The pattern of fibrillin deposition correlates with microfibril-associated glycoprotein 1 (MAGP-1) expression in cultured blood and lymphatic endothelial cells.

Fibrillins constitute the major structural components of 10-12nm microfibrils of the extracellular matrix of several elastic and non elastic tissues and of initial lymphatic vessel anchoring filaments. Microfibril-Associated Glycoprotein-1 (MAGP-1) binds fibrillin to tropoelastin during elastogenesis. We recently reported that cultured blood endothelial cells deposit fibrillin in a honeycomb pattern, whereas lymphatic endothelial cells form an irregular web. The aim of this immunohistochemical study was to verify whether the deposition pattern of fibrillin is related to the expression of MAGP-1 in confluent and postconfluent cultures of bovine aortic (AEC), pulmonary artery (PAEC) and lymphatic endothelial cells (LEC). In AEC and PAEC, MAGP-1 and fibrillin co-localized and their deposition increased with time in culture. In AEC, both proteins formed a honeycomb pattern. In LEC, MAGP-1 deposition was still negligible when fibrillin formed an irregular web covering the entire surface. PAEC, which in vivo are exposed to physiological conditions intermediate between AEC and LEC, had an intermediate pattern of deposition of fibrillin and MAGP-1. Assuming that early elastogenesis is an intrinsic functional need for the aorta, but not for the thoracic duct, we propose that delayed appearance of MAGP-1 in LEC may correlate with their irregular fibrillin deposition. Different fibrillin scaffolds could in turn account for the specificity of elastic fibers in compliance with the specific functional requirements of the tissue.

Specific adhesion molecules bind anchoring filaments and endothelial cells in human skin initial lymphatics.

Anchoring filaments are a characteristic feature of initial lymphatic vessels. They connect the abluminal membrane of endothelial cells to the surrounding elastic fibers. The main molecular component of anchoring filaments is fibrillin. Initial lymphatic vessels of human skin were stained with monoclonal antibodies to fibrillin, integrins alpha 2 beta 1, alpha 3 beta 1 and alpha v beta 3, vinculin, talin, beta-actin and focal adhesion kinase (FAK). A double-labeling immunofluorescence method was used to simultaneously stain fibrillin and alpha 3 beta 1 integrin or FAK. Close contiguities between integrins and anchoring filaments were observed. These results suggest that the anchoring filaments connect the extracellular matrix and the endothelial cell cytoskeleton through the transmembrane integrin and FAK molecule. The results also demonstrate the presence of focal adhesions in the wall of initial lymphatic vessels. These connections possibly enable transmission of chemical and/or mechanical stimuli from the extracellular matrix to the endothelial cells. Here, they are transformed in cytoskeleton rearrangements and intracellular signaling events, some of which may contribute to the initial formation of lymph.

An immunological correlation between the anchoring filaments of initial lymph vessels and the neighboring elastic fibers: a unified morphofunctional concept.

Little has been published on the histochemical and cytochemical properties of anchoring filaments of initial lymph vessels. Previous research suggests that the microfibrils of the anchoring filaments have ultrastructural, histochemical and cytochemical characteristics similar to those of the microfibrils associated with elastic fibers. With the aim of further investigating the histological identity of anchoring filaments, we performed an immunohistochemical study with human skin lymphatics, using antibody HB8, specific for elastic fiber microfibrils. The findings suggested strong molecular similarities between elastic fibers and the fibrils of anchoring filaments of the initial lymph vessels. A comparison of these fibrils showed both constitutional homogeneity and structural continuity from the abluminal surface of the initial lymph vessel to the perivascular elastic fibers and to the adjacent elastic network of connective tissue. In conjunction with previous findings, we propose a unified hypothesis that the elastic fiber system composed of anchoring filaments, perilymphatic sheath and adjacent connective tissue acts by alternating stretching and relaxation to propel lymph towards lymph collectors and draining lymph nodes.

Phospholipase A2 activity and calpactin I levels in rat lymphokine-activated killer cells: correlation with the cytotoxic activity.

In the present paper we have shown evidence for a significant increase of type II sPLA2 activity in A-LAK cells. The A-LAK-mediated cytotoxicity against YAC-1 target cells was strongly inhibited by two inhibitors of sPLA2, p-BPB and mepacrine, suggesting the involvement of this enzyme in the lytic mechanism of A-LAK. On the other hand, stimuli such as A23187 ionophore and TPA, which were able to induce in control cells an increased AA release, failed to cause this effect in IL-2-treated cells, suggesting that PLA2 was not active in these cells. Thus, we analyzed the levels of calpactin I, which is considered to be involved in the down-regulation of PLA2 activity. HrIL-2 treatment led to an increased expression of calpactin I at both the RNA and the protein level. A substantial portion of calpactin I was associated with the external surface of A-LAK and was able to exert a strong inhibitory effect on a purified porcine pancreatic PLA2 activity in vitro. Our results suggest that the role of calpactin I could be relevant to regulate PLA2 activity, and to protect the effector cells against a possible toxic effect which this enzyme could exert if present at high levels.