Extracellular Matrix In Patients Research Articles

Content and tissue expression of Collagen I, Collagen IV, and Laminin in the Extracellular Matrix in Prostate Adenocarcinoma.

Prostate cancer is one of the most common neoplasm in the male population. It is not known why some tumors become more aggressive than others. Although most studies show changes in the expression of cell adhesion molecules and the extracellular matrix correlated with the Gleason score, no study has objectively measured the tissue content of these molecules. This study aims to measure the content and tissue expression of collagen type I and IV and laminin in the extracellular matrix of patients with prostate adenocarcinoma and correlate these findings with the Gleason score and clinical characteristics. Forty-one patients who underwent radical prostate surgery at the Urology Department of a reference Hospital in Brazil between January 2015 and December 2020 were studied. The tissue protein content was estimated under light microscopy at a final magnification of 200 × . The mean collagen I score in prostate adenocarcinoma tissue samples was 7.16 ± 1.03 pixels/field. The mean type IV collagen score was 3.44 ± 0.61 pixels/field. The mean laminin score was 5.19 ± 0.79 pixels/field. The total Gleason score was correlated with both collagen and laminin. All the correlations were negative, which shows that the higher the collagen/laminin expression was, the lower the total Gleason score (p-value < 0,05). According to the Pearson correlation analysis, age has no statistical relationship with collagen and laminin content. PSA, in turn, showed a correlation only with laminin, but r = -0.378 (p = 0.015). Among the associated diseases and lifestyle habits, there is only statistical significance in the comparison of alcoholism for collagen I. For collagen IV and laminin, no statistical significance was obtained with the clinical variables analyzed.

Long-Term Course of Circulating Elastin, Collagen Type I, and Collagen Type III in Patients with Spontaneous Cervical Artery Dissection: a Prospective Multicenter Study.

An impaired integrity of vascular elements and the extracellular matrix (ECM) has been discussed to play a critical role in the pathophysiology of spontaneous cervical artery dissection (sCAD). This study aimed to explore the temporal course of circulating elastin, collagen type I, and collagen type III in patients with sCAD and evaluated their eligibility as diagnostic biomarkers. Patients with sCAD were prospectively enrolled in four German stroke centers. Blood samples were collected at baseline (acute phase), at day 10 ± 3 (subacute phase), and after 6 ± 1months (chronic phase). Patients with acute ischemic stroke not related to sCAD, healthy probands, and patients undergoing thromboendarterectomy of the carotid artery served as control groups. Serum levels of elastin and collagen types I and III were determined by ELISAs. Fifty-seven patients with sCAD were enrolled. Compared to all three control groups, patients with sCAD had significantly lower levels of elastin and collagen type III at baseline and after 6months. Compared to healthy probands, patients with sCAD showed similar collagen type I levels at baseline and in the subacute phase, but significantly increased levels after 6months. As serum levels of elastin, collagen types I and III were not elevated in the acute phase, they do not appear eligible as biomarkers for the diagnosis of sCAD. Persisting low serum levels of elastin and collagen type III towards the chronic phase of sCAD strengthens the hypothesis of a subtle, in most cases clinically inapparent affection of the ECM in patients with sCAD.

Biochemical composition of the glomerular extracellular matrix in patients with diabetic kidney disease.

In the glomeruli, mesangial cells produce mesangial matrix while podocytes wrap glomerular capillaries with cellular extensions named foot processes and tether the glomerular basement membrane (GBM). The turnover of the mature GBM and the ability of adult podocytes to repair injured GBM are unclear. The actin cytoskeleton is a major cytoplasmic component of podocyte foot processes and links the cell to the GBM. Predominant components of the normal glomerular extracellular matrix (ECM) include glycosaminoglycans, proteoglycans, laminins, fibronectin-1, and several types of collagen. In patients with diabetes, multiorgan composition of extracellular tissues is anomalous, including the kidney, so that the constitution and arrangement of glomerular ECM is profoundly altered. In patients with diabetic kidney disease (DKD), the global quantity of glomerular ECM is increased. The level of sulfated proteoglycans is reduced while hyaluronic acid is augmented, compared to control subjects. The concentration of mesangial fibronectin-1 varies depending on the stage of DKD. Mesangial type III collagen is abundant in patients with DKD, unlike normal kidneys. The amount of type V and type VI collagens is higher in DKD and increases with the progression of the disease. The GBM contains lower amount of type IV collagen in DKD compared to normal tissue. Further, genetic variants in the α3 chain of type IV collagen may modulate susceptibility to DKD and end-stage kidney disease. Human cellular models of glomerular cells, analyses of human glomerular proteome, and improved microscopy procedures have been developed to investigate the molecular composition and organization of the human glomerular ECM.

