Stress proteins (Hsp72/73, Grp94) expression pattern in rat organs following metavanadate administration. Effect of green tea drinking.

Expression pattern of heat shock proteins (Hsp) 72/73 and glucose regulated protein (Grp) 94 was studied in liver, kidney and testis of rats injected with sublethal doses of ammonium metavanadate (5 mg/kg/day). In addition, some batches of animals were given green tea decoction, known to be rich in anti-oxidative compounds, as sole beverage in order to evaluate its protective properties. In control animals, the stress proteins expression was found to be organ-dependent: anti-Grp94 antibody revealed two bands at 96 and 98 kDa in kidney and liver whereas the 98 kDa band only was found in testis; anti-Hsp72/73 antibody revealed that the constitutive Hsp73 was present in all organs whereas the inducible Hsp72 was only present in kidney and testis. In kidney of vanadium-treated rats, Hsp73 was over-expressed by about 50% whereas Hsp72 was down-regulated by 50-80%. No such effects were observed in liver and testis. In liver and kidney of vanadium-treated rats, Grp94 was over-expressed by 50% and 150% respectively whereas no change was found in testis. In rats given green tea as sole beverage, the 96 kDa protein expression level in liver was reduced both in controls and in vanadium-treated animals. However, green tea drinking failed to prevent the vanadium-induced Hsp72 under-expression in kidney of vanadium-treated rats.

Effects of nickel poisoning on expression pattern of the 72/73 and 94 kDa stress proteins in rat organs and in the COS-7, HepG2, and A549 cell lines.

The present study deals with the effects of Ni on the expression level of three stress proteins, namely, the cytosolic HSP72 and HSP73, and the reticulum-associated GRP94. Experiments were carried out on "Wistar'' female rats daily injected with 4 mg NiCl2 per kg body weight for 1, 3, 5, and 10 days. Another set of experiments were carried out using cell lines, derived from the monkey kidney (COS-7), and from human tumors of the lung (A549) and liver (HepG2). Cells were cultured for 4 days in the permanent presence of 100, 200, or 400 microM NiCl2. In control rats, stress proteins pattern was found to be tissue specific: two protein bands of 96 and 94 kDa were immunodetected with the anti-GRP94 antibody in kidney and liver extracts, whereas only the 96 kDa band was present in ovary extracts. HSP73 was present in kidney, liver, and ovary whereas HSP72 was only found in kidney. In kidney of nickel-treated animals, HSP73 and the 96 kDa proteins were overexpressed whereas HSP72 was strongly down regulated. No such effect was observed in liver or ovary. Similarly, in nickel-treated cell lines, HSP72 was downregulated and GRP94 (96 kDa protein) was overexpressed. HSP73 expression appeared moderately increased in A549 cells but decreased in COS-7 cells. Because long-term caloric restriction was reported to reduce free radical generation in cells, the effect of 1 month food restriction (50%) was tested in rats as a possible way to lower oxidative damages induced by Ni. No significant effect on HSP expression was observed.

Volatile organic compounds cytotoxicity and expression of HSP72, HSP90 and GRP78 stress proteins in cultured human cells.

The aim of this study was to determine whether overexpression of stress proteins (SPs) could be a sensitive biomarker for cell injury due to exposure to low doses of volatile organic compounds (VOCs) such as benzene, ethylbenzene, toluene, xylene, and chlorinated derivatives (ClB). Sublethal and cytotoxic threshold concentrations of the VOCs were determined by studying the growth rate of normal (fibroblasts) or tumor-derived human cell lines (A549, HepG2) exposed for 4 days to VOCs. Changes in SP expression as a function of concentrations were investigated by Western blotting.VOC toxicity was found to be correlated with their degree of chlorination and their hydrophobicity. Cytotoxic threshold concentrations (no-observed effect concentration, NOEC) were found to be similar for the three cell lines. It was observed that using a mixture of VOCs, each of them at concentration below the NOEC, resulted in an actual toxicity to the cells. This finding reveals a synergistic effect and should be taken into account when assessing threshold risk and exposure limit values in the worker's environment when several pollutants may be present. HSP72 and HSP90 expression levels were not affected whereas GRP78 expression was increased by all the VOCs. Taking into account the specific molecular function of GRP78, it suggests that VOC exposure results in misfolded or underglycosylated protein accumulation in the endoplasmic reticulum. GRP78 overexpression was closely related to the magnitude of growth inhibition due to increasing concentrations of each VOC. The overexpression was found to be significant for concentrations 5 to 30 times higher than NOEC, indicating that, under our experimental conditions, GRP78 expression cannot be considered as a sensitive biomarker of exposure to environmental VOCs.

