Posttranslational regulation of keratins: degradation of mouse and human keratins 18 and 8.

Human keratin 18 (K18) and keratin 8 (K8) and their mouse homologs, Endo B and Endo A, respectively, are expressed in adult mice primarily in a variety of simple epithelial cell types in which they are normally found in equal amounts within the intermediate filament cytoskeleton. Expression of K18 alone in mouse L cells or NIH 3T3 fibroblasts from either the gene or a cDNA expression vector results in K18 protein which is degraded relatively rapidly without the formation of filaments. A K8 cDNA containing all coding sequences was isolated and expressed in mouse fibroblasts either singly or in combination with K18. Immunoprecipitation of stably transfected L cells revealed that when K8 was expressed alone, it was degraded in a fashion similar to that seen previously for K18. However, expression of K8 in fibroblasts that also expressed K18 resulted in stabilization of both K18 and K8. Immunofluorescent staining revealed typical keratin filament organization in such cells. Thus, expression of a type I and a type II keratin was found to be both necessary and sufficient for formation of keratin filaments within fibroblasts. To determine whether a similar proteolytic system responsible for the degradation of K18 in fibroblasts also exists in simple epithelial cells which normally express a type I and a type II keratin, a mutant, truncated K18 protein missing the carboxy-terminal tail domain and a conserved region of the central, alpha-helical rod domain was expressed in mouse parietal endodermal cells. This resulted in destabilization of endogenous Endo A and Endo B and inhibition of the formation of typical keratin filament structures. Therefore, cells that normally express keratins contain a proteolytic system similar to that found in experimentally manipulated fibroblasts which degrades keratin proteins not found in their normal polymerized state.

[Dynamic bimaxillary osteotomy: the new occlusal base]. / L'ostéotomie bimaxillaire dynamique: "la nouvelle donne occlusale".

The authors present an ortho-surgical method which unites occlusion and aesthetic without compromise and without stopping orthodontic work during the immediate post-operative period. The occlusal preparation permits us a global and simultaneous mobilisation of the two maxillars which are ostesynthezed in posterior skeletal disclosing. This disposition allows a lingual replacement and gives more facility to an eventual immediate post-operative occlusal replacement. The stiff osteosynthesis with immovable plates realize a therapeutic dissociation between the skeletal stage and the basal alveolo-dental stage. The "proprioceptive amnesia" and the "muscular sideration" permit a proprioceptive reorganisation and a new neuro-muscular fonctionnement elaborated from a new occlusal base. The free movements of the T.M.J. facilitate these acquisitions and allow a perspective supervision of the occlusion in a dynamic perspective.

Transformation of human keratinocytes by SV40 virus alters their response to the tumor promoter TPA.

Recently, we have shown that transformation of human keratinocytes by SV40 virus induces the re-expression of characters found in fetal epidermis and monostratified epithelia. In the present study, TPA was found to alter some features of the human keratinocyte phenotype in a similar manner to SV40 transformation. Indeed, 12-O-tetradecanoyl-13-phorbol acetate (TPA) treatment induced the expression of cytokeratin no. 8, recognized by the monoclonal antibody TROMA-1, which is present in fetal epidermis and/or monostratified epithelia only, and fibronectin expression. However, TPA and SV40 transformation had specific non-overlapping effects. For example, TPA did not prevent terminal differentiation and stratification, as SV40-transformation did, but stimulated these processes. Moreover, TPA was found to induce additional changes in SV40-transformed keratinocytes. In particular it provoked the individualization of cells within the colonies. This effect was seen as little as 1 h after treatment and was reversible. This cellular alteration was accompanied by the reorganization of actin and by a decrease in the number of desmosomes; these changes were not observed after treatment of normal keratinocytes with TPA. These observations lead to the conclusion that TPA is able to trigger cellular responses which cannot be induced by SV40 products alone, but that TPA and some SV40 products can cooperate to elicit new responses, which could reflect a higher state of malignancy.

Precocious appearance of involucrin and epidermal transglutaminase during differentiation of psoriatic skin.

We compared the distribution in psoriatic skin of three different markers usually found in the stratum granulosum of normal skin. Using dansylcadaverine, we demonstrate that epidermal transglutaminase activity can be detected in most of the suprabasal layers of involved psoriatic skin and that the epidermal transglutaminase activity closely matches involucrin distribution. The glycoprotein GP37 was not detected in involved psoriatic skin of stable lesions. These results suggest that the integrated control of several independent pathways of terminal differentiation is lost in psoriasis, resulting in the classical feature of parakeratosis with absence of the stratum granulosum.

