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Hereditary cystatin C amyloid angiopathy (HCCAA) is an Icelandic disease that belongs to a disease class called cerebral amyloid angiopathy, a group of heterogenous diseases presenting with aggregation of amyloid complexes and deposition predominantly in the central nervous system. HCCAA is dominantly inherited, caused by L68Q mutation in the cystatin C gene, leading to aggregation of the cystatin C protein. HCCAA is a very progressive and severe disease, with widespread cerebral and parenchymal cystatin C and collagen IV deposition within the central nervous system (CNS) but also in other organs in the body, for example, in the skin. Most L68Q carriers have clinical symptoms characterized by recurrent hemorrhages and dementia, between the age of 20–30 years. If the carriers survive the first hemorrhage, the frequency and severity of the hemorrhages tend to increase, resulting in death at average of 30 years with mean number of major hemorrhages ranging from 3.2 to 3.9 over a 5‐year average life span. The pathogenesis of the disease in carriers is very similar in the CNS and in the skin based on autopsy studies, thus skin biopsies can be used to monitor the progression of the disease by quantifying the cystatin C immunoreactivity. The cystatin C deposition always colocalizes with collagen IV and fibroblasts in the skin are found to be the main cell type responsible for the deposition of both proteins. No therapy is available for this devastating disease. [ABSTRACT FROM AUTHOR]

BACKGROUND: Type IV collagen is an abundant component of basement membranes in all multicellular species and is essential for the extracellular scaffold supporting tissue architecture and function. Lower organisms typically have two type IV collagen genes, encoding α1 and α2 chains, in contrast with the six genes in humans, encoding α1-α6 chains. The α chains assemble into trimeric protomers, the building blocks of the type IV collagen network. The detailed evolutionary conservation of type IV collagen network remains to be studied. RESULTS: We report on the molecular evolution of type IV collagen genes. The zebrafish α4 non-collagenous (NC1) domain, in contrast with its human ortholog, contains an additional cysteine residue and lacks the M93 and K211 residues involved in sulfilimine bond formation between adjacent protomers. This may alter α4 chain interactions with other α chains, as supported by temporal and anatomic expression patterns of collagen IV chains during the zebrafish development. Despite the divergence between zebrafish and human α3 NC1 domain (endogenous angiogenesis inhibitor, Tumstatin), the zebrafish α3 NC1 domain exhibits conserved antiangiogenic activity in human endothelial cells. CONCLUSIONS: Our work supports type IV collagen is largely conserved between zebrafish and humans, with a possible difference involving the α4 chain.

Abstract Riemerella anatipestifer infection is characterized by meningitis with neurological symptoms in ducklings and has adversely affected the poultry industry. R. anatipestifer strains can invade the duck brain to cause meningitis and neurological symptoms, but the underlying mechanism remains unknown. In this study, we showed that obvious clinical symptoms, an increase in blood‒brain barrier (BBB) permeability, and the accumulation of inflammatory cytokines occurred after intravenous infection with the Yb2 strain but not the mutant strain Yb2ΔsspA, indicating that Yb2 infection can lead to cerebrovascular dysfunction and that the type IX secretion system (T9SS) effector SspA plays a critical role in this pathological process. In addition, we showed that Yb2 infection led to rapid degradation of occludin (a tight junction protein) and collagen IV (a basement membrane protein), which contributed to endothelial barrier disruption. The interaction between SspA and occludin was confirmed by coimmunoprecipitation. Furthermore, we found that SspA was the main enzyme mediating occludin and collagen IV degradation. These data indicate that R. anatipestifer SspA mediates occludin and collagen IV degradation, which functions in BBB disruption in R. anatipestifer-infected ducks. These findings establish the molecular mechanisms by which R. anatipestifer targets duckling endothelial cell junctions and provide new perspectives for the treatment and prevention of R. anatipestifer infection.

Simple Summary: Basement membranes (BMs) are thin sheets of connective tissue that form boundary layers in the organs of most animal bodies. There is increasing interest in how changes in the strength and stiffness of BMs contribute to the normal processes by which fertilized eggs become mature adults and the abnormal processes associated with disease, such as the spread of cancerous tumors. The spread of cancer can be helped by the weakening and disruption of BMs, which is thought to result mainly from BM components being broken down by specific enzymes, although other factors may be involved. Brittlestars are marine invertebrate animals related to starfish and sea urchins. The BMs of their muscles act as tendons that link the muscles to the skeleton. Brittlestars are able to jettison their arms when they are attacked by predators. Such arm loss depends partly on the ability of muscles to detach from the skeleton due to the sudden weakening of their tendons. This contribution provides an overview of current knowledge of the structure and mechanical behavior of BMs in non-echinoderm animals and of brittlestar tendons and discusses the possible usefulness of brittlestar tendons as a model for understanding mechanisms of BM weakening in normal and disease-related processes. Basement membranes (BMs) are thin layers of extracellular matrix that separate epithelia, endothelia, muscle cells, and nerve cells from adjacent interstitial connective tissue. BMs are ubiquitous in almost all multicellular animals, and their composition is highly conserved across the Metazoa. There is increasing interest in the mechanical functioning of BMs, including the involvement of altered BM stiffness in development and pathology, particularly cancer metastasis, which can be facilitated by BM destabilization. Such BM weakening has been assumed to occur primarily through enzymatic degradation by matrix metalloproteinases. However, emerging evidence indicates that non-enzymatic mechanisms may also contribute. In brittlestars (Echinodermata, Ophiuroidea), the tendons linking the musculature to the endoskeleton consist of extensions of muscle cell BMs. During the process of brittlestar autotomy, in which arms are detached for the purpose of self-defense, muscles break away from the endoskeleton as a consequence of the rapid destabilization and rupture of their BM-derived tendons. This contribution provides a broad overview of current knowledge of the structural organization and biomechanics of non-echinoderm BMs, compares this with the equivalent information on brittlestar tendons, and discusses the possible relationship between the weakening phenomena exhibited by BMs and brittlestar tendons, and the potential translational value of the latter as a model system of BM destabilization. [ABSTRACT FROM AUTHOR]

