Brita von Gaudecker

Ontogeny and organization of the stationary non-lymphoid cells in the human thymus

SummaryOntogenetic differentiation of the human thymus was investigated in 50 embryos by means of light and electron microscopic methods in an attempt to clarify the morphogenesis of the complicated microecology of thymic tissue. At the 8th gestational week (g.w.), the primordium of the thymus contains almost exclusively undifferentiated epithelial cells. At the 10th g.w., the epithelial cells in the central part are spindle-shaped. During the subsequent weeks the cortical region of the thymus becomes separated into lobes by mesenchymal septa containing hemopoietic precursor cells and large electronlucent cells with irregularly shaped nuclei. The latter cells are also found in the deeper presumptive medullary regions of the thymus; they differentiate into interdigitating reticulum cells (IDC). The permeation of the medulla of the thymus by non-epithelial IDC occurs concurrently with the formation of cortical and medullary epithelial cells. Between the 12th and 14th g.w. the cortical and medullary differentiation is completed. At this time-stage cortical small lymphocytes differ in morphological shape from medullary lymphocytes, the latter acquiring the appearance of immunocompetent T cells and establishing intimate contact with the IDC.These findings indicate that the thymic cortex and medulla contain different epithelial cells. In addition, the thymic medulla displays cells characterized by the morphology of typical interdigitating reticulum cells of peripheral lymphoid tissue. The structural pattern of the thymus is correlated to morphologically differing lymphoid cell populations in the cortical and medullary regions.

ber den Formwechsel einiger Zellorganelle bei der Bildung der Reservestoffe im Fettkrper von Drosophila-Larven