The majority of epidermal Merkel cells are non-proliferative: a quantitative immunofluorescence analysis.

Although epidermal Merkel cells (MC) are able to form synapses and synthetize neuromediators, they can be considered as being of epithelial nature because of the presence of cytokeratins in their cytoskeleton and desmosomes on their membranes. Since epidermis is an epithelium undergoing permanent renewal, it was important to determine whether MC were able to renew, as neighbouring keratinocytes do. This was investigated by studying whether S phase nuclei could be found in cells bearing a specific MC marker. The technique consisted of injecting rabbits with bromodeoxyuridine (BrdUrd) and performing double immunofluorescence on skin sections with the antikeratin number 8 monoclonal antibody (MAb) TROMA-I and anti-BrdUrd MAb. The results show that, in contrast to the neighbouring epidermal cells, the great majority of MC were found to be devoid of BrdUrd labelling, indicating that most of these cells are unable to divide, or divide very rarely.

Sequence analysis of murine cytokeratin endo A (no. 8) cDNA. Evidence for mRNA species initiated upstream of the normal 5' end in PCC4 cells.

Keratin 8 (Endo A) is expressed in simple epithelia, together with keratin 18 (Endo B). Filaments formed by this keratin pair are the first cytoskeletal elements induced during mouse embryogenesis. We have isolated Endo A cDNA clones from lambda gt11 libraries prepared with mRNA isolated from PCC4 embryonal carcinoma (EC) cells. Sequencing of three overlapping cDNAs and of a genomic clone allowed us to determine the complete sequence of the Endo A message. Analysis of the protein sequence deduced showed that the Endo A protein presented all the characteristics of intermediate filaments, including an alpha-helical central rod domain and nonhelical N- and C-termini. In the rod domain, the degree of similarity to the other members of the basic keratin family was high. A high degree of homology to keratin 8 of other species was observed, even in the non-helical domains. During these analyses, we found clones extending upstream of the normal 5' end of the mRNA. Sequence comparison between these cDNAs and the 5' upstream region of the Endo A gene suggested that they corresponded to transcripts initiated at an upstream alternative promoter. These observations supported previous results showing the presence of Endo A transcripts initiated upstream of the normal 5' end in mouse morulae and blastocysts.

Intraepidermal leakage of plasma proteins after tape stripping of normal skin and uninvolved psoriatic skin.

Following tape stripping of normal skin and uninvolved psoriatic skin, there was an early leakage of plasma proteins such as fibrinogen, fibronectin and immunoglobulins. This was accompanied by migration of mononuclear and polymorphonuclear cells into the epidermis. Both events appeared earlier, were more pronounced and lasted longer in psoriatic subjects than in controls.

Atherogen lipid profile in HIV-1-infected patients with lipodystrophy syndrome.

Background: Cases of lipodystrophy syndrome and metabolic disorders have been described since the onset of highly active antiretroviral therapy in HIV-infected patients. The aim of our study was to estimate the prevalence of lipodystrophy (LD) and to define the associated lipid profile of these patients. Methods: The following were determined for each patient: lipid profile (cholesterol and its subfractions, atherogenicity ratios, and triglycerides), blood glucose, and immunovirological markers (CD4(+) cell count and plasma viral load). Patients were classified into two groups on the basis of whether or not they presented with clinical signs of LD. Results: Among 233 HIV-infected patients included in the study, 61 cases (26.1%) of lipodystrophy (LD) were noted. Compared with non-LD patients (NLD), LD patients were older men (P<10(-4)) with a lower CD4(+) lymphocyte cell count (P<0.007) and more often at the AIDS stage (P<10(-3)) (OR=3.2 (95% CI: 1.47-6.2)). Multivariate analysis showed a correlation between LD cases and age (10 years older) (OR=1.78 (95% CI: 1.23-2.57), P<0.002) and the decrease in CD4(+) cell count (100 CD4(+)/mm(3) lower) (OR=1.31 (95% CI: 1.09-1.58), P<0.004). An analysis of lipid subfractions and atherogenicity ratios clearly indicated a proatherogenic lipid profile for the LD patients. Conclusions: The underlying physiopathological mechanism of LD is still unknown. However, the lipid profile of HIV-1-infected patients with a LD syndrome appears to place these patients at an increased risk of progression of atherosclerosis.