mitochondria. CRF-induced lesions showed ultrastructural changes of sciatic nerve fibres that were irreversible. PRF-induced pain relief may be due to temporary blockage of nerve signals through the nerve pathway responsible for reversible neuronal depression. However, CRF-induced pain relief may be due to permanent blockage of nerve signals through other nerve pathways. Therefore, CRF could be applied to chronic inflammatory models used to study the mechanism of neuropathic pain.

声帯粘膜の細胞外マトリックスの加齢的変化

声帯粘膜の細胞外マトリックスの加齢的変化

The effect of the platelet derived wound healing formula and the nerve growth factor on the experimentally injured spinal cord.

The main purpose of this study is to investigate the effect of platelet derived wound healing formula (PDWHF) and nerve growth factor (NGF) in the treatment of experimental spinal cord injury. PDWHF is a conglomerate of growth factors which include platelet derived growth factor (PDGF), platelet derived angiogenesis factor (PDAF), transforming growth factor-beta (TGF beta) and platelet factor IV (PF4). Complete spinal cord transection was performed at T12 in rats and the treatment of the spinal cord injury was achieved by filling the dead space with type 1 collagen gel impregnated with PDWHF, or with 2.5S-NGF. Controls were treated with only type 1 collagen gel. Animals were sacrificed at 1, 2 or 3 months. Histopathologically, tissue autolysis and cavity formation by phagocytosis expanded 1-3 mm into the cord stumps and the volume of cavitation was less in the two treated groups. In the NGF group, a greater number of surviving nerve cells were observed in this region. Most of the control animals formed only thin, short axonal bundles, however, increased axonal regrowth was noted in animals treated with trophic factors, especially in the NGF group. The NGF group formed thick axonal bundles and abundant neuroma. Increased angiogenesis was observed in the collagen gel matrix and the injured spinal cord parenchyma, in the PDWHF group. Recent studies have shown that mammalian adult CNS possesses the ability for structural and/or functional plasticity following injury under appropriate circumstances. In this in vivo study, exogenous NGF appeared to induce axomal outgrowth and nerve cell survival. PDWHF produced notable angiogenesis which seemed to improve the extracellular microenvironment. This may be important for the delivery of exogenous trophic factors, nutrients and for the changes of extracellular matrices to support nerve cells and axons.

Proteoglycans in haemodialysis-related amyloidosis.

Changes in extracellular matrices of articular tissue, intervertebral discs and systemic organs in patients with haemodialysis-related amyloidosis were investigated by immunohistochemical and biochemical examination of proteoglycans. Increased staining for chondroitin sulfate (CS) was detected in the amyloid deposits of all patients, ranging from early to advanced stages. Degenerative tissue changes around early-stage amyloid deposits in the intervertebral discs also showed positive staining for CS. Heparan sulfate (HS) was detected in amyloid deposits, especially in the synovial membrane. Biochemical analysis of connective tissues containing amyloid supported the immunohistochemical studies; CS was the major glycosaminoglycan species in these tissues, accounting for 55-81% of the total glycosaminoglycans. Although previous studies have stressed the importance of HS in amyloidogenesis, the present study showed that CS, which increased significantly in articular tissues associated with mechanical stress, also has a close relationship with amyloidogenesis.

Changes of cytoskeletons, extracellular matrices and extracellular matrix receptors in cutaneous squamous cell carcinoma

Changes of cytoskeletons, extracellular matrices and extracellular matrix receptors in cutaneous squamous cell carcinoma

Glomerular extracellular matrices in rat diabetic glomerulopathy by scanning electron microscopy.

Characteristic pathological changes in the glomeruli in diabetic nephropathy include expansion of the mesangial matrix and thickening of the glomerular basement membrane (GBM). Using an acellular digestion technique combined with scanning electron microscopy, the three-dimensional ultrastructural changes in glomerular extracellular matrices were studied in rats with diabetic glomerulopathy. Diabetes was induced by the intravenous injection of streptozotocin and morphological analyses were performed 3, 6 and 11 months after the injection. Expansion of mesangial area and GBM thickening became evident with time. After treatment with the series of detergents, all cellular components were completely removed leaving the extracellular matrices intact. In normal controls, the mesangial matrix appeared as fenestrated septa with oval or round stomata between the glomerular capillaries. In diabetic glomerulopathy, expansion of mesangial matrix and narrowing of the mesangial fenestrae were observed. These changes in the mesangial matrices seem to play a vital role in the progression of glomerulosclerosis in rat diabetes. A subendothelial thin layer of the GBM was continuous with the mesangial matrix. One cause of GBM thickening in streptozotocin diabetes may be expansion of the mesangial matrix into the peripheral GBM.

Heparan sulfate proteoglycans are lost in patients with diabetic nephropathy.

The pathogenesis of diabetic nephropathy relative to the changes in the glomerular extracellular matrices was investigated. Renal tissues from 10 diabetic patients were immunostained with antibodies directed against heparan sulfate proteoglycans (HS-PGs), laminin, type IV collagen and fibronectin. Seven patients were nephrotic and had advanced glomerulosclerosis with nodular lesion, while the other 3 had no renal manifestations or minor glomerular tissue alterations. Controls included kidneys removed from patients with renal tumors and specimens obtained by renal biopsy from patients with IgA nephropathy. Relationships among proteinuria, intensity of fluorescence and glomerular changes were studied. In diabetes 3 patients with minor glomerular lesions were found to have no changes in various components of extracellular matrices. A marked reduction in the intensity of staining with anti-HS-PG antibodies was observed in renal specimens from patients with nodular glomerulosclerosis and proteinuria, while a mild decrease in the intensity of fluorescence was observed in tissues stained with antilaminin antibodies. An increase compared to normal control sample findings in type IV collagen and fibronectin was observed in the mesangium of sclerosing glomeruli. No loss of HS-PG was observed in patients with IgA nephropathy. These results indicate that glomerular extracellular matrix HS-PG is lost in association with diabetic nephropathy; this loss results in alteration of the charge-selective properties of glomerular capillaries. This alteration may, in part, be the cause of the proteinuria associated with diabetic nephropathy.

Three-dimensional architecture of glomerular extracellular matrices in diabetic glomerulosclerosis

The three-dimensional ultrastructural changes of the glomerular extracellular matrices in diabetic glomerulosclerosis were studied in acellular rat and human diabetic glomeruli by scanning electron microscopy. The mesangial matrix appeared as fenestrated septa with oval stomata between the glomerular capillaries in normal control specimens. In diabetic glomerulosclerosis, both in humans and rats, expansion of mesangial matrix and narrowing of the mesangial fenestrae were observed. A thin layer of the mesangial matrix extended into the peripheral glomerular basement membrane (GBM) subendothelially. Thickening of the GBM in diabetic nephropathy might be due to expansion of the mesangial matrix into the peripheral GBM.