High endothelial venules in involved and uninvolved psoriatic skin: recognition by homing receptors on cytotoxic T lymphocytes.

We report the presence of tannic acid staining material in the intercellular spaces between adjacent endothelial cells of the high endothelial venules (HEVs) in psoriatic skin, which appears to be recognized by T8 (cytotoxic/suppressor) lymphocytes, as the presence of HEVs was found to be related to the presence of T8 lymphocytes in the epidermis. As HEVs with similar tannic acid staining material were also observed in peripheral lymph nodes, we suggest that this material may serve as a marker for HEVs recognized by the T8 lymphocyte subset. T8 + lymphocytes were found in the epidermis at 5 min and I h after tape-stripping of uninvolved skin of psoriatic patients with active disease, but not until 24 h in a psoriatic patient in remission. The prior existence of HEVs in uninvolved psoriatic skin could account for the rapid egress of T8 lymphocytes from the vasculature to the epidermis in response to trauma.

Ultrastructural evidence for the presence of bacteria, viral-like particles, and mycoplasma-like organisms associated with Giardia spp.

Giardia trophozoites and cysts, isolated from mammalian and avian hosts, were examined by transmission electron microscopy (TEM), scanning electron microscopy (SEM), and by fluorescent light microscopy for the presence of microbial symbionts. Mycoplasma-like organisms were observed on the surfaces of trophozoites isolated from the prairie vole, laboratory rat, and beaver. Intracellular bacteria were observed by TEM in the trophozoites and cysts of G. microti and by fluorescence microscopy in trophozoites and cysts of Giardia spp. isolated from beaver, muskrat, great-blue heron, and the green heron. Trophozoites of G. muris from rat small intestine contained viral-like particles measuring 60 nm in diameter. These observations suggest that biological associations between Giardia spp. and diverse microbes may be more common than formerly appreciated. It also raises the possibility of transmission of these apparent symbionts, via the Giardia cyst, to other mammalian hosts including man.

Erythroderma associated with mixed lymphocyte-endothelial cell interaction and Staphylococcus aureus infection.

We report 11 consecutive cases of erythroderma, a high percentage of which were associated with the growth of Staphylococcus aureus in cultures from blood, joint fluid or skin. In biopsies from all the patients we found close morphological associations between lymphocytes and endothelial cells, with some of the lymphocytes showing features of blastoid transformation to T helper lymphocytes. In extreme cases, sheets of T cells, including T helper lymphocytes, formed a syncytium with endothelial cells in the dermis. Marked capillary proliferation was noted both on light and electron microscopy. We suggest that erythroderma is precipitated by antigens such as protein A, a potent T cell mitogen present on the cell surface of Staphylococcus aureus, or by drugs, such as phenytoin. These antigens induce antigen presentation by individual endothelial cells, leading to T helper transformation and lymphocyte proliferation. Endothelial proliferation resulting from lymphocyte-endothelial interaction results in the vascular proliferation associated with this syndrome.

Significance and pathogenesis of basal keratinocyte herniations in psoriasis.

Using transmission electron microscopy, we studied, quantitatively, basal keratinocyte herniations (BKH) in relation to the other basement membrane zone changes in psoriatic lesions of varying clinical activity, and in psoriasiform skin diseases. BKH appears to correlate with disease activity. They do not occur passively as a result of the formation of gaps in the basal lamina. BKH in active psoriasis are associated with electron-lucent areas suggestive of proteolytic enzyme release. Their apparent association with Langerhans cells, neutrophils, macrophages, and endothelial cells may point to these cells as the source of proteolytic enzymes in psoriasis. BKH may prove to be a useful marker for clinical psoriasis.

Rod photoreceptor cells dissociated from mature mice retinas.

Intact rod photoreceptors were dissociated from pronase-treated whole retinas of adult mice by repeated passage through a plastic pipette tip. Hemocytometer counts of the cell suspensions indicate that, during a series of ten dissociation steps, a total of about 1-2 million intact photoreceptor cells are dissociated from one adult mouse retina, with less than 5% contamination from Müller cells and neurons of the inner retina. Visual cells with rod outer segments (ROS) and synaptic terminals are released in each step, but they occur in the greatest number during the sixth to ninth steps; detached ROS are released most frequently in the early steps, and neurons of the inner retinal layers appear in the later steps of dissociation. Nuclei are found in each step. Cell intactness was estimated by Trypan blue and Erythrosin B exclusion and by microscopic analysis using differential interference optics or scanning electron microscopy. The cells bind lectins (concanavalin A, Ricinis communis, and wheat germ agglutinin but not peanut agglutinin), displaying surface topography like that observed in situ. The metabolic capacity of dissociated cells was assessed by measuring the utilization of 32P inorganic phosphate for the synthesis of phospholipids and for the light-dependent phosphorylation of rhodopsin. Mature photoreceptor cells were estimated to contain, on average, 6.4 X 10(-12) g DNA, 2.3 X 10(-12) g RNA and 42-64 X 10(-12) g protein. The dissociation procedure provides a population of photoreceptor cells that appears suitable for microscopic, electrophysiological, and biochemical analysis.

