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Introduction: IgA nephropathy is the most common primary glomerulonephritis among adults in clinic. Thin basement membrane nephropathy is often underestimated or even omitted if it coincides with IgA nephropathy. Therefore, it is necessary to study the epidemiological, clinical, and molecular characteristics of the concurrence of this entity., Methods: Eight patients with concurrent IgA nephropathy and thin basement membrane nephropathy (IgA-T) were retrospectively analyzed based on their clinicopathological characteristics. Genetic analysis was performed using whole-exome sequencing and Sanger's sequencing. Data of the patients with IgA nephropathy and normal basement membrane (IgA-N) and variants in the local in-house database were used as controls. All candidate variants were assessed in silico., Results: The clinical manifestations of patients with IgA-T were hematuria, proteinuria, and renal insufficiency. Histopathological analysis showed mild mesangial hyperplasia, focal segmental glomerulosclerosis, podocyte activation, and foot process fusion. Crescent was rarely seen. COL4A and/or podocyte cytoskeleton and mitochondria-related gene variants were detected in seven IgA-T patients. Three patients exhibited pathogenic variants of COL4A, including a new variant. All IgA-T and one IgA-N patient possessed ITGB4 and/or PLEC variants, but there was no corresponding genotype-phenotype relationship. Six patients possessed other podocyte cytoskeleton and mitochondria-related gene variants such as NPHS2, SRGAP1, MYO1E, MYO1C, WT1, and COQ9, which were first reported in patients with IgA-T and were not in controls. Altogether, there were no significant differences in the degrees of proteinuria, serum creatinine, and eGFR during the follow-up period of 5-10 years, but there was a significant difference in the degree of proteinuria between IgA-T patients with podocyte-related gene variants and IgA-N patients. In the IgA-T group, patients with podocyte-related gene variants seemed predisposed to progress than patients without those variants, with higher proteinuria and serum creatinine and reduced eGFR., Conclusion: Concurrent thin basement membrane nephropathy and/or heterozygous COL4A gene pathogenic variants do not necessarily predict the short-term progress of sporadic IgA nephropathy in adults. Predisposition factors for this disease progression should be considered for detecting the variants of COL4A and podocyte cytoskeleton and mitochondria-related genes simultaneously, which also manifests the complexity and heterogeneity of IgA nephropathy with concurrent thin basement membrane nephropathy., (© 2022 The Author(s). Published by S. Karger AG, Basel.)

Breakthrough bleeding (BTB) is the cause of 50% of discontinuations of use by long-acting progestogen-only contraceptive users. BTB appears to be associated with capillaries of abnormal morphology and fragility. Disturbances in vessels integrity, intercellular junctions or alteration of the basement membrane (BM) composition has been proposed to contribute toward the fragility of these capillaries. In a clinical trial study, 34 healthy fertile volunteers were studied before and during use of levonorgestrel releasing subdermal contraceptive implant Norplant. Three endometrial biopsies were taken from each volunteer, one before insertion and 2 at different time intervals after exposure to Norplant. Then 3 major components of BM of endometrial vessels; Collagen IV (CIV), laminin and heparan sulphate proteglycan (HSPG) were immunostained and studied. The density of immunostained vessels by each antibody were measured and compared with density of vessels stained by CD34. Double immunostain of CD34 and laminin was also used for demonstration of vessels without BM. Significant decrease in vascular density showing BM components were shown in the study. The density of laminin, CIV, and HSPG-positive vessels were decreased from 176±8, 166±6.6 and 61±6 vessels/mm² (Mean±SEM) in the endometrium of Norplant acceptors before use control samples to 156±6, 144±6.2 and 33±2.9 in the endometrium of Norplant acceptors after 2 to 13 weeks Norplant exposure, respectively (P

Synopsis Objective Acetyl aspartic acid (A-A-A) was discovered through gene array analysis with corresponding Cmap analysis. We found that A-A-A increased keratinocyte regeneration, inhibited dermal matrix metalloprotease ( MMP) expression and relieved fibroblast stiffness through reduction of the fibroblast stiffness marker F-actin. Dermal absorption studies showed successful delivery to both the epidermal and dermal regions, and in-use trial demonstrated that 1% A-A-A was well tolerated. In this study, the aim was to investigate whether A-A-A could stimulate the synthesis of extracellular matrix supporting proteins in vivo and thereby improving the viscoelastic properties of human skin by conducting a dual histological and biophysical clinical study. Method Two separate double-blind vehicle-controlled in vivo studies were conducted using a 1% A-A-A containing oil-in-water (o/w) emulsion. In the histological study, 16 female volunteers (>55 years of age) exhibiting photodamaged skin on their forearm were included, investigating the effect of a 12-day treatment of A-A-A on collagen IV ( COLIV) and fibrillin-1. In a subsequent pilot study, 0.1% retinol was used for comparison to A-A-A (1%). The biomechanical properties of the skin were assessed in a panel of 16 women (>45 years of age) using the standard Cutometer MPA580 after topical application of the test products for 28 days. The use of multiple suction enabled the assessment of F4, an area parameter specifically representing skin firmness. Results Twelve-day topical application of 1% A-A-A significantly increased COLIV and fibrillin with 13% and 6%, respectively, compared to vehicle. 1% A-A-A and 0.1% retinol were found to significantly reduce F4 after 28 days of treatment by 15.8% and 14.7%, respectively, in the pilot Cutometer study. No significant difference was found between retinol and A-A-A. However, only A-A-A exhibited a significant effect vs. vehicle on skin firmness which indicated the incremental benefit of A-A-A as a skin-firming active ingredient. Conclusion In this study, we showed the in vivo efficacy of 1% A-A-A both on a protein level (fibrillin and collagen IV) and on a clinical end point, specifically skin firmness, providing proof that, acetyl aspartic acid has a strong potential as an anti-ageing 'cosmeceutical' ingredient answering the needs of our key consumer base. [ABSTRACT FROM AUTHOR]