Alport syndrome is a genetic kidney disease that causes worsening of kidney function over time, often progressing to kidney failure. Some types of Alport syndrome cause other symptoms and signs, including hearing loss and eye abnormalities. Research now indicates that Alport syndrome (autosomal dominant inheritance) is the most common form. Alport syndrome can have X-linked or a rare form of autosomal recessive inheritance. Traditionally, a kidney biopsy was used to diagnose Alport syndrome, but genetic testing provides a more precise and less invasive means of diagnosis and reveals the underlying pattern of inheritance. At present, there are no specific curative treatments for Alport syndrome however there is a strong international effort in pursuit of future therapies. Currently, angiotensin-converting enzyme inhibitors (ACEi), or an angiotensin receptor blocker (ARB) if a patient cannot tolerate an ACEi, slow down the progression of kidney disease and can delay the onset of kidney failure by years. There are other potential treatments in research that potentially can help delay the onset of kidney issues. Early treatment of patients and identification of their at-risk relatives is a priority. People living with Alport syndrome and their doctors now benefit from an active international research community working on translating further treatments into clinical practice and providing up-to-date clinical guidelines. [ABSTRACT FROM AUTHOR]

Background: In recent years, anti-angiogenic peptides have received considerable attention as candidates for cancer treatment. Arresten is an angiogenesis inhibitor that cleaves from the α1 chain of type IV collagen and stimulates apoptosis in endothelial cells. We have recently indicated that a peptide corresponding to the amino acid 78 to 86 of arresten, so-called Ars, prevented the migration and tube formation of HUVECs and the colon carcinoma growth in mice significantly. The current study aimed to determine whether induction of apoptotic cell death in endothelial cells is one of the biochemical mechanisms of this anti-angiogenic peptide. Methods and results: This hypothesis was assessed using the MTT assay, cell cycle analysis, Annexin V-FITC/PI staining, BCL2, CASP8, CASP9, p53, and CDKN2A gene expression studies as well as evaluating apoptosis in tumor tissues by TUNEL assay. Results demonstrated that 40 µM of Ars significantly stimulated 46.2% of early and late apoptosis in HUVECs compared to 13.6% in the untreated cells and did not significantly alter the cell cycle distribution. Moreover, BCL2 and CASP8 were down-regulated, while CASP9 and p53 were up-regulated in endothelial cells. CDKN2A gene expression, the regulator of G1 cell cycle arrest, was not significantly altered. Conclusions: It might be suggested that Ars induced apoptosis in endothelial cells through the mitochondrial pathway and had no effect on the cell cycle. Besides, Ars induced apoptosis significantly in vivo. However, further studies are required to confirm the detailed molecular mechanism of Ars, this peptide has the potential to be optimized for clinical translations. [ABSTRACT FROM AUTHOR]

Basement membranes are among the most widespread, non‐cellular functional materials in metazoan organisms. Despite this ubiquity, the links between their compositional and biophysical properties are often difficult to establish due to their thin and delicate nature. In this article, we examine these features on a molecular level by combining results from proteomics, elastic, and nanomechanical analyses across a selection of human basement membranes. Comparing results between these different membranes connects certain compositional attributes to distinct nanomechanical signatures and further demonstrates to what extent water defines these properties. In all, these data underline BMs as stiff yet highly elastic connective tissue layers and highlight how the interplay between composition, mechanics and hydration yields such exceptionally adaptable materials. [ABSTRACT FROM AUTHOR]

The aim of this research was to select the fragments that make up the outer layer of the collagen IV (COL4A6) protein and to assess their potential usefulness for regenerative medicine. It was expected that because protein–protein interactions take place via contact between external domains, the set of peptides forming the outer sphere of collagen IV will determine its interaction with other proteins. Cellulose‐immobilized protein fragment libraries treated with polyclonal anti‐collagen IV antibodies were used to select the peptides forming the outer sphere of collagen IV. In the first test, 33 peptides that strongly interacted with the polyclonal anti‐collagen IV antibodies were selected from a library of non‐overlapping fragments of collagen IV. The selected fragments of collagen IV (cleaved from the cellulose matrix) were tested for their cytotoxicity, their effects on cell viability and proliferation, and their impact on the formation of reactive oxygen species (ROS). The studies used RAW 264.7 mouse macrophage cells and Hs 680.Tr human fibroblasts. PrestoBlue, ToxiLight™, and ToxiLight 100% Lysis Control assays were conducted. The viability of fibroblasts cultured with the addition of increasing concentrations of the peptide mix did not show statistically significant differences from the control. Fragments 161–170, 221–230, 721–730, 1331–1340, 1521–1530, and 1661–1670 of COL4A6 were examined for cytotoxicity against BJ normal human foreskin fibroblasts. None of the collagen fragments were found to be cytotoxic. Further research is underway on the potential uses of collagen IV fragments in regenerative medicine. [ABSTRACT FROM AUTHOR]

To explore the renal protection mechanism of targeted regulating of transforming growth factor beta receptor type 1 expression by let-7a in diabetic nephropathy rats, diabetic nephropathy rats were created through high-sugar and high-fat diet combined with a small dose of streptozotocin. Normally reared rats were taken as control group. Diabetic nephropathy rats were divided into two groups, one group was treated with let-7a analogs group, and the other group was not treated (diabetic nephropathy group). Experimental results showed that diabetic nephropathy rats had symptoms such as increased dietary frequency, weight loss, proteinuria, and decreased creatinine clearance. After let-7a treatment, the proteinuria symptoms of rats in let-7a analogs group were notably improved, and the creatinine clearance rate increased. The expression level of collagen IV in let-7a analogs group was higher than that of the control, but the difference was not significant. The expression level of fibronectin increased significantly in the diabetic nephropathy group (p<0.05), and that of the let-7a analogs group was significantly lower than that of the diabetic nephropathy group (p<0.05). The immunofluorescence expression result was consistent with the expression trend detected by Western blot. Compared with the control, the expressions of transforming growth factor-beta 1, transforming growth factor beta receptor type 1, and mothers against decapentaplegic homolog 2 in the diabetic nephropathy group were evidently increased (p<0.05), and the expression of mothers against decapentaplegic homolog 7 decreased obviously (p<0.05). The protein expression level was consistent with the messenger ribonucleic acid expression result. In summary, by regulating the levels of collagen IV and fibronectin and targeting the transforming growth factor beta receptor type 1 pathway, let-7a inhibited extracellular matrix synthesis to protect kidney function in patients with diabetic nephropathy. [ABSTRACT FROM AUTHOR]

