Extensive congenital bilateral epidermal naevus syndrome—a case report from Nigeria with ultrastructural observations

A case of an extensive congenital bilateral verrucous epidermal naevus syndrome is reported from Nigeria. No naevus cells were detected by both tight and electron microscopes. There was an exuberant epidermis with ultrastructural abnormalities of Leratinocuc degeneration in upper stratum spinosum, accumulation of highly electron–dense amorphous keratohyaline granules, and absolute deficiency of fibrous substance of intermediate-cell type in the stratum granulosum and the cornified layer.     

The Keratinization Disorder in Collodion Babies Evolving into Lamellar Ichthyosis*

Two collodion baby girls with disorder evolving into lamellar ichthyosis were followed by light and electron microscopy. Light microscopically, the neonatal collodion skin was characterized by a thick compact stratum corneum which was PAS positive in its upper two thirds, by a thin stratum granulosum and by a non-acanthotic stratum spinosum with normal mitotic activity. Electron microscopically, the upper stratum corneum appeared pathological, whereas the lower part was normal except for some minor parakeratosis. The main alterations in the underlying stratum granulosum were diminished tonofibrils and keratohyalin. Biopsy specimens taken at the age of 2 weeks were typical for lamellar ichthyosis and showed hyperkeratosis with focal parakeratosis, a thickened stratum granulosum in which the cellular content of keratohyalin and tonofibrils was moderately diminished, and acanthosis with increased mitotic activity. It appears that the ultrastructural changes of the stratum granulosum, seen in lamellar ichthyosis, are already present in the collodion skin of the newborn, at a time when the epidermis does not yet show an increase in mitotic activity.     

The action of sodium lauryl sulphate on rat skin-an ultrastructural study

An assessment was made of the ultrastructural changes in the epidermis of clipped rat skin following six washings over 3 days with 1% sodium lauryl sulphate. Findings included deposition of lipid in most cells, marked spongiosis and vesiculation, partial detachment of basal cells from the basal lamina, isolated rupture of the basal lamina, condensation of tonofibrils within cells of the stratum basale and stratum spinosum, protein deposition between the deeper epidermal cells (epidermal oedema/blister formation) and thickening of the epidermis and stratum corneum. There was a reduc-tion in the number of keratohyalin granules, with no change in their structure or average size. A 1% solution of sodium lauryl sulphate can cause marked general damage to the epidermis of clipped rat skin under the conditions described above.     

Embryology of the epidermis: ultrastructural aspects. II. Period of differentiation in the mouse with mammalian comparisons.

A detailed light and electron microscopic study of the cellular morphology of the epidermis in the 13 through 16 day mouse fetus reveals that an occasional intermediate cell is interposed between the basal and peridermal layers on day 13. All layers are mitotically active. Tonofilaments, unassociated with desmosomes, are present within the basal cell cytoplasm and the mitotic axis of the basal cells has changed from a parallel to a perpendicular plane with respect to the epidermal surface. At 14 days, a complete stratum intermedium, composed of one or two cell layers, is present. Rarely, developing hemidesmosomes are observed. Pools of glycogen are present in all cells below the periderm. The periderm is dense and no longer mitotically active. The skein of filaments, present in the inferior cytoplasmic region of the basal cells on days 12 and 13, is now absent. In the 15 day fetus, numerous developing hemidesmosomes are present. The stratum intermedium contains three to four layers of cells, and filaments are located deep within the cytoplasm of these intermediate cells. Rarely, a few developing keratohyalin granules and keratinosomes are present. A stratum intermedium is no longer present in the 16 day fetus. This region is now composed of a stratum spinosum and a stratum granulosum. Numerous keratinosomes are located in the upper stratum spinosum and lower stratum granulosum. The cells in the stratum granulosum are nucleated and the uppermost cells contain large keratohyalin granules. Three heterogeneous and one homogeneous type of keratohyalin granule is described. Dense bodies are present within mitochondria, nuclei, glycogen pools and the peridermal cytoplasm. The periderm is no longer dense and glycogen and keratohyalin granules are not observed in this layer.     

Embryology of the epidermis: ultrastructural aspects. III. Maturation and primary appearance of dendritic cells in the mouse with mammalian comparisons.

