Marion D. Kendall

The Anatomy of the Tarsi of Schistocerca gregaria Forskål

SummaryThe tarsus of S. gregaria is divided into three units (here called segments) and an arolium set between a pair of claws. The first segment bears three pairs of pulvilli in the fore and middle legs, and one pair and two single pulvilli in the hind legs. Segment two bears a pair of pulvilli, segment three one long pulvillus and the arolium a similar pad on the undersurface. The outer layers of the arolium pad differ from those of the pulvilli in possibly lacking an epicuticle and in having a layer of cuticle which, unlike the corresponding layer in the pulvilli, does not stain with protein stains. The claws and dorsal surfaces bear trichoid sensilla, basiconic sensilla and campaniform sensilla. Smaller basiconic sensilla and canal sensilla occur on the proximal part of the pulvilli, and basiconic sensilla on the arolium undersurface. Internally the cuticle is modified in the arolium and pulvilli so that rods of probably chitin and resilin are formed. This would impart flexibility to the undersurfaces whilst retaining some degree of rigidity which might prevent damage to the small and delicate sense organs on the pulvilli. The tip of the arolium is specialised for adhesion, and there are two large neurones internally which could conceivably monitor attachment or detachment of the tip. There are chordotonal organs in segment three, and several other large neurones throughout the tarsus, some of which are associated with the slings of tissue holding the apodeme in a ventral position. Gland cells occurring in the dorsal epidermis of the adult mature male are also briefly described.

Reversal of ageing changes in the thymus of rats by chemical or surgical castration

SummaryDifferences in the thymus of young and old male CSE Wistar rats were examined by use of routine histological stains on paraffin-embedded sections. There was a highly significant loss of thymic weight and disruption of architecture with age. Both surgical castration and chemical castration induced by a luteinizing hormone-releasing hormone analogue (Goserelin) caused a significant increase in thymic weight and the reappearance of a well-defined cortex and medulla in ageing rats. Cell surface antigens were detected on cryosections after incubation with a range of monoclonal antibodies. The Pan T cell marker (detected with antibody W3/13) showed fewer positive cells in ageing rats, and an increase after chemical castration. The smaller glands of old rats had fewer positive T cells with CD4 (MRC OX35) and CD8 (MRC OX8) antigens, and more after chemical castration in both young and ageing rats, but the greatest changes were seen in the intensity of Class II major histocompatibility complex (MRC OX6) immunoreactivity. In both young and ageing chemically-castrated rats, the numbers of cells and the intensity of immunoreactivity were greatly increased in the medulla.

The thymus in the mouse changes its activity during pregnancy: a study of the microenvironment.

The mouse thymus changes dramatically during pregnancy. It shrinks in size, and the cortex is extensively reduced from midpregnancy onwards. Despite this, there is surprisingly little evidence for any increase in apoptosis, and considerable evidence that mitosis of thymocytes continues throughout pregnancy. In spite of overall involution the thymic medulla actually expands in midpregnancy due to a combination of mitosis of epithelial cells and an accumulation of lymphocytes. The extent and nature of these changes are examined in this study at the ultrastructural level. The epithelial cells of the subcapsular cortex (type 1 cells) become wrinkled and exhibit powers of phagocytosis, whilst the other cortical epithelial cells are relatively unchanged, although the formation of epithelial/thymocyte rosettes and thymic nurse cells is more clearly seen in midpregnancy than usual. Other changes associated with pregnancy involve the medullary epithelial cells that undergo an increased level of mitosis. Their greater numbers surround accumulations of lymphocytes to form the characteristic medullary epithelial rings. Cell movement through blood vessel walls was clearly observed in midpregnancy, but not at other times. Interdigitating cells in the medulla become more conspicuous as pregnancy proceeds and the cells become phagocytic. The endoplasmic reticulum in plasma cells becomes expanded, indicating increased secretory activity. These results highlight the active nature of the thymus in pregnancy in spite of its involution. This picture contradicts the conventional notion that an involuted thymus is inactive.

Immuno‐electron microscopy of the thymic epithelial microenvironment

Normal T cell development depends upon interactions between progenitor cells and the thymic microenvironment. Monoclonal antibodies (Mabs) have been used to define subtypes of thymic epithelium by light microscopy (clusters of thymic epithelial staining [CTES]). We have now used a range of these Mabs together with gold‐coupled reagents in immuno‐electron microscopy to study the fine cellular distribution of the molecules to which the antibodies bind. Anti‐cytokeratin antibodies were used to identify all thymic epithelial cells, while the distribution of MHC class II molecules was revealed with Mabs to shared nonpolymorphic determinants. MR6, a CTES III Mab, shows strong surface labelling of cortical epithelial cells and thymic nurse cells and very weak surface staining of thymocytes, medullary macrophages, and interdigitating cells. Mab 8.18 (CTES V) also labels a cell surface molecule; this is present on Hassalls corpuscles and associated medullary epithelial cells. The molecules detected by Mabs MR6 and 8.18 are therefore located in a position where they are available to interact with external cellular and soluble signals within the thymus. In contrast, Mabs MR10 and MR19 (CTES II) recognise intracellular molecules within subcapsular, perivascular, and medullary epithelium. A similar distribution was seen with Mab 4β, directed against the thymic hormone thymulin, although, in addition to the expected intracellular epithelial staining, large lymphoblasts in the subcapsular zone showed surface positivity, indicating the presence of thymulin bound to surface receptors on these early lymphoid cells. As expected, MHC class II molecules were expressed on some medullary and essentially all cortical epithelial cells. However, although most subcapsular epithelium was class II–negative, some cells did express these MHC molecules on their apical surface and on the surface of their cytoplasmic extensions into the cortex. Interestingly, some cortical epithelial cells surrounding capillaries were positive for both MR6 (CTES III) and for MR10, MR19, and 4β (CTES II). Double‐labelling experiments, using MR6 and MR19 simultaneously, revealed a double‐positive perivascular epithelial cell population in the thymic cortex. The possibility that these cells represent a thymic epithelial progenitor population is discussed. Microsc. Res. Tech. 38:237–249, 1997.

