Mitoses in normal and psoriatic epidermis

The morning count of mitoses in uninvolved epidermis of psoriatics is not significantly different from normal skin. The afternoon count is higher than the morning in both, and significantly higher in uninvolved than in normal epidermis. These results may be interpreted as favouring the existence of a non-cycling G2 population in human epidermis. This G2 population could be released into mitosis by the factors involved in diurnal variation, and might be larger and more labile in psoriatics than in normals. The pathogenesis and aetiology of psoriasis may be related to such a G2 population.     

Alpha-L-fucosidase in normal and psoriatic epidermis

Alpha-L-fucosidase levels in psoriatic epidermis rule out the concept of a primary defect of this enzyme in psoriasis.     

Effect of enzyme digestion on anionic sites and charge-selective permeability of dermo-epidermal junction

To study components of anionic sites on the lamina densa of the dermo-epidermal junction (DEJ) and to assess the effect of removal of sialic acid or glycosaminoglycans on its charge-selective permeability, epidermal sheets, whose dermis had been removed by treatment with dithiothreitol, were digested with heparitinase, chondroitinase ABC, hyaluronidase, or neuraminidase. They were then stained with polyethyleneimine for demonstration of the anionic sites or incubated in a medium containing native anionic ferritin for tracer experiments. The anionic sites were completely removed after heparitinase digestion. Although the numerical density of the sites was not altered, their electron density was decreased after chondroitinase ABC digestion. The other enzymes had no effect on the sites. In the tracer experiments, heparitinase or neuraminidase increased the number of tracer molecules penetrating into the lamina lucida of the epidermal sheet, while the other enzymes had no effect on it. These data indicate that heparan sulfate, which is a main component of the anionic sites, plays an important role in the charge-selective permeability of the DEJ, whereas chondroitin sulfate, which seems to be contained in the sites, does not, probably because of its small amount. These data also indicate that sialic acid, which is not a main component of the anionic sites demonstrated with the cationic probe, has a role in the permeability function.     

Localization of transglutaminase 1 mRNA in normal and psoriatic epidermis by non-radioactive in situ hybridization

We examined the expression of the transglutaminase 1 (TGase 1) gene IN frozen sections of normal and psoriatic epidermis by means of non-radioactive in situ hybridization with digoxigenin-labelied cRNA probes. TGase l mRNA was expressed in the granular layer of normal epidermis. regardless of ortho-or hyperkeratosis. However, in psoriatic epidermis. TGasel mRNA was detected in the suprabasal spinous layer, but not in the subcorneal layer. These results indicate that TGase 1 gene expression is limited to the last stage of keratinization in normal epidermis and this regulation is disturbed in psoriasis.     

Cyclophilin content of normal and psoriatic epidermis

The unique, immunosuppressive agent cyclosporine A has been shown to be of important therapeutic value in the treatment of psoriasis and other inflammatory dermatoses. To investigate the basis for its therapeutic efficacy, the tissue and cellular content of cyclophilin, a cytosolic receptor for cyclosporine A, has been determined in keratome biopsies from normal and psoriatic epidermis and in cultured, adult human keratinocytes. The mean cyclophilin content of normal (n = 10), involved (n = 10), and uninvolved (n = 10) psoriatic epidermal samples was not significantly different (0.126 +/- 0.016, 0.106 +/- 0.009, and 0.153 +/- 0.018 microgram cyclosporine A bound/mg cytosolic protein). Similarly, the cyclophilin content of keratinocytes cultured from normal and psoriatic epidermis (0.21 +/- 0.016 and 0.18 +/- 0.024 microgram/mg protein, respectively) did not differ significantly. Western blot analysis of normal and psoriatic epidermal extracts with monospecific rabbit anti-cyclophilin antisera revealed a single band of 17,000 daltons, which co-migrated with highly purified bovine cyclophilin. The intensity of this band was similar in normal and psoriatic samples, indicating that immunoreactive cyclophilin content was similar. Cyclophilin mRNA was readily detected in normal and involved psoriatic epidermis by RNA blot hybridization, and expression was not significantly different in normal and psoriatic epidermis. These findings indicate that cyclophilin is present in human keratinocytes and epidermis, where it may account for the accumulation of cyclosporine during therapy. However, the similar cyclophilin content of normal and diseased tissue suggests that differences in cyclosporine uptake probably do not account for its therapeutic efficacy in psoriasis.     

Localization of elevated transforming growth factor-alpha in psoriatic epidermis

Biopsies of involved and uninvolved skin from five patients with plaque psoriasis and normal skin from four healthy volunteers were investigated for steady-state quantities of TGF-alpha RNA and protein by in situ hybridization and immunohistochemistry. Increased levels of TGF-alpha RNA were found only in the high-level keratinocytes of involved psoriatic skin (p less than 0.001). Elevated levels of TGF-alpha protein were seen in both the high-level and basal layers of involved psoriatic skin compared to uninvolved psoriatic and normal control skin. Elevated TGF-alpha gene expression is thus implicated in the hyperproliferation of keratinocytes and possibly the altered maturation pattern seen in psoriasis.     

Membrane-bound phospholipase C activity in normal and psoriatic epidermis

We report the quantification of a membrane-bound phospholipase C in human epidermis which is active against the physiologically relevant substrate, phosphatidylinositol 4,5-bisphosphate. The level of this enzyme is significantly increased in the psoriatic lesion, both on a weight and protein basis. Etiological implications of this observation are discussed.     

