Constituent Cell Types Research Articles

Nucleolar organizer regions and image analysis nuclear morphometry of small cell (nevoid) melanoma

Small cell (nevoid) melanomas may provide difficulties in diagnosis as their constituent cell type resembles a benign nevoid melanocyte. In the present study, 10 small cell melanomas were analyzed for the silver staining of their nucleolar organizing regions (AgNORs), and their nuclear area and perimeter were measured by computerized digital image analysis and compared with 10 superficial spreading melanomas lacking small cell differentiation and 10 dermal nevi. The average number of AgNORs per nucleus was 5.83 (SD +/- 1.69) for small cell melanomas and was significantly different when compared with 8.49 (SD +/- 1.58) for superficial spreading melanomas (p < 0.05) and 2.71 (SD +/- 0.50) for dermal nevi (p < 0.05). Digital image analysis confirmed that the nuclear perimeter and nuclear area of cells in nevoid melanomas did not significantly differ from those of ordinary dermal nevi (p > 0.05), but both group were significantly different from superficial spreading melanomas lacking a small cell morphology (p < 0.05). Counting AgNOR numbers may be useful in evaluating small cell (nevoid) melanomas and provides a technique for differentiating their constituent cell from ordinary nevus cells. Nuclear morphometry determined by digital image analysis may help better define the nuclear size in small cell melanomas.

Susceptibility of human foetal brain tissue to cool- and freeze-storage

Human second trimester foetal brain tissue was stored for a period of 1–6 weeks under various conditions in an attempt to evaluate factors influencing its susceptibility (cell loss) and survivability. Post-storage viability of mesencephalon, striatum, cerebellum and occipital cortex was assessed by a protocol combining vital staining with cell density counts so that tissue viability and cell loss could be evaluated simultaneously; tissue survivability was evaluated by cell culture. A significant amount of cell loss occurred after 24 h storage at room temperature, after one week at 4°C and by two weeks at −20°C in all structures; storage at −196°C resulted in 17–21% cell loss at the end of a 6 week period. At −20°C the cryoprotective effect of 20% FCS was equivalent to that of 15% FCS + 7% DMSO combined, suggesting potential use of serum in replacement of chemical additives. The procedure for removal of DMSO was critical to cell viability and survivability: single step dilution led to 27–39% greater cell loss than slow, multi-step dilutions. In comparison to fresh, non-stored tissue, immunocytochemical characterization of in vitro propagated stored tissue revealed no changes in the populations of major constituent cell types including neurones, dopaminergic neurones, glial and fibroblast cells. These results provide information on possible conditions under which transplant tissue can be satisfactorily stored depending on the prevailing requirements.

Adenosine receptors and signaling in the kidney.

It is now generally accepted that adenosine is capable of regulating a wide range of physiological functions. Nowhere is the diversity of this action better illustrated than in the kidney. When adenosine binds to plasma membrane receptors on a variety of cell types in the kidney, it stimulates functional responses that span the entire spectrum of renal physiology, including alterations in hemodynamics, hormone and neurotransmitter release, and tubular reabsorption. These responses to adenosine appear to represent a means by which the organ and its constituent cell types can regulate their metabolic demand such that it is maintained at an appropriate level for the prevailing metabolic supply. Extracellular adenosine, produced from the hydrolysis of adenosine 5'-monophosphate and stimulated by increased substrate availability and enzyme induction, acts on at least two types of cell surface receptors to stimulate or inhibit the production of cyclic adenosine-3',5'-monophosphate and also acts in some renal cells to stimulate the production of inositol phosphates and elevation of cytosolic calcium concentration. To understand when and why this complicated system becomes activated, how it interacts with other known extracellular effector systems, and ultimately how to manipulate the system to therapeutic advantage by selective agonists or antagonists, requires a detailed knowledge of renal adenosine receptors and their signaling mechanisms. The following discussion attempts to highlight our knowledge in this area, to present a modified hypothesis for adenosine as a feedback regulator of renal function, and to identify some important questions regarding the specific cellular mechanisms of adenosine in renal cell types.