Cervical cancer is one of the most commonly diagnosed cancers and a significant cause of mortality in women worldwide. Although cervical cancer is fully treatable in the early stages, once it has metastasized, patient outcome is poor. The objective of the present study was to investigate the effect of dietary supplementation with a nutrient mixture (NM) containing lysine, ascorbic acid, proline, green tea extract and other micronutrients on the expression of extracellular matrix (ECM) proteins in HeLa cell xenografts in nude female mice. After housing for 1 week, female athymic nude mice between 5 and 6 weeks of age (n=12) were inoculated subcutaneously with 3x106 HeLa cells in phosphate-buffered saline and Matrigel and randomly divided into two groups. These were the control group, in which the mice were fed with regular mouse chow, and the NM group, in which the mice were fed with the regular diet supplemented with 0.5% NM (w/w). After 4 weeks, the tumors were excised and processed for histology. Tumor growth was evaluated and the tumors were stained for the ECM proteins collagen I, collagen IV, fibronectin, laminin, periodic acid-Schiff (PAS) and elastin. NM strongly inhibited (by 59%, P=0.001) the growth of HeLa xenografts in nude mice. Tumors from control mice exhibited little to no collagen I expression either internally or in the fibrous capsule, while tumors from the NM group expressed collagen I in the fibrous capsule and within the tumor. Tumors from the control group showed diffuse cytoplasmic and capsular collagen IV with abundant nucleated cells. NM treatment substantially increased collagen IV production and induced a dense fibrous network of collagen IV with chambers that surrounded live nucleated cells and large amounts of necrotic cell debris. Tumors from the mice fed with the NM exhibited a well-defined border of fibronectin in the capsule and intense areas of staining internally whereas control group tumors showed less overall fibronectin with sporadic internal staining and little in the fibrous capsule. Although laminin appeared abundantly in control and NM-treated tumors, the NM group tumors exhibited a chamber-like network of laminin internally. Tumors from the control group exhibited internal areas of intense PAS staining, whereas tumors from the NM-treated group exhibited a more uniform diffuse pattern of PAS staining. In conclusion, NM supplementation of HeLa xenograft-bearing female nude mice demonstrated a potent inhibition of tumor growth and enhancement of ECM proteins, suggesting the therapeutic value of this specific nutrient complex in the treatment of cervical cancer. [ABSTRACT FROM AUTHOR]

Basement membrane collagens or derived fragments are measured in biological fluids such as blood and urine of patients and appear to be useful for diagnosis, prognostication, or treatment monitoring as proposed for endostatin, a fragment of collagen XVIII, or tumstatin, a fragment of collagen IV. Tetrastatin, the NC1 alpha 4 collagen IV domain, was previously reported to inhibit tumor growth and angiogenesis. The aim of this study was to develop and validate a method to measure tetrastatin concentrations in human fluids. We developed a competitive enzyme-linked immunosorbent assay (ELISA). It allowed measuring tetrastatin levels in human serum, bronchial aspiration and bronchoalveolar lavage fluids, and lung tissue extracts. The tetrastatin level was significantly higher in tumor tissues than in healthy lung tissues. Tetrastatin competitive ELISA could be useful to quantify tetrastatin in tissues and biological fluids for the diagnosis or prognostication of diseases in which basement membrane metabolism may be altered, especially tumor progression. [ABSTRACT FROM AUTHOR]

Peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists have beneficial effects on the kidney diseases through preventing microalbuminuria and glomerulosclerosis. However, the mechanisms underlying these effects remain to be fully understood. In this study, we investigate the effects of PPAR-γ agonist, rosiglitazone (Rosi) and pioglitazone (Pio), on collagen IV production in mouse podocytes. The endogenous expression of PPAR-γ was found in the primary podocytes and can be upregulated by Rosi and Pio, respectively, detected by RT-PCR and Western blot. PPAR-γ agonist markedly blunted the increasing of collagen IV expression and extraction in podocytes induced by TGF-β. In contrast, adding PPAR-γ antagonist, GW9662, to podocytes largely prevented the inhibition of collagen IV expression from Pio treatment. Our data also showed that phosphorylation of Smad2/3 enhanced by TGF-β in a time-dependent manner was significantly attenuated by adding Pio. The promoter region of collagen IV gene contains one putative consensus sequence of Smad-binding element (SBE) by promoter analysis, Rosi and Pio significantly ameliorated TGF-β-induced SBE4-luciferase activity. In conclusion, PPAR-γ activation by its agonist, Rosi or Pio, in vitro directly inhibits collagen IV expression and synthesis in primary mouse podocytes. The suppression of collagen IV production was related to the inhibition of TGF-β-driven phosphorylation of Smad2/3 and decreased response activity of SBEs of collagen IV in PPAR-γ agonist-treated mouse podocytes. This represents a novel mechanistic support regarding PPAR-γ agonists as podocyte protective agents. [ABSTRACT FROM AUTHOR]

To explore the preventative effects of prostaglandin E1 (PGE1) on a rabbit model of CCl4-induced liver fibrosis after transcatheter arterial chemoembolization (TACE), we generated a rabbit model of CCl4-induced liver fibrosis by treatment with 40% CCl4 in iodized olive oil for 16 weeks. Body mass and serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total protein (TP), albumin (ALB), albumin:globulin ratio (A:G), total bilirubin (TBIL), and direct bilirubin (DBIL) were measured. After TACE, the levels of hyaluronic acid (HA), procollagen III (PC III), laminin (LN), and collagen IV (IV-C) were measured, and the severity of liver fibrosis as well as the morphology of liver tissues were determined. Body mass in the model group was significantly decreased from 10 to 16 weeks, and the serum levels of ALT, AST, TP, TBIL, and DBIL levels were significantly increased while the model was being generated; the levels of ALB and A:G were significantly decreased. After TACE, serum levels of HA, PC III, and LN in the group injected with 1.0 mL iodized olive oil (Group B) were higher than in the group that were injected with 1.0 mL iodized olive oil + 0.2 mL PGE1 (Group C), whereas the serum levels of IV-C were lower. The severity of liver fibrosis was ameliorated in Group C. The combination of PGE1 and iodized olive oil prevented the development of liver fibrosis following TACE. [ABSTRACT FROM AUTHOR]

Basement membranes provide structural support and convey regulatory signals to cells in diverse tissues. Assembly of collagen IV into a sheet-like network is a fundamental mechanism during the formation of basement membranes. Peroxidasin (PXDN) was recently described to catalyze crosslinking of collagen IV through the formation of sulfilimine bonds. Despite the significance of this pathway in tissue genesis, our understanding of PXDN function is far from complete. In this work we demonstrate that collagen IV crosslinking is a physiological function of mammalian PXDN. Moreover, we carried out structure–function analysis of PXDN to gain a better insight into its role in collagen IV synthesis. We identify conserved cysteines in PXDN that mediate the oligomerization of the protein into a trimeric complex. We also demonstrate that oligomerization is not an absolute requirement for enzymatic activity, but optimal collagen IV coupling is only catalyzed by the PXDN trimers. Localization experiments of different PXDN mutants in two different cell models revealed that PXDN oligomers, but not monomers, adhere on the cell surface in “hot spots,” which represent previously unknown locations of collagen IV crosslinking. [ABSTRACT FROM AUTHOR]