Effect of chronic lead exposure on kidney function in male and female rats: determination of a lead exposure biomarker.

Several cytotoxic chemical pollutants inducing peroxidative damages are liable to induce kidney failure. Among these pollutants we find heavy metals such as: lead, nickel, cadmium, vanadium and mercury. Lead is one of the most dangerous metals because it is widely spread in the environment, and because it may be a source of several nervous diseases. The aim of this study is to provide evidence concerning the effect of this metal on the renal function and to try to determine a storage corner in the organism which serves as an indicator of a lead intoxication. Lead acetate was administered by oral route in the drinking water to adult rats aged three months at the rate of 0.3% (P1) and 0.6% (P2). Reference rats received distilled water to drink under the same conditions. The treatment continued for 15, 30, 45, 60 and 90 days. The creatinemia, uremia, glycemia and creatinuria are determined by colorimetric techniques. Lead concentration in blood as well as the lead content of the tail are determined by atomic absorption after nitroperchloric mineralization at the liquid stage. The results showed an increase of creatinemia on the 30th day of the experiment for both sexes in (P1 and P2). The same happened for ureamia. The increase of these two parameters would indicate a renal deficiency which is confirmed by a decrease of creatinuria and urinary pH observed mainly on and after the 45th day of the experiment. An increase of the renal relative weight was noticed in P1 and P2 on the 30th day of the treatment. The determination of the concentration of lead in the blood shows that this factor increases among treated subjects in a constant way, independently of the dose and the duration of the treatment. Nevertheless, the rate increase of lead in the tail seems to be dose-dependent. In conclusion, lead administered by oral route causes a renal deficiency to the rat without distinction between males and females. In addition, the tail seems to be a reliable exposure biomarker that demonstrates lead intoxication. The tail seems to be a dosimeter of lead bio-accumulation. It constitutes an endogenous source of lead impregnation. The concentration of lead in the blood is only an indicator of recent exposure.

Cellular method for evaluation of noxiousness of inorganic pollutants in industrial wastes: calculation of a safety index for monitoring sludge discharge.

This article deals with a biological test of safety applicable to industrial wastes. The test is based on the measurement of the growth rate of cultured human cells exposed to waste samples with different dilutions. As a first approach, 15 chemicals in which discharge concentrations are submitted to sanitary regulations were tested one by one. For Zn, Cu, Ni, Cd, Ag, Co, Mg, sulfates, and fluorides, it was possible to detect concentrations that are below the allowed limit. For Hg, Al, As(V), Cr(III), Fe, and Pb, the concentrations that affect cell growth are higher than the allowed limit. Tests were also performed using actual samples (liquid effluent from a laundry and sludge from waste-water treatment plants). Results indicate that, in contrast to chemical analyses, the current biological test has the advantage of providing an indication of global toxicity, integrating all substances and factors that can be harmful to life processes. From the sludge data and the observed threshold of concentration that does not affect cell growth, a numeric safety index has been calculated which indicates the amount of sludge that could be dispersed, as a fertilizer, per hectare of agricultural soil. Such an index could be conveniently used for designing sewage sludge disposal strategies.

Pattern of stress protein expression in human lung cell-line A549 after short- or long-term exposure to cadmium.

