Arginine-vasopressin in anterior pituitary cells: in situ hybridization of mRNA and ultrastructural localization of immunoreactivity.

The hypothalamic nonapeptide arginine-vasopressin (AVP) exerts several distinct receptor-mediated actions on pituitary cells. Although hypothalamic AVP reaches the anterior pituitary via well-defined pathways, there is now accumulating evidence that AVP may also be produced endogenously in anterior pituitary cells. Using in situ hybridization, we demonstrate here the presence of AVP mRNA in the anterior pituitary of the rat. The observed grain density over pituitary cells was, however, greater than 10-fold lower than the one observed over AVP producing neurons present in the supraoptic and paraventricular nuclei of the hypothalamus. Immunoelectron microscopic analysis using two different AVP-specific antibodies revealed that the distribution of AVP-like immunoreactivity (AVP-LI) in the anterior pituitary is cell-specific. AVP-LI is most abundant in corticotrophs, followed by lactotrophs, gonadotrophs and thyrotrophs. On the other hand, there is complete absence of AVP-LI from somatotrophs. Interestingly, all pituitary cells in which AVP-LI is detected also represent potential target sites for AVP action. A minor fraction of AVP-LI was found to be membrane-associated and may originate, at least in part, from extrapituitary sources. This fraction likely represents receptor-bound peptide. The bulk of AVP-LI, however, was present in the cellular cytoplasm, not associated with any specific ultracellular structure. Specifically in corticotrophs, AVP-LI was excluded from secretory granules. However, our finding of AVP mRNA in anterior pituitary cells indicates that intracellular AVP-LI includes endogenously produced peptide, suggesting a paracrine and/or autocrine action.

Human immunodeficiency virus type 1 infection of U937 cells promotes cell differentiation and a new pathway of viral assembly.

The differentiation of U937 monoblastoid cells after human immunodeficiency virus type 1 (HIV-1) infection was studied using the following approaches: reverse transcriptase activity measurement, immunofluorescence labeling, and electron microscopy. For comparison, uninfected U937 cells were induced to differentiate from monocyte to macrophage by phorbol 12-myristate 13-acetate (PMA) or retinoic acid (RA) treatment. Both infected and drug-treated cells showed important and similar ultrastructural cell modifications, with a phenotype that decreased in monocyte specificity and increased in that of macrophages. When U937 cells were induced to differentiate upon HIV-1 infection, a very different pathway of viral production was observed. Production and accumulation of the virus in a vacuolar compartment of intracytoplasmic origin and escape to the antiviral lysosomal activity could explain virus persistence. This makes the cell system a good model with which to study the relationship between HIV-1 production and cell differentiation.

Outer root sheath cells of human hair follicle are able to regenerate a fully differentiated epidermis in vitro.

During wound healing, interfollicular epidermis can be regenerated from the outer root sheath of hair follicles, showing that the cells of this structure can shift toward an interfollicular epidermal phenotype. Similarly, it has been shown that a multilayered epithelium originating from outer sheath cells can be obtained in vitro by culturing hair follicles. However, in the culture systems developed so far, the phenotypical shift was incomplete since the cells retained some of their original characteristics and did not acquire several key markers of terminally differentiated epidermis. In this paper, we describe a new tissue culture method for obtaining a multilayered epithelium from outer sheath cells. This is performed by implanting human hair follicles vertically into dermal equivalents and then raising the culture at the air-liquid interface. The morphological, immunological, and biochemical features of the in vitro reconstructed tissue are very similar to those observed in normal interfollicular epidermis, including those specific for terminally differentiated keratinocytes. Thus, under appropriate in vitro conditions, outer root sheath cells are able to express an interfollicular epidermal phenotype as occurs in vivo during wound healing.

Normal Merkel cells express a synaptophysin-like immunoreactivity.