The sequence of events in psoriatic plaque formation after tape-stripping.

The sequence of events leading to the formation of a psoriatic plaque induced by tape-stripping was studied using light and electron microscopy. Among the earliest changes noted were increased mobility of the epidermal Langerhans cells across the basement membrane, evidence of Langerhans cell-lymphocyte interaction, and increased Langerhans cells 'activity' or 'cytotoxicity'. These changes were seen as early as 2 min after stripping and remained until the development of clinical psoriasis. Collections of epidermal lymphocytes showing the features of blastoid transformation while in contact with processes from activated Langerhans cells, suggest the involvement of Ia antigens in this process. We postulate that these findings are a manifestation of an increased immune responsiveness to trauma, controlled by genes located at the HLA-D locus of the major histocompatibility complex, and mediated by enhanced cellular interactions. The appearance of basal keratinocyte herniations (BKH) at 1-3 weeks after stripping, coincided with the development of clinical psoriasis. Neutrophils made their appearance at the same time as, or slightly before, the appearance of BKH, and are suspected to play a role in the development of these structures. We believe that BKH maintain epidermal proliferation through the persistence of the epidermal-stromal interaction.

Cone visual cell degeneration in ground squirrel retina: disruption of morphology and cyclic nucleotide metabolism by lodoacetic acid.

Photoreceptor degeneration was induced in the cone-dominant retina of the ground squirrel by intracardiac injection of iodoacetic acid. Morphology was examined and cyclic nucleotide levels were determined in retinas taken at various times between 35 min and 11 days after injection. Degenerating cone cells were first detected at day 1 and all cone cells were reduced to compact dense masses by day 4. Cellular debris was removed by macrophages that entered the photoreceptor layer from the inner neuroretina. Cyclic AMP levels of dark-adapted retinas were doubled 24 hrs after injection and were reduced to approximately 50% of the dark-adapted level of control retina between days 1 and 3. The concentration of cyclic GMP was 3 to 4 times higher than normal at 4 to 5 hrs postinjection, dropped to 9% of normal at day 4, and was 2% at day 11. Since these changes were coincident with the loss of morphologic integrity of cone cells, they imply that at least 50% of cyclic AMP and most of the cyclic GMP in the cone-dominant retina of the ground squirrel is present in cone cells. The elevation of cyclic AMP and cyclic GMP levels prior to pathological morphology suggests that the iodoacetic acid-induced disruption of cyclic nucleotide metabolism may be associated with the degeneration of the cone photoreceptors.

Cyclic nucleotides of cone-dominant retinas. Reduction of cyclic AMP levels by light and by cone degeneration.

Dark-adapted retinas or whole eyes of 13-line ground squirrels (Citellus tridecemlineatus) and western fence lizards (Sceloporus occidentalis) contain higher levels of cyclic AMP than of cyclic GMP. In these cone-dominant retinas, light reduces cyclic AMP content selectively. Freezing of dark- or light-adapted retinas or eyes also reduces cyclic AMP content, with only minimal changes in cyclic GMP levels. In addition, exposure of frozen retinas of dark-adapted ground squirrel to light results in a significant decrease in cyclic AMP content. The destruction of cone visual cells of ground squirrel retina by iodoacetic acid injection decreases the cyclic nucleotide content of the dark-adapted retina. Considering the relative loss of cyclic nucleotides from cone degeneration, we estimate that the content of cyclic AMP in visual cells of ground squirrel retina is about four times greater than that of cyclic GMP.

Cyclic nucleotides in rod- and cone-dominant retinas.

Rod-and cone-dominant retinas differ in their relative content of cyclic AMP and cyclic GMP. Cyclic GMP is concentrated in retinas dominated by rods and is responsive to light; cyclic AMP is enriched in those having a majority of cones. Light reduces by about 50% the content of cyclic AMP in cone-dominant retinas, but the levels of cyclic GMP are affected only minimally. Microdissection of rod-dominant retinas shows that most of the cyclic GMP is localized in photoreceptor cells whereas cyclic AMP is evenly distributed throughout the retina. In contrast, our studies of cyclic nucleotides and the morphological changes that occur in cone visual cells of the ground squirrel retina, during hibernation and iodoacetate-induced cone degeneration, suggest that cyclic AMP is localized in cone visual cells. By analogy with the rod system, cyclic AMP may modulate the intracellular metabolism of cones.

Evidence for a complex life cycle and endospore formation in the attached, filamentous, segmented bacterium from murine ileum.