Studies on rat and human thymus to demonstrate immunoreactivity of calcitonin gene‐related peptide, tyrosine hydroxylase and neuropeptide Y

The peptidergic and noradrenergic innervation of rat and human thymus was investigated by immunohistochemistry at the light and electron microscopical level (avidin‐biotin‐complex, sucrose‐phosphate‐glyoxylic‐acid, and immunogold techniques). The distribution of noradrenergic neural profiles, and positive immunoreactivity for calcitonin gene‐related peptide (CGRP), tyrosine hydroxylase (TH) and neuropeptide Y (NPY) is described in female rats during ageing, and in human children. In the neonatal rat thymus, the arteries and septa are well supplied by fine varicose nerves. In older animals (2 wk–1 y) the number of septa and blood vessels increase and consequently also the innervation. No nerves were found in the cortex. Apart from the innervation of the septal areas, immunoreactivity for CGRP and TH was present in thymic cells. Except for the young rats (neonatal–14 d), all rats showed CGRP positivity in subcapsular/perivascular epithelial cells (type 1 cells). All rat thymuses also contained a few TH positive cells in the medulla, which could only be confirmed as epithelial cells (type 6 cells) in children. Type 1 cells in the human thymus were not CGRP positive, but as in the rat, there were similar TH positive cells in the medulla. It was concluded that in addition to nerves containing CGRP, noradrenaline or dopamine, epithelial cells also contain these transmitters. They could therefore act on different cells (compared with neural targets) in a paracrine manner.

The brain and thymus have much in common: a functional analysis of their microenvironments

Research into the neural and immune systems has begun to converge. Since the first reports that interleukins play important roles in both systems and that lymphocytes secrete neuronal factors, scientists have been surprised by the ever-increasing list of interactions. Here, Rolf Mentlein and Marion Kendall examine the major supporting cells of the brain and thymus - astrocytes and thymic epithelial cells - the similar neuroectodermal origin of which could explain such fundamental analogies.

Neuropeptides Exert Direct Effects on Rat Thymic Epithelial Cells in Culture

To determine if major thymic neuropeptides and neurotransmitters can directly influence the functional activity of cultured rat thymic epithelium, neuropeptides and neurotransmitters were applied, and intercellular communication, proliferation, and thymulin secretion assessed. After injections of a mixture of lucifer yellow dextran (too large to pass gap junctions) and cascade blue (which does) into single cells, some neuropeptides decrease dye coupling: 0.1 mM GABA (P < 0.0001), 100 nM NPY (P < 0.0001), 100 nM VIP (P < 0.001), 100 nM CGRP (P < 0.001), 100 nM SP (P < 0.01), and 0.1 mM histamine (P < 0.01), whereas 0.1 mM 5-HT, mM acetylcholine, and 1 μM isoproterenol (β-adrenergic agonist) had no effect. Proliferation (incorporation of tritiated thymidine) was increased by CGRP (P = 0.004) and histamine (P < 0.02), but decreased by isoproterenol (P = 0.002), 5-HT (P = 0.003), and acetylcholine (P < 0.05). The percentage of multinucleate cells was decreased after isoproterenol (2.5%), and increased after 5-HT (21.3%), GABA (15%), and histamine (15.1%). Compared to controls, thymulin in the supernatant was decreased after challenge with acetylcholine (52%), isoproterenol (71%), 5-HT (73%), and histamine (84%). This study demonstrates direct effects of neuropeptides and neurotransmitters on functional aspects of cultured thymic epithelial cells.

