Basement membrane deposition of nidogen 1 but not nidogen 2 requires the nidogen binding module of the laminin gamma1 chain

The nidogen-laminin interaction is proposed to play a key role in basement membrane (BM) assembly. However, though there are similarities, the phenotypes in mice lacking nidogen 1 and 2 (nidogen double null) differ to those of mice lacking the nidogen binding module (γ1III4) of the laminin γ1 chain. This indicates different cell- and tissue-specific functions for nidogens and their interaction with laminin and poses the question of whether the phenotypes in nidogen double null mice are caused by the loss of the laminin-nidogen interaction or rather by other unknown nidogen functions. To investigate this, we analyzed BMs, in particular those in the skin of mice lacking the nidogen binding module. In contrast to nidogen double null mice, all skin BMs in γ1III4-deficient mice appeared normal. Furthermore, although nidogen 1 deposition was strongly reduced, nidogen 2 appeared unchanged. Mice with additional deletion of the laminin γ3 chain, which contains a γ1-like nidogen binding module, showed a further reduction of nidogen 1 in the dermoepidermal BM; however, this again did not affect nidogen 2. This demonstrates that in vivo only nidogen 1 deposition is critically dependent on the nidogen binding modules of the laminin γ1 and γ3 chains, whereas nidogen 2 is independently recruited either by binding to an alternative site on laminin or to other BM proteins.     

A heterogeneous immunofluorescence staining for laminin-1 and related basal lamina molecules in the dorsal root ganglia following constriction nerve injury

The bodies of primary sensory neurons and their satellite glial cells (SGCs) are limited by the basal laminae from extracellular matrix of the dorsal root ganglia (DRG). The basal laminae displayed uniform immunofluorescence staining for laminin-1 in the sections of rat intact (naive) DRG. A proximal or distal ligature of the spinal nerves resulted in a heterogeneous immunostaining for laminin-1 around neuron-SGC units in the sections of the corresponding DRG. The pattern of irregular laminin-1 immunofluorescence was more extensive in the ipsilateral than the contralateral DRG of the operated rats. The immunofluorescence for laminin-1 exactly coincided with binding of Concanavalin-A as well as immunostaining for type IV collagen in both naive DRG and DRG affected by nerve ligature. Nidogen immunostaining decreased or fully disappeared at the surface of the SGCs consistently with immunofluorescence staining for laminin-1, but retained or increased in the endothelial cells and ED-1 positive cells invaded the DRG affected by nerve ligature. The results indicate an alteration of the content of basal laminae surrounding the bodies of primary sensory neurons and their SGSs following nerve constriction injury. A modulation of the basal laminae may be related with other cellular and molecular alterations related with peripheral neuropathic pain, for example, expansion of sympathetic sprouts.     

Proteomic profiling of cardiomyopathic tissue from the aged mdx model of Duchenne muscular dystrophy reveals a drastic decrease in laminin,nidogen and annexin

The majority of patients afflicted with Duchenne muscular dystrophy develop cardiomyopathic complications, warranting large‐scale proteomic studies of global cardiac changes for the identification of new protein markers of dystrophinopathy. The aged heart from the X‐linked dystrophic mdx mouse has been shown to exhibit distinct pathological aspects of cardiomyopathy. In order to establish age‐related alterations in the proteome of dystrophin‐deficient hearts, cardiomyopathic tissue from young versus aged mdx mice was examined by label‐free LC‐MS/MS. Significant age‐dependent alterations were established for 67 proteins, of which 28 proteins were shown to exhibit a lower abundance and 39 proteins were found to be increased in their expression levels. Drastic changes were demonstrated for 17 proteins, including increases in Ig chains and transferrin, and drastic decreases in laminin, nidogen and annexin. An immunblotting survey of young and old wild‐type versus mdx hearts confirmed these proteomic findings and illustrated the effects of natural aging versus dystrophin deficiency. These proteome‐wide alterations suggest a disintegration of the basal lamina structure and cytoskeletal network in dystrophin‐deficient cardiac fibres, increased levels of antibodies in a potential autoimmune reaction of the degenerating heart, compensatory binding of excess iron and a general perturbation of metabolic pathways in dystrophy‐associated cardiomyopathy.     

Interleukin-1β and tumor necrosis factor-α have opposite effects on fibroblasts and epithelial cells during basement membrane formation

Interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) are typical proinflammatory cytokines that influence various cellular functions, including metabolism of the extracellular matrix. We examined the roles of IL-1beta and TNF-alpha in basement membrane formation in an in vitro model of alveolar epithelial tissue composed of alveolar epithelial cells and pulmonary fibroblasts. Formation of the basement membrane by immortalized rat alveolar type II epithelial (SV40-T2) cells, which ordinarily do not form a continuous basement membrane, was dose-dependently upregulated in the presence of 2 ng/ml IL-1beta or 5 ng/ml TNF-alpha. IL-1beta or TNF-alpha alone induced increased secretion of type IV collagen, laminin-1, and nidogen-1/entactin, all of which contributed to this upregulation. In contrast, while SV40-T2 cells cultured with a fibroblasts-embedded type I collagen gel were able to form a continuous basement membrane, they failed to form a continuous basement membrane in the presence of IL-1beta or TNF-alpha. Fibroblasts treated with IL-1beta or TNF-alpha secreted matrix metalloproteinase (MMP)-9 and MMP-2, and these MMPs inhibited basement membrane formation and degraded the basement membrane architecture. Neither IL-1beta- nor TNF-alpha-treated SV40-T2 cells increased the secretion of MMP-9 and MMP-2. These results suggest that IL-1beta participates in basement membrane formation in two ways. One is the induction of MMP-2 and MMP-9 secretion by fibroblasts, which inhibits basement membrane formation, and the other is induction of basement membrane component secretion from alveolar epithelial cells to enhance basement membrane formation.     

Basement membranes in skin: unique matrix structures with diverse functions?

The view of extracellular matrix (ECM) has evolved from a merely scaffolding and space filling tissue element to an interface actively controlling cellular activities and tissue functions. A highly specialized form of ECM is the basement membrane (BM), an ubiquitous sheet-like polymeric structure composed of a set of distinct glycoproteins and proteoglycans. In this review we are largely focusing on function and assembly of BM in skin (1) at the dermo-epidermal interface and (2) in the resident micro-vasculature. The role of the non-polymeric components perlecan and particularly nidogen is exemplified by reviewing experiments based on genetic approaches and adequate experimental skin models in vivo and in vitro. While in mice total deficiency of one of these components is eventually developmentally lethal, the severity of the defects varies drastically between tissues and also the skin models recapitulating BM formation in vitro. There is accumulating evidence that this relies on the mechanical properties, the molecular composition of the BM, the adjacent ECM or connective tissue, the dynamics of molecular assembly, and ‘minor’ tissue-specific modifier or adapter components. Though the role of nidogen or perlecan is still remaining a controversial issue, the statements ‘being essential for BM/or not’ should be consequently referred to the developmental, tissue, and functional (e.g., repair) context.