Peroxidasin (PXDN) is an extracellular peroxidase, which generates hypobromous acid to form sulfilimine cross-links within collagen IV networks. We have previously demonstrated that mouse and human renal basement membranes (BM) are enriched in bromine due to PXDN-dependent post-translational bromination of protein tyrosine residues. The goal of the present study was identification of specific brominated sites within renal BM. A comprehensive analysis of brominated proteome of mouse glomerular matrix had been performed using liquid chromatography-tandem mass spectrometry. We found that out of over 200 identified proteins, only three were detectably brominated, each containing a single distinct brominated tyrosine site i.e., Tyr-1485 in collagen IV α2 chain, Tyr-292 in TINAGL1 and Tyr-664 in nidogen-2. To explain this highly selective bromination, we proposed that these proteins interact with PXDN within the glomerular matrix. Experiments using purified proteins demonstrated that both TINAGL1 and nidogen-2 can compete with PXDN for binding to collagen IV and that TINAGL1 can directly interact with PXDN. We propose that a protein complex, including PXDN, TINAGL1, nidogen-2 and collagen IV, may exist in renal BM. • Hypobromous acid produced by peroxidasin (PXDN) can brominate bystander proteins in renal extracellular matrix (ECM). • We identified specific sites of PXDN-dependent bromination in mouse renal ECM using mass spectrometry. • Bromination was detected at a single tyrosine site in each of the three ECM proteins: α2 (IV) chain, TINAGL1 and nidogen-2. • This highly specific bromination consistent with interactions of the three proteins with PXDN within renal ECM. [ABSTRACT FROM AUTHOR]

Immune response to several kidney self-antigens (KSAg) such as Collagen IV (Col-IV), Perlecan (PL), and Fibronectin (FN) have been associated with antibody-mediated damage and poor allograft survival. Thus, the aim of this study was to determine if humoral immune responses to KSAg correlates with progression of chronic immune injury (CII) changes at 1 year or 2 years. Kidney transplant recipients who underwent 1- or 2-year biopsies, with chronic interstitial inflammation (ci > 1) and/or glomerular membrane double contouring (cg > 0) were analyzed with matched controls. Sera were analyzed retrospectively for antibodies against KSAg using ELISA. The presence of antibodies to KSAg were compared at 0, 4, 12, and 24 months using logistic regression. We identified a cohort of 214 kidney transplant recipients. Of these, we identified 33 cases and matched 66 controls. Logistical regression showed an odds ratio of 1 with the confidence interval crossing 1 for the presence of response to KSAg at all the time points. Humoral immune responses to either KSAg alone or in combination with donor-specific anti-HLA antibodies are not associated with progression to CII at 1 and 2 years after kidney transplantation. [ABSTRACT FROM AUTHOR]

Pierson syndrome and X-linked Alport syndrome result from pathogenic variants in LAMB2 and COL4A5, respectively, and both affect basement membranes in the kidney and the eye. This study describes the ocular features in an individual with a homozygous LAMB2 pathogenic variant and compares the reported abnormalities in Pierson syndrome with those in Alport syndrome. A 28-year-old man who developed kidney failure 10 years previously and subsequently had an atrial septal defect repair was suspected of having genetic kidney disease on the basis of his likely diagnosis of Focal and Segmental Glomerulosclerosis (FSGS), his young age at presentation, and his cardiac anomaly. He then underwent Whole Exome Sequencing and a formal ophthalmological examination. The patient was found to have a homozygous Likely Pathogenic missense variant (p.(Arg1719Cys)) in LAMB2 consistent with the diagnosis of Pierson syndrome. He had normal visual acuity, normal optic globe and cornea size, and normal lens appearance on direct examination. Upon further testing, his cornea demonstrated central thinning. There was also increased corneal endothelial pleomorphism, a reduced foveal reflex, and a blunted foveal curvature, similar to the features seen in X-linked Alport syndrome. Our patient had a later onset form of Pierson syndrome or "FSGS type 5, with or without ocular abnormalities," consistent with his "milder" LAMB2 missense variant. The resemblance of the ocular features in Pierson syndrome and X-linked Alport syndrome suggests that mutations in LAMB2 and COL4A5 have similar effects on basement membranes and the pathogenesis of ocular damage. [ABSTRACT FROM AUTHOR]

The crystalline lens is an important structure in the eye that starts to develop as early as the 22nd day of gestation, with further differentiation that continues after the induction. Congenital anomalies of the lens may involve the size, shape, and position of the lens. They may sometimes be associated with anterior segment dysgenesis or persistence of the tunica vasculosa lentis and hyperplastic vitreous and hyaloid system. Manifestations of anomalies of the lens shape are usually seen in early or late childhood however may sometimes be delayed into adulthood based on the level of visual impairment or the presence or absence of any syndromic associations. While lens coloboma has more often been reported in isolation, the more commonly implicated genes include the PAX6 gene, lenticonus in particular anterior is often part of Alport syndrome with extra-ocular manifestations in the kidneys and hearing abnormalities due to mutations in the alpha 5 chain of the Type IV collagen gene. Recognition of these manifestations and obtaining a genetic diagnosis is an important step in the management. The level of visual impairment and amblyopia dictates the outcomes in patients managed either conservatively with optical correction as well as surgically where deemed necessary. This review discusses the various anomalies of the lens shape with its related genetics and the management involved in these conditions. [ABSTRACT FROM AUTHOR]

Basement membranes (BMs) are thin layers of extracellular matrix that separate epithelia, endothelia, muscle cells, and nerve cells from adjacent interstitial connective tissue. BMs are ubiquitous in almost all multicellular animals, and their composition is highly conserved across the Metazoa. There is increasing interest in the mechanical functioning of BMs, including the involvement of altered BM stiffness in development and pathology, particularly cancer metastasis, which can be facilitated by BM destabilization. Such BM weakening has been assumed to occur primarily through enzymatic degradation by matrix metalloproteinases. However, emerging evidence indicates that non-enzymatic mechanisms may also contribute. In brittlestars (Echinodermata, Ophiuroidea), the tendons linking the musculature to the endoskeleton consist of extensions of muscle cell BMs. During the process of brittlestar autotomy, in which arms are detached for the purpose of self-defense, muscles break away from the endoskeleton as a consequence of the rapid destabilization and rupture of their BM-derived tendons. This contribution provides a broad overview of current knowledge of the structural organization and biomechanics of non-echinoderm BMs, compares this with the equivalent information on brittlestar tendons, and discusses the possible relationship between the weakening phenomena exhibited by BMs and brittlestar tendons, and the potential translational value of the latter as a model system of BM destabilization.