The epidermis of mice ranging in age from prenatal day 17 through postnatal day 4 and, in addition, postnatal day 18, was studied with the electron microscope. In the 17 day fetus, the periderm may or may not be present and a stratum corneum is developing in the latter case. The cells of the strata spinosum and granulosum contain much glycogen and many keratinosomes and homogeneous keratohyalin granules which appear to line up in a row near the distal cell membranes, fuse and form keratinized cells which contain material similar in density to that of the individual keratohyalin granules. In the 18 to 20 day fetuses, no periderm is present. The stratum corneum becomes well-developed. The quantity of glycogen decreases but the number of keratinosomes and keratohyalin granules increases. Some cells in the basal region contain necrotic phagocytosed cells. Few changes occur in the epidermis of the neonate, although the stratum corneum increases in thickness. In the early postnatal period, the mouse epidermis is mature in appearance and resembles that of man. By postnatal day 18, the whole epidermis is much thinner, although all classical strata can usually be identified. The primary appearance and subsequent maturation of epidermal dendritic cells was also studied in the mouse during the embryonic, fetal, early postnatal and 18 day postnatal periods. Melanocytes which lacked cell processes were first identified in the 15 day fetus. Melanocyte cell processes and an increasing number of melanosomes, as well as melanocytes, are present between prenatal day 18 and postnatal day 3. On postnatal day 4, fewer melanocytes are present and they contain vacuoles. Phagocytosed melanocytes are also present within basal keratinocytes. Undifferentiated dendritic cells or indeterminate cells are observed beginning on prenatal day 16. These cells do not appear to increase significantly in number subsequently. On postnatal day 18, Langerhans cells and indeterminate cells are present but no melanocytes can be identified. No Merkel cells were observed. Mitotic dendritic cells or dendritic cells traversing the basal lamina were not observed.     

An immunohistochemical study of abnormal keratinocyte differentiation in molluscum contagiosum

To investigate abnormalities in the keratinization process in lesional epidermis of molluscum contagiosum, production of filaggrin, loricrin, Ted-H-1 antigen, involucrin, cystatin A and CD95 ligand (CD95L) was investigated using specific antibodies. Anti-filaggrin monoclonal antibody (MoAb) did not react with keratohyalin granules (KHG), but with the substance around virus particles in the stratum corneum. KHG reacted with anti-loricrin polyclonal antibody (PoAb) and anti-Ted-H-1 MoAb. Anti-involucrin PoAb and anti-cystatin A PoAb reacted with materials in the cytoplasm of the middle stratum spinosum to the stratum granulosum. CD95L was expressed in the cell membrane region of the living cell layers in lesional epidermis. These observations suggest that the keratinization process may be altered in molluscum contagiosum.     

Development and characterization of a canine skin equivalent

The development of a complex cellular model, which incorporates the basic cell components of the dog skin, would be a useful tool to investigate the biology and pathology of canine skin and also to replace animal testing partially. The aim of the present study was to develop and characterize a canine skin equivalent. Epidermal keratinocytes and dermal fibroblasts were freshly isolated from skin biopsies from healthy dogs. Fibroblasts were embedded into a bio-matrix from collagen type I matrix protein; this built the scaffold where the keratinocytes were seeded, at air exposed conditions. At 3, 7, 15 and 21 days of culture in special growth media, skin equivalents were analysed by histological, immunohistochemical and electron microscopical techniques. At 15 days, keratinocytes underwent differentiation to a multilayer epidermis with stratum basal, stratum spinosum, stratum granulosum and stratum corneum. Expression of epidermal cytokeratins in keratinocytes was detected by immunhistochemistry, and followed the same pattern than in the normal canine epidermis. Fibroblasts from the skin equivalent expressed vimentin as dermal fibroblasts do. A basement membrane (BM) was observed underneath the epidermis; ultrastructurally, it was similar to the normal canine BM and collagen IV and laminin 5 were detected immunohistochemically as major components of this structure. Skin equivalents developed from canine cutaneous cells presented a similar morphological structure than healthy canine skin. Moreover, the immunohistochemical analysis revealed the expression of the major markers of the epidermis (keratins), dermis (vimentin) and BM (collagen type IV, laminin 5).     