The present study investigated whether Qianliening capsules (QC) affected the apoptosis of benign prostatic hyperplastia epithelial (BPH-1) cells by regulating the extracellular matrix (ECM). The levels of fibronectin (FN) and collagen IV were determined in the culture medium of BPH-1 cells maintained in normal medium and of BPH-1 cells maintained in an environment rich in FN and collagen IV using an enzyme-linked immunosorbent assay. Reverse transcription quantitative polymerase chain reaction and western blot analysis were performed to determine the mRNA and protein expression levels of FN, collagen IV, B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax) and cyclin D1, respectively. The cell morphology and viability were determined using light microscopy and an MTT assay and cell apoptosis was detected by annexin V staining. The results demonstrated that FN and collagen IV affected the apoptotic response of the BPH-1 cells, QC treatment significantly reduced the levels of FN and collagen IV secreted by the cells into the culture medium (P<0.01), inhibited the mRNA and protein expression levels of FN, collagen IV, Bcl-2 and cyclin D1 and promoted the mRNA and protein expression of Bax. Therefore, one of the mechanisms underlying the anti-BPH action of QC involves promoting apoptosis by regulating the expression of the extracellular matrix. [ABSTRACT FROM AUTHOR]

Breast cancer survival rates decrease from 99% for patients with local disease to 25% for those with distant metastases. Matrix metalloproteinases (MMPs), including MMP2, are associated with metastatic progression. We found that loss of host MMP2 reduces the proliferation of experimental metastases in the lungs and identified fibroblasts in tumour-bearing lungs as the major source of MMP2. In vitro, spheroidal mammary tumour growth was increased by co-culture with control fibroblasts isolated from tumour-bearing lungs, but not when fibroblasts with stable Mmp2 knockdown were used. This result prompted us to assess whether MMP2 was responsible for a tumour-proliferative, activated fibroblast phenotype. To test this, we evaluated: (a) fibroblasts from wild-type tumour-bearing lungs, with or without shRNA-mediated MMP2 knockdown; and (b) normal, quiescent fibroblasts isolated from either WT or Mmp2-/- mice. Quantitative PCR revealed that Mmp2 knockdown attenuated expression of two markers of activation (-smooth muscle actin and vimentin), but there was minimal expression in quiescent WT or Mmp2-/- fibroblasts, as expected. Placing quiescent fibroblasts under activating conditions led to increases in activation-associated transcripts in WT but not Mmp2-/- fibroblasts. Additionally, Mmp2 knockdown fibroblasts showed significantly decreased expression of the matrix transcripts collagen I, collagen IV and fibronectin. Addition of active TGF was sufficient to rescue the MMP2-dependent collagen I and IV expression, while MMP2-induced collagen expression was blocked by the addition of TGF1-neutralizing antibody. Gene expression data in stromal cells of human breast cancers reveal that MMP2 expression is also positively correlated with activation and matrix transcripts. Thus, we present a model whereby MMP2 production in tumour fibroblasts is important for TGF1 activity and subsequent activation of fibroblasts to a matrix-producing, proliferation-supportive phenotype. Overall, our results reveal a previously undefined role for MMP2 in metastatic outgrowth mediated by fibroblasts, and extend the mechanisms by which MMPs contribute to tumour progression. [ABSTRACT FROM AUTHOR]

Background: SPARC is a collagen-binding glycoprotein whose functions during early development are unknown. We previously reported that SPARC is expressed in Drosophila by hemocytes and the fat body (FB) and enriched in basal laminae (BL) surrounding tissues, including adipocytes. We sought to explore if SPARC is required for proper BL assembly in the FB. Results: SPARC deficiency leads to larval lethality, associated with remodeling of the FB. In the absence of SPARC, FB polygonal adipocytes assume a spherical morphology. Loss-of-function clonal analyses revealed a cell-autonomous accumulation of BL components around mutant cells that include collagen IV (Col lV), Laminin, and Perlecan. Ultrastructural analyses indicate SPARC-deficient adipocytes are surrounded by an aberrant accumulation of a fibrous extracellular matrix. Conclusions: Our data indicate a critical requirement for SPARC for the proper BL assembly in Drosophila FB. Since Col IV within the BL is a prime determinant of cell shape, the rounded appearance of SPARC-deficient adipocytes is due to aberrant assembly of Col IV. Developmental Dynamics 244:540-552, 2015. © 2014 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Diabetic nephropathy becomes the single most frequent cause of end-stage renal disease. The present study aimed therefore to investigate possible protective effect of eprosartan, an angiotensin II type 1 receptor blocker, on high-fat diet/streptozotocin-induced early diabetic nephropathy in rats and various mechanisms underlie this effect. Male Wistar rats with type 2 diabetes induced by high-fat diet/streptozotocin were treated with eprosartan at the dose levels of 30 and 60 mg/kg daily for 5 weeks. Eprosartan induced a nephroprotective effect as evident by the significant decrease in serum creatinine, urea, total cholesterol, triglycerides and glucose levels, urinary albumin excretion and kidney index as well as renal levels of malondialdehyde and nitric oxide products (nitrite/nitrate), in addition to angiotensin II, inducible nitric oxide synthase, transforming growth factor-β 1 and collagen IV expressions with a concurrent increase in renal levels of reduced glutathione and catalase activity compared to diabetic untreated rats. Histopathological examination confirmed the renoprotective effect of eprosartan. In conclusion, eprosartan protects rats against high-fat diet/streptozotocin-induced early diabetic nephropathy possibly, in part, through its antioxidant effect as well as by abrogating the overexpression of angiotensin II, inducible nitric oxide synthase, transforming growth factor-β 1 and collagen IV. [ABSTRACT FROM AUTHOR]

Background & Aims: The incidence of non-alcoholic fatty liver disease (NAFLD) is increasing worldwide. We evaluated serum collagen IV as a direct non-invasive marker of severe liver fibrosis in NAFLD. Methods: The study included 148 NAFLD and 187 chronic hepatitis C patients in whom histological severity of liver fibrosis was evaluated. The utility of serum collagen IV measured by immune-mediated agglutination using two types of monoclonal antibodies for distinguishing severe fibrosis (⩾ stage 3 and ⩾ F3) from nonto- moderate fibrosis in NAFLD or chronic hepatitis C was assessed in comparison to serum hyaluronic acid or other indirect fibrosis markers. Results: Multiple logistic regression analysis showed that serum collagen IV was significantly associated with severe fibrosis in NAFLD (odds ratio: 1.21, p<0.001) but not in chronic hepatitis C. For distinguishing severe fibrosis in NAFLD, collagen IV showed the largest area under the receiver-operating characteristic curve (0.827, 95%CI: 0.746-0.908) followed by FIB-4 (0.805, 95%CI: 0.728-0.890); in chronic hepatitis C, those for FIB-4 (0.813, 95%CI: 0.748-0.878) and collagen IV (0.770, 95%CI: 0.683-0.857) were the largest and smallest, respectively. To detect severe fibrosis in NAFLD, a cutoff of collagen IV ⩾ 177 exhibited 77.1% sensitivity, 84.0% specificity, 76.5% positive predictive value, and 84.0% negative predictive value. Combined with a cutoff of FIB-4 ⩾ 2.09, the negative and positive predictive values, and specificity for detecting severe fibrosis in NAFLD increased further. Conclusion: Collagen IV is a reliable marker for distinguishing severe liver fibrosis from non-to-moderate fibrosis in NAFLD but not chronic hepatitis C. [ABSTRACT FROM AUTHOR]