Mouse models of hyper- and hypothyroidism were used to examine the effects of thyroid hormone (TH) dyshomeostasis on the aging mammalian brain. 13–14 month-old mice were treated for 4 months with either levothyroxine (hyperthyroid) or a propylthiouracil and methimazole combination (PTU/Met; hypothyroid). Hyperthyroid mice performed better on Morris Water Maze than control mice, while hypothyroid mice performed worse. Brain weight was increased in thyroxine-treated, and decreased in PTU/Met-treated animals. The brain weight change was strongly correlated with circulating and tissue T4. Quantitative measurements of microvessels were compared using digital neuropathologic methods. There was an increase in microvessel area in hyperthyroid mice. Hypothyroid mice showed a trend for elevated glial fibrillary acidic protein-immunoreactive astrocytes, indicating an increase in neuroinflammation. Gene expression alterations were associated with TH perturbation and astrocyte-expressed transcripts were particularly affected. For example, expression of Gli2 and Gli3 , mediators in the Sonic Hedgehog signaling pathway, were strongly impacted by both treatments. We conclude that TH perturbations produce robust neurobehavioral, pathological, and brain gene expression changes in aging mouse models. [ABSTRACT FROM AUTHOR]

The crystalline lens is an important structure in the eye that starts to develop as early as the 22nd day of gestation, with further differentiation that continues after the induction. Congenital anomalies of the lens may involve the size, shape, and position of the lens. They may sometimes be associated with anterior segment dysgenesis or persistence of the tunica vasculosa lentis and hyperplastic vitreous and hyaloid system. Manifestations of anomalies of the lens shape are usually seen in early or late childhood however may sometimes be delayed into adulthood based on the level of visual impairment or the presence or absence of any syndromic associations. While lens coloboma has more often been reported in isolation, the more commonly implicated genes include the PAX6 gene, lenticonus in particular anterior is often part of Alport syndrome with extra-ocular manifestations in the kidneys and hearing abnormalities due to mutations in the alpha 5 chain of the Type IV collagen gene. Recognition of these manifestations and obtaining a genetic diagnosis is an important step in the management. The level of visual impairment and amblyopia dictates the outcomes in patients managed either conservatively with optical correction as well as surgically where deemed necessary. This review discusses the various anomalies of the lens shape with its related genetics and the management involved in these conditions.

Abstract Diabetic nephropathy (DN) is a prevalent diabetic microvascular condition. It is the leading cause of kidney disease in the advanced stages. There is no currently effective treatment available. This research aimed to investigate the curative potentials of exosomes isolated from mesenchymal stem cells affecting DN. This study was performed on 70 male adult albino rats. Adult rats were randomized into seven groups: Group I: Negative control group, Group II: DN group, Group III: Balanites treated group, Group IV: MSCs treated group, Group V: Exosome treated group, Group VI: Balanites + MSCs treated group and Group VII: Balanites + exosome treated group. Following the trial period, blood and renal tissues were subjected to biochemical, gene expression analyses, and histopathological examinations. Results showed that MDA was substantially increased, whereas TAC was significantly decreased in the kidney in the DN group compared to normal health rats. Undesired elevated values of MDA levels and a decrease in TAC were substantially ameliorated in groups co-administered Balanites aegyptiacae with MSCs or exosomes compared to the DN group. A substantial elevation in TNF-α and substantially diminished concentration of IGF-1 were noticed in DN rats compared to normal health rats. Compared to the DN group, the co-administration of Balanites aegyptiacae with MSCs or exosomes substantially improved the undesirable elevated values of TNF-α and IGF-1. Furthermore, in the DN group, the mRNA expression of Vanin-1, Nephrin, and collagen IV was significantly higher than in normal healthy rats. Compared with DN rats, Vanin-1, Nephrin, and collagen IV Upregulation were substantially reduced in groups co-administered Balanites aegyptiacae with MSCs or exosomes. In DN rats, AQP1 expression was significantly lower than in normal healthy rats. Furthermore, the groups co-administered Balanites aegyptiacae with MSCs or exosomes demonstrated a substantial increase in AQP1 mRNA expression compared to DN rats.

Obesity and aging are known to affect the skeletal muscles. Obesity in old age may result in a poor basement membrane (BM) construction response, which serves to protect the skeletal muscle, thus making the skeletal muscle more vulnerable. In this study, older and young male C57BL/6J mice were divided into two groups, each fed a high-fat or regular diet for eight weeks. A high-fat diet decreased the relative gastrocnemius muscle weight in both age groups, and obesity and aging individually result in a decline in muscle function. Immunoreactivity of collagen IV, the main component of BM, BM width, and BM-synthetic factor expression in young mice on a high-fat diet were higher than that in young mice on a regular diet, whereas such changes were minimal in obese older mice. Furthermore, the number of central nuclei fibers in obese older mice was higher than in old mice fed a regular diet and young mice fed a high-fat diet. These results suggest that obesity at a young age promotes skeletal muscle BM formation in response to weight gain. In contrast, this response is less pronounced in old age, suggesting that obesity in old age may lead to muscle fragility. [ABSTRACT FROM AUTHOR]

The development of uremic cardiomyopathy (UC) and cardiac fibrosis in chronic kidney disease (CKD) was comparatively analyzed in model experiments on SPF males of Sprague–Dawley and Wistar rats. CKD was induced by left subtotal nephrectomy (NE) followed after 2 weeks by right radical NE. Sham-operated (Sham) animals served as controls. Four weeks after radical NE, blood pressure (BP) was measured, and the expression of pro-fibrotic factors (Fli1, pro-collagen I, collagen I, collagen IV) in left ventricular myocardial tissue was assessed by real-time PCR and immunoblotting. After NE, Sprague–Dawley rats exhibited a significant increase in systolic BP, as well as left ventricular hypertrophy. In Wistar rats, the difference between BP in NE and Sham animals was smaller, while the left ventricular mass to body weight ratio did not change. Experimental CKD in Sprague–Dawley rats was accompanied by a 1.5–2.5-fold suppression of Fli1 gene expression and a decrease in Fli1 protein content in cardiac muscle tissue, while no significant differences were observed in Wistar rats. Pro-collagen I and collagen I levels in the heart of rats of both strains changed at neither transcription nor translation levels. Such a difference in the development of pathological processes indicates an ineffectiveness of the applied type of NE to induce UC and investigate pro-fibrotic processes in Wistar vs. Sprague–Dawley rats. [ABSTRACT FROM AUTHOR]