Extracellular Matrix in Heart Failure: Role of ADAMTS5 in Proteoglycan Remodeling.

Remodeling of the extracellular matrix (ECM) is a hallmark of heart failure (HF). Our previous analysis of the secretome of murine cardiac fibroblasts returned ADAMTS5 (a disintegrin and metalloproteinase with thrombospondin motifs 5) as one of the most abundant proteases. ADAMTS5 cleaves chondroitin sulfate proteoglycans such as versican. The contribution of ADAMTS5 and its substrate versican to HF is unknown. Versican remodeling was assessed in mice lacking the catalytic domain of ADAMTS5 (Adamts5ΔCat). Proteomics was applied to study ECM remodeling in left ventricular samples from patients with HF, with a particular focus on the effects of common medications used for the treatment of HF. Versican and versikine, an ADAMTS-specific versican cleavage product, accumulated in patients with ischemic HF. Versikine was also elevated in a porcine model of cardiac ischemia/reperfusion injury and in murine hearts after angiotensin II infusion. In Adamts5ΔCat mice, angiotensin II infusion resulted in an aggravated versican build-up and hyaluronic acid disarrangement, accompanied by reduced levels of integrin β1, filamin A, and connexin 43. Echocardiographic assessment of Adamts5ΔCat mice revealed a reduced ejection fraction and an impaired global longitudinal strain on angiotensin II infusion. Cardiac hypertrophy and collagen deposition were similar to littermate controls. In a proteomics analysis of a larger cohort of cardiac explants from patients with ischemic HF (n=65), the use of β-blockers was associated with a reduction in ECM deposition, with versican being among the most pronounced changes. Subsequent experiments in cardiac fibroblasts confirmed that β1-adrenergic receptor stimulation increased versican expression. Despite similar clinical characteristics, patients with HF treated with β-blockers had a distinct cardiac ECM profile. Our results in animal models and patients suggest that ADAMTS proteases are critical for versican degradation in the heart and that versican accumulation is associated with impaired cardiac function. A comprehensive characterization of the cardiac ECM in patients with ischemic HF revealed that β-blockers may have a previously unrecognized beneficial effect on cardiac chondroitin sulfate proteoglycan content.

Transcriptome genetic differences between responders and non-responders before bronchial thermoplasty

Background Bronchial thermoplasty (BT) is an endoscopic therapy used for the treatment of refractory asthma. Some predictive factors, for example the number of activations and severity of disease at baseline, have been used to determine the effectiveness of BT in treating patients with asthma. The aim of the present study was to comprehensively analyze RNA samples from the airway bronchial tissues of patients with severe asthma treated by BT, and to characterize each patient as a BT responder or non-responder. Methods Eight patients with severe asthma scheduled to undergo BT and bronchus biopsies were recruited before the procedures were conducted. Extracted RNA samples from bronchial tissues were sequenced and differential gene expression analysis was carried out. Results/discussion: Subjects with Asthma Quality of Life Questionnaire score changes ≥0.5 for a period of 12 months were considered BT responders. Non-responders had score changes <0.5 for 12 months. Histopathology findings were similar to those reported previously, and no significant differences in the expression of α-smooth muscle actin and protein gene product 9.5 were observed between responders and non-responders. Transcriptome analysis at baseline identified 67 genes that were differentially expressed between responders and non-responders, including SLPI, MMP3, and MUC19, which were upregulated in responders. Although the differentially expressed gene products may have conflicting effects, genes in the airway epithelium and extracellular matrix of patients with severe asthma may determine the BT response. Our results identified possible transcriptomic changes that could be used to identify BT responders.