Ultrastructural localization of calcium in psoriatic and normal human epidermis

With ion capture cytochemistry, we previously demonstrated the distribution of calcium ions in murine epidermis, a pattern consistent with a role for this ion in the regulation of epidermal differentiation. Because of the known proliferation and differentiation defects in psoriasis, we compared the calcium distribution of involved vs uninvolved psoriatic lesions and normal human epidermis. Whereas normal human and uninvolved psoriatic epidermis revealed increased calcium-containing precipitates in the uppermost stratum granulosum, in contrast the basal layer of psoriatic lesions contained less extracellular calcium, a condition that favored enhanced proliferation. Moreover, all psoriatic suprabasal cell layers displayed heavier than normal concentrations of calcium, indicating loss of the normal calcium gradient that programs terminal differentiation. This abnormal profile may account for the differentiation defects (eg, parakeratosis) that occur in psoriasis. Finally, psoriatic lesions displayed retained ionic Ca in intercellular domains of the upper stratum granulosum with absence of normal intercellular bilayers, findings that may underlie the abnormal desquamation and permeability barrier in psoriasis.     

A novel histone-stimulated protein kinase in normal and psoriatic epidermis

During the course of studies on protein kinases in psoriatic epidermis, a novel histone-activated protein kinase activity was identified. This activity (referred to as PK-II because it was the second peak of protein kinase activity eluted from a DEAE column) was partially purified from the supernatant of an epidermal homogenate by DEAE-cellulose column chromatography. Although histone was not a substrate for phosphorylation, in the presence of histone, endogenous proteins of Mr 105 and 95 kDa were phosphorylated. Activity was not affected by Ca2+/phospholipid, cAMP, cGMP, cAMP-dependent kinase inhibitor, spermine, spermidine, calmodulin, EGF, or phorbol ester. Phosphorylation was specific for serine and threonine residues. A major peak of PK-II activity eluted from sepharose 6B with an apparent Mr of 100 kDa, suggesting that histone may stimulate autophosphorylation. The properties of PK-II resemble those recently described for a class of polypeptide-dependent protein kinases isolated from placenta, Ehrlich ascites tumor cells, and bakers' yeast. PK-II was significantly higher in psoriatic involved epidermis (32.6 +/- 11.6 pmol/min/mg protein) compared to psoriatic uninvolved epidermis (5.7 +/- 0.6 pmol/min/mg; p = 0.03) and normal epidermis (9.5 +/- 2.2 pmol/min/mg; p = 0.05). The function of histone stimulated protein kinase in epidermal function and its role in the pathophysiology of psoriasis remain to be explored.     

Low density lipoprotein receptor expression on keratinocytes in normal and psoriatic epidermis

Biochemical and morphologic studies on the interaction of low density lipoprotein (LDL) with cultured normal keratinocytes and squamous carcinoma cells have shown a negative correlation between LDL receptor activity and terminal differentiation of the epidermal cells [Ponec M et al, J Invest Dermatol 83:436-440, 1984 and Vermeer, BJ et al, J Invest Dermatol 86:195-200, 1986]. Whether such in vitro studies pertain to the epidermis in vivo is not known. To obtain information on the distribution of LDL receptors in the epidermis in situ, morphologic studies were performed using LDL-gold as an ultrastructural marker. When freshly isolated mouse and human epidermal cells were incubated with LDL-gold complexes, only keratinocytes with the morphologic characteristics of basal cells showed binding and uptake of LDL-gold. No LDL receptor activity was found on Langerhans cells, melanocytes or highly differentiated keratinocytes. Since cell separation techniques can destroy receptors, the staphylococcal epidermolytic toxin was utilized to produce intercellular and intra-epithelial splitting of the epidermis. In preparations of both normal mouse and human epidermis, LDL-gold binding was restricted to basal cells and a few suprabasal keratinocytes. In contrast, in psoriatic epidermis, and to a lesser extent, essential fatty acid-deficient mouse epidermis, cells in the stratum spinosum showed abundant LDL-gold binding. Thus LDL-gold may be a useful marker for epidermal differentiation.     

comparison between DNA synthesis and mitosis in uninvolved and involved psoriatic epidermis and normal epidermis

Biopsies have been taken from the involved and uninvolved skin of eleven patients with psoriasis and the normal skin of six patients without psoriasis, 1 h after in vivo injection of 10 μCi of ³H-thymidine. Mitotic counts and counts of labelled cells (synesizing DNA) after autoradiography have been carried out. The ratio of mitoses to labelled cells was 1: 25 in clinical psoriasis, 1: 190 in uninvolved psoriatic epidermis and 1: 450 in normal epidermis. The number of labelled cells in clinical psoriasis was approximately 12 times greater than normal epidermis. However, the number of labelled cells in uninvolved psoriatic epidermis was approximately twice that of normal. These results imply an alteration in the components of the cell cycle in both involved and uninvolved psoriatic epidermis.     

Histochemistry of the intercellular substance of the normal and psoriatic human epidermis

Summary The intercellular substance of skin samples obtained from normal subjects and from psoriatic patients has been studied with histochemical methods for carbohydrate containing substances and checked with enzymatic extractions. The surface coat which makes up most of the intercellular substance was stained with colloidal iron and with Alcian Blue solutions containing up to 0.20 M magnesium chloride; the stainings were heavily affected by the previous treatment of the sections with testicular hyaluronidase, but not with neuraminidase. The staining of the intercellular substance with Alcian Blue solutions containing up to 0.20 M magnesium chloride and the action of the hyaluronidase gives strength to the hypothesis that hyaluronic acid is contained in the substance. In the skin of psoriatic patients intercellular spaces wider than in normal skin and a reduced surface coat, particularly in the higher layers, has been observed.Paper presented at the V th International Congress of Histochemistry and Cytochemistry, Bucarest, 1976