Aims Renal ischemia/reperfusion (I/R) injury is highly associated with morbidity and mortality. Oxidative stress, inflammation, and apoptosis play pivotal roles in the development of renal dysfunction following renal I/R. Experimental studies have reported the effectiveness of many antioxidant and anti-inflammatory compounds against renal I/R injury. On the other hand, açaí ( Euterpe oleracea Mart. Palmae, Arecaceae) has recently gained considerable appreciation as a natural source of antioxidants. However, the effect of açaí extract has not been studied before on renal I/R. Therefore, the present study was carried out to investigate the possible mechanisms of renal injury attenuation by açaí extract in a rat renal I/R model. Main methods To achieve the aim of the study, rats were administered açaí extract at two dose levels (500 and 1000 mg/kg) for 15 consecutive days before bilateral renal I/R induction. Serum and kidneys were isolated and used for subsequent biochemical analysis. Key findings The present data showed that açai extract significantly and dose-dependently attenuated I/R-induced renal damage. It suppressed the levels of blood urea nitrogen (BUN), serum creatinine, and renal tissue content of kidney injury molecule-1 (KIM-1). In addition, it inhibited serum lactate dehydrogenase (LDH) activity. Moreover, renal contents of malondialdehyde (MDA), myeloperoxidase (MPO), interferon-gamma (IFN-γ), caspase-3, collagen IV, and endothelin-1 were reduced, while renal interleukin-10 (IL-10) content was increased by açaí extract administration to rats before renal I/R induction. Significance Açaí extract ameliorated bilateral I/R-induced renal injury in rats in a dose-dependent manner. [ABSTRACT FROM AUTHOR]

Collagen IV has been described as a structural protein of the basement membrane, which as a whole forms a specialized extracellular matrix. Recent studies have indicated a possible relationship between collagen IV and the innate immune response of invertebrate organisms. The present study characterized the alpha-1 chain of collagen IV in the red abalone Haliotis rufescens (Hr-ColIV) and evaluated its association with the innate immune response against Vibrio anguillarum . To further evidence the immune response, the matrix metalloproteinase-1 (Hr-MMP-1) and C-type lectin (Hr-CLEC) genes were also assessed. The complete sequence of Hr-ColIV was composed of 6658 bp, with a 5′UTR of 154 bp, a 3′UTR of 1177 bp, and an ORF of 5327 bp that coded for 1776 amino acids. The innate immune response generated against V. anguillarum resulted in a significant increase in the transcript levels of Hr-ColIV between 3 and 6 hpi, whereas Hr-MMP-1 and Hr-CLEC had the highest transcript activity 6 and 12 hpi, respectively. The results obtained in this study propose a putative biological function for collagen IV involved in the early innate immune response of the red abalone H. rufescens . [ABSTRACT FROM AUTHOR]

Human auditory nerve afferents consist of two separate systems; one is represented by the large type I cells innervating the inner hair cells and the other one by the small type II cells innervating the outer hair cells. Type I spiral ganglion neurons (SGNs) constitute 96% of the afferent nerve population and, in contrast to other mammals, their soma and pre- and post-somatic segments are unmyelinated. Type II nerve soma and fibers are unmyelinated. Histopathology and clinical experience imply that human SGNs can persist electrically excitable without dendrites, thus lacking connection to the organ of Corti. The biological background to this phenomenon remains elusive. We analyzed the pre- and post-somatic segments of the type I human SGNs using immunohistochemistry and transmission electron microscopy (TEM) in normal and pathological conditions. These segments were found surrounded by non-myelinated Schwann cells (NMSCs) showing strong intracellular expression of laminin-β2/collagen IV. These cells also bordered the perikaryal entry zone and disclosed surface rugosities outlined by a folded basement membrane (BM) expressing laminin-β2 and collagen IV. It is presumed that human large SGNs are demarcated by three cell categories: (a) myelinated Schwann cells, (b) NMSCs and (c) satellite glial cells (SGCs). Their BMs express laminin-β2/collagen IV and reaches the BM of the sensory epithelium at the habenula perforata. We speculate that the NMSCs protect SGNs from further degeneration following dendrite loss. It may give further explanation why SGNs can persist as electrically excitable monopolar cells even after long-time deafness, a blessing for the deaf treated with cochlear implantation. [ABSTRACT FROM AUTHOR]

Background Although several approaches for identification and isolation of carcinoma cells with tumour initiating properties have been established, enrichment of a population of pure and viable tumour-initiating cells (TICs) is still problematic. This study investigated possibilities to isolate a population of cancer cells with tumour initiating properties based on their adherence properties, rather than expression of defined markers or clonogenicity. Methods Several human cell lines derived from oral dysplasia and oral squamous cell carcinoma (OSCC), as well as primary cells derived from patients with OSCC were allowed to adhere to collagen IV-coated dishes sequentially. Rapid adherent cells (RAC), middle adherent cells (MAC) and late adherent cells (LAC) were then harvested and further investigated for their morphology, stem cell-like properties and molecular profile while grown in vitro and tongue xenotransplantation in NOD-SCID mice at serial dilutions. Results RAC showed significantly higher colony forming efficiency ( p < 0.05), sphere forming ability, greater migration ability ( p < 0.05), exhibited longer G2 phase and displayed higher expression of integrin β1 and other stem-cell related molecules as compared to MAC and LAC. RAC induced tongue tumours in NOD-SCID mice with the highest incidence. These tumours were also bigger and metastasised more frequently in loco-regional lymph nodes than MAC and LAC. Conclusions These findings prove for the first time that OSCC cells with tumour initiating properties can be enriched based on their rapid adhesiveness to collagen IV. This separation procedure provides a potentially useful tool for isolating TICs in OSCC for further studies on understanding their characteristics and drug-resistant behaviour. [ABSTRACT FROM AUTHOR]