Serum‐free culture of normal human melanocytes: Growth kinetics and growth factor requirements

Normal human epidermal melanocytes were selectively propagated from mixed (keratinocyte-melanocyte) cultures and primary epidermal cell suspensions in serum-free medium, MCDB 153 containing insulin, bovine pituitary extract (BPE), phorbol-12-myristate-13-acetate (PMA), ethanolamine, phosphoethanolamine, and hydrocortisone. Neonatal foreskin melanocytes (NFMs) replicated more readily than adult melanocytes in culture. Early passage NFMs grown in serum-free medium exhibited a population generation time of 24-48 hours. NFMs assumed a less dendritic appearance and were less pigmented than adult melanocytes. PMA or other protein kinase C-activating phorbol esters significantly enhanced mitogenesis of NFMs; however, cAMP-elevating agents were not required for efficient replication of NFMs. Basic fibroblast growth factor (bFGF) was a potent mitogen for NFMs and replaced the requirement for BPE in the culture medium. NFMs expressed a single class of specific, high-affinity receptors for bFGF, exhibiting a Kd = 3 x 10(-11) M and approximately 76,500 receptors/cell. Neither EGF nor TGF-alpha were mitogenic for NFMs, and TGF-beta reversibly inhibited NFM growth. Rapidly growing, early passage NFMs were shown to have cell cycle times of 19.5, 7.5, and 9 hours for G1, S, and G2/M phases of the cell cycle, respectively. Culture of NFMs to confluence or depletion of growth factors from the culture medium caused reversible, G1 phase-specific, cell cycle growth arrest. Senescence of NFMs was associated with irreversible growth arrest in the G1 phase after 40-45 population doublings in culture. Our data demonstrate that basal medium MCDB 153 can be supplemented with defined factors to cultivate selectively two major constituent cell types of the epidermis, the melanocyte and the keratinocyte.

TGF beta gene transcription in normal and neoplastic liver growth.

TGF beta is a potent, nontoxic inhibitor of mitogen-induced DNA synthesis in primary cultures of adult rat hepatocytes. Using a cDNA probe, we investigated TGF beta gene expression in quiescent, regenerating, and neoplastic liver, and several hepatoma lines by Northern gel analysis. We found that regenerating liver had increased TGF beta gene transcripts beginning at about 8 h, with a broad peak of 48-120 h and return to normal after 9 days. Separation of the regenerating liver into its constituent cell types, followed by RNA extraction and reprobing, revealed that increased TGF beta gene transcripts were confined to the enriched endothelial-cell population and not the hepatocytes. Increased hepatic TGF beta expression was also found in fetal liver and in rats immediately after birth. Elevated TGF beta mRNA levels were also found in primary cultures of oval cells and an established bile ductular cell line, as well as in carcinogen-altered liver epithelial cell lines. Transcripts were undetectable in normal human liver but were abundant in the human hepatoma lines Hep G2, Hep 3B, PLC/PRF/5, and SK-Hep-1. Elevated levels were also found in the normal rat liver-derived lines BRL-3A and clone 9 and the H4IIE rat hepatoma, but not in the HTC, MH1C1, and MH7777 rat hepatomas. The hepatocarcinogen diethylnitrosamine induced high transcript levels after single injections in a time- and dose-dependent manner. These results suggest that the liver may be a paracrine organ with respect to TGF beta gene expression, which can be induced by carcinogens and by growth stimulation.

Effect of cepharanthin on superoxide anion (O2-) production by macrophages.

AbstractOur previous report showed the inhibitory action of cepharanthin on oxygen‐derived free radical production by polymorphonuclear leukocytes (PMN). In the present study, we examined the effects of cepharanthin on production of superoxide anion (O2–), one oxygen radical, by macrophages in vitro and in vivo.Phorbol myristate acetate‐induced O2– production by mouse peritoneal exudate cells (PEC), whose constituent cell types were identified as macrophages, lymphocytes, and PMN (75, 20, and less than 5%, respectively), was measured by ferricytochrome C reduction assay. Superoxide anion production by PEC, which depended mainly on the macrophage component, was inhibited 34% by 5 μg/ml cepharanthin and 85% by 50 μg/ml. These results indicate that cepharanthin suppresses O2– production by macrophages as well as by PMN. The fact that no inhibition of O2– by a xanthine‐xanthine oxidase system was observed indicates that cepharanthin is not a scavenger of O2–.Carrageenan‐induced paw edema is due to the activity of macrophages. Participation of O2– and other oxygen radicals is implicated in this edema, because the swelling is inhibited by administration of superoxide dismutase. The effects of cepharanthin on O2– production by macrophages was examined using this experimental system in mice. However, no significant inhibition of the edema by cepharanthin was observed.

Sclerosing haemangiomas of the lung.

A series of 29 pulmonary sclerosing haemangiomas is analysed. Included is one case with multiple tumours and another with metastatic growth in a hilar lymph node. Histochemical and electron microscopic studies show that type 2 pneumocytes are an important constituent cell type though bronchial structures, tumorlets and angiomas also occur within these tumours. They are considered to be pulmonary hamartomas formed from distal lung structures. They are of slow growth and have been confused in the past with pulmonary histiocytomas and plasma cell granulomas.

Functional anatomy of the ovaries of pregnant and lactating Cape porcupines, Hystrix africaeaustralis.