ZusammenfassungDer Fettkörper von Larven von Drosophila melanogaster (Stamm Berlin normal) wurde zu verschiedenen definierten Zeitpunkten zwischen der letzten Larvenhäutung und der Puppenhäutung licht- und elektronenmikroskopisch untersucht. Dabei ergab sich:1.Der Fettkörper ist von einer zarten Basalmembran umhüllt. Die darunterliegenden Zellen besitzen eine Reihe von tief einschneidenden verzweigten Falten. Ihre Oberfläche wird außerdem durch zahlreiche kurze, hakenförmig in das Cytoplasma vorspringende Tubuli vergrößert. Verschiedene Befunde sprechen dafür, daß diese Strukturen der Pinocytose dienen.2.Gruppen von offenbar pinocytotisch entstandenen 300 Å großen Bläschen treten zum granabesetzten Endoplasmatischen Reticulum in innige Beziehung.3.Fettkörperzellen von Drosophila enthalten nur granabesetztes Endoplasmatisches Reticulum.4.Fett wird intrazisternal im Lumen von granatragenden ER-Lakunen angereichert und gespeichert.5.Das granatragende ER bleibt auch dann mit den Fettvakuolen in Verbindung, wenn, wie in der zweiten Hälfte des letzten Larvenstadiums, die Fettmenge pro Zelle absinkt (Planimetrie).6.Glykogentröpfchen von 300–800 Å Durchmesser werden in unmittelbarem Kontakt mit ribosomenartigen Grana abgeschieden.7.Die bis zu 6 μ großen Eiweißgrana enthalten vereinzelt ribosomenartige Grana, granabesetzte ER-Lakunen, Glykogentröpfchen, stets Mitochondrienreste und sehr viel amorphe dunkle Substanz (OsO4-Methacrylat). Sie sind von einer einfachen Membran begrenzt.8.Wachsende Eiweißgrana enthalten Wickel aus dünnsten Membranen oder Lamellen.9.Hinweise sprechen dafür, daß die jüngsten Eiweißgrana durch Vermittlung von Substanzen, die in Form von 200–300 Å großen rundlichen Körpern im Zellkern entstehen, unter Beteiligung von Mitochondrien gebildet werden. Es wird vermutet, daß die Eiweißgrana mit Lysosomen verwandt sind.10.Auch granatragende ER-Lakunen falten sich von der äußernen Kernmembran ab.11.Die Kernmembran wird in späten dritten Larven auch neben den Poren so stark von fibrillärem Material durchsetzt, daß ihre scharfen Umrisse verschwinden.12.Das Problem der Steuerung dieses Vorganges wird besprochen. Es wird gefolgert, daß ER, Eiweißgrana und wohl auch Glykogen unter unmittelbarer Beteiligung von Genom und Kernmembran gebildet werden.13.Über die Steuerung der Fettkörperentwicklung innerhalb des Organismus liegen noch keine eigenen Befunde vor.Der Fettkorper von Larven von Drosophila melanogaster (Stamm Berlin normal) wurde zu verschiedenen definierten Zeitpunkten zwischen der letzten Larvenhautung und der Puppenhautung licht- und elektronenmikroskopisch untersucht. Dabei ergab sich: 1. Der Fettkorper ist von einer zarten Basalmembran umhullt. Die darunterliegenden Zellen besitzen eine Reihe von tief einschneidenden verzweigten Falten. Ihre Oberflache wird auserdem durch zahlreiche kurze, hakenformig in das Cytoplasma vorspringende Tubuli vergrosert. Verschiedene Befunde sprechen dafur, das diese Strukturen der Pinocytose dienen. 2. Gruppen von offenbar pinocytotisch entstandenen 300 A grosen Blaschen treten zum granabesetzten Endoplasmatischen Reticulum in innige Beziehung. 3. Fettkorperzellen von Drosophila enthalten nur granabesetztes Endoplasmatisches Reticulum. 4. Fett wird intrazisternal im Lumen von granatragenden ER-Lakunen angereichert und gespeichert. 5. Das granatragende ER bleibt auch dann mit den Fettvakuolen in Verbindung, wenn, wie in der zweiten Halfte des letzten Larvenstadiums, die Fettmenge pro Zelle absinkt (Planimetrie). 6. Glykogentropfchen von 300–800 A Durchmesser werden in unmittelbarem Kontakt mit ribosomenartigen Grana abgeschieden. 7. Die bis zu 6 μ grosen Eiweisgrana enthalten vereinzelt ribosomenartige Grana, granabesetzte ER-Lakunen, Glykogentropfchen, stets Mitochondrienreste und sehr viel amorphe dunkle Substanz (OsO4-Methacrylat). Sie sind von einer einfachen Membran begrenzt. 8. Wachsende Eiweisgrana enthalten Wickel aus dunnsten Membranen oder Lamellen. 9. Hinweise sprechen dafur, das die jungsten Eiweisgrana durch Vermittlung von Substanzen, die in Form von 200–300 A grosen rundlichen Korpern im Zellkern entstehen, unter Beteiligung von Mitochondrien gebildet werden. Es wird vermutet, das die Eiweisgrana mit Lysosomen verwandt sind. 10. Auch granatragende ER-Lakunen falten sich von der ausernen Kernmembran ab. 11. Die Kernmembran wird in spaten dritten Larven auch neben den Poren so stark von fibrillarem Material durchsetzt, das ihre scharfen Umrisse verschwinden. 12. Das Problem der Steuerung dieses Vorganges wird besprochen. Es wird gefolgert, das ER, Eiweisgrana und wohl auch Glykogen unter unmittelbarer Beteiligung von Genom und Kernmembran gebildet werden. 13. Uber die Steuerung der Fettkorperentwicklung innerhalb des Organismus liegen noch keine eigenen Befunde vor.