Exposure to cadmium is associated with the development of pulmonary damage such as emphysema and lung cancer. This metal is also a powerful inducer of stress proteins in many biologic models. The present study was undertaken to evaluate whether an overexpression of the heat shock protein (hsp)72 stress protein, which indicates repair of damaged proteins, could be a sensitive and early biomarker of environmental pollution by Cd. In comparative studies, we examined the effects of exposure to Cd (as CdCl(2)) on the growth rate of the A549 pulmonary cell line, and (by Western blot analyses) on the induction of the hsp72 stress protein and metallothioneins (MTs). CdCl(2) exposure was studied for periods of 2 hr to 1 month. For short-term exposure (2-6 hr) to Cd concentrations higher than 50 microM, an overexpression of hsp72 appeared 6 hr later, suggesting that hsp72 might be considered an early biomarker of acute exposure to Cd. For exposures lasting more than 4 days, lower doses of Cd (0.1-10 microM) similar to levels encountered in occupational exposure induced a significant increase of the hsp72 level. Because the increase of hsp72 occurs for doses that did not affect cell proliferation, our work supports the idea that its overexpression might be used as a sensitive indicator of occupational exposure to Cd. However, increased resistance to Cd appeared in A549 cells exposed for 1 month and overexpression of hsp72 disappeared simultaneously. It is possible that, in vivo, cell adaptation also occurs throughout chronic exposure to Cd, with a decrease of hsp induction as a consequence. A dose-related increase of MTs was found after 4 days of exposure to Cd concentrations ranging from 0.1 to 10 microM without change of overexpression during chronic exposure, suggesting that MT expression could be a more constant indicator of Cd pollution. Because 0.1 microM Cd (11 microg/L) induces hsp72 expression, showing the presence of damaged proteins, our work suggests that the maximum allowable biologic exposure limit should be lowered.

Comparative evaluation of the potential noxiousness in domestic sludge used in agriculture and in commercial fertilizers.

The noxiousness of actual sludge collected in eight water treatment plants around the city of Toulouse, France, was evaluated using a biological test based on the growth rate of cultured human cells. Results were compared with those obtained from 18 fertilizers and culture supports that are commercially available in gardening shops. Surprisingly, it was found that sludge extracts, at low concentrations (below 5 g of dry material/liter), were improving the cell growth rate, which suggests the presence of useful oligoelements. At higher concentrations, a noxious effect, expressed as inhibition of cell growth, was observed. However, this negative effect was of the same order of magnitude as that obtained, under the same experimental conditions, with commercial garden fertilizers which are available and used without any restriction. It is concluded that discarding the sludge, after submission to the biological test, in controlled amount as an agricultural fertilizer should not be hazardous to the environment.

Supernatant cytotoxicity and proteolytic activity of selected oral bacteria against human gingival fibroblasts in vitro.

The purpose of this study was to determine if endodontic bacterial act in vitro on human gingival fibroblast functions via extracellular products. The bacteria used were Prevotella nigrescens, Capnocytophaga ochracea, Peptostreptoccocus micros and Actinobacillus actinomycetemcomitans. Supernatants were collected from bacterial cultures at the beginning of the stationary phase when their density was similar. Toxins that inhibited fibroblast proliferation were found in all culture supernatants of Gram-positive or Gram-negative bacterial strains, except for Prev. nigrescens. The cytotoxicity of A. actinomycetemcomitans supernatant was about 1000 fold higher than the others. This supernatant diluted to 1/1000 led to total fibroblast growth inhibition whereas only 25% growth inhibition was obtained with Capn. ochracea and Pept. micros diluted to 1/10. Bacterial supernatant proteolytic activity was investigated in confluent fibroblast cultures that were incubated for 48 hr with each of the supernatants diluted to 1/2 except for A. actinomycetemcomitans supernatant diluted to 1/20. Indirect immunofluorescence studies of extracellular-matrix molecules, followed by immunoelectrophoretic analysis of extracts of whole-cell layers, demonstrated that only conditioned medium of Prev. nigrescens had a proteolytic activity capable of degrading the greater part of type I collagen and fibronectin fibres in the extracellular matrix.

Modulation by hypergravity of extracellular matrix macromolecules in in vitro human dermal fibroblasts.