SIGNIFICANCE OF THE STRUCTURE AND METABOLISM OF THE EXTRACELLULAR MATRIX IN THE PATHOGENESIS OF ABDOMINAL HERNIAS. REVIEW

: ventral hernias remain one of the most frequent reason for scheduled and emergency surgeries. Despite the widespread use of various methods and technologies of hernias repair, including traditional non-invasive and laparoscopic methods popular in recent years, the problem of recurrent hernias remains relevant to this day. At the end of the last century, studies have shown that patients with inguinal hernias, compared with patients operated on for other intraabdominal reasons, have pronounced quantitative and qualitative changes in collagen proteins. This, in turn, led to increased interest in the exchange of connective tissue and the structure of the extracellular matrix in patients with hernia disease and other similar diseases, as well as the biology of wound healing and scar formation to this day. Significant number of experimental and clinical studies of the structure and function of the extracellular matrix in patients with hernia disease have been accumulated. The review is designed to reveal current understanding of relationships of metabolism of connective tissues and structure of extracellular matrix to pathogenesis of ventral hernias in order to identify key points, which will, in the future, develop a personalized approach to treatment of patients with these disorders.

Cytokine Production by Blood Immune Cells, Tumor and its Microenvironment, Characteristics of Extracellular Matrix in Patients with Invasive Ductal Carcinoma of No Special Type

Cytokine Production by Blood Immune Cells, Tumor and its Microenvironment, Characteristics of Extracellular Matrix in Patients with Invasive Ductal Carcinoma of No Special Type

P.383Antisense oligonucleotides therapy for COL6-related congenital muscular dystrophy

Collagen VI-related congenital muscular dystrophies (COL6-CMD) are caused by recessive and dominant mutations in the three Collagen VI genes (COL6A1, COL6A2 and COL6A3). COL6-CMD has a wide clinical spectrum ranging from the severe early-onset Ullrich congenital muscular dystrophy, to the intermediate phenotypes and the milder Bethlem myopathy. Currently there is no effective treatment for COL6-CMD. Antisense oligonucleotides (AON) can interfere with gene expression by modulating pre-mRNA splicing or inducing gene silencing. Splice-switching AONs have been successfully used in neuromuscular conditions, with drugs now available for Duchenne muscular dystrophy and spinal muscular atrophy. We have recently explored two different therapeutic strategies of AONs for dominant COL6-CMD. For one group of patients carrying a common splice site mutation that results in exon 16 deletion in COL6A3, we have developed an allele-specific silencing AON approach. For another group of patients affected by a recurrent deep intronic mutation that induces a pseudo-exon in intron 11 in COL6A1, we have developed an exon-skipping splice-switching approach. Both approaches are being investigated in skin fibroblasts derived from patients carrying the target heterozygous mutations. Regarding the allele-specific silencing approach, we show that gapmer AONs can selectively silence the mutant COL6A3 transcripts. Regarding the exon-skipping approach, the AONs we designed efficiently correct the aberrant splicing of the mutant pre-mRNA and produce wild-type COL6A1 transcripts. This approach also successfully restore the deposition of functional collagen VI protein in the extracellular matrix in patients' skin fibroblasts. In conclusion, our study provides the proof of concept of different AON approaches as therapeutic strategy for COL6-related CMD. The lead AONs will be further validated in relevant mouse models in collaboration with the team at NIH lead by Prof Bönnemann.

Long noncoding RNA TM1P3 is involved in osteoarthritis by mediating chondrocyte extracellular matrix degradation.