The development of the human tonsilla palatina

SummaryTonsils of human fetuses at the 8th to the 28th gestational week (g.w.) were investigated by electron microscopy, enzyme histochemistry, and immunohistochemistry on cryostat sections. The development of the tonsilla palatina starts during the 14th g.w. when the mesenchyme underlying the mucous membrane of the tonsillar cavity becomes invaded by mononuclear wandering cells. In fetuses of about the 16th g.w. epithelial crypts grow down into the connective tissue and are infiltrated by T-lymphocytes. At the same time, precursors of interdigitating cells (IDC) can be identified among the epithelial cells. Frequently, lymphocytes and IDC-like cells are in close contact. From these findings it is concluded that the infiltrated crypt epithelium in the human tonsilla palatina represents a T-cell region. Primary follicles develop in earlier fetal stages than in all other secondary lymphoid organs. They contain precursors of dendritic reticulum cells and lymphoid cells that belong to the B-cell line. These primary follicles may be considered as the first assemblage of B-cell regions in human fetal lymphoid tissue. The present findings indicate that the formation of different stationary elements during the development of B-cell regions and T-cell regions is an important factor for the homing and antigen-dependent maturation of different subpopulations of immunocompetent lymphoid cells.

Similarities between Hassall's corpuscles of the human thymus and the epidermis

SummarySubstrate-histochemical, enzyme-hystochemical and ultrastructural investigations were performed on thymic tissue from children, obtained in heart operations. β-Amylaseresistant, PAS-positive and Hale-positive substrates presumably neutral and acid mucosubstances, can be demonstrated in the central concentric lamellae of Hassalls corpuscles (HC). These lamellae also give positive reactions for sulphydryl groups and disulphide groups. Some flattened cell elements gave strong reactions for phospholipids, and small sudanophilic droplets, presumably neutral fats, are scattered throughout the HC. All investigated hydrolases and dehydrogenases either give no or only very weak reactions in the central part of progressive HC, but react strongly positive in their peripheral hypertrophic epithelial cells. In the central part of regressive HC, positive reactions for acid phosphatase and β-D-glucuronidase were recognized. These lysosomal enzymes may indicate degenerative processes.By electron microscopy progressive HC show central concentric lamellae with an amorphous matrix tightly filled with tonofilaments. They are surrounded by a thickened plasma membrane (200 Å), and do not contain nuclei. These central lamellae resemble the horny cells of the epidermis. The peripheral hypertrophic epithelial cells have pale nuclei with one or two nucleoli. Their cytoplasm contains numerous tonofibrils. These cells resemble stratum spinosum cells of the epidermis. In regressive HC the central concentric lamellae loose their intercellular contacts. The widened intercellular spaces are filled with cellular debris, and are invaded by macrophages.Similarities between the ultrastructure and the patterns of the histochemically investigated substrates and enzymes in human HC and epidermis are discussed.Substrate-histochemical, enzyme-hystochemical and ultrastructural investigations were performed on thymic tissue from children, obtained in heart operations. β-Amylaseresistant, PAS-positive and Hale-positive substrates presumably neutral and acid mucosubstances, can be demonstrated in the central concentric lamellae of Hassalls corpuscles (HC). These lamellae also give positive reactions for sulphydryl groups and disulphide groups. Some flattened cell elements gave strong reactions for phospholipids, and small sudanophilic droplets, presumably neutral fats, are scattered throughout the HC. All investigated hydrolases and dehydrogenases either give no or only very weak reactions in the central part of progressive HC, but react strongly positive in their peripheral hypertrophic epithelial cells. In the central part of regressive HC, positive reactions for acid phosphatase and β-D-glucuronidase were recognized. These lysosomal enzymes may indicate degenerative processes.

Substrate-histochemical investigations and ultrahistochemical demonstrations of acid phosphatase in larval and prepupal salivary glands of Drosophila melanogaster.