Synaptophysin (SY), a specific component of the membrane of presynaptic vesicles, has been reported as a novel marker for neurons, certain neuroendocrine cells and their neoplasms including neuroendocrine carcinomas of the skin. The origin of the Merkel cells (MC) being far from clear, this study was performed to establish if normal MC express SY. It is demonstrated by immunofluorescence and immunoelectron microscopy using a monoclonal antibody SY38 to this glycoprotein that normal MC in man, rabbit and pigs express an SY-like reactivity. Although immunoblotting identification of the immunoreactive material gave negative results, it is likely that normal MC contain SY. By immunoelectron microscopy, the staining was located at the surface of cytoplasmic vesicles. In view of the possible involvement of SY in the Ca2+-dependent neurotransmitter release, the observation of an SY-like immunoreactivity in MC supports the view that they are epithelial neuroendocrine cells and that they may possess a neurosecretory function.

Herpes gestationis factor reacts with the amniotic epithelial basement membrane.

Sera from five patients with clinically and immunopathologically proven herpes gestationis were studied by complement fixing immunofluorescence and complement fixing immuno-electron microscopy using specimens of skin, amniochorion and placenta. The results demonstrated that the complement fixation antibody (herpes gestationis factor) could bind to the basement membrane zone of skin, amnion and chorion laeve but not to that of the placental syncytiotrophoblast. These data suggest that the herpes gestationis factor may be induced by the basement membrane zone antigens of extra-villous cytotrophoblasts.

Ultrastructural aspects of skin after cryoultramicrotomy.

Skin was studied by transmission electron microscopy after cryoultramicrotomy. Global and ultrastructural preservation of the tissue was obtained with satisfactory cohesion between dermis and epidermis. The main constitutive elements could be observed with good definition allowing easy recognition of all the organelles and therefore absolute identification of each cellular type encountered. The methodology provided a comparable quality to the conventional technique. Ice crystal artefacts were minimized but slight enlargement of intercellular spaces persisted. One of the main advantages of this procedure is that neither lipidic solvents nor embedding resins, are used, so that one can envisage cytochemical studies.

Lamellar cells of sensory receptors and perineural cells of nerve endings of pig skin contain cytokeratins.

The lamellar cells of the sensory corpuscles of the pig dermis must be considered to be epithelial cells as they contain cytokeratins. The cytokeratins detected are similar to those found in simple epithelia. Moreover, lamellar cells are embedded in an extracellular matrix reminiscent of the basement membrane of epithelium since it contains laminin and collagen IV. The perineural cells surrounding the nerves of pig dermis present the same features. These results suggest that lamellar cells and perineural cells have the same origin. The nature of the lamellar and perineural cells of the rabbit or human dermis is not as clear since cytokeratins were not detected in those cells. These results, together with recent observations on Merkel cells, may indicate that epithelio-neuronal junctions are a general feature of cutaneous sensory receptors.

Identification of a 37 kilodalton protein at the epidermal basement membrane by an antiserum to human amnion.

A protein which is recognized by an antibody to human amniotic epithelial basement membrane was identified at the basal lamina of human epidermis by immunohistology. This protein was localized at the lamina lucida of human epidermal basement membrane by immunoelectron microscopy. Studies of normal human keratinocyte cultures and epidermal wound healing suggested that the protein was probably produced by keratinocytes. By immunoblotting, a basic apparent isoelectric pH (pHiapp = 7.3) protein band of 37 kD was seen. These data indicate that this 37 kD protein, clearly different from other known basement membrane components, is present in simple and stratified epithelia of ectodermal origin, and is associated with hemidesmosomes.

High-yield purification of plasma membranes from transformed human keratinocytes in culture.

The density pertubation technique with cationic silica microbeads was applied to prepare highly purified plasma membranes from cultured human keratinocytes. Trypsinized cells were coated successively with the beads (diameter approximately 50 nm, gravity greater than 2 g/cm3) and polyacrylic acid before they were lysed by osmotic shock and mechanical shear. The plasma membranes remained in the form of large open sheets which could easily be separated from other cell organelles and the cytosol by low-speed centrifugation. The membrane preparation was characterized by scanning and transmission electron microscopy, marker enzyme activities, one-dimensional sodium dodecyl sulfate polyacrylamide electrophoresis, and the specific beta-adrenergic receptor count. A yield of 79 +/- 9% was calculated by comparing the amount of beta-adrenoceptors in the purified membrane preparation with that of a crude cellular particulate fraction. The specific beta-adrenoceptor count of these two preparations was 1.2 +/- 0.02 and 0.2 +/- 0.05 pmol/mg protein, respectively, indicating a 6-fold improved purification with this microbead technique. The purified membranes were essentially free from contamination of other cell organelles.