Multiple tumor exacerbations and treatment procedures, such as extracellular matrix remodeling, metabolic reprogramming, immunological evasion, and resistance to chemotherapy and radiotherapy, are influenced by intratumoral hypoxia. It is becoming increasingly clear how hypoxia interacts with the extracellular matrix and how this affects the growth of cancer. We analyzed the published sequencing results of hypoxia-stressed mouse kidney tumor cells and found that the expression of miR-29b was significantly downregulated. There are several sites that are complementary to the miR-29 seed sequence in the 3' non-coding regions (3'UTRs) of various extracellular matrix-related genes, including collagen IV. We analyzed the sequences of the 3'UTRs of different subunits of collagen IV in different species and constructed the corresponding phylogenetic trees. We found that the 3'UTRs of Col4a1 and Col4a4 may have been subjected to particular evolutionary pressures. By cloning the 3'UTRs of collagen IV subunits into the psiCHECK TM -2 vector, we found that seven of the eight sites in the Col4a3 - Col4a6 gene complementary to miR-29 were significantly repressed by miR-29a, b (except for the 7774-7781 of Col4a3 gene). The inhibitory efficiency of miR-29a, b on these seven sites was between 27% and 57%. The research on the regulation of miR-29 and extracellular matrix by hypoxia can provide a theoretical basis for tumor and fibrosis research and treatment.

Laparoscopy as a diagnostic tool for patients with suspected endometriosis is associated with several potentially life-threatening complications. Therefore, it is imperative to identify reliable, non-invasive biomarkers of the disease. The aim of this study was to analyse the concentrations of fibronectin and type IV collagen in peritoneal fluid and plasma to assess their role as potential biomarkers in the diagnosis of endometriosis. Fibronectin and collagen IV protein levels were assessed by surface plasmon resonance imaging (SPRi) biosensors with the usage of monoclonal antibodies. All patients enrolled in the study were referred for laparoscopy for the diagnosis of infertility or chronic pelvic pain (n = 84). The study group included patients with endometriosis confirmed during surgery (n = 49). The concentration of fibronectin in the plasma (329.3 ± 98.5 mg/L) and peritoneal fluid (26.8 ± 11.1 μg/L) in women with endometriosis was significantly higher than in the control group (251.2 ± 84.0 mg/L, 7.0 ± 5.9 μg/L). Fibronectin levels were independent of endometriosis stage ( p = 0.874, p = 0.469). No significant differences were observed in collagen IV levels ( p = 0.385, p = 0.465). The presence of elevated levels of fibronectin may indicate abnormalities in cell-ECM signalling during the course of endometriosis, and may be a potential biomarker for early detection.

Lichen sclerosus (LS) is an acquired chronic inflammatory dermatosis predominantly affecting the anogenital area with recalcitrant itching and soreness. Progressive or persistent LS may cause urinary and sexual disturbances and an increased risk of local skin malignancy with a prevalence of up to 11%. Investigations on lipoid proteinosis, an autosomal recessive genodermatosis caused by loss-of-function mutations in the extracellular matrix protein 1 ( ECM1 ) gene, led to the discovery of a humoral autoimmune response to the identical molecule in LS, providing evidence for an autoimmune and genetic counterpart targeting ECM1. This paper provides an overview of the fundamental importance and current issue of better understanding the immunopathology attributed to ECM1 in LS. Furthermore, we highlight the pleiotropic action of ECM1 in homeostatic and structural maintenance of skin biology as well as in a variety of human disorders possibly associated with impaired or gained ECM1 function, including the inflammatory bowel disease ulcerative colitis, Th2 cell-dependent airway allergies, T-cell and B-cell activation, and the demyelinating central nervous system disease multiple sclerosis, to facilitate sharing the concept as a plausible therapeutic target of this attractive molecule.

Context: Kirenol possesses anti-inflammatory, antifibrotic and anti-arthritic effects. However, its reno-protective effects against diabetic nephropathy (DN) have not been evaluated., Objective: This study explores the reno-protective effects of kirenol against DN and clarifies the potential mechanisms., Materials and Methods: The mesangial cells were treated with 20 µM kirenol and 10 ng/mL human recombinant TGF-β1 or 30 mM glucose for 24 h. Then the cells were harvested to assay the expression of the target genes or proteins. Thirty C57BL/6J male mice were given high-fat diet with streptozotocin injection to induce diabetes and then were randomized into three groups ( n  = 10): vehicle administration (DM group), 2 mg/kg kirenol (DM + kirenol group) and 200 mg/kg metformin (Met group) for 3 months, orally. A healthy group (Con, n  = 10) was included as the control., Results: Compared to the DM group, kirenol treatment decreased the phosphorylation of Smad2/3 and NF-κB (0.64- and 0.43-fold) as well as the accumulation of FN and Col IV (0.58- and 0.35-fold); moreover, the expression of IκBα was restored to normal level by kirenol treatment both in vivo and in vitro . After kirenol treatment, IL-6 expression was decreased 0.35- and 0.57-fold, and TNF-α expression was decreased 0.34- and 0.46-fold, in vitro and in vivo , respectively. Furthermore, kirenol alleviated the glomerular basement membrane thickness and foot process fusion., Discussion and Conclusions: Kirenol could alleviate DN by downregulating the TGF-β/Smads and the NF-κB signal pathway. Our study provides a potential mechanism for the treatment of DN with kirenol.

The basement membrane (BM) underlying epithelial tissue is a thin layer of extracellular matrix that governs tissue integrity and function. Epithelial BMs are generally assembled using BM components secreted from two origins: epithelium and stroma. Although de novo BM formation involves self-assembly processes of large proteins, it remains unclear how stroma-derived macromolecules are transported and assembled, specifically in the BM region. In this study, we established an in vitro co-culture model of BM formation in which DLD-1 human colon epithelial cells were cultured on top of collagen I gel containing human embryonic OUMS-36T-2 fibroblasts as stromal cells. A distinct feature of our system is represented by OUMS-36T-2 cells which are almost exclusively responsible for synthesis of collagen IV, a major BM component. Exploiting this advantage, we found that collagen IV incorporation was significantly impaired in culture conditions where OUMS-36T-2 cells were not allowed to directly contact DLD-1 cells. Soluble collagen IV, once diluted in the culture medium, did not accumulate in the BM region efficiently. Live imaging of fluorescently tagged collagen IV revealed that OUMS-36T-2 cells deposited collagen IV aggregates directly onto the basal surface of DLD-1 cells. Collectively, these results indicate a novel mode of collagen IV deposition in which fibroblasts proximal to epithelial cells exclusively contribute to collagen IV assembly during BM formation., (© 2022 Federation of European Biochemical Societies.)