SummaryA major function of the larval salivary glands of Drosophila melanogaster is known to be the production of a mucopolysaccharide that serves as an adhesive during puparium formation. In order to localize the mucosubstances during development substrate histochemical methods were used, and the site of acid phosphatase was demonstrated by the ultrahistochemical lead-salt method. It could be shown that the “glue”-granules in the corpus cells of larval salivary glands as well as the large secretion vacuoles in the prepupal corpus cells give a positive β-amylase-resistent PAS-reaction, which indicates neutral mucosubstances. Granular PAS-positive deposits in the larval and prepupal collum cells were reduced after preincubation with β-amylase and may represent glycogen, which has also been seen in electron micrographs of these cells. The Hale-reaction gave a weak indication that acid mucosubstances are present in the larval “glue” granules and in the large prepupal secretory vacuoles. After digestion of sialic acid with α-neuraminidase the weak indication was absent showing that the acid mucosubstances had been sialomucines. Ultrahistochemical demonstration of acid phosphatase indicated the presence of this enzyme in Golgi fields and lysosomal structures. Acid phosphatase seems to be missing in the large secretion vacuoles of the prepupal salivary gland.It is concluded, that the large vacuoles in the corpus cells of prepupal salivary glands represent a secretion product, obviously a mucosubstance. The lysosomal structures, containing acid phosphatase, may be accumulated in preparation for the autolysis of the gland which begins about two hours after the pupal moult, i.e. 15 hours after puparium formation.