Light and electron microscope observations showed that the filamentous, segmented bacterium commonly found attached to the ileal epithelium of rats and mice undergoes a complex life cycle. Filaments comprising up to 90 segments were attached to the microvillous border of absorptive epithelial cells by a specialized terminal holdfast segment. Starting at the free end of the filament and progressing toward the attached end, undifferentiated segments were converted into reproductive or mother segments. Within each mother cell two new holdfast segments developed. As the holdfasts matured, their mother cells degenerated and released them into the intervillar space where they attached, grew, and divided to produce new segmented filaments. Alternately, in some filaments, newly formed but not yet released holdfasts were converted into endospores, which were released in the same manner as holdfasts, presumably to spread the bacterial colony to other members of the rodent population.

Plasmid-controlled colonization factor associated with virulence in Esherichia coli enterotoxigenic for humans.

An enterotoxin-producing strain of Escherichia coli isolated from a case of cholera-like diarrhea (E. coli strain H-10407) was found to possess a surface-associated colonization factor. Colonization was manifested as the ability of small inocula (10(5) bacteria) to attain large (10(9)) populations in the infant rabbit intestine with a concomitant diarrheal response. A laboratory-passed derivative of E. coli H-10407, designated H-10407-P, failed to exhibit an increase in population in the infant rabbit and also failed to induce diarrhea. Cell-free culture supernatant fluids of E. coli H-10407 and H-10407-P produced equivalent enterotoxic responses in infant and in adult rabbits. Specific anti-colonization factor antiserum was produced by adsorbing hyperimmune anti-H-10407 serum with both heat-killed and living cells E. coli H-10407-P. This specific adsorbed serum protected infant rabbits from challenge with living E. coli H-10407 although the serum did not possess bactericidal activity. The anti-colonization factor serum did not agglutinate a strain of E. coli K-12 possessing the K88 colonization factor peculiar to E. coli enterotoxigenic for swine. By electron microscopy it was demonstrated that E. coli H-10407, but not H10407-, possessed pilus-like surface structures which agglutinated with the specific adsorbed (anti-colonization factor) antiserum. E. coli H-10407 possessed three species of plasmid deoxyribonucleic acid, measuring 60 X 10(6), 42 X 10(6), and 3.7 X 10(6) daltons, respectively. E. coli H-10407-P possessed only the 42 X 10(6)- and the 3.7 X 10(6)-dalton plasmid species. Spontaneous loss of the specific H-10407 surface-associated antigen was accompanied by loss of the 60 X 10(6)-dalton species of plasmid deoxyribonucleic acid and loss of colonizing ability. Thus, it is concluded that the E. coli colonization factor described here is a virulence factor which may play an important and possibly essential role in naturally occurring E. coli enterotoxic diarrhea in man.

Bacteriophage-associated spherical bodies in Bacteroides fragilis.

Unique spherical bodies with multilayered walls were observed by electron microscopy in cells of a single strain of Bacteroides fragilis subsp. fragilis. Phage-like particles were present in the same cells, both free in the cytoplasm and within the spheres. The proportion of cells containing the phage-associated spherical structures ranged from less than 0.01% to about 7% depending on the culture conditions. Phage particles of morphological type B and spherical bodies were also found free in the medium surrounding the cells. Spherical bodies with discontinuities in their walls, through which phage-like particles sometimes appeared to be escaping, were also found both intra- and extracellularly. The biological significance of these distinctive spherical structures is a matter of conjecture.

Role of the mitochondrial genome during early development in mice. Effects of ethidium bromide and chloramphenicol.

The role of the mitochondrial genome in early development and differentiation was studied in mouse embryos cultured in vitro from the two to four cell stage to the blastocyst (about 100 cells). During this period the mitochondria undergo morphological differentiation: progressive enlargement followed by an increase in matrix density, in number of cristae, and in number of mitochondrial ribosomes. Mitochondrial ribosomal and transfer RNA synthesis occurs from the 8 to 16 cell stage on and contributes to the establishment of a mitochondrial protein-synthesizing system. Inhibition of mitochondrial RNA- and protein-synthesis by 0.1 microg/ml of ethidium bromide or 31.2 microg/ml of chloramphenicol permits essentially normal embryo development and cellular differentiation. Mitochondrial morphogenesis is also nearly normal except for the appearance of dilated and vesicular cristae in blastocyst mitochondria. Such blastocysts are capable of normal postimplantation development when transplanted into the uteri of foster mothers. Higher concentrations of these inhibitors have general toxic effects and arrest embryo development. It is concluded that mitochondrial differentiation in the early mouse embryo occurs through the progressive transformation of the preexisting mitochondria and is largely controlled by the nucleocytoplasmic system. Mitochondrial protein synthesis is required for the normal structural organization of the cristae in blastocyst mitochondria. Embryo development and cellular differentiation up to the blastocyst stage are not dependent on mitochondrial genetic activity.