Osteoarthritis (OA) is a widespread degenerative joint disease characterized by articular cartilage degradation and is the leading cause of physical disability. Noncoding RNAs, especially long noncoding RNAs (lncRNAs) and microRNAs, are involved in the degradation of the chondrocyte extracellular matrix (ECM) in patients with OA. The present study was aimed to investigate the effects of lncRNA and miR-22 on the degradation of the chondrocyte ECM and underlying mechanisms. To simulate conditions found in OA, primary cultured chondrocytes were treated with IL-1, TGF-β, or sb525334. Real-time PCR and Western blot analysis were performed to detect expressions of miR-22, lncRNA-TM1P3, ALK1, MMP13, pSMAD1/5, SMAD1, and pSMAD5. Small interfering RNAs and a miR-22 mimic or inhibitor were utilized to determine lncRNA-TM1P3 knockdown and miR-22 overexpression or inhibition. The lncRNA-TM1P3 significantly upregulated in patients with OA, accompanied by the downregulation of miR-22 and upregulation of pSMAD1/5 and MMP13, which ultimately resulted in the degradation of the chondrocyte ECM in patients with OA. Bioinformatics analysis predicted miR-22 as a target of both lncRNA-TM1P3 and MMP13. The lncRNA-TM1P3 knockdown significantly increased the expression of ALK1, a corresponding increase in ECM degradation was observed by affecting the phosphorylation of SMAD1/5 and the expression of MMP13, which did not affect the expression of ALK1. These findings demonstrated that the lncRNA-TM1P3/miR-22/TGF-β signaling/MMP13 axis is involved in the degradation of chondrocyte ECM in patients with OA, which could provide novel therapies for OA treatment.

Functional Assessment of Decellularized Extracellular Matrix in Patients Undergoing LVAD Implantation

Purpose While changes in the extracellular matrix (ECM) have been demonstrated in the failing heart, changes in mechanical ECM properties are unknown. We aimed to determine the biomechanical properties of ECM obtained from myocardial tissue samples at the time of left ventricular assist device (LVAD) implantation. Methods Myocardial tissue samples obtained at the time of LVAD implantation underwent decellularization and determination of the stress-strain relationship. Storage modulus (G’, an assessment of elastic properties) and loss modulus (G’’, an assessment of viscous properties) were compared against the likelihood of clinical recovery as computed using an established predictive model, the INTERMACS Cardiac Recovery Score (I-CARS). Results 14 myocardial samples were obtained and analyzed at the time of LVAD implantation. Mean age was 48.8 years (range: 17-74). 11 subjects were male (79%) with ischemic etiology in 3 (21%). Mean ejection fraction was 19.2% ± 7.2%. Storage and loss modulus of the ECM samples were grouped into tertiles based on the likelihood of myocardial recovery as predicted by I-CARS score. The group with the lowest recovery likelihood (0.0%, I-CARS score of 0-3) had the highest average value of storage modulus (1312 ±1553 Pa at 1 rad/s). The group with intermediate recovery likelihood (6.3%, I-CARS score of 4-6) had intermediate average value (781 ±517 Pa at 1 rad/s). The group with the highest recovery likelihood (38.9%, I-CARS score of 7-9) had the lowest average value (450 ±308 Pa at 1 rad/s). Similar trends were seen with loss modulus (figure). Conclusion Analysis of ECM in myocardial tissue samples obtained at the time of LVAD implantation showed a trend toward lower storage modulus and loss modulus in those with a higher predicted likelihood of myocardial recovery based on the I-CARS recovery score. The functional properties of decellularized ECM can be assessed ex vivo and may represent an independent parameter for myocardial recoverability.

Cytokine production by blood immune cells, tumor and its microenvironment, characteristic of extracellular matrix in patients with invasive ductal carcinoma of no special type

The aim of this research was to study cytokine production by blood immune cells, tumor, and its microenvironment, and characterize extracellular matrix of patients with invasive ductal carcinoma of no special type and lymphatic metastases. Spontaneous and polyclonal activators stimulated production of cytokines by blood immune cells, tumor and its microenvironment were studied in 95 patients with invasive ductal carcinoma of no special type. The concentration of IL-2, IL-4, IL-6, IL-8, IL-10, IL-17, IL-18, IL-1β, IL-1Ra, TNF-α, IFN-γ, G-CSF, GM-CSF, VEGF and MCP-1 was determined by the solid-phase enzyme-linked immunosorbent assay. The condition of fibrous component and presence of neutral glycoproteins and sulfated glycosaminoglycans were evaluated during the research of extracellular matrix. Regional lymphatic metastases were detected in 35 of 95 patients. It was shown that in the presence or absence of lymphatic metastases index of polyclonal activators influence on the production of cytokines by blood immune cells was different for IL-6, IL-8, and IL-1β; while in the case of cytokine production by tumor and its microenvironment the index of influence was different for IL-2 and IL-17. The presence of lymphatic metastases corresponded with the rise of cytokines spontaneous production, while the absence of lymphatic metastases corresponded with the rise of cytokines production stimulated by polyclonal activators. The value of indices of polyclonal activators influence on the production of cytokines by blood immune cells pointed to the highly stimulating effect of polyclonal activators while the value of indices of polyclonal activators influence on cytokines production by tumor and its microenvironments pointed to the low and sometimes even absent effect of polyclonal activators. Basing on these data we propose a ratio of indices of polyclonal activators influence for the better evaluation of the probability of lymphatic metastases during preoperative period. After characterizing extracellular matrix we found out a point threshold, which, in 100% of cases, predicted the presence of lymphatic metastases basing on the condition of extracellular matrix. Using the data acquired, we are proposing a risk group for metastasis among women with no lymphatic metastases in the moment of check-up.