Bromine and peroxidasin (an extracellular peroxidase) are essential for generating sulfilimine cross-links between a methionine and a hydroxylysine within collagen IV, a basement membrane protein. The sulfilimine cross-links increase the structural integrity of basement membranes. The formation of sulfilimine cross-links depends on the ability of peroxidasin to use bromide and hydrogen peroxide substrates to produce hypobromous acid (HOBr). Once a sulfilimine cross-link is created, bromide is released into the extracellular space and becomes available for reutilization. Whether the HOBr generated by peroxidasin is used very selectively for creating sulfilimine cross-links or whether it also causes oxidative damage to bystander molecules (e.g., generating bromotyrosine residues in basement membrane proteins) is unclear. To examine this issue, we used nanoscale secondary ion mass spectrometry (NanoSIMS) imaging to define the distribution of bromine in mammalian tissues. We observed striking enrichment of bromine (79Br, 81Br) in basement membranes of normal human and mouse kidneys. In peroxidasin knockout mice, bromine enrichment of basement membranes of kidneys was reduced by ∼85%. Proteomic studies revealed bromination of tyrosine-1485 in the NC1 domain of α2 collagen IV from kidneys of wild-type mice; the same tyrosine was brominated in collagen IV from human kidney. Bromination of tyrosine-1485 was reduced by >90% in kidneys of peroxidasin knockout mice. Thus, in addition to promoting sulfilimine cross-links in collagen IV, peroxidasin can also brominate a bystander tyrosine. Also, the fact that bromine enrichment is largely confined to basement membranes implies that peroxidasin activity is largely restricted to basement membranes in mammalian tissues.

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.

Objective To explore the phenotype-genotype correlation of Alport syndrome in children. Methods Retrospectively analyze the clinical and pathological features of 55 patients with Alport syndrome with COL4A mutations detected by second-generation sequencing, who were treated at Beijing Children's Hospital from January 2016 to December 2020. Results A total of 55 children with Alport syndrome were included. All cases had hematuria, including 31 cases (56.4%) with gross hematuria and 24 cases (43.6%) with microscopic hematuria. A total of 39 (70.9%) patients also had proteinuria. Extrarenal manifestations were pres- ent in 12 patients (21.8%). 36(65.4%) patients had a family history of Alport syndrome. 32 patients underwent pathological examination and 23 of them had the specific pathological changes of Alport syndrome. In 55 cases, 36 (65.4%) were diagnosed as X-linked Alport syndrome, 5(9.1%) were diagnosed as autosomal recessive Alport syndrome, 10(18.2%) were diagnosed as autosomal dominate Alport syndrome, and 4(7.3%) were diagnosed as digenic Alport syndrome. Missense mutations in COL4A genes accounted for 62.5%, 67.5% of missense mutations resulted in glycine substitution. There were statistical significances in proteinuria degree and hearing loss between male and female patients with XLAS (P < 0.05) as well as statistical significance in the degree of proteinuria between autosomal recessive Alport syndrome and autosomal dominate Alport syndrome (P=0.044), and there was critical statistical significance in the age of onset. There was statistical significance in hearing loss between children with renal impairment and children with normal renal function (P=0.001). Conclusions Most of the pathogenic variants in COL4A genes that cause Alport syndrome result in glycine substitutions. The degree of proteinuria and hearing loss of males with XLAS were greater than those of females. The degree of proteinuria in autosomal recessive Alport syndrome was greater than that of children with autosomal dominate Alport syndrome, and the age of onset was earlier than that of autosomal dominate Alport syndrome. Renal manifestation was more severe in children with hearing loss. The early clinical manifestations of Alport syndrome are diverse and pathological manifestations may be atypical. The application of next-generation sequencing can reduce misdiagnosises of Alport syndrome.

The basement membrane (BM), mainly composed of collagen IV, plays an important role in the maintenance, protection, and recovery of muscle fibers. Collagen IV expression is maintained by the balance between synthetic and degradative factors, which changes depending on the level of muscle activity. For example, exercise increases collagen IV synthesis, whereas inactivity decreases collagen IV synthesis. However, the effects of stretching on the BM structure remain unclear. Therefore, to investigate the effects of stretching on the BM of the skeletal muscle, we continuously applied stretching to the rat soleus muscle and examined the altered expression of BM-related factors and structure using quantitative polymerase chain reaction (qPCR), western blotting, zymography, immunohistochemistry, and electron microscopy. The results show that stretching increased the matrix metalloproteinase 14 (MMP14) expression and MMP2 activity, and decreased the collagen IV expression and width of the lamina densa in the soleus muscle. These results suggest that stretching promotes BM degradation in the rat soleus muscle. The findings of this study indicate a new influence of stretching on skeletal muscles, and may contribute to the new use of stretching in rehabilitation and sports fields. [ABSTRACT FROM AUTHOR]

Scalp-Ear-Nipple syndrome is caused by pathogenic KCTD1 variants and characterised by a scalp defect, prominent ears, and rudimentary breasts. We describe here further clinical associations in the eye and kidney. Fifteen affected members from two unrelated families with p.(Ala30Glu) or p.(Pro31Leu) in KCTD1 were examined for ocular and renal abnormalities. The relevant proteins were studied in the eye and kidney, and the mutation consequences determined from mouse knockout models. Five males and 10 females with a median age of 40 years (range 1–70) with pathogenic variants p.(Ala30Glu) (n = 12) or p.(Pro31Leu) (n = 3) in KCTD1 were studied. Of the 6 who underwent detailed ophthalmic examination, 5 (83%) had low myopic astigmatism, the mean spherical equivalent of 10 eyes was 2.38D, and one (17%) had hypermetropic astigmatism. One female had a divergent strabismus. Five individuals had renal cysts (5/15, 33%), with renal biopsy in one demonstrating a thinned glomerular basement membrane identical to that seen in Thin basement membrane nephropathy (AD Alport syndrome). In the eye, KCTD1 and its downstream targets, TFAP2, and the collagen IV α3 and α4 chains localised to the cornea and near the retinal amacrine cells. In the kidney, all these proteins except TFAP2 were expressed in the podocytes and distal tubules. TFAP2B and COL4A4 knockout mice also had kidney cysts, and COL4A3 and COL4A4 knockout mice had myopia. Individuals with a pathogenic KCTD1 variant may have low myopic astigmatism and represent a further rare genetic cause for a thinned glomerular basement membrane. [ABSTRACT FROM AUTHOR]