In vitro human dermal fibroblasts were submitted to normal gravity (1 g) or to chronic hypergravity (20 g) over a period of 8 days. Changes in organization of extracellular matrix molecules were seen by indirect immunofluorescence. In the fibronectin layer, bundles of fibrils were gathered together leading to a disorganisation of the normal parallel pattern of fibers seen in control cultures. Type I collagen fibrils appeared with wooly outlines in controls whereas thick fibers were closely packed in 20-g cultures. A moderate increase of type III collagen fibril density was observed. No elastic fibers were seen in control or in 20-g cultures. In the culture medium, the release of soluble elastin (ELISA) and type I and III collagens (RIA) was undisturbed. Assays of enzymes involved in the remodeling of extracellular matrix showed an increase of cellular elastase activity (10%) and a decrease of the spontaneously active collagenase. Nevertheless, the total collagenase activity, (activated by trypsin), was increased by up to 30%. These data show a significant rise of the latent collagenase activity and suggest that release of the tissue inhibitor of metalloproteinase (TIMP1) was enhanced by hypergravity.

Effects of hypergravity on the cell shape and on the organization of cytoskeleton and extracelluar matrix molecules of in vitro human dermal fibroblasts.

In vitro human dermal fibroblasts were submitted to normal gravity (1 g) or to chronic hypergravity ranging from 2 to 20 g for 8 days. Changes only appeared above 15 g. The majority of 20 g-subjected cells showed fine filipods in the shape of a star whereas most control cells had rounded shapes and spread by forming lamellipodia. Indirect immunofluorescence staining of vinculin, alpha-actinin and actin stress fibers showed changes of the arrangement anchoring points of stress fibers under hypergravity. Tubulin staining showed that the centrosomal material generally located above the nucleus in control cells had migrated to the nucleus side in 20 g-exposed cells. After 8 d of culture under 20 g hypergravity the thickness of fibronectin network seemed to be increased and bundles of fibrils appeared linking ordered arrays of fibers. The fibrils of collagen I formed better delimited and thicker bundles of fibers. We may assume that 20 g hypergravity can induce changes in fibroblast cell shape, migration way, and anchorage leading to a reorganization of extracellular matrix without concomitant change of cell proliferation.

[Typing of human papillomaviruses in intra-epithelial malpighian lesions of the cervix uteri: correlation with cytohistologic grade and progression of the lesions]. / Typage des papillomavirus humains (PVH) dans les lésions malpighiennes intra-épithéliales du col utérin: corrélations avec le grade cytohistologique et l'évolution des lésions.

The clinical course of condylomas of the cervix, and more generally of all cervical dysplasias whether associated with koilocytosis or not, remains unpredictable. Based on a series of earlier publications, we undertook a routine typing for human papillomaviruses in intraepithelial malpighian lesions of the cervix. We examined the real usefulness of such typing to determine prognosis of the lesions and correct clinical management. We determined the correlations between human papillomaviruses, as detected by the in situ hybridization method, with the histological grade of 125 cervical lesions (in 106 patients) and the clinical course (mean follow-up = 24 months) in 56% of them. Viruses 16/18 and/or 31/33/55, considered to be potential oncogenes, predominated, whatever the grade, in in situ hybridization positive intraepithelial malpighian lesions (57 out of 59 cases; 97%). In the follow-up cases, there was no difference in the clinical course of the lesions in terms of grade (simple condyloma mild or moderate dysplasia) based on the hybridization results (detection and typing). These findings suggest that routine viral typing cannot be proposed to determine the therapeutic attitude (abstention, destruction) for the management of intraepithelial lesions of the cervix.

[Comparative study of the expression of CEA and a myelomonocytic antigen (CD15) in serous effusions using two monoclonal antibodies NEO 723 and Leu M1]. / Etude Comparative de l'expression de l'ACE et d'un antigène myélomonocytaire (CD15) dans les épanchements des séreuses à l'aide de deux anticorps monoclonaux NEO 723 et Leu M1.