S&T-51 EXTRACELLULAR MATRIX CHARACTERISTICS OF TUNICA ALBUGINEA IN MEN WITH AND WITHOUT PEYRONIE'S DISEASE

INTRODUCTION AND OBJECTIVES: Peyronie’s disease (PD) is characterized by the formation of fibrous plaques in the tunica albuginea (TA) of the penis, causing pain and penile curvature during erection, with great impact on quality of life of patients. There is a lack of highly effective medical treatments for PD, mainly because it’s pathophysiology is not completely understood. The aim of this study was to compare possible markers related to alterations and remodeling of extracellular matrix which can be involved on the development of the disease in men with and without PD METHODS: Samples of TA of 10 patients with PD were obtained at the moment of surgical correction of penile curvature. Samples of TA of 10 corpses with normal penis at the physical exam were obtained (control group). Tissues were processed and stained with Hematoxilin, submitted to immunohistochemistry (IHQ) with the following primary antibodies: heparanase (HPSE), heparanase2 (HPSE2), metalloproteinases 2 and 9 (MMP2 and MMP9) and TGF-s. IHQ reaction used peroxidase kit LSAB -peroxidase (DakoCytomation , Copenhagen, Dinamarca). Genic expression of heparanase-1 (HPSE), heparanase2 (HPSE2), metalloprotease 9 (MMP9), syndecan 1 (SYN1), interleukin 6 (IL-6), hyaluronic acid sintetase 1 e 2 (HAS1 e HAS2), hyaluronidases 1 e 2 (HYL1 e HYL2), was done by quantitative RTPCR (qRT-PCR). Statystical analyses were done using software Graph PRISM 5 . Project was reviewed and approved by the Research Ethical Committee of Faculdade de Medicina do ABC, (# 242/2010). RESULTS: Histopathological evaluation showed higher number of cells, reduction of apoptosis events, alteration of the distribution of collagen fibers and diminished number of vessels on TA of patients with PD compared to the controls TA. qRT-PCR analysis showed increased concentration of TGF-s and interleucine1 reduction on HSPE-1 expression and decreased concentration of dermatan sulfate and chondroitin sulfate in patients with PD (p<0.05). However, there were no alterations on HPSE2, MMP2 e MMP9 expressions. Those findings suggest a higher remodeling of extracellular matrix in patients with PD. CONCLUSIONS: In comparison with the controls the TA of patients with PD showed lesser number of vessels and apoptosis events; increased concentration of TGF-s and interleukin-1, reduction of heparanase-1 expression and decreased concentration of dermatan sulfate and chondroitin sulfate.

S&T-52 EARLY CHANGES IN SKELETAL MUSCLE AS A STRONG PROGNOSTIC BIOMARKER IN PATIENTS WITH METASTATIC RENAL CELL CARCINOMA