The constituent cell types of the ovary of the porcupine were similar to those of New World hystricomorph rodents and accessory corpora lutea and luteal bodies were formed through the luteinization of the membrana granulosa or theca interna of antral follicles. All luteal bodies were histologically similar. The total volume of luteal tissue per female was not affected by fetal age and was unrelated to circulating concentrations of maternal plasma progesterone. Maternal plasma progesterone concentrations were correlated with fetal age. Follicular activity occurred throughout pregnancy but was not affected by fetal age or related to circulating values of oestradiol-17 beta. The formation of accessory corpora lutea during pregnancy is regarded as important in supplementing progesterone during pregnancy.

Localization of neurons innervating masticatory muscle spindle and periodontal receptors in the mesencephalic trigeminal nucleus and their reflex actions.

The mesencephalic trigeminal nucleus was studied in anaesthetized and curarized rabbits by recording the unitary activity through extracellular microelectrodes and identifying the constituent cell types. Two types of units were found, namely primary afferents supplying jaw raising muscle spindles and periodontal or gingival mechanoreceptors. These two groups of neurons exhibited a rostrocaudal somatotopy: the former occupied the entire rostral portion of the nucleus (A7-P2.3; trochlear decussation being taken as an arbitrary 0 level), the latter was located caudally (P3-P4.5) while the somata of both types of afferent fibres were present between P2.2 and P3. No evidence was found for representation of both tendon organs of jaw muscles and joint receptors. Among the units innervating muscle spindles, secondary afferents were largely more numerous than the primary ones. Among periodontal and gingival mechanoreceptor afferents, incisors were the most widely represented, followed by interalveolar gingiva and molars; the axonal conduction velocity ranged between 9 and 40 m/sec and between 8 and 16 m/sec for ipsilaterally and contralaterally projecting neurons, respectively. The motor responses obtained by electrical stimulation of discrete areas of the MTN confirmed the presence of a high degree of segregation between the two different populations of neurons. In fact, jaw raising movements are obtained when stimulating the area within A7 and P2 containing the somata of spindle afferent neurons, while only jaw opening movements are elicited by stimulation of the caudal levels of the nucleus. These data also show that the periodontal neurons whose somata are located in the MTN participate in the jaw opening reflex, just as the more numerous periodontal mechanoreceptors whose somata are located in the Gasser ganglion. Soma-somatic and soma-axon hillock gap junctions were found among the neurons of the MTN, particularly in the caudal third of the nucleus.

Characterization of monolayer and organ cultures of cloned and enriched lymphohematopoietic stroma cell populations.

The role of hematopoietic microenvironments in the regulation of maturation and differentiation of hematopoietic cells, although heavily debated, remains uncertain. Several investigators have suggested that the adherent "stromal" cell populations, which grow as colonies in cultures of lymphomyeloid tissues, include the cells involved in such regulatory processes. Grossly, the colonies described by several investigators appear similar morphologically, and the cells giving rise to them have been variously termed 1) fibroblast colony forming cells (FCFC), 2) plaque forming units-culture (PFU-C), 3) macrophage colonies, and 4) marrow stromal cells. FCFC have been reported to re-establish their parent microenvironment when transplanted in an allogeneic system. In this study, cloned and enriched cell populations obtained from such colonies in cultures of murine lymphomyeloid tissues have been characterized by their growth in culture and using morphological, histochemical, and electron microscopic techniques. The results demonstrated that, although the initial stromal colonies appeared to be identical, the constituent cell types varied considerably. Some colonies were comprised primarily of macrophages, while others appeared to contain predominantly fibroblasts; two additional cell types that established colonies have not yet been satisfactorily identified. These results demonstrate the heterogeneity of lymphomyeloid stromal colonies. There is a need for caution in the analysis of experiments in which uncharacterized stromal cell colonies are transplanted or employed as supporting monolayers in culture systems in experiments designed to evaluate the origins and functions of lymphohematopoietic stroma.

Benign nerve sheath myxoma: Light and electron microscopic features of two cases

Two new cases of the rare benign nerve sheath myxoma (NSM) are presented. The light microscopic findings are compared with previously reported cases. In addition, the ultrastructural features of three constituent cell types are described. The lesion is confirmed as a neural sheath neoplasm, and its similarities to other myxomas are discussed.

The cytology and occurrence of granulocytomas in mussels

A non-neoplastic inflammatory cellular condition in the marine mussel Mytilus edulis is described and a possible relationship drawn between its incidence and water quality as effected by chronic insult by domestic and industrial waste products. This condition, termed a granulocytoma owing to its dominant constituent cell type, is capable of overcoming host encapsulation and inducing atrophy and autolysis of the digestive gland and may therefore represent a terminal condition.