A comparative study of the reactivity of two monoclonal antibodies (MAb), NEO 723 (anti-CEA) and Leu M1 (CD15) was performed by immunocytochemistry on sixty five reactive effusions and sixty two neoplastic effusions, fifty eight due to metastases from carcinomas, two due to disseminations of sarcoma and two due to malignant mesotheliomas. The study of the expected reactivity of NEO 723 and the cross-reactivity of Leu M1 on exfoliated neoplastic cells in effusion fluids showed that the sensitivity of NEO 723 was superior to that of Leu M1 for the detection of carcinomatous metastases, as 78% reacted with NEO 723 versus 38% with Leu M1. Among the positive cases, the mean number of reactive cells was twice as high with NEO 723, while only three of the carcinomas no expressing CEA reacted with Leu M1. The study of the reactivity of benign and malignant mesothelial cells with these two antibodies also confirmed the absence of labelling of these cells. Thus, despite a good specificity for carcinoma, the combination of these two antibodies provides only a minor gain in diagnostic sensitivity (+5%) compared with the use of an anti-CEA antibody alone and a loss of sensitivity (-5%) compared with the combination of an anti-CEA and an anti-EMA antibodies. These results appear to justify the suppression of Leu M1 from the first panel of antibodies screening for carcinomatous cells in favour of a combination of anti-CEA and an anti-EMA antibodies. However, Leu M1 may be useful as a second-line test in order to define the primary tumour responsible for the effusion.

Identification of late differentiation antigens of human cornified epithelia, expressed in re-organized desmosomes and bound to cross-linked envelope.

Little is known about the process leading to desquamation in cornified epithelia. We describe late differentiation antigens (Ag) specific for human cornified squamous epithelia, defined by two murine monoclonal antibodies (MoAb), G36-19 and B17-21, produced after immunization with plantar stratum corneum (SC). Histologically, in epidermis both Ag are cytoplasmic in the lower stratum granulosum (SG), become pericellular in the upper SG, and progressively disappear in the lower SC. In contrast, they persist up to the desquamating corneocytes in the palmoplantar epidermis and hard palate epithelium, as well as in the three cornified epithelial components of the inner root sheath (IRS) of the hair follicle (HF). Cytologically, both Ag are expressed as surface spots only on rough corneocytes. They are largely preserved on cross-linked envelopes (CLE) of the fragile type. Ultrastructurally, both Ag appear in keratinosome-like cytoplasmic vesicles in the upper stratum spinosum (SS) and the SG keratinocytes, then are found in both the regular and reorganizing desmosomes of the SG keratinocytes, and lastly in the corneocyte-specific reorganized desmosomes we propose to name corneodesmosomes. On CLE, the Ag are located on fibrils gathered over the external side of the envelope. Immunochemically, the G36-19--defined epitope is sequential and shared by five non-cytokeratin protein antigens of molecular weight 33.5, 36.5, 40, 49, and 52 kD, the higher molecular weight polypeptides being possibly precursors of the 33.5-kD protein. In contrast, the B17-21 epitope, unaccessible by immunoblotting, is probably conformational. In long-term cultured keratinocytes, the Ag are only expressed when epidermal sheets are morphologically differentiated. The expression is enhanced in the absence of fetal calf serum (FCS) and of epidermal growth factor (EGF). G36-19 and B17-21 Ag participate in a corneodesmosome-CLE superstructure that is probably involved in corneocyte cohesiveness and partly responsible for the mechanical resistance of the SC. These Ag are relevant markers for studying desmosomal maturation during epidermal differentiation and desquamation.

Natural IgG to epidermal cytokeratins vs IgG to the stratum corneum of the rat oesophagus epithelium, so-called 'antikeratin antibodies', in rheumatoid arthritis and other rheumatic diseases.