All epithelia have their basal side in contact with a specialized extracellular matrix, the basement membrane (BM). During development, the BM contributes to the shaping of epithelial organs via its mechanical properties. These properties rely on two core components of the BM, collagen type IV and perlecan/HSPG2, which both interact with another core component, laminin, the initiator of BM assembly. While collagen type IV supplies the BM with rigidity to constrain the tissue, perlecan antagonizes this effect. Nevertheless, the number of organs that has been studied is still scarce, and given that epithelial tissues exhibit a wide array of shapes, their forms are bound to be regulated by distinct mechanisms. This is underscored by mounting evidence that BM composition and assembly/biogenesis is tissue-specific. Moreover, previous reports have essentially focused on the mechanical role of the BM in morphogenesis at the tissue scale, but not the cell scale. Here, we took advantage of the robust conservation of core BM proteins and the limited genetic redundancy of the Drosophila model system to address how this matrix shapes the wing imaginal disc, a complex organ comprising a squamous, a cuboidal and a columnar epithelium. With the use of a hypomorphic allele, we show that the depletion of Trol (Drosophila perlecan) affects the morphogenesis of the three epithelia, but particularly that of the squamous one. The planar surface of the squamous epithelium (SE) becomes extremely narrow, due to a function for Trol in the control of the squamous shape of its cells. Furthermore, we find that the lack of Trol impairs the biogenesis of the BM of the SE by modifying the structure of the collagen type IV lattice. Through atomic force microscopy and laser surgery, we demonstrate that Trol provides elasticity to the SE's BM, thereby regulating the mechanical properties of the SE. Moreover, we show that Trol acts via collagen type IV, since the global reduction in the trol mutant context of collagen type IV or the enzyme that cross-links its 7S -but not the enzyme that cross-links its NC1- domain substantially restores the morphogenesis of the SE. In addition, a stronger decrease in collagen type IV achieved by the overexpression of the matrix metalloprotease 2 exclusively in the BM of the SE, significantly rescues the organization of the two other epithelia. Our data thus sustain a model in which Trol counters the rigidity conveyed by collagen type IV to the BM of the SE, via the regulation of the NC1-dependant assembly of its scaffold, allowing the spreading of the squamous cells, spreading which is compulsory for the architecture of the whole organ., (Copyright © 2022 Elsevier B.V. All rights reserved.)

In glaucoma, astrocytes within the optic nerve head (ONH) rearrange their actin cytoskeleton, while becoming reactive and upregulating intermediate filament glial fibrillary acidic protein (GFAP). Increased transforming growth factor beta 2 (TGF β2) levels have been implicated in glaucomatous ONH dysfunction. A key limitation of using conventional 2D culture to study ONH astrocyte behavior is the inability to faithfully replicate the in vivo ONH microenvironment. Here, we engineer a 3D ONH astrocyte hydrogel to better mimic in vivo mouse ONH astrocyte (MONHA) morphology, and test induction of MONHA reactivity using TGF β2. Primary MONHAs were isolated from C57BL/6J mice and cell purity confirmed. To engineer 3D cell-laden hydrogels, MONHAs were mixed with photoactive extracellular matrix components (collagen type I, hyaluronic acid) and crosslinked for 5 minutes using a photoinitiator (0.025% riboflavin) and UV light (405-500 nm, 10.3 mW/cm2). MONHA-encapsulated hydrogels were cultured for 3 weeks, and then treated with TGF β2 (2.5, 5.0 or 10 ng/ml) for 7 days to assess for reactivity. Following encapsulation, MONHAs retained high cell viability in hydrogels and continued to proliferate over 4 weeks as determined by live/dead staining and MTS assays. Sholl analysis demonstrated that MONHAs within hydrogels developed increasing process complexity with increasing process length over time. Cell processes connected with neighboring cells, coinciding with Connexin43 expression within astrocytic processes. Treatment with TGF β2 induced reactivity in MONHA-encapsulated hydrogels as determined by altered F-actin cytoskeletal morphology, increased GFAP expression, and elevated fibronectin and collagen IV deposition. Our data sets the stage for future use of this 3D biomimetic ONH astrocyte-encapsulated hydrogel to investigate astrocyte behavior in response to injury. • Optic nerve head astrocytes (ONHA) retain viability and proliferate in photo-crosslinked biopolymer hydrogels. • The hydrogel system allows for astrocytes to achieve stellate morphology and develop process complexity. • TGFB2 induces astrogliosis in ONHA-hydrogels consistent with 2D models. • This novel 3D biomimetic hydrogel system can be used to further investigate ONHA behavior under glaucomatous insult. [ABSTRACT FROM AUTHOR]

Cerebral amyloid angiopathy or CAA is a type of vascular dementia that can cause neuroinflammation, ischemia and hemorrhage, among other complications. CAA results from the deposition of amyloid beta (Aβ) in blood vessels and is frequently observed in individuals with Alzheimer's disease (AD). One functional output of those pathological changes is measurable cognitive decline. Still not well understood, however, is the impact of gender or sex on the pathology of CAA, as well as CAA-induced cognitive decline. Here, we studied how sex impacts deposition of CAA-related pathology and the associated cognitive decline. We observed differential hippocampal pathology as far as regions of deposition, type of morphology, and total amount of pathology when assessing CAA pathology via (E,E)-1-fluoro-2,5-bis-(3-hydroxycarbonyl-4-hydroxy)styrylbenzene (FSB)-labeling, as well as neurodegeneration via Fluoro Jade C (FJC)-labeling, and lysosomal associated membrane protein deposition via LAMP-1 labeling. In accordance with other studies, our data suggest female TG-SwDI mice exhibit more severe pathological alterations in CAA pathology. Additionally, behavioral assessments revealed an impact of genotype that was more pronounced in TG-SwDI females. While the primary measure of learning and memory, the water maze, suggests an overall effect of genotype, effects in measures of locomotor activity and anxiety-like behavior suggest reduced habituation in females. This could be due to a lower retention for the tasks. Results of this study offer significant insight into the importance of examining effects of sex on CAA. • Sex-dependent response to cerebral amyloid angiopathy. • Hippocampal deposition of cerebral amyloid angiopathy. • Sex difference in Tg-SwDI mouse model. • Behavioral response to cerebral amyloid angiopathy. • Cerebral amyloid angiopathy causes cognitive deficits. [ABSTRACT FROM AUTHOR]