INTRODUCTION AND OBJECTIVES: Peyronie’s disease (PD) is characterized by the formation of fibrous plaques in the tunica albuginea (TA) of the penis, causing pain and penile curvature during erection, with great impact on quality of life of patients. There is a lack of highly effective medical treatments for PD, mainly because it’s pathophysiology is not completely understood. The aim of this study was to compare possible markers related to alterations and remodeling of extracellular matrix which can be involved on the development of the disease in men with and without PD METHODS: Samples of TA of 10 patients with PD were obtained at the moment of surgical correction of penile curvature. Samples of TA of 10 corpses with normal penis at the physical exam were obtained (control group). Tissues were processed and stained with Hematoxilin, submitted to immunohistochemistry (IHQ) with the following primary antibodies: heparanase (HPSE), heparanase2 (HPSE2), metalloproteinases 2 and 9 (MMP2 and MMP9) and TGF-s. IHQ reaction used peroxidase kit LSAB -peroxidase (DakoCytomation , Copenhagen, Dinamarca). Genic expression of heparanase-1 (HPSE), heparanase2 (HPSE2), metalloprotease 9 (MMP9), syndecan 1 (SYN1), interleukin 6 (IL-6), hyaluronic acid sintetase 1 e 2 (HAS1 e HAS2), hyaluronidases 1 e 2 (HYL1 e HYL2), was done by quantitative RTPCR (qRT-PCR). Statystical analyses were done using software Graph PRISM 5 . Project was reviewed and approved by the Research Ethical Committee of Faculdade de Medicina do ABC, (# 242/2010). RESULTS: Histopathological evaluation showed higher number of cells, reduction of apoptosis events, alteration of the distribution of collagen fibers and diminished number of vessels on TA of patients with PD compared to the controls TA. qRT-PCR analysis showed increased concentration of TGF-s and interleucine1 reduction on HSPE-1 expression and decreased concentration of dermatan sulfate and chondroitin sulfate in patients with PD (p<0.05). However, there were no alterations on HPSE2, MMP2 e MMP9 expressions. Those findings suggest a higher remodeling of extracellular matrix in patients with PD. CONCLUSIONS: In comparison with the controls the TA of patients with PD showed lesser number of vessels and apoptosis events; increased concentration of TGF-s and interleukin-1, reduction of heparanase-1 expression and decreased concentration of dermatan sulfate and chondroitin sulfate.

Abstract 16611: Impact of Mechanical Unloading on Structural and Non-structural Components of the Extracellular Matrix in Patients With Dilated Cardiomyopathy

Introduction: Extracellular matrix (ECM) is the non-cellular component of the myocardium and its remodeling in cardiomyopathy is a major contributor to disease outcomes. While the impact of left ventricular assist device (LVAD) on remodeling of fibrillar ECM has been studied, its effects on the non-fibrillar ECM and matricellular proteins within the ECM that can influence cardiac structure and function are unknown. Hypothesis: Mechanical unloading preserves the non-fibrillar ECM composition, thereby contributing to reversal of adverse myocardial remodeling. Methods: Hearts were obtained from heart failure patients with idiopathic dilated cardiomyopathy (DCM) with LVAD support (&gt;3mths; LVAD), without LVAD (no-LVAD), and non-failing control hearts (NFC) (n=10/group). LV free wall were fixed or flash-frozen for molecular and IHC evaluations. Results: Assessment of fibrillar ECM showed greater total and insoluble collagen content in no-LVAD hearts, consistent with an increase in collagen type I mRNA, and collagen type I-to-type III ratio. Proteolytic activities (collagenase &amp; gelatinase) were significantly higher in no-LVAD (by 45% and 40%) suggesting greater ECM turnover in these hearts. TIMP1 mRNA decreased in both DCM groups (by 60%), but its protein decreased only in no-LVAD hearts (by 30%). TIMP3 and TIMP4 mRNA (by 30% and 55%) and protein (by 60% and 55%) decreased in both LVAD and no-LVAD hearts. ADAM-TS2, the N-terminal endopeptidase for collagen, was markedly lower in no-LVAD, while LOX1, the cross-linking enzyme showed a small increase in no-LVAD hearts. Evaluation of the basement membrane revealed that laminin arrangements were dispersed in no-LVAD hearts. Integrin β1, a cardiomyocyte cell-surface receptor for laminin, was reduced in no-LVAD but preserved in LVAD hearts. Other basement membrane proteins (fibronectin-1, collagen IV) showed similar trends, whereas differences in matricellular proteins (osteopontin, SPARC) were less striking between LVAD and no-LVAD hearts. Conclusions: Mechanical unloading positively impacts multiple aspects of myocardial ECM, the fibrillar ECM network and the less explored basement membrane, although the matricellular proteins residing within the ECM seem to be less affected.