Der Strukturwandel der larvalen Speicheldrüse vonDrosophila melanogaster

SummaryThe development and differentiation of the larval salivary glands ofDrosophila melanogaster have been investigated with light and electron microscopical methods. The organ has been dissected out of exactly dated stages of the III. instar larva, the prepupa and the early pupa. In order to avoid great variations in the physiological age of the animals a culture method has been developed, enabling the larval molts to be observed and used for identification of the age. The results are as follows: 1.The salivary gland of the early larva up to the middle of the III. instar period is a homogenous sack consisting of one sort of cells, in which very small secretion granules (∅ 0,3 μm) are synthesized. These secretion granules concentrate near the cellular apex. They are supposed to contain digestion enzymes.2.In the second half of the III. larval instar period three cell types are differentiated, which are called corpus cells, transitional cells and collum cells. A gradient of differentiation from distal to proximal can be observed.3.Thecorpus cells, located at the distal part of the gland, stop the production of digestion enzymes in the second half of the III. larval instar period and begin to synthesize a cement substance. This cement first is stored in grana (∅ up to 10 μm) inside the corpus cells. Shortly before puparium formation it is extruded into the lumen of the gland. Shortly after puparium formation it is expectorated out of the mouth, runs along the body wall and affixes the puparium to the substrate. The cement is PAS-positive, probably being a mucoproteid. In the corpus cells large vacuoles are formed during the prepupal instar period. On the basis of these electron microscopical results the vacuoles are interpreted to represent another form of a secretory product and not an equivalent of beginning degeneration. The possible function of this substance is discussed.4.Thetransitional cells are located between the corpus cells and the collum cells. They also synthesize cement at a delayed rate, through shortly before puparium formation they are filled with cement like the corpus cells and cannot be distinguished from the latter.Thecollum cells form the most proximal part of the salivary gland. They do not produce cement but continue to synthesize digestion enzyme granules in the second half of the III. instar period. The large secretion vacuoles, found in the corpus cells during the prepupal instar period, are not synthesized in the collum cells.5.The involution of the larval salivary gland begins after pupation and is indicated by autolytic processes, which begin at the distal end of the gland. One hour later all cells exept the imaginalanlage show signs of degeneration.6.The course of development of the salivary glands investigated in the present study inDrosophila melanogaster is compared with similar investigations onDrosophila virilis, robusta andhydei. It is pointed out that the development of the larval salivary gland in different species ofDrosophila shows close parallels.The relationships between metabolic activities in the cytoplasm and gene physiological activities (pattern of puffs) on the giant chromosomes, as known so far, are discussed.ZusammenfassungDie Entwicklung bzw. Differenzierung der larvalen Speicheldrüse vonDrosophila melanogaster wurde an genau datierten Altersstadien aus dem III. Larvenstadium, der Vorpuppe und der Puppe mit lichtmikroskopischen und elektronenmikroskopischen Methoden untersucht. Zur Vermeidung großer Streuung im physiologischen Alter der Tiere wurde eine Kulturmethode entwickelt, die es erlaubt, die Häutungen zu beobachten und zur Altersbestimmung heranzuziehen. Folgende Ergebnisse wurden erzielt: 1.Die Speicheldrüse besteht bis zur Mitte des III. Larvenstadiums morphologisch aus einem einheitlichen Zelltypus, der sehr kleine Sekretgrana (∅ 0,3 μm) bildet. Diese sammeln sich am Zellapex. Die Vermutung liegt nahe, daß es sich um ein Verdauungssekret handelt.In der 2. Hälfte des III. Larvenstadiums differenzieren sich drei Zelltypen, die hier Corpuszellen, Übergangszellen und Halszellen genannt werden. Dabei ist ein Differenzierungsgradient von distal nach proximal zu beobachten. Die distal gelegenenCorpuszellen stellen die Bildung des Verdauungssekretes in der 2. Hälfte des III. Larvenstadiums ein und bilden stattdessen ein Klebesekret. Dieses Sekret wird in Form großer Grana (∅ bis zu 10 μm) zunächst in den Zellen gespeichert und kurz vor der Pupariumbildung ins Lumen der Drüse abgegeben. Kurz nach der Pupariumbildung wird das Klebesekret aus dem Körper entlassen und dient dazu, die Tönnchenpuppe an einem trockenen Substrat anzuheften. Das Klebesekret ist PAS-positiv. Wahrscheinlich handelt es sich um ein Mucoproteid. Während des Vorpuppenstadiums bilden sich in den Corpuszellen große Vakuolen, die auf Grund der elektronenmikroskopischen Befunde als Ausdruck einer weiteren Sekretionsphase und nicht als beginnende Degeneration gedeutet werden. Die mögliche Bedeutung dieses Sekretes wird diskutiert.DieÜbergangszellen liegen zwischen den Corpuszellen und den Halszellen. Sie bilden ebenfalls Klebesekret, jedoch mit zeitlicher Verzögerung. Kurz vor der Pupariumbildung sind sie wie die Corpuszellen mit ausgereiften Klebesekretgrana beladen und von diesen nicht mehr zu unterscheiden.Die proximal gelegenenHalszellen bilden kein Klebesekret, sondern setzen die Bildung des Verdauungssekretes in der 2. Hälfte des III. Larvenstadiums fort. Während des Vorpuppenstadiums bilden sich in den Halszellen nicht die großen Vakuolen wie in den Corpuszellen.2.Die Involution der larvalen Speicheldrüse erfolgt nach der Puppenhäutung durch Autolyseprozesse, die am distalen Ende der Drüse beginnen und innerhalb 1 Std alle Zellen mit Ausnahme der Imaginalanlage erfassen.3.Die in dieser Untersuchung erhobenen entwicklungsgeschichtlichen Befunde anDrosophila melanogaster werden mit Beobachtungen anDrosophila virilis, D. robusta undD. hydei verglichen. Dabei wird aufgezeigt, daß die Entwicklung der larvalen Speicheldrüsen von verschiedenenDrosophila-Arten enge Parallelen aufweist. Die bisher bekannten Zusammenhänge zwischen Stoffwechselaktivitäten im Zytoplasma und Genaktivitäten (Puffmuster) an den Riesenchromosomen dieser Zellen werden diskutiert.

Immunohistochemical characterization of the thymic microenvironment

SummaryThe epithelial framework of the human thymus has been studied in parallel by immunohistochemical methods at the light- and electron-microscopic levels. Different monoclonal antibodies were used, reacting with components of the major histocompatibility complex, keratins, thymic hormones and other as yet antigenically undefined substances, which show specific immunoreactivities with