COMPARATIVE STUDIES ON TISSUE CORTICOSTEROID CONTENT AND MORPHOLOGICAL FEATURE IN FUNCTIONING ADRENOCORTICAL ADENOMA AND HYPERPLASIA

For the purpose of clarifying the functional significance of clear- and compact-type cells, correlative hormonal and morphological studies were per-formed on adrenocortical tumors associated with primary aldosteronism, Cushing's syndrome or non-dyshormonal symptoms. The content of aldosterone, corticosterone and cortisol in adrenal tissue was estimated by radioimmunoassay, and, on the surface of a quantitative sample, the ratio of its constituent cell-type was examined in sections with oil red O stain and/or H-E stain.The content of aldosterone and corticosterone was significantly higher in primary aldosteronism (P<0.001) than in Cushing's syndrome and in non-functioning tumor, with a mean value of 1.22±0.15 and 7.52±1.05ng/mg tissue, respectively. In Cushing's syndrome, cortisol content showed a high value, 9.27±1.60ng/mg tissue. The steroid content was different in each case, and varied with parts of the quantitative sample even in the same case.Though the correlation of aldosterone content and compact-type cell population in primary aldosteronism showed a negative trend and that of cortisol content and compact-type cell in Cushing's syndrome had a positive, neither correlation was statistically significant.

Plasminogen activator in early embryogenesis: Enzyme production by trophoblast and parietal endoderm

We have surveyed the early stages in the development and differentiation of cultured mouse embryos for plasminogen activator production. This enzyme is first detectable by the sixth equivalent gestation day. Thereafter, cultured blastocysts produce plasminogen activator with a biphasic time course: in the first phase, enzyme secretion rises to a maximum at about the eighth day and then decreases; a second phase, during which more enzyme accumulates, begins somewhat later and continues to at least the fifteenth day. By fractionating the blastocyst into its constituent cell types, we have identified the trophoblast as the cells responsible for the first phase of enzyme synthesis. The pattern of enzyme production by the trophoblast is closely correlated with the invasive period of these cells in vivo and implies that plasminogen activator is involved in embryo implantation. The second phase of plasminogen activator production is due to parietal endoderm, which initiates enzyme synthesis upon differentiation from the inner cell mass. The properties of the parietal endoderm suggest that plasminogen activator may participate in the migration of these cells and/or in the metabolism of Reichert's membrane which accompanies embryo growth. These results are consistent with the concept, developed from work on other cell types, that plasminogen activator may represent a generalized mechanism for tissue remodeling and cell migration.

Histological changes associated with enlargement and regression of the thymus glands of the red-billed quelea Quelea quelea L. (Ploceidae: weaver-birds).

Thymic lobes form over 200 red-billed queleas, Quelea quelea L., were examined histologically. Samples were taken from embryos about to hatch, juveniles and adults. The lobes varied in size from very small to very enlarged (1- greater than 5 mm long). The constituent cell types are described in detail and the occurrence of these cells in different sized lobes is discussed. A cycle of events is proposed which accounts for the observations presented here. It is suggested that the large numbers of erythroid cells found in the cortex of some individuals were developing in situ. The significance of erythropoiesis within the thymus is discussed.

A functional analysis of the components of the mesencephalic nucleus of the fifth nerve in the cat.

1. The mesencephalic nucleus of the trigeminal nerve has been studied using extracellular micro-electrode recording and the constituent cell types identified.2. Two types of unit were found, namely, muscle spindle first order afferents of ipsilateral jaw-closing muscles and mechanoreceptor afferents of ipsilateral maxillary and mandibular teeth.3. No evidence was found for representation of extra-ocular muscle stretch receptors, of temporo-mandibular joint receptors or of tendon organs of jaw muscles.4. Spindle units of each of the jaw-closing muscles were recorded in all parts of the nucleus and there was no evidence of their segregation according to muscle of origin.5. Attempts to classify spindle units by their dynamic response to ramp stretches, their following of high frequency vibration and their interspike interval variability at constant length gave no indication of two populations when fusimotor activity was suppressed.6. Following the injection of suxamethonium, however, units fell into two groups according to their dynamic index. Their behaviour resembled that described for primary and secondary spindle afferents. In data pooled from all of the jaw-closing muscles there were approximately equal numbers of units in each group.

The fine structure of the axial organ of the feather star Nemaster rubiginosa (echinodermata: crinoidea)

The fine structure of all the constituent cell type of the crinoid axial organ is described (coelomic epithelial cells, muscles, free cells, gland cells and neurons) ; also described is the fine structure of the extracellular haemal fluid. The gland cells of the glandular tubules of the axial organ have the characteristic fine structure of protein exporting cells and may produce granular and filamentous components of the haemal fluid. The neurons (perikarya and axons) of the axial organ may possibly be neurosecretory, since they are filled with electron-dense granules. Homologies between the axial organs of different echinoderm classes are discussed.