Development of the epidermis from fish to human]

The epidermis, the outermost layer of all individuals, is a point of contact between individuals and their surroundings, especially insofar as it mediates sensory stimuli, but it also has the function of separating them from their surroundings and providing protection from harmful environmental influences. This paper traces the adaptation of the epidermis of vertebrates to these multiple functions via various methods of differentiation. In the human, epidermal differentiation is divided into synthesis, transformation, and terminal stages, and the stratum corneum is the final product of this differentiation. The differentiation products (tonofilaments, keratohyalin granules, membrane coating granules, cornified envelopes) are discussed with reference to their molecular bases (especially cytokeratin polypeptides, filaggrin, involucrin, lipids). The importance of these new cell-biological results for the pathogenesis of disorders of keratinization is discussed.     

Autosomal dominant ichthyosis exfoliativa

An unusual ichthyosiform dermatosis with circumscribed areas of blistering hyperkeratoses as well as scaly areas was observed in five consecutive generations of a German family. Light and electron microscopical studies revealed oedema of the keratinocytes in the upper epidermis but no defects of tonofilaments and keratohyalin granules. We suggest that this type of ichthyosis exfoliativa inherited as an autosomal dominant trait represents a new and distinct entity.     

Study of normal and pathological keratinization using anti-keratin polypeptide sera (author's transl)

Anti-keratin polypeptide sera were obtained against the different bands of polyacrylamide gel electrophoresis of fibrous proteins of stratum corneum derived from normal human epidermis. The sera were tested by indirect immunofluorescence of immunoperoxidase techniques. It was demonstrated that antibodies against P1 and P2 polypeptides, of MW 67,000 and 62,000 daltons respectively, were directed towards cytoplasmic antigens of keratinocytes of the upper Malpighian layers, while no labelling could be detected in the basal cell layer. Anti-P3 polypeptide sera and the anti-whole keratin serum labelled the whole epidermis, including the basal cell layer. Ultrastructural immuno-labelling performed on free epidermal cells obtained after trypsinization demonstrated that receptors for anti-P1 polypeptide sera were tonofilaments. These results showed that some keratin components (P1 and P2 polypeptides) might be absent in basal cell tonofilaments. This in in favour of various differentiation stages of the keratinizing cells. Keratin polypeptide 1 could be a useful marker of keratinization. According to preliminary studies, the expression of this keratin antigens was markedly disturbed in tumors such as basal and squamous cell carcinoma, in warts and in ichthyosis.     

Ultrastructural localization of lectin-binding sites in normal skin

The distribution of carbohydrate residues in keratinocytes of normal epidermis was studied. Normal skin was embedded in Lowicryl. Thin sections were incubated with concanavalin A (Con A), peanut agglutinin (PNA), wheat germ agglutinin (WGA), Ulex europaeus agglutinin I (UEA I), dolichos biflorus agglutinin (DBA), and soybean agglutinin (SBA). A positive reaction in the dermis, in the basal lamina (lamina densa, lamina lucida), intracellularly and within the plasma membrane including the desmosomes was obtained after incubation with Con A and WGA. PNA binding sites were found predominantly in the plasma membrane between the desmosomes. The labeling with Con A, WGA, and PNA was most pronounced in the upper stratum spinosum and granulosum. Incubation with UEA revealed heavy labeling of the keratohyalin granules and the cytoplasm of the corneocytes. Incubation with DBA and SBA revealed weak labeling of the keratinocytes. The study of the distribution of carbohydrate residues in normal epidermis is important, since alterations in this distribution might be linked to autoimmunity or malignant transformation.     