Discoidin domain receptor 1 (DDR1) is a tyrosine kinase receptor expressed in epithelial cells from different tissues in which collagen binding activates pleiotropic functions. In the brain, DDR1 is mainly expressed in oligodendrocytes (OLs), the function of which is unclear. Whether collagen can activate DDR1 in OLs has not been studied. Here, we assessed the expression of DDR1 during in vitro OL differentiation, including collagen IV incubation, and the capability of collagen IV to induce DDR1 phosphorylation. Experiments were performed using two in vitro models of OL differentiation: OLs derived from adult rat neural stem cells (NSCs) and the HOG16 human oligodendroglial cell line. Immunocytofluorescence, western blotting, and ELISA were performed to analyze these questions. The differentiation of OLs from NSCs was addressed using oligodendrocyte transcription factor 2 (Olig2) and myelin basic protein (MBP). In HOG16 OLs, collagen IV induced DDR1 phosphorylation through slow and sustained kinetics. In NSC-derived OLs, DDR1 was found in a high proportion of differentiating cells (MBP+/Olig2+), but its protein expression was decreased in later stages. The addition of collagen IV did not change the number of DDR1+/MBP+ cells but did accelerate OL branching. Here, we provide the first demonstration that collagen IV mediates the phosphorylation of DDR1 in HOG16 cells and that the in vitro co-expression of DDR1 and MBP is associated with accelerated branching during the differentiation of primary OLs. [ABSTRACT FROM AUTHOR]

Safety concerns associated with foetal bovine serum (FBS) have restricted its translation into clinics. We hypothesised that platelet lysate (PL) can be utilised as a safe alternative to produce serum-free 3D-engineered skin. PL supported a short-term expansion of fibroblasts, with negligible replication-induced senescence and directed epidermal stratification. PL-expanded fibroblasts were phenotypically separated into three subpopulations of CD90+FAP+, CD90+FAP− and CD90−FAP+, based on CD90 (reticular marker) and FAP (papillary marker) expression profile. PL drove the expansion of the intermediate CD90+ FAP+ subpopulation in expense of reticular CD90+FAP−, which may be less fibrotic once grafted. The 3D-engineered skin cultured in PL was analysed by immunofluorescence using specific markers. Detection of ColIV and LMN-511 confirmed basement membrane. K10 confirmed near native differentiation pattern of neo-epidermis. CD29- and K5-positive interfollicular stem cells were also sustained. Transmission and scanning electron microscopies detailed the ultrastructure of the neo-dermis and neo-epidermis. To elucidate the underlying mechanism of the effect of PL on skin maturation, growth factor contents in PL were measured, and TGF-β1 was identified as one of the most abundant. TGF-β1 neutralising antibody reduced the number of Ki67-positive proliferative cells, suggesting TGF-β1 plays a role in skin maturation. Moreover, the 3D-engineered skin was exposed to lucifer yellow on days 1, 3 and 5. Penetration of lucifer yellow into the skin was used as a semi-quantitative measure of improved barrier function over time. Our findings support the concept of PL as a safe and effective serum alternative for bioengineering skin for cell therapies. [ABSTRACT FROM AUTHOR]

Runx2 is a key transcription factor involved in valvular interstitial cells (VIC) osteodifferentiation, a process actively entwined with the calcific aortic valve disease (CAVD). We hypothesize that a strategy intended to silence Runx2 could be a valuable novel therapeutic option for CAVD. To this intent, we aimed at (i) developing targeted nanoparticles for efficient delivery of short hairpin (sh)RNA sequences specific for Runx2 to the aortic valve employing a relevant mouse model for CAVD and (ii) investigate their therapeutic potential in osteoblast-differentiated VIC (oVIC) cultivated into a 3D scaffold. Since collagen IV was used as a target, a peptide that binds specifically to collagen IV (Cp) was conjugated to the surface of lipopolyplexes encapsulating shRNA-Runx2 (Cp-LPP/shRunx2). The results showed that Cp-LPP/shRunx2 were (i) cytocompatible; (ii) efficiently taken up by 3D-cultured oVIC; (iii) diminished the osteodifferentiation of human VIC (cultured in a 3D hydrogel-derived from native aortic root) by reducing osteogenic molecules expression, alkaline phosphatase activity, and calcium concentration; and (iv) were recruited in aortic valve leaflets in a murine model of atherosclerosis. Taken together, these data recommend Cp-LPP/shRunx2 as a novel targeted nanotherapy to block the progression of CAVD, with a good perspective to be introduced in practical use.

Age-related hearing loss (ARHL) is the most common sensory degenerative disease and can significantly impact the quality of life in elderly people. A previous study using GeneChip miRNA microarray assays showed that the expression of miR-29a changes with age, however, its role in hearing loss is still unclear. In this study, we characterized the cochlear phenotype of miR-29a knockout (miR-29a–/–) mice and found that miR-29a-deficient mice had a rapid progressive elevation of the hearing threshold from 2 to 5 months of age compared with littermate controls as measured by the auditory brainstem response. Stereocilia degeneration, hair cell loss and abnormal stria vascularis (SV) were observed in miR-29a–/– mice at 4 months of age. Transcriptome sequencing results showed elevated extracellular matrix (ECM) gene expression in miR-29a–/– mice. Both Gene Ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis revealed that the key differences were closely related to ECM. Further examination with a transmission electron microscope showed thickening of the basilar membrane in the cochlea of miR-29a–/– mice. Five Col4a genes (Col4a1-a5) and two laminin genes (Lamb2 and Lamc1) were validated as miR-29a direct targets by dual luciferase assays and miR-29a inhibition assays with a miR-29a inhibitor. Consistent with the target gene validation results, the expression of these genes was significantly increased in the cochlea of miR-29a–/– mice, as shown by RT-PCR and Western blot. These findings suggest that miR-29a plays an important role in maintaining cochlear structure and function by regulating the expression of collagen and laminin and that the disturbance of its expression could be a cause of progressive hearing loss.

