Production of epidermal sheets in a serum free culture system: a further appraisal of the role of extracellular calcium.

Rhenwald and Green's technique is currently the standard method for growing stratifying epidermal cell cultures. The serum free system developed in Ham's laboratory (MCDB 153) was designed to grow keratinocyte monolayers in clonogenic conditions. Our aim was to optimize conditions in serum-free MCDB 153 for culturing epidermal sheets from adult normal skin, and to assess the effect of extracellular calcium and temperature on proliferation and differentiation of cultured keratinocytes. Sixteen strains derived from plastic surgery specimens (mean age of donors 37 years; range 5-89) were used. Primary cultures were seeded at an optimal density of 8 x 10(4) cells/cm2 in primary cultures and 10(4) cells/cm2 in secondary cultures in complete medium including EGF, insulin, hydrocortisone and bovine pituitary extract, supplemented with isoleucine, tyrosine, methionine, phenylalanine, tryptophane and histidine. Amino acid (AA) supplementation allows a 5.8-fold increase in cell counts at confluency and monolayers with densely packed cells are obtained. In AA supplemented cultures, confluency is obtained in 16 +/- 3 days in primary cultures and in 13 +/- 0.5 days at first passages. Switches to 1.1 mM calcium at first or second passages resulted in a significant increase in cell counts (P less than 0.001), when compared with AA supplemented low calcium cultures. Low temperature/low calcium cultures resulted in a 50% decrease in cell counts. Low temperature/high calcium cultures gave similar cell counts as the 37 degrees C controls. AA and calcium supplemented cultures were evaluated for differentiation markers: involucrin expression was increased, keratins 5, 6, 14, 17 were expressed, and the sheets were 6-10 layers thick by electron microscopy, with keratohyalin granules and cornified envelopes appearing at layers 3-6 (from basal layer). Dispase treatment allowed an easy detachment of these sheets. These results show that the culture medium MCDB 153 can be adapted without serum supplementation to batch culture of human adult keratinocytes to produce epidermal sheets suitable for grafting. They also indicate that extracellular calcium in physiological range of concentration is not a sufficient signal for growth arrest when other growth conditions are optimized.

[Cardiac involvements in scleroderma: study with echocardiography and cardiac Doppler in 38 patients]. / Atteintes cardiaques au cours des sclérodermies: étude en écho-cardiographie et Doppler cardiaque chez 38 patients.

A study of heart lesions has been conducted in 38 patients with systemic sclerosis using echocardiography, doppler and colour doppler. Abnormalities were found in 66% patients. Valvulopathies (mainly mitral regurgitation) were the most frequent lesions (45%). The presence of anti-SCL70 antibody was correlated to right cavities dilatation. Pulmonary hypertension was found in 6 patients.

Perineal ecthyma gangrenosum in infancy and early childhood: septicemic and nonsepticemic forms.

BACKGROUND: Ecthyma gangrenosum is characterized by necrotic ulcerations surrounded by an erythematous halo. It is secondary to Pseudomonas aeruginosa infection. Most lesions are located in the anogenital and axillary areas, but the route of infection is generally difficult to establish. OBJECTIVE: We report six children with perineal ecthyma gangrenosum and discuss predisposing factors, origin, and route of infection. METHODS: This was a retrospective clinical study. RESULTS: Three children had blood cultures positive for P. aeruginosa, and one died. Predisposing factors were present in all cases; two had received chemotherapy (neuroblastoma, acute lymphoblastic leukemia), and two had idiopathic granulocytopenia. The last two patients previously had received treatment with systemic antibiotics and had abnormal granulocyte killing several months later. CONCLUSION: Septicemic ecthyma gangrenosum can be rapidly fatal in young children and requires aggressive antibiotic therapy. Benign ecthyma gangrenosum in healthy infants may result from a modification of bowel microflora after antibiotic therapy in conjunction with maceration in the diaper area. However, careful evaluation and long-term follow-up must be done to detect neutropenia, functional abnormalities of granulocytes, or a possible immune deficiency.

[Gangrenous ecthyma of the diaper area in infants]. / Ecthyma gangréneux du siège du nourrisson.

Ecthyma gangrenosum due to Pseudomonas aeruginosa is a skin infection in which necrotic ulcerations surrounded by a red areola develop. The diaper area is the region most often involved in infants. Typically, ecthyma gangrenosum occurs in patients with septicemia and risk factors (chemotherapy, neutropenia). However, transient bacteremia or an infection confined to the skin may be the cause in some patients, with maceration in the diaper area and previous antibiotic therapy as risk factors.

Porphyria cutanea tarda associated with human immunodeficiency virus infection. A study of four cases and review of the literature.

We report the cases of 4 male subjects, 29, 32, 41 and 44 years old, presenting isolated seropositivities for the human immunodeficiency virus (HIV), or full-blown acquired immunodeficiency syndrome, associated with a typical porphyria cutanea tarda (PCT). The 4 patients are in the usual risk groups for HIV infection. Viral hepatitis was observed in 3 of the 4 cases. Over the past 3 years, 15 cases associating HIV infection and PCT have been reported; almost all had the usual risk factors for HIV infection and hepatopathy. We speculate that HIV infection may have favored the occurrence of early PCT in these cases by altering the metabolism of the porphyrins, either directly or by means of the associated hepatopathy.