E‐FABP induces differentiation in normal human keratinocytes and modulates the differentiation process in psoriatic keratinocytes in vitro

Epidermal fatty acid‐binding protein (E‐FABP) is a lipid carrier, originally discovered in human epidermis. We show that E‐FABP is almost exclusively expressed in postmitotic (PM) keratinocytes, corresponding to its localization in the highest suprabasal layers, while it is barely expressed in keratinocyte stem cells (KSC) and transit amplifying (TA) keratinocytes. Transfection of normal human keratinocytes with recombinant (r) E‐FABP induces overexpression of K10 and involucrin. On the other hand, E‐FABP inhibition by siRNA downregulates K10 and involucrin expression in normal keratinocytes through NF‐κB and JNK signalling pathways. E‐FABP is highly expressed in psoriatic epidermis, and it is mainly localized in stratum spinosum. Psoriatic PM keratinocytes overexpress E‐FABP as compared to the same population in normal epidermis. E‐FABP inhibition in psoriatic keratinocytes markedly reduces differentiation, while it upregulates psoriatic markers such as survivin and K16. However, under high‐calcium conditions, E‐FABP silencing downregulates K10 and involucrin, while survivin and K16 expression is completely abolished. These data strongly indicate that E‐FABP plays an important role in keratinocyte differentiation. Moreover, E‐FABP modulates differentiation in psoriatic keratinocytes.     

Expression of the l-fucose moiety on epidermal keratinocytes in psoriasis induced by the Koebner phenomenon: a sequential study

The expression of Ulex europaeus agglutinin (UEA I) binding sites on cell-surface glycoproteins has been used as a marker for terminal differentiation. Increased number of UEA I binding sites of L-fucose specificity have been demonstrated in psoriatic epidermis. The results of lectin-binding studies in a series of biopsies taken sequentially (0 min, 5 min, 24 h, 7 days and 8 weeks) after tape-stripping of uninvolved skin in 12 psoriatic patients (three of whom were taking diltiazem, a calcium blocker at the time of the study) and six controls are presented. UEA I binding sites, which were expressed on the granular layer and upper layers of the stratum spinosum of pre-tape stripped uninvolved skin in psoriatic individuals, were progressively more numerous, with the expression of the L-fucose moiety on the lower stratum spinosum keratinocytes in the 7-day post-tape-stripping biopsies and 8-week biopsies, correlating with a moderate and marked increase in the proliferative index, respectively. In the Koebner-negative and non-psoriatic individuals who failed to develop psoriasis after tape-stripping, the UEA I binding sites were not expressed on keratinocytes of the lower stratum spinosum in any of the biopsies, although a mild increase in the proliferative index was noted in the 7-day biopsies. Our data suggest that the increased commitment of keratinocytes to terminally differentiate may be involved in the psoriatic process.(ABSTRACT TRUNCATED AT 250 WORDS)     

ULTRASTRUCTURE OF SENILE KERATOSIS

Three cases of a hypertrophic type of senile keratosis were examined by an electron microscope. All three showed similar changes, although some variability from case to case was also observed. Lesional epidermis showed widened intercellular spaces with microvilli-formation, a general increase in the bulk of the basal cells, a decrease in number of desmosomes, a decrease in the amount of tonofilaments, various degrees of aggregations of tonofibrils, binucleated keratinocytes, mitotic figures, prominent nucleoli, and an increase in number of nuclear bodies. In lesional dermoepidermal junctions, discontinuity of the basal lamina with cytoplasmic projections of the basal cells to the upper dermis was not infrequently observed. These changes are similar to those found in Bowen's disease, but are somewhat milder. In addition, in one case, keratinocytes of lesional epidermis showed lamellar structures closely associated with the rough endoplasmic reticulum. Significance of these structures is discussed.     

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.     

Localization of anionic sites in normal and psoriatic epidermis: the effect of enzyme digestion on these anionic sites

Cell surface anionic charge is known to be related to various cellular functions. Therefore, we ultrastructurally localized anionic sites in normal and psoriatic human epidermis, using poly-l -lysine-gold complex (cationic gold), to assess their possible participation in the differentiation of keratinocytes and the pathogenesis of psoriasis. In normal and psoriatic epidermis, the cell membrane of keratinocytes showed positive staining at pH 2.0. At pH 7.4 the cytoplasm and nucleus were diffusely stained, in addition to the cell membrane. In normal epidermis, the intensity of labelling on the cell membrane at pH 2.0 was strong in the basal layer and lower stratum spinosum, and decreased in parallel with differentiation of keratinocytes. In psoriatic epidermis, the intensity of labelling on the cell membrane at pH 2.0 was stronger than in normal epidermis. In normal epidermis, heparitinase digested 63% and chondroitinase ABC digested 80% of cationic labelling. This suggests that heparan sulphate and chondroitin sulphate (and/or dermatan sulphate) constitute anionic sites in normal epidermis. In psoriatic epidermis, chondroitinase ABC-sensitive anionic sites were greatly increased, whereas heparitinase-sensitive anionic sites were the same, when compared with normal epidermis. This suggests that chondroitin sulphate and/or dermatan sulphate constitute anionic sites which are increased in psoriatic epidermis.     