Regulation of extracellular matrix In patients with Q-wave myocardial infarction after thrombolytic therapy

Aims. In order to study the levels of matrix metlloproteinase-9, tissue inhibitor of metalloproteinase-1, intracardiac hemodynamics, frequency of acute cardiac aneurysm development in patients with Q-wave myocardial infarction after thrombolysis, 74 patients were examined.Methods and results. All participants underwent clinical and laboratory examination, echocardiography, determination of serum levels of matrix metlloproteinase-9 and tissue inhibitor of metalloproteinase-1.Conclusion. Patients, who got thrombolysis in 6 to 12 hours period, had excessive activation of the proteolysis system on the background of relative deficit of tissue inhibitor of metalloproteinase-1, the prevalence of dilatation and restrictive diastolic dysfunction, cardiac aneurysm was detected frequently and thrombogenic potential was higher.

The Prostaglandin F2α Analog Fluprostenol Attenuates the Fibrotic Effects of Connective Tissue Growth Factor on Human Trabecular Meshwork Cells

Abstract Purpose: The trabecular meshwork (TM) outflow pathways of the aqueous humor show an increase in extracellular matrix in patients with primary open-angle glaucoma (POAG). The increase in TM extracellular matrix appears to be caused by transforming growth factor-β signaling and its downstream mediator connective-tissue growth factor (CTGF). Here we studied whether treatment with the prostaglandin F2α analog fluprostenol modulates the CTGF-mediated increase of the TM extracellular matrix. Human TM cells from 3 different donors were treated with CTGF (50 ng/mL) and/or fluprostenol (10(-6) M and 10(-7) M) and were analyzed by real-time reverse transcription polymerase chain reaction and Western blotting. Cell supernatants of the treated cells were analyzed by zymography. Treatment with CTGF induced the expression and synthesis of CTGF, fibronectin, collagen type IV and VI, while treatment with fluprostenol alone had no effects. The effects of CTGF were blocked by 1-h pretreatment with fluprostenol in a dose-dependent manner. Treatment with fluprostenol or combined fluprostenol/CTGF induced the activity of matrix metalloproteinase 2 (MMP2) in TM cells, whereas treatment with CTGF alone had no effects on MMP2 activity. Fluprostenol blocks the fibrotic effects of CTGF on human TM cells and increases the activity of MMP2. Both effects have the distinct potential to attenuate a CTGF-mediated increase in TM extracellular matrix in patients with POAG and any effects on TM outflow resistance that may result from that.

Systemic connective tissue involvement and cardiac involvement: the 2010 revised Gent nosology in the Marfan syndrome diagnostics

The author presents his view on the problem of Marfan syndrome-related hereditary connective tissue disorders, in particular, primary mitral valve prolapse and Marfanoid habitus, as the most prevalent manifestations. The changes in the assessment of external and visceral manifestations of Marfan syndrome, according to the originalGentnosology and its 2010 revision, are discussed. The importance of the evaluation of systemic connective tissue involvement, with the systemic score calculation and cardiac extracellular matrix assessment, is emphasised. The author presents the evidence justifying the need for a wider use of the cardiac involvement concept, in order to assess the severity of structural and functional disturbances of cardiac extracellular matrix in patients with primary mitral valve prolapse and Marfanoid habitus. The cardiac involvement assessment should be based on the systemic score, minor cardiac criteria, the number and type of minor cardiac abnormalities, the results of clinical examination, and the serum levels of transforming growth factors (TGF)

Advanced Glycation Endproducts and Alendronate Differentially Inhibit early and Late Osteoclastogenesis In vitro