Glomerular diseases involving podocyte/glomerular epithelial cell (GEC) injury feature protein misfolding and endoplasmic reticulum (ER) stress. Inositol-requiring enzyme 1α (IRE1α) mediates chaperone production and autophagy during ER stress. We examined the role of IRE1α in selective autophagy of the ER (reticulophagy). Control and IRE1α knockout (KO) GECs were incubated with tunicamycin to induce ER stress and subjected to proteomic analysis. This showed IRE1α-dependent upregulation of secretory pathway mediators, including the coat protein complex II component Sec23B. Tunicamycin enhanced expression of Sec23B and the reticulophagy adaptor reticulon-3-long (RTN3L) in control, but not IRE1α KO GECs. Knockdown of Sec23B reduced autophagosome formation in response to ER stress. Tunicamycin stimulated colocalization of autophagosomes with Sec23B and RTN3L in an IRE1α-dependent manner. Similarly, during ER stress, glomerular α5 collagen IV colocalized with RTN3L and autophagosomes. Degradation of RTN3L and collagen IV increased in response to tunicamycin, and the turnover was blocked by deletion of IRE1α; thus, the IRE1α pathway promotes RTN3L-mediated reticulophagy and collagen IV may be an IRE1α-dependent reticulophagy substrate. In experimental glomerulonephritis, expression of Sec23B, RTN3L, and LC3-II increased in glomeruli of control mice, but not in podocyte-specific IRE1α KO littermates. In conclusion, during ER stress, IRE1α redirects a subset of Sec23B-positive vesicles to deliver RTN3L-coated ER fragments to autophagosomes. Reticulophagy is a novel outcome of the IRE1α pathway in podocytes and may play a cytoprotective role in glomerular diseases. • Production of endoplasmic reticulum vesicles is a non-canonical outcome IRE1α. • IRE1α promotes expression of Sec23B and the reticulophagy adaptor RTN3L. • IRE1α promotes reticulophagy during endoplasmic reticulum stress. • Glomerular type IV collagen may be a reticulophagy substrate in podocytes. [ABSTRACT FROM AUTHOR]

Objective: To investigate the effect of urokinase-type plasminogen activator (uPA) on mesangial matrix in the kidney of diabetic rats and its related mechanisms. Methods: Diabetic Sprague-Dawley (SD) rats induced by intraperitoneal injection of streptozotocin (STZ) were randomly and evenly divided into two groups: DM + vehicle, and DM + uPA (2500 U kg−1 uPA via tail vein once a day for four weeks). The normal SD rats without diabetes were considered as control group. Rats in the three groups were executed and the heart blood was sampled for determination of blood glucose and serum creatinine. Meanwhile, kidney tissues of rats were also harvest for measurement of glomerular area, volume, and mesangial area by periodic acid silver methenamine (PASA) staining. The expression of urokinase-type plasminogen activator receptor (uPAR), plasminogen activator inhibitor-1 (PAI-1), and collagen IV in renal tissues was tested with immunohistochemistry. Results: Compared with control, the DM rats had obvious albuminuria, significantly ( p < 0.01) increased glomerular volume and mesangial matrix area, and significantly ( p < 0.05) higher expression of uPAR, PAI-1 and collagen IV in mesangial matrix, significantly up-regulated ( p < 0.05) glomerular uPAR, PAI-1, and collagen IV expression. After treated with uPA, the diabetic rats had significantly ( p < 0.05) reduced albuminuria, significantly ( p < 0.01) improved glomerular volume and mesangial matrix, significantly ( p < 0.05) down-regulated PAI-1 and collagen IV expression in mesangial matrix. However, the uPAR expression in renal tissues were unchangeable ( p > 0.05) and PAI-1 and collagen IV expression were significantly ( p < 0.05) reduced when diabetic rats were treated with uPA. Conclusion: uPA can down-regulate glomerular PAI-1 expression in the DM rats but not significantly influence uPAR expression, suggesting that uPA might regulate the mesangial cell (MC) and its matrix expression and improve diseased diabetic mesangial matrix via its combination with uPAR to uptake PAI-1 and accelerate its degradation. [ABSTRACT FROM AUTHOR]

Purpose To detect by immunohistochemical means the changes in bleb capsules at the cellular level between functioning and non-functioning glaucoma drainage implants ( GDIs). Methods Three samples each of functioning (1 Baerveldt and 2 Molteno implants) and non-functioning filtration blebs (1 Molteno and 2 Ahmed implants) were studied. A non-functioning bleb was defined as an intra-ocular pressure ( IOP) >21 mmHg or a <20% reduction in IOP from baseline on three consecutive follow-up visits with maximal tolerated medication. The capsules were obtained between 6 and 108 months after GDI insertion for medical reasons only. Primary antibodies were used to stain fibronectin, tenascin, laminin, collagen IV and smooth muscle actin ( SMA). The samples were graded on the basis of the intensity and quantity of immunohistochemical staining into four categories as follows: no staining or a mild, moderate or marked staining. Results The non-functioning blebs expressed more fibronectin, tenascin and SMA through the whole capsule wall than the functioning blebs. In the functioning blebs, tenascin was found mainly in the inner layer of the capsule. More type IV collagen and laminin were also found in the non-functioning bleb capsules than in the functioning blebs. No difference was found between the bleb capsules irrespective of whether they had been perfused with aqueous humour immediately after surgery ( Ahmed) or after a delay ( Molteno, Baerveldt). Conclusion Accumulation of extracellular matrix components and activated fibroblasts in the bleb capsules of non-functioning GDI indicates the presence of an active wound healing process, suggesting a possible reduction in filtration through the bleb wall. [ABSTRACT FROM AUTHOR]

Background The majority of sera from patients with primary membranous nephropathy have autoantibodies against the M-type phospholipase A2 receptor ( PLA2 R) which is expressed on human podocytes. The rabbit variant of PLA2 R attaches to collagen type IV via the fibronectin type II domain, which is also present in the human variant of PLA2 R. Design To assess whether the human PLA2R variant is also involved in attachment to collagen type IV, we conducted a cell adhesion assay on a collagen-coated surface using PLA2 R-transfected and mock-transfected human embryonic kidney ( HEK) cells. To test the hypothesis that sera from patients containing anti- PLA2 R antibodies interfere with the adhesion of podocytes to collagen, we performed cell adhesion assays on a collagen type IV-coated surface using positive and negative serum samples from patients and cultured human podocytes in vitro expressing PLA2 R. Results The HEK cell adhesion assay confirmed an enhanced attachment of PLA2R-transfected cells to collagen type IV. We confirmed diminished podocyte adhesion in the presence of serum with anti- PLA2 R antibodies. The concentration of anti- PLA2 R antibodies correlated with proteinuria and to the degree of diminished adhesion of podocytes. Conclusions We demonstrated that serum of patients containing autoantibodies directed to PLA2 R interferes with the ability of podocytes to attach to collagen type IV in vitro, providing evidence of a serum soluble pathogenic factor interfering with podocyte adhesion in membranous nephropathy. [ABSTRACT FROM AUTHOR]