Collagen type IV alpha 1 and alpha 2 chains form heterotrimers ([α1(IV)]2α2(IV)) that represent a fundamental basement membrane constituent. Dominant COL4A1 and COL4A2 mutations cause a multisystem disorder that is marked by clinical heterogeneity and variable expressivity and that is generally characterized by the presence of cerebrovascular disease with ocular, renal, and muscular involvement. Despite the fact that muscle pathology is reported in up to one-third of individuals with COL4A1 and COL4A2 mutations and in animal models with mutations in COL4A1 and COL4A2 orthologs, the pathophysiological mechanisms underlying COL4A1-related myopathy are unknown. In general, mutations are thought to impair [α1(IV)]2α2(IV) secretion. Whether pathogenesis results from intracellular retention, extracellular deficiency, or the presence of mutant proteins in basement membranes represents an important gap in knowledge and a major obstacle for developing targeted interventions. We report that Col4a1 mutant mice develop progressive neuromuscular pathology that models human disease. We demonstrate that independent muscular, neural, and vascular insults contribute to neuromyopathy and that there is mechanistic heterogeneity among tissues. Importantly, we provide evidence of a COL4A1 functional subdomain with disproportionate significance for tissue-specific pathology and demonstrate that a potential therapeutic strategy aimed at promoting [α1(IV)]2α2(IV) secretion can ameliorate or exacerbate myopathy in a mutation-dependent manner. These data have important translational implications for prediction of clinical outcomes based on genotype, development of mechanism-based interventions, and genetic stratification for clinical trials. Collectively, our data underscore the importance of the [α1(IV)]2α2(IV) network as a multifunctional signaling platform and show that allelic and tissue-specific mechanistic heterogeneities contribute to the variable expressivity of COL4A1 and COL4A2 mutations.

Cutaneous collagenous vasculopathy is a rare clinicopathological entity, first described in 2000. Cutaneous collagenous vasculopathy has been considered a form of microangiopathy of superficial dermal vessels and produce lesions that appear as telangiectasia. We present a patient with histopathologic features of cutaneous collagenous vasculopathy and scattered erythematous papules on the trunk with a striking dermatoscopic finding. We propose the term of 'cutaneous papular collagenous vasculopathy' as a new clinical manifestation of this disease.

This article discusses cutaneous collagenous vasculopathy (CCV), a rare microangiopathy that affects the skin. The article presents a case series of 19 new CCV cases from five hospitals in northern Spain over a period of five years. The clinical and anatomopathological findings of these cases are compared to those previously reported in the literature. CCV is characterized by the appearance of fine, arborizing telangiectasias, typically starting on the lower limbs and slowly progressing upward. The diagnosis requires ruling out other systemic conditions and conducting a biopsy. The article concludes that CCV is an underdiagnosed disorder that should be considered in the differential diagnosis of generalized telangiectasias. Further research is needed to better understand its pathogenesis. [Extracted from the article]

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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

• Trol, the Drosophila orthologue of the basement membrane component HSPG2/perlecan, controls the morphogenesis of the three distinct types of epithelia of a complex organ, the wing imaginal disc. • In the basement membrane of the squamous epithelium of the wing imaginal disc, trol regulates the organization of the collagen type IV network and provides elasticity. • This role of trol in basement membrane biogenesis is critical for the squamous shape of the cells. • Trol-directed morphogenesis of the squamous epithelium promotes the relaxation of the two other epithelia, cuboidal and columnar, ensuring proper architecture of the wing imaginal disc. 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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Alport syndrome (AS) is the most frequent monogenic inherited glomerulopathy and is also genetically and clinically heterogeneous. It is caused by semi-dominant pathogenic variants in the X-linked COL4A5 (NM_000495.5) gene or recessive variants in the COL4A3/COL4A4 (NM_000091.4/NM_000092.4) genes. The disease manifests in early childhood with persistent microhematuria and can progress to proteinuria and kidney failure in adolescence or early adulthood if left untreated. On biopsy, pathognomonic features include alternate thinning, thickening and lamellation of the glomerular basement membrane (GBM), in the presence of podocyte foot process effacement. Although previous studies indicate a prevalence of AS of about 1/50,000, a recent publication reported a predicted rate of pathogenic COL4A5 variants of 1/2320. We herewith present 98 patients (40 M/58 F) from 26 Greek families. We are selectively presenting the families segregating the X-linked form of AS with pathogenic variants in the COL4A5 gene. We found 21 different pathogenic variants, 12 novel: eight glycine and one proline substitutions in the collagenous domain, one cysteine substitution in the NC1 domain, two premature termination of translation codons, three splicing variants, one 5-bp insertion/frameshift variant, one indel-frameshift variant and four gross deletions. Notably, patients in six families we describe here and three families we reported previously, carried the COL4A5-p.G624D substitution, a founder defect encountered all over Europe which is hypomorphic with mostly milder symptomatology. Importantly, on several occasions, the correct genetic diagnosis reclassified patients as patients with AS, leading to termination of previous immunosuppressive/cyclosporine A therapy and a switch to angiotensin converting enzyme inhibitors (ACEi). With the understanding that all 98 patients span a wide range of ages from infancy to late adulthood, 15 patients (11 M/4 F) reached kidney failure and 11 (10 M/1 F) received a transplant. The prospects of avoiding lengthy diagnostic investigations and erroneous medications, and the advantage of delaying kidney failure with very early administration of renin-angiotensin-aldosterone system (RAAS) blockade, highlights the importance of timely documentation of AS by genetic diagnosis. [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 psiCHECKTM-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. [ABSTRACT FROM AUTHOR]