In order to study the relationships between the circulating IgG autoantibodies to epidermal cytokeratins (AECK), which were described in normal human sera as well as in sera from patients with various diseases, and the so-called 'antikeratin' IgG antibodies ('AKA'), which are highly specific for rheumatoid arthritis (RA), we simultaneously investigated AECK by a specific ELISA using cytokeratins from human stratum corneum (SC) and 'AKA' by semiquantitative indirect immunofluorescence assay on rat oesophagus epithelium, in a large series of 595 rheumatic sera including 229 RA. AECK were found to be present in all the 595 sera, with large inter-individual variations in titre. Whatever the titre chosen as threshold, the autoantibodies (auto-Ab) were never found to be specific for any rheumatic disease. Moreover, in RA, they were found to vary independently of IgM rheumatoid factor (IgM-RF), erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP), while they were found to vary in parallel with the total serum IgG concentration. In contrast, although 568 of the 595 rheumatic sera contained antibodies that labelled the rat oesophagus SC, the highest titre-like values were obtained with RA sera. At a convenient threshold, 95 (41.5%) of the 229 RA were detected while only three false positives (0.08%) remained among the 366 non-RA sera. Moreover, in RA, 'AKA' were found to be related to IgM-RF, ESR and CRP, while their titre was found to be independent of the total serum IgG concentration. Lastly, no statistical correlation was found between the antibodies, either in the whole sample of 595 sera or in any diagnostic group. In conclusion, the simultaneous investigation of AECK and 'AKA' showed that they differ from each other in all the aspects explored. AECK belong to the widely explored family of natural auto-Ab against cytoskeleton components and do not constitute a diagnostic marker while, on the other hand, 'AKA' confirmed their high diagnostic specificity for RA. It can also be asserted that, in spite of their name, 'AKA' do not recognize human epidermal cytokeratins, at least in the denatured form they present in ELISA. Therefore, they recognize either conformational epitope(s) appearing on cytokeratins during the late stages of the cornification process, or epitope(s) borne by rat cytokeratins but absent on human cytokeratins, or lastly a non-cytokeratin SC antigen.

Effects of hypergravity on adherent human cells.

In recent years, accumulating evidence has shown that microgravity or hypergravity may affect cell growth and differentiation. Since it is not easy to carry out researches in space or to simulate weightlessness on earth, we conducted experiments on simulated hypergravity (2 to 15 g) by using a centrifuge (radius: 80 cm; speed motor: 180 rpm). We looked for the effects of chronic hypergravity (7 to 10 days) on cultures of three human cell lines: lung or dermic fibroblasts and lung adenocarcinoma A 549 cells. The results showed a significant decrease (10-20%, P<0.05) in cell proliferation connected to a significant decrease (20-50%, P<0.01) in culture DNA content under hypergravity, but only for lung fibroblasts. The protein content was never disturbed. Dermic fibroblast elastase activity was enhanced (8-13%, P<0.02) under 15 g. Total phospholipid content as well as relative amounts of phospholipid components, analysed by thin layer chromatography, were unchanged in A 549 cells.

Immunophenotyping of mesothelial cells and carcinoma cells with monoclonal antibodies to cytokeratins, vimentin, CEA and EMA improves the cytodiagnosis of serous effusions.

This paper presents an immunocytochemical study performed on cytocentrifuged deposits from 109 peritoneal and pleural effusions including 20 transudates, 43 malignant metastatic effusions and 46 effusions containing atypical cells, unidentifiable as reactive mesothelial or malignant epithelial cells on the classical morphological criteria. A panel of four monoclonal antibodies (MAb) was used, including KL1 directed to cytokeratins (KER), V9 to vimentin (VIM), NEO 723 to carcinoembryonic antigen (CEA) and E29 to epithelial membrane antigen (EMA). In most transudates the reactive mesothelial cells coexpressed VIM and KER with a ring-like pattern for the latter proteins. In contrast, they were unreactive to anti-CEA and weakly and inconsistently reactive to anti-EMA. In malignant effusions, most carcinoma cells coexpressed EMA, CEA and KER with a predominant diffuse cytoplasmic pattern for the latter. Only a few malignant epithelial cells from five metastatic adenocarcinomas weakly expressed VIM. When used on the 46 effusions with unidentifiable cells, the panel of MAb allowed reactive mesothelial cells and malignant epithelial cells to be distinguished from each other in 39 of 46 cases (85%).

Effects of hypergravity on lung carcinoma cells maintained in continuous organotypic culture.