The basement membranes (BMs) of the nervous system include (a) the pial BM that surrounds the entire CNS, (b) the BMs that outline the vascular system of the CNS and PNS and (c) the BMs that are associated with Schwann cells. We previously found that isolated BMs are bi-functionally organized, whereby the two surfaces have different compositional, biomechanical and cell adhesion properties. To find out whether the bi-functional nature of BMs has an instructive function in organizing the tissue architecture of the developing nervous system, segments of human BMs were inserted into (a) the parasomitic mesoderm of chick embryos, intersecting with the pathways of axons and neural crest cells, or (b) into the midline of the embryonic chick spinal cord. The implanted BMs integrated into the embryonic tissues within 24 h and were impenetrable to growing axons and migrating neural crests cells. Host axons and neural crest cells contacted the epithelial side but avoided the stromal side of the implanted BM. When the BMs were inserted into the spinal cord, neurons, glia cells and axons assembled at the epithelial side of the implanted BMs, while a connective tissue layer formed at the stromal side, resembling the tissue architecture of the spinal cord at the pial surface. Since the spinal cord is a-vascular at the time of BM implantation, we propose that the bi-functional nature of BMs has the function of segregating epithelial and connective cells into two adjacent compartments and participates in establishing the tissue architecture at the pial surface of the CNS. [ABSTRACT FROM AUTHOR]

Highlights: [•] The optimal pH of cathepsin B activity for degrading collagen IV is 4. [•] The lower stability of triple helical conformation increases cathepsin B activity. [•] The endo- and exopeptidase activity participates in degrading collagen IV at pH 4. [•] The affinity of cathepsin B to collagen IV at pH 4 is higher than that at pH 5. [ABSTRACT FROM AUTHOR]

Background Gallbladder carcinoma (GBC) is highly lethal, and effective treatment will require synergistic anti-tumor management. The study is aimed at investigating the oncolytic value of myxoma virus (MYXV) infection against GBC and optimizing MYXV oncolytic efficiency. Methods We examined the permissiveness of GBC cell lines to MYXV infection and compared the effects of MYXV on cell viability among GBC and control permissive glioma cells in vitro and in vivo after MYXV + rapamycin (Rap) treatment, which is known to enhance cell permissiveness to MYXV by upregulating p-Akt levels. We also assessed MYXV + hyaluronan (HA) therapy efficiency by examinating Akt activation status, MMP-9 expression, cell viability, and collagen distribution. We further compared hydraulic conductivity, tumor area, and survival of tumor-bearing mice between the MYXV + Rap and MYXV + HA therapeutic regimens. Results MYXV + Rap treatment could considerably increase the oncolytic ability of MYXV against GBC cell lines in vitro but not against GBC xenografts in vivo. We found higher levels of collagen IV in GBC tumors than in glioma tumors. Diffusion analysis demonstrated that collagen IV could physically hinder MYXV intratumoral distribution. HA-CD44 interplay was found to activate the Akt signaling pathway, which increases oncolytic rates. HA was also found to enhance the MMP-9 secretion, which contributes to collagen IV degradation. Conclusions Unlike MYXV + Rap, MYXV + HA therapy significantly enhanced the anti-tumor effects of MYXV in vivo and prolonged survival of GBC tumor-bearing mice. HA may optimize the oncolytic effects of MYXV on GBC via the HA-CD44 interaction which can promote viral infection and diffusion. [ABSTRACT FROM AUTHOR]

Background Bronchial asthma often starts in early childhood. Clinical manifestation of the disease is likely due to inflammatory processes in the airways initiated by various stimuli. Developed remodelling is regularly observed in the bronchial mucosa of adult asthmatics but we still lack information about its onset and latter development with the natural course of the disease. In this study, we analysed histological findings in bronchial biopsies obtained from very young children (under 4 yr of age). We hypothesized that initial undetectable changes in the airway epithelium of children predisposed to asthma may be one of the first mechanisms leading to morphological changes in the bronchial mucosa. Methods We measured the thickness of the basement membrane using a light microscope and analysed the presence of its three basic structural glycoproteins: laminin, tenascin and collagen IV, using immunohistochemical techniques. We compared these findings in children predisposed to asthma according to the selected clinical criteria of the Asthma Predictive Index and in a control group of children. Results We found a significant difference in the thickness of the basement membrane between the two groups. We also found a difference in the subepithelial deposition of laminin and collagen IV in the basement membrane but no difference in the deposition of tenascin. Conclusions We conclude that initial changes leading to further remodelling may start at a very early age even before clinical manifestation of the disease. [ABSTRACT FROM AUTHOR]

Aims/hypothesis: Increased extracellular matrix (ECM) collagen is a characteristic of muscle insulin resistance. Matrix metalloproteinase (MMP) 9 is a primary enzyme that degrades collagen IV (ColIV). As a component of the basement membrane, ColIV plays a key role in ECM remodelling. We tested the hypotheses that genetic deletion of MMP9 in mice increases muscle ColIV, induces insulin resistance in lean mice and worsens diet-induced muscle insulin resistance. Methods: Wild-type ( Mmp9) and Mmp9-null ( Mmp9) mice were chow or high-fat (HF) fed for 16 weeks. Insulin action was measured by the hyperinsulinaemic-euglycaemic clamp in conscious weight-matched surgically catheterised mice. Results: Mmp9 and HF feeding independently increased muscle ColIV. ColIV in HF-fed Mmp9 mice was further increased. Mmp9 did not affect fasting insulin or glucose in chow- or HF-fed mice. The glucose infusion rate (GIR), endogenous glucose appearance (EndoRa) and glucose disappearance (Rd) rates, and a muscle glucose metabolic index (Rg), were the same in chow-fed Mmp9 and Mmp9 mice. In contrast, HF-fed Mmp9 mice had decreased GIR, insulin-stimulated increase in Rd and muscle Rg. Insulin-stimulated suppression of EndoRa, however, remained the same in HF-fed Mmp9 and Mmp9 mice. Decreased muscle Rg in HF-fed Mmp9 was associated with decreased muscle capillaries. Conclusions/interpretation: Despite increased muscle ColIV, genetic deletion of MMP9 does not induce insulin resistance in lean mice. In contrast, this deletion results in a more profound state of insulin resistance, specifically in the skeletal muscle of HF-fed mice. These results highlight the importance of ECM remodelling in determining muscle insulin resistance in the presence of HF diet. [ABSTRACT FROM AUTHOR]