Radiation-induced brain injury (RIBI) is one of the most common long-term complications for patients with malignant brain tumors after radiotherapy. At present, there is no effective treatment for RIBI. Recent studies have also confirmed that polysaccharides from laminaria japonica (LJP) display potential neuroprotective function. However, its mechanisms of neuroprotection remain unclear. In this study, we aimed to explore the effect and underlying mechanism of LJP on neurogenesis in radiation-induced brain injury mice. SPF two-month-old male mice were randomly divided into control group (Con), LJP treatment group (LJP), irradiation group (IR), and irradiation with LJP treatment group (IR + LJP). LJP (40 mg/kg/day) was intraperitoneally injected at one day before radiation for seven consecutive days (once daily). The mice were exposed to 10 Gy × 2 fractionated doses, once every other day, with a total dose of 20 Gy. Changes in cognitive function of mice following radiation were evaluated by the Morris water maze test. Furthermore, body weight and general status of mice were measured throughout the experiment. Immunohistochemical staining for neural proliferating cells (Ki67+ cells) and immature neurons (DCX + cells) was utilized to assay changes of neurogenesis in hippocampus. Microglial activation and collagen IV deposition within the neurogenic microenvironment were observed respectively by immunohistochemical staining for Iba-1 and Collagen IV in the hippocampus. Levels of pro-inflammatory cytokines (TNF-α and IL-1β) in the hippocampus were detected by ELISA kits post-radiation. Morris water maze test showed that LJP therapy markedly reduced the escape latency and increased the times of crossing platform and percent time of the target quadrant in the radiated mice. In addition, the decrease of the neural proliferating cells (Ki67+ cells) and immature neurons (DCX + cells) in the hippocampus of mice following irradiation was significantly mitigated by the LJP treatment, suggesting that LJP could prevent from neurogenesis damage after irradiation. LJP injection significantly attenuated degradation of collagen IV, activation of microglia, and increase of pro-inflammatory cytokines (TNF-α and IL-1β) levels in the neurogenic microenvironment of the hippocampus after radiation. These findings suggest that LJP early treatment may mitigate radiation-induced cognitive impairments and that its mechanism may relate to its protection of neurogenesis by alleviating neuroinflammation and collagen IV degradation within the neurogenic microenvironment. [ABSTRACT FROM AUTHOR]

Alport syndrome is a hereditary kidney disease caused by mutations in the three genes encoding for collagen IV: COL4A3, COL4A4, and COL4A5. Several mouse models have been created for the study of this disease with variable phenotypic outcomes. This review is an up-to-date presentation of the current mouse models existing in the literature with a detailed comparison of the phenotypic features characterizing each model. Although in humans it is primarily a glomerulopathy, data suggest that in some mouse models, the initial symptoms appear in the tubule-interstitial region rather than the glomerulus. Additionally, in some other models, the severity of disease in the tubule-interstitial region is affected by the genetic background. In conclusion, the phenotypic spectrum of each model appears to be affected by the model's genetic background, the position of the genetic alteration within the gene, and the type of the genetic alteration. Despite these disparities, mouse models recapitulate with relatively high fidelity several features of the human disease, which makes them useful for studies aimed at better understanding cellular pathomechanisms and for finding new treatments. [ABSTRACT FROM AUTHOR]

Objectives: Dysfunction of the thyroid gland has a negative effect on the male reproductive system. Studies also show that extracellular matrix (ECM) components play an essential role in testicular development and function. In hypothyroidism, there is a significant disruption in the ECM structure of mammalian tissues. In addition, notable changes have been reported in the germ cell population under a hypothyroid state. This study aimed to investigate the relationship between ECM proteins and apoptosis of testicular germ cells due to hypothyroidism. Materials and Methods: In the present experimental study, 20 male Balb/C mice were divided into control and hypothyroid groups. The hypothyroid group received 0.05% 6-n-propyl-2-thiouracil (PTU) through drinking water for 35 days. Finally, real-time polymerase chain reaction, immunohistochemistry, periodic acid-Schiff (PAS) staining, terminal transferase-mediated dUTP nick-end labelling (TUNEL) assay, and biochemical measurements were performed after hypothyroidism confirmation. Results: Laminin α5 and collagen IV mRNA levels were upregulated in the hypothyroid group compared to the controls (P

Canstatin, the NC1 domain of the α2 chain of collagen IV, prevents tumor growth through angiogenesis inhibition and apoptosis induction. N-terminal 1–89 amino acid fragment of canstatin induces apoptosis much higher than the C-terminal fragment. Recently, we demonstrated that the amino acids 78–86 of canstatin, so-called Cans, have more anti-migration, anti-tube formation, and anti-tumor activities than other collagen IV derived peptides. Here, we evaluated HUVEC, MCF10A, and L929 cell viability, the percentage of apoptotic cells by Annexin V-FITC/PI staining, caspase-3 activity, Bcl-2, and caspase-8 gene expression using RT-qPCR in endothelial cells, and Bax and Bcl-2 expression in tumors by immunohistochemistry to investigate the apoptotic effect of Cans. Results showed that the peptide reduced the percentage of viable HUVE cells with EC50 of 21 μM and was not toxic for normal cell lines. 30 and 50 μM of Cans induced 34.6% and 50.7% early and late apoptosis in HUVECs compared to 16.5% in control. In addition, caspase-3 activity was amplified up to threefold compared to the untreated cells. Cans down-regulated Bcl-2 and caspase-8 gene expression. This result may show that this peptide acts through the intrinsic pathway and cannot affect the extrinsic pathway of apoptosis. However, this hypothesis requires more investigation. Besides, Bcl-2 reduction and Bax elevation in tumor sections indicated that this peptide could stimulate apoptosis in vivo. In conclusion, we showed that the short canstatin peptide induces apoptosis in endothelial and tumor cells as one of its anti-angiogenic and anti-tumor mechanisms. [ABSTRACT FROM AUTHOR]

Kidney podocytes and endothelial cells assemble a complex and dynamic basement membrane that is essential for kidney filtration. Whilst many components of this specialised matrix are known, the influence of fluid flow on its assembly and organisation remains poorly understood. Using the coculture of podocytes and glomerular endothelial cells in a low-shear stress, high-flow bioreactor, we investigated the effect of laminar fluid flow on the composition and assembly of cell-derived matrix. With immunofluorescence and matrix image analysis we found flow-mediated remodelling of collagen IV. Using proteomic analysis of the cell-derived matrix we identified changes in both abundance and composition of matrix proteins under flow, including the collagen-modifying enzyme, prolyl 4-hydroxylase (P4HA1). To track collagen IV assembly, we used CRISPR-Cas9 to knock in the luminescent marker HiBiT to the endogenous COL4A2 gene in podocytes. With this system, we found that collagen IV was secreted and accumulated consistently under both static and flow conditions. However knockdown of P4HA1 in podocytes led to a reduction in the secretion of collagen IV and this was more pronounced under flow. Together, this work demonstrates the effect of fluid flow on the composition, modification, and organisation of kidney cell-derived matrix and provides an in vitro system for investigating flow-induced matrix alteration in the context of kidney development and disease., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)