Aqueous Lumican Correlates with Central Retinal Thickness in Patients with Idiopathic Epiretinal Membrane: A Proteome Study.

Idiopathic epiretinal membrane (iERM) is a pathological fibrocellular change in the vitreoretinal junction over the macular area; however, possible pathogenic mechanisms remain unclear. Changes in the differential protein composition of the aqueous humor (AH) may represent potential molecular changes associated with iERM. To gain new insights into the molecular mechanisms of iERM pathology, a sensitive label-free proteomics analysis was performed to compare AH protein expressions in patients with cataracts with or without iERM. This study employed nanoflow ultra-high-performance liquid chromatography-tandem mass spectrometry to investigate protein compositions of the AH obtained from individual human cataract eyes from 10 patients with iERM and 10 age-matched controls without iERM. Eight proteins were differentially expressed between the iERM and control samples, among which six proteins were upregulated and two were downregulated. A gene ontology (GO) analysis revealed that iERM was closely associated with several biological processes, such as immunity interactions, cell proliferation, and extracellular matrix remodeling. Additionally, multiple proteins, including lumican, cyclin-dependent kinase 13, and collagen alpha-3(VI) chain, were correlated with the central retinal thickness, indicating a multifactorial response in the pathogenic process of iERM. Changes in the AH level of lumican between iERM and control samples were also confirmed by an enzyme-linked immunosorbent assay. In conclusion, several pathological pathways involved in iERM were identified in the AH by a proteomic analysis, including immune reactions, cell proliferation, and remodeling of the extracellular matrix. Lumican is a potential aqueous biomarker for predicting iERM development and monitoring its progression. More clinical parameters also need to be identified to complete the analysis, and those could provide additional targets for treating and preventing iERM.

Proteomic Signature of Dysfunctional Circulating Endothelial Colony-Forming Cells of Young Adults.

Background A subpopulation of endothelial progenitor cells called endothelial colony-forming cells (ECFCs) may offer a platform for cellular assessment in clinical studies because of their remarkable angiogenic and expansion potentials in vitro. Despite endothelial cell function being influenced by cardiovascular risk factors, no studies have yet provided a comprehensive proteomic profile to distinguish functional (ie, more angiogenic and expansive cells) versus dysfunctional circulating ECFCs of young adults. The aim of this study was to provide a detailed proteomic comparison between functional and dysfunctional ECFCs. Methods and Results Peripheral blood ECFCs were isolated from 11 subjects (45% men, aged 27±5 years) using Ficoll density gradient centrifugation. ECFCs expressed endothelial and progenitor surface markers and displayed cobblestone-patterned morphology with clonal and angiogenic capacities in vitro. ECFCs were deemed dysfunctional if <1 closed tube formed during the in vitro tube formation assay and proliferation rate was <20%. Hierarchical functional clustering revealed distinct ECFC proteomic signatures between functional and dysfunctional ECFCs with changes in cellular mechanisms involved in exocytosis, vesicle transport, extracellular matrix organization, cell metabolism, and apoptosis. Targeted antiangiogenic proteins in dysfunctional ECFCs included SPARC (secreted protein acidic and rich in cysteine), CD36 (cluster of differentiation 36), LUM (lumican), and PTX3 (pentraxin-related protein PYX3). Conclusions Circulating ECFCs with impaired angiogenesis and expansion capacities have a distinct proteomic profile and significant phenotype changes compared with highly angiogenic endothelial cells. Impaired angiogenesis in dysfunctional ECFCs may underlie the link between endothelial dysfunction and cardiovascular disease risks in young adults.

Identification of Lumican and Fibromodulin as Hub Genes Associated with Accumulation of Extracellular Matrix in Diabetic Nephropathy.

INTRODUCTION: Diabetic nephropathy (DN) remains a major cause of end-stage renal disease. The development of novel biomarkers and early diagnosis of DN are of great clinical importance. The goal of this study was to identify hub genes with diagnostic potential for DN by weighted gene co-expression network analysis (WGCNA). METHODS: Gene Expression Omnibus database was searched for microarray data including distinct types of CKD. Gene co-expression network was constructed, and modules specific for DN were identified by WGCNA. Gene ontology (GO) analysis was performed, and the hub genes were screened out within the selected gene modules. In addition, cross-validation was performed in an independent dataset and in samples of renal biopsies with DN and other types of glomerular diseases. RESULTS: Dataset GSE99339 was selected, and a total of 179 microdissected glomeruli samples were analyzed, including DN, normal control, and 7 groups of other glomerular diseases. Twenty-three modules of the total 10,947 genes were grouped by WGCNA, and a module was specifically correlated with DN (r = 0.54, p = 9e-15). GO analysis showed that module genes were mainly enriched in the accumulation of extracellular matrix (ECM). LUM, ELN, FBLN1, MMP2, FBLN5, and FMOD were identified as hub genes. Cross verification showed LUM and FMOD were higher in the DN group and were negatively correlated with estimated glomerular filtration rate (eGFR). In renal biopsies, expression levels of LUM and FMOD were higher in DN than IgA nephropathy, membranous nephropathy, and normal controls. CONCLUSION: By using WGCNA approach, we identified LUM and FMOD related to ECM accumulation and were specific for DN. These 2 genes may represent potential candidate diagnostic biomarkers of DN.