The effects of hypergravity levels ranging from 1 to 15 g were studied on A549 lung adenocarcinoma cell line, cultivated as nodules. This organotypic culture model preserves as closely as possible the cellular structures and differentiation functions of the in vivo situation. Nodules submitted to hypergravity conditions for 27 d did not show any change of cell growth, protein and DNA contents, compared with controls. Also, cellular differentiation, as regards intracellular phospholipid composition and more particularly phosphatidylcholine content, appeared undisturbed. The only obvious effect of hypergravity was a modification of the structural organization, with a disappearance of the large alveoli present at the surrounding of control nodules and the development of a dense cellular mass instead.

Subclass distribution of IgG antibodies to the rat oesophagus stratum corneum (so-called anti-keratin antibodies) in rheumatoid arthritis.

Serum IgG, labelling the stratum corneum of the rat oesophagus epithelium, so-called anti-keratin antibodies (AKA) constitute the most specific marker for the diagnosis of rheumatoid arthritis. In this study, we investigated 31 IgG AKA-positive rheumatoid sera and 21 control sera from patients with non-rheumatoid inflammatory rheumatic diseases. The serum level of IgG1,2,3 and 4 was determined by radial immunodiffusion and the subclass distribution of IgG AKA by a three-step semi-quantitative immunofluorescence assay using standard monoclonal antibodies specific for each of the four human IgG subclasses. In the rheumatoid sera, the serum level of IgG1 was found to be significantly increased and the level of IgG2 significantly decreased with regard to the control sera, while the levels of IgG3 and 4 as well as total IgG were in the normal range. IgG1,2,3, and 4 AKA were detected in 27 (87%), 6 (19%), 4 (13%) and 11 (35%) of the 31 rheumatoid sera, respectively, and were found to be independent of the clinical and biological indices of the disease. In spite of inter-individual heterogeneity, two predominant profiles were distinguished: IgG1 (alone) and IgG(1 + 4), which together represented 18 sera (58%). The large predominance of IgG1 AKA and the quasi-absence of IgG2 AKA suggest that the recognized antigen may be partly comprised of protein. Moreover, the high frequency of occurrence of IgG4 AKA might result from chronic exposure to the eliciting antigen, which could be a genuine autoantigen since we demonstrated that it is also present in the stratum corneum of human epidermis.

High diagnostic value in rheumatoid arthritis of antibodies to the stratum corneum of rat oesophagus epithelium, so-called 'antikeratin antibodies'.

Serum antibodies to the stratum corneum of rat oesophagus epithelium, so-called 'antikeratin antibodies', have been largely demonstrated in rheumatoid arthritis (RA). IgM and IgG antibodies to this epithelium were studied by semiquantitative immunofluorescence in 528 patients with perfectly characterised rheumatic diseases, including 178 with classical or definite RA. Histological analysis of IgG antibodies showed that only antibodies which produce a linear laminated pattern restricted to the stratum corneum (IgG antikeratin antibodies) are highly specific for RA; all the other labelling patterns are not disease specific. By a semiquantitative evaluation of the stratum corneum fluorescence intensity it was shown that the diagnostic value of IgG antikeratin antibodies closely depends on their titre and it was established in objective conditions that the sensitivity is 43.26% when the specificity reaches 99.14%. A high titre of IgG antikeratin antibodies was actually pathognomonic for RA. Both the histological and semi-quantitative analyses showed that IgM antibodies to rat oesophagus epithelium, though frequently detected, are of no diagnostic value, either for RA or for any other rheumatic disease that was studied. From a review of all the international reports on IgG antikeratin antibodies it was found that, to date, 4080 patients, including 1694 with RA, have been assayed for antikeratin antibodies by 11 different research groups. Analysis of all the results obtained under comparable technical conditions showed that IgG antikeratin antibodies constitute the most specific serological criterion for the diagnosis of RA. Furthermore, it was found that their incidence does not depend on disease duration: they are present in one third of rheumatoid factor negative patients with RA, and they seem to be related to disease severity or activity, or both. Their detection in the diagnosis of rheumatic diseases should become systematic.