Advanced Glycation Endproducts (AGEs) are greatly elevated in bone extracellular matrix of patients with Diabetes mellitus, and this has been associated with the increased incidence of fractures observed in these patients. AGEs affect the homeostasis of bone cells such as osteoblasts and osteoclasts. Bisphosphonates are first-line anti-osteoporotic drugs that principally exert their effects by inhibiting osteoclastic activity. However, the effect of bisphosphonate treatment on bone quality in patients with Diabetes is uncertain. In the present work we have evaluated the action of AGEs (50-200 μg/ml), with or without Alendronate (10-8-10-4M), on osteoclastogenesis induced by co-cultures of Raw 264.7 macrophages and UMR106 osteoblasts. We determined the effects of different culture conditions on osteoclastic recruitment, tartrate-resistant acid phosphatase (TRAP) activity and expression of RAGE (receptor for AGEs); and on the osteoblastic expression of RANK ligand (RANKL). AGEs and Alendronate inhibited the recruitment and TRAP activity of osteoclasts, with an additive effect of both agents at high concentrations of Alendronate. While AGEs prevented the early and late stages of osteoclastogenesis, Alendronate (alone or in co-incubation with AGEs) only inhibited its later stages. In addition, both AGEs and Alendronate increased the osteoclastic expression of RAGE and decreased the osteoblastic expression of RANKL, correlating closely with their inhibition of osteoclastogenesis. If these in vitro results can be extrapolated to a clinical setting, they may be indicating a potentiation of the anti-resorptive effects of Alendronate in the context of bone extracellular matrix with excess accumulation of AGEs, as might occur in a patient with Diabetes mellitus.

Characteristics of aortic wall extracellular matrix in patients with acute myocardial infarction: tissue microarray detection of collagen I, collagen III and elastin levels

Extracellular matrix (ECM) remodelling of the vessel wall is hypothesized to be an important step in atherosclerosis. Changes of the ECM are associated with the gradual progression of an atherosclerotic lesion from a lipid streak to complicated unstable plaque, leading to a complete vessel occlusion and eventually myocardial infarction (MI). Understanding of this process is critical in the treatment and prevention of ischaemic heart disease (IHD). We investigated the histopathological characteristics of aortic wall ECM in IHD patients. Collagen I, collagen III and elastin were assessed immunohistochemically in patients with acute MI and those with stable angina, using aortic punch tissues obtained from coronary artery bypass graft surgery. Fluorescence tissue images were analysed using the tissue microarray technique. The results showed that collagen III expression was found to be significantly lower in the acute MI group (P < 0.001). As a result of this change, the patients with MI also revealed a significant reduction in the collagen III/collagen I ratio. The elastin/collagen III ratio was significantly higher in the MI group (P < 0.001). Our study provided evidence of a decrease in collagen III content in patients with MI, which could possibly explain the mechanism of plaque vulnerability and weakening of the plaque cap. A reduction in collagen III content, particularly away from the atherosclerotic lesions, might be explained by the systemic vascular changes in patients with MI, and inflammation and immune responses could be potential causes of these systemic transformations. The biochemical mechanisms and factors regulating collagen III production might be potential markers to predict possible cardiovascular events.

Matrix Proteins from Smoke-Exposed Fibroblasts Are Pro-proliferative

Airway remodeling decreases lung function in chronic obstructive pulmonary disease (COPD). Extracellular matrix (ECM) deposition is increased in remodeled airways and drives cellular processes of proliferation, migration, and inflammation. We investigated the role of cigarette smoke in altering the ECM deposited from human lung fibroblasts. Lung fibroblasts isolated from patients with COPD or other lung disease were exposed to cigarette smoke extract (CSE) and 5 ng/ml transforming growth factor-β1 for 72 hours; in some experiments, inhibitors of signaling molecules were added. Deposition of perlecan, fibronectin, and elastin were measured by ELISA, as was release of IL-8 and IL-13. Unstimulated fibroblast cells were reseeded onto deposited matrix and assessed for proliferation and cytokine release. CSE (5%) increased deposition of fibronectin and perlecan from only COPD fibroblasts. Fibronectin and perlecan deposition was attenuated by addition of the NF-κB inhibitor, BMS-345541, and the signal transduction and activator of transcription-1/3 inhibitor, pyridone 6, respectively. CSE (5%) increased IL-8 release from COPD fibroblasts more than non-COPD fibroblasts. This increase was attenuated by BMS-345541. Matrix deposited after 5% CSE stimulation increased proliferation of fibroblasts, but did not alter cytokine release. ECM produced from COPD fibroblasts after CSE exposure has proproliferative effects. Thus, the ECM in patients with COPD may create an environment that promotes airway remodeling.