Lumican is upregulated in osteoarthritis and contributes to TLR4-induced pro-inflammatory activation of cartilage degradation and macrophage polarization.

OBJECTIVE: Lumican (LUM) is a major extracellular matrix glycoprotein in adult articular cartilage and its expression is known to be upregulated upon cartilage degeneration. LUM is associated with the pathogen-associated molecular pattern (PAMP) activation of the TLR4 signalling cascade, with TLR4 being highly associated with inflammation in rheumatic diseases. However, the main role of the LUM structural molecule in osteoarthritis (OA) remains elusive. The aim of this study was, therefore, to understand the role of LUM during TLR4-mediated activation in OA. METHODS: After measuring LUM levels in synovial fluid (SF) of OA patients and lipopolysaccharide (LPS)-induced TLR4 activation, the role of LUM in the expression of pro-inflammatory molecules and cartilage degradation was assessed in vitro and ex vivo in a cartilage explant model. Primary macrophage activation and polarization were studied upon LUM co-stimulation with LPS. RESULTS: We demonstrate that LUM is not only significantly upregulated in SF from OA patients compared to healthy controls, but also that LUM increases lipopolysaccharide (LPS)-induced TLR4 activation. Furthermore, we show that a pathophysiological level of LUM augments the LPS-induced TLR4 activation and expression of downstream pro-inflammatory molecules, resulting in extensive cartilage degradation. LUM co-stimulation with LPS also provided a pro-inflammatory stimulus, upregulating primary macrophage activation and polarization towards the M1-like phenotype. CONCLUSIONS: These findings strongly support the role of LUM as a mediator of PAMP-induced TLR4 activation of inflammation, cartilage degradation, and macrophage polarization in the OA joint and potentially other rheumatic diseases.

Extracellular Matrix Components HAPLN1, Lumican, and Collagen I Cause Hyaluronic Acid-Dependent Folding of the Developing Human Neocortex.

Neocortical expansion, thought to underlie the cognitive traits unique to humans, is accompanied by cortical folding. This folding starts around gestational week (GW) 20, but what causes it remains largely unknown. Extracellular matrix (ECM) has been previously implicated in neocortical expansion and here we investigate the potential role of ECM in the formation of neocortical folds. We focus on three specific ECM components localized in the human fetal cortical plate (CP): hyaluronan and proteoglycan link protein 1 (HAPLN1), lumican and collagen I (collectively, HLC). Addition of HLC to cultures of human fetal neocortex (11-22 GW) caused local changes in tissue stiffness, induced CP folding, increased CP hyaluronic acid (HA), and required the HA-receptor CD168 and downstream ERK signaling. Importantly, loss of HA reduced HLC-induced and 22 GW physiological nascent folds. This was altered in samples with neurodevelopmental disorders, indicating it may be a useful system to study such disorders.

Rheumatic Heart Disease and Myxomatous Degeneration: Differences and Similarities of Valve Damage Resulting from Autoimmune Reactions and Matrix Disorganization.

Autoimmune inflammatory reactions leading to rheumatic fever (RF) and rheumatic heart disease (RHD) result from untreated Streptococcus pyogenes throat infections in individuals who exhibit genetic susceptibility. Immune effector mechanisms have been described that lead to heart tissue damage culminating in mitral and aortic valve dysfunctions. In myxomatous valve degeneration (MXD), the mitral valve is also damaged due to non-inflammatory mechanisms. Both diseases are characterized by structural valve disarray and a previous proteomic analysis of them has disclosed a distinct profile of matrix/structural proteins differentially expressed. Given their relevance in organizing valve tissue, we quantitatively evaluated the expression of vimentin, collagen VI, lumican, and vitronectin as well as performed immunohistochemical analysis of their distribution in valve tissue lesions of patients in both diseases. We identified abundant expression of two isoforms of vimentin (45 kDa, 42 kDa) with reduced expression of the full-size protein (54 kDa) in RHD valves. We also found increased vitronectin expression, reduced collagen VI expression and similar lumican expression between RHD and MXD valves. Immunohistochemical analysis indicated disrupted patterns of these proteins in myxomatous degeneration valves and disorganized distribution in rheumatic heart disease valves that correlated with clinical manifestations such as valve regurgitation or stenosis. Confocal microscopy analysis revealed a diverse pattern of distribution of collagen VI and lumican into RHD and MXD valves. Altogether, these results demonstrated distinct patterns of altered valve expression and tissue distribution/organization of structural/matrix proteins that play important pathophysiological roles in both valve diseases.