Scanning electromicroscopy of suction blister-roofs from psoriatic lesions and normal skin.

Suction blisters were raised on psoriatic lesions and normal appearing skin. The epidermis was separated at the epidermal-dermal junction. Scanning electronmicroscopy of the dermal side of the blister-roofs from normal looking skin and almost healed psoriatic lesions showed stellate cells probably formed by cytoplasmic extensions ending at desmosomes. In non-treated psoriatic plaques the cells were rounded and lacked the stellate extensions.

Familial rolled and spiral hairs with palmoplantar keratoderma.

A 59-year-old man with palmoplantar keratoderma and rolled spiral hairs on the abdomen and extremities is reported. His father had the same skin manifestations but his brother and sister only keratoderma palmoplantare. Scanning electron microscopy of the rolled hairs showed that they were coiled in a spiral around their own axis. These spiral hairs had lower cysteine than the normal appearing hairs on the body. The scalp hair appeared normal but was low in cysteine which was compensated by an increase in threonine. Urine analysis showed a decrease of cysteine.

Actin containing filaments in sheep choroid plexus cells infected in vitro by Visna virus: an immunofluorescent and ultrastructural study.

The in vitro infection of sheep choroid plexus cells by Visna virus induces changes in the distribution and the orientation of actin containing filament bundles. During the cell fusion the stellate shape of cells with several interdigitations is associated with radial spanning of the contractile system. The intercellular contacts are characterized by the occurrence of a discontinuous line of rich actin containing dots which could be a junction of several short filament bundles.

[Ultrastructural aspects of sheep choroid plexus cells during exogenous cell fusion induced by Visna virus in vitro]. / Aspects ultrastructuraux de cellules de plexus choroïdes de mouton en voie de fusion cellulaire exogène induite in vitro par le virus Visna.

During the first moments of the in vitro infection of Sheep choroid plexus cells with high multiplicities of infection of Visna virus, the authors observed: a strong development of GERL, some signs of an intensive protein synthesis and numerous filament bundles. This infection leads to an exogenous cell fusion.

[Technic for the morphometric study of cells in culture, about the in vitro stimulation of ovarian stroma cells by H.C.G]. / Mise au point d'une technique d'étude morphométrique de cellules en culture à propos de la stimulation in vitro de cellules du stroma ovarien par H.C.G.

Sizes for ovarian stroma cells in culture are according to a log normal distribution. Cell stimulation by H.C.G. promotes a significative decrease of these sizes.

[Evidence for the production of a fusion factor during in vitro infection of sheep choroid plexus cells by visna virus]. / Mise en évidence du facteur de fusion produit au cours de l'infection in vitro de cellules de plexus choroïdes de mouton par le virus visna.

One of the most important early events during the infection of sheep choroïd plexus cells in culture by Visna virus is the synthesis by these cells of a fusion factor distinct from the virus. The cell fusion activity of this factor does not seem transmissible. It promotes cell shape changes and a migration of the nucleus towards the cell wall.

Inorganic cytoplasmic inclusions in alveolar macrophages. The role of cigarette smoking.

A case of tobacco-associated pulmonary fibrosis, with the results of histological, ultrastructural, and spectrometric analysis is reported. Abnormalities of the alveolar macrophages, which are particularly affected by tobacco inhalation were found. The size of the macrophages was increased and many large, polymorphous inclusions, including fat vacuoles and granular deposits, which were either homogeneous or electron lucent vacuoles, were seen in the cytoplasm. A few laminar structures were observed. All of these lesions are frequently found in cigarette smokers. Still more interesting was the discovery of numerous fiber-, needle-, or laminar-like inclusions that varied in size from 0.2 to more than 2 mu. The digestions of the inclusions with potassium hydroxide confirmed the presence of various metals, such as sodium, magnesium, potassium, iron, sulfur, and especially, aluminum, and silicon; these last two elements correspond to the presence of kaolinite in the tissue, as has been previously described, and can be considered as evidence of the use of tobacco.