Elastin changes during chronological and photo-ageing: the important role of lysozyme.

Cutaneous ageing, as a result of combined chronological and photo-ageing in sun-exposed areas, is accompanied by major modifications of the elastic fibres. We aimed to investigate qualitative and quantitative changes of dermal elastin fibres during cutaneous chronological and photo-ageing and the involvement of lysozyme in these processes. Morphological, age-related changes and variations in the relative elastin content in sun-protected (buttock) and sun-exposed (forearm and face) skin of healthy volunteers were studied (145 samples). The deposition of lysozyme in elastin fibres was studied using light and immuno-electron microscopy and taking into consideration the relative efficacy of different UV wavebands (UVA or SSR (solar simulated radiation)). Our studies also included the proteolytic degradation of elastin by human leucocyte elastase (HLE) in situ. Our results indicate a reduction of elastin content with age in sun-protected and sun-exposed skin, associated for the latter with high elastin content, resulting in elastosis. Total UVA (320-400 nm), and in particular long wave UVA (UVA-1, 340-400 nm), induces lysozyme deposition in elastin fibres to a higher extent than solar simulated radiation (SSR, 280-400 nm). Immuno-electron microscopy revealed lysozyme association with the electron-dense granular amorphous elastin structures, corresponding to a basophilic degeneration induced by sun exposure. Lysozyme has no elastolytic activity in situ; however, its binding to elastin limits elastin degradation by human leucocyte elastase (HLE). In addition, a direct inhibitory effect of lysozyme on HLE was observed. Our data suggest that lysozyme prevents elastin degradation by HLE after binding to the damaged parts of the elastin network and by direct lysozyme-HLE interaction, which reduces HLE proteolytic activity. These observations contribute to a better understanding of the chronological and photo-induced changes of the dermal elastic network.

Vascular wall remodeling in patients with supravalvular aortic stenosis and Williams Beuren syndrome.

Supravalvular aortic stenosis (SVAS) and Williams Beuren syndrome (WBS) can be considered as inherited diseases affecting the whole arterial tree and causing narrowing of the vessels. It has been reported that abnormal deposition of elastin in arterial walls of patients with SVAS and WBS leads to increased proliferation of arterial smooth muscle cells (SMC), which result in the formation of hyperplastic intimal lesions. In this work, we conducted morphological and morphometrical analysis with stenotic aortas from patients suffering from SVAS and WBS and from healthy control subjects and demonstrated that the amount of elastic fibers and the loss of integrity of vascular elastic fibers in the aortas reflect similar changes in the skin of patients with SVAS or WBS, as reported in our previous work conducted on skin in these pathological states. On the other hand, we conducted investigations on metalloproteinases (MMP2, MMP9, MMP7) and their specific tissue inhibitors TIMP1 and TIMP2 to verify their possible involvement in the etiopathogeny of SVAS and WBS. We particularly evidenced an altered MMP9/TIMP1 balance in favor of matrix degradation which could facilitate SMC migration and neointimal hyperplasia. Our findings suggest that elastinolytic enzymes secreted by arterial SMC, possibly including matrilysin 1, are critical for the development of arterial lesions in SVAS and WBS and contribute to perpetuate arterial stenosis in either SVAS or WBS.

Possible involvement of gelatinase A (MMP2) and gelatinase B (MMP9) in toxic epidermal necrolysis or Stevens-Johnson syndrome.

Toxic epidermal necrolysis (TEN) and Stevens-Johnson syndrome (SJS) are considered to be drug-induced diseases, and are characterized by extensive mucocutaneous disorder and epidermal necrosis which result in the detachment of the epidermis. Inactive and active forms of metalloproteinases (MMP2 and MMP9) secreted by skin explants maintained in organ culture for 72 h and in blister fluid from two TEN and three SJS patients were investigated. Interestingly, lesional skin from both the TEN and the SJS patients cultured for 3 days in conditioned medium showed high levels of both 72 kDa progelatinase A and 66 kDa activated gelatinase A, and the 66 kDa activated form was not observed in cultures of skin from control individuals. Furthermore, indirect immunodetection showed the presence of MMP2 and MMP9 in TEN and SJS patients' skin. Increased gelatinase activity in the culture medium of TEN and SJS skin maintained in organ culture and in blister fluid indicates that these gelatinases may be responsible for the detachment of the epidermis in these drug-induced necrolyses.

Structural alterations of the vascular wall in magnesium-deficient mice. A possible role of gelatinases A (MMP-2) and B (MMP-9).

Vascular alterations during magnesium deficiency have long been known but the implicated mechanisms have so far been poorly documented. In this preliminary assay, we compared the thoracic aortic histology in Swiss OF1 mice fed a severe magnesium-deficient diet (50 +/- 5 ppm) for 42 days to that of controls fed a standard diet (1700 +/- 100 ppm magnesium). It appeared (eosin-haematoxylin coloration) that, in magnesium-deficient mice, the aortic wall was thinner than in controls. Specific colorations of the two of main fibers vascular tissue (collagens and elastin) showed severe structural alterations of both components. These changes were consecutive to the expression of matrix metalloproteinases (MMP) -2 and -9 which were present as zymogens (inactive forms) in controls and supposed to be present in their active and inactive forms in magnesium-deficient mice (zymography). These changes which have not been reported so far would explain, at least in part, the sensitivity of magnesium-deficient mice to various stress or xenobiotics.

Marfan syndrome, magnesium status and medical prevention of cardiovascular complications by hemodynamic treatments and antisense gene therapy.

The medical management of Marfan Syndrome (MFS) mainly relies on early prevention of the aortic complications. Hemodynamic treatments try to diminish the forcefulness of cardiac contractions and to reduce blood pressure: for example long term administration of propranolol may significantly reduce the rate of increase in aortic ratio (aortic diameter/expected aortic diameter). Retardation of aortic dilatation may be most often observed by early treatment started when the baseline end-diastolic aortic root diameter is < 40 mm. It seems better to use beta-blockers without intrinsic sympathomimetic activity. Successful acceptance of beta-blockers may be limited by side-effects, but the efficiency of alternative hypotensive agents (calcium channel inhibitors, ACE inhibitors) is not yet validated. Gene therapy might constitute an etiologic specific treatment of MFS. FBN1-RZ1 hammerhead antisense ribozyme is able to suppress expression of the mutant FBN1 allele. The use of ribozymes as systemic therapeutic agents will depend on efficient delivery to its target, but the various proposed vectors raise yet unsolved problems. A hydrogel angioplasty balloon might be a possible vector for delivering an antisense ribozyme in the aortic wall specifically. Ribozymes--as deoxyribonucleotides--may be taken up by tissue upon local application. Further research should study ex vivo local application of antisense ribozyme on human aortic wall, before assessing in vivo efficiency and tolerance of this aortic local vectorisation. It is always necessary to maintain a balanced magnesium intake in patients with MFS. Firstly to prevent the multiple noxious effects of magnesium deficiency on cardiovascular targets. Secondly to ensure the best efficiency and the least toxicity of the hemodynamic drugs used as long term prophylactic treatment for cardiovascular complications and of the etiologic antisense magnesium-dependent gene therapy, in the future.