The growth and differentiation of cultured newborn rat keratinocytes

Keratinocytes were cultured from adult and newborn rat epidermis using the 3T3 feeder cell technique. By modifying culture conditions a long-lived line of newborn rat keratinocytes was developed which showed a plating efficiency of 40% and a doubling time of 16 h. The cells produced stratified colonies with tonofilaments, desmosomes, cell envelopes, and keratohyaline granules. When the cells were grown on a collagen gel they formed a thick stratum corneum and many keratohyaline granules. The fibrous proteins synthesized by the newborn rat cultured keratinocytes were different than those of newborn rat epidermis but similar to those of adult rat cultured keratinocytes. A histidine-rich basic protein was identified by immunologic techniques but it appeared to be more heterogeneous than that of newborn rat epidermis. A cell envelope precursor protein was identified by dansyl cadaverine incorporation studies and was identical to a major envelope precursor of newborn rat epidermis. The growth characteristics, colony morphology, and biochemical markers did not change for up to 40 passages and there was no evidence of malignant transformation. Because of their case of growth and long-term survival these cells are useful for studying a variety of problems related to keratinization.     

Richner-Hanhart's syndrome: ultrastructural abnormalities of epidermal keratinization indicating a causal relationship to high intracellular tyrosine levels

Richner-Hanhart's syndrome (corneal dystrophies, palmoplantar keratoses, and mental retardation) is caused by high levels of L-tyrosine in the blood, probably due to a defect of soluble tyrosine aminotransferase. Biopsies of skin lesions of 3 cases revealed peculiar ultrastructural changes that were not found in controls and have not been recorded before. Thickening of the granular layer and increased synthesis of tonofibrils and keratohyalin occurred in all cases. In the ridged palmar or plantar skin large numbers of microtubules and unusually tight packing of tonofibrillar masses were regularly demonstrable, the latter containing tubular channels or inclusions of microtubules. It is assumed that increased cohesion and tight packing of tonofilaments could prevent normal spreading of keratohyalin and result in its globular appearance. No crystal formation was observed in epidermal keratinocytes nor was there lysosomal damage. A biochemical model to correlate these ultrastructural findings to known biochemical and clinical features is proposed. It is suggested that excessive amounts of intracellular tyrosine enhance cross-links between aggregated tonofilaments and modulate the number and stability of microtubules.     

Location of HLA-A,B,C antigens in dendritic cells of normal human skin: an immunoelectron microscopic study

The distribution of the major histocompatibility antigens HLA-A,B,C, (HLA) on dendritic cells of normal human skin was studied by immunoelectron microscopy and a 4-step immunoperoxidase technique utilizing monoclonal antibodies. Light microscopy revealed peripheral staining for HLA of epidermal and pilar infundibular keratinocytes. In the epidermis, the staining was present from the basal layer to the upper stratum spinosum. In the follicles below the level of the infundibulum, HLA was detected only on rare intraepithelial dendritic cells. These dendritic cells could not be identified in the epidermis due to the HLA staining of the surrounding keratinocytes. Similar cells stained diffusely with anti-T6 antibody; the keratinocytes did not. Immunoelectron microscopy demonstrated: (1) the presence of HLA staining of keratinocyte membranes from the stratum basalis to the level of the upper stratum spinosum and in the pilar infundibulum, (2) the possible absence of HLA on melanocytes, (3) the presence of focal HLA staining of the membranes of epidermal and follicular dendritic cells that contained Birbeck granules and were, therefore, Langerhans cells, (4) dendritic mononuclear cells within the follicular epithelium, which although devoid of Birbeck granules, exhibited similar reactivity with anti-HLA antibody. These findings suggest that HLA antigens are present on the membranes of Langerhans cells, but are not demonstrable on melanocytes in normal human skin.