Dietary minerals in the gastrointestinal tract: hydroxypolymerisation of aluminium is regulated by luminal mucins

The regulation of mineral absorption in the gastrointestinal tract is poorly understood. Recent work has identified an intracellular metal-ion transporter but considerable evidence suggests that both soluble and mucosally associated luminal metal-binding ligands regulate initial uptake. Molecules ranging from low molecular weight organic acids to large glycoproteins have been suggested but a definite role for any such species has remained elusive. Here, a series of analytical techniques, allowing for this wide variation in potential binding ligands, was applied to the study of intestinal contents and tissue of rats following different feeding protocols. Aluminium, that has a low endogenous background and maintains a high concentration in the gastrointestinal tract, was investigated as a suitable dietary metal with hydrolytic behaviour similar, for example, to copper, iron and zinc. High resolution nuclear magnetic resonance spectroscopy identified a number of endogenous low molecular weight weak ligands that are secreted into the intestinal lumen. These may slow the rate of hydroxy-polymerisation of hydrolytic metals, allowing their effective donation to less mobile, higher molecular weight binding ligands. Histochemical staining suggested that such species may be soluble mucins as these were consistently associated with luminal aluminium. Significantly, this interaction prevented hydroxy/phosphate precipitation of aluminium, even at supraphysiological levels of the element. This was confirmed with X-ray micro-analysis investigations of ex vivo luminal contents. Nevertheless, from phase distribution experiments, the majority (60-95%) of luminal aluminium was associated with the intestinal solid phase and further histochemistry confirmed this to be gelatinous mucus, chiefly as the mucosally adherent layer. All results suggest a major role for mucus in regulating the gastrointestinal absorption of aluminium. It is proposed that, initially, soluble luminal mucus prevents the hydroxy-precipitation of hydrolytic metals at intestinal pH, allowing their effective donation to the mucus layer. Based on the differing reported metal-mucus interactions, elements that bind well to mucus (Al3+, Fe3+), with kinetically slow rates of ligand exchange (Al3+ < Fe3+) will be less well absorbed than poorly bound elements with kinetically faster rates of ligand exchange (Cu2+, Zn2+ etc.). This mechanism would readily explain many of the reported observations on mineral availability, including the marked variation in absorption of different elements, the differential effects of dietary ligands on mineral uptake and the competition for absorption between different metals.

Implantation of cultured thymic fragments in congenitally athymic (nude) rats

Cultured thymic fragments correspond to the thymic microenvironment depleted of lymphocytes and dendritic cells. When these fragments are implanted under the kidney capsule of congenitally athymic rats, lymphocytes and dendritic cells of host origin enter the graft and induce thymus-dependent immunity in the recipient. This paper describes the ultrastructure of the fragments and the changes that occur during the restoration of normal thymic architecture. At the end of the culture period of 6-9 days and in the early stages after implantation, the grafts consist of keratin-containing epithelial cells of unusual morphology that can be labelled with antibodies raised against the epithelium of the mid/deep cortex and the subcapsule/medulla. Normal thymic architecture develops, including nerves and blood vessels, as lymphocytes populate the environment, and by 4-6 weeks the epithelial cells are the same phenotypically and ultrastructurally as those found in normal rat thymus. However, some areas without lymphocytes still contain the atypical epithelial cells seen before implantation. Large multinucleated giant cells are also present with a few associated epithelial cells of subcapsular/medullary phenotype. In conclusion, the cultured thymic fragments contain a hitherto unknown precursor epithelial cell with an atypical ultrastructure and phenotype that is not seen in normal development.SummaryCultured thymic fragments correspond to the thymic microenvironment depleted of lymphocytes and dendritic cells. When these fragments are implanted under the kidney capsule of congenitally athymic rats, lymphocytes and dendritic cells of host origin enter the graft and induce thymus-dependent immunity in the recipient. This paper describes the ultrastructure of the fragments and the changes that occur during the restoration of normal thymic architecture. At the end of the culture period of 6–9 days and in the early stages after implantation, the grafts consist of keratin-containing epithelial cells of unusual morphology that can be labelled with antibodies raised against the epithelium of the mid/deep cortex and the subcapsule/medulla. Normal thymic architecture develops, including nerves and blood vessels, as lymphocytes populate the environment, and by 4–6 weeks the epithelial cells are the same phenotypically and ultrastructurally as those found in normal rat thymus. However, some areas without lymphocytes still contain the atypical epithelial cells seen before implantation. Large multinucleated giant cells are also present with a few associated epithelial cells of subcapsular/medullary phenotype. In conclusion, the cultured thymic fragments contain a hitherto unknown precursor epithelial cell with an atypical ultrastructure and phenotype that is not seen in normal development.

Thymic microenvironment at the light microscopic level.

The thymus is a primary lymphoid organ that serves the immune system by providing an optimal microenvironment for developing T cells to rearrange the genes encoding the T‐cell receptor and to undergo positive and negative selection in shaping the peripheral T‐cell repertoire. The microenvironment of the organ is peculiar among lymphoid organs, as the supporting stroma consists of reticular epithelial cells. Bone marrow‐derived interdigitating cells and macrophages are the main accessory cell populations. The epithelium, interdigitating cells, and macrophages each contribute to the T‐cell selection process. During the last decade knowledge has been gathered that these cell populations show a considerable heterogeneity, as documented for subcellular features and immunologic phenotype. This heterogeneity may reflect various stages in differentiation, but may otherwise be linked to the functional activity of the cells. The authors survey the major cell populations, i.e., epithelial cells and lymphocytes. Macrophages and interdigitating cells are briefly discussed. Emphasis is given to functional aspects of histologic/cytologic features. Microsc. Res. Tech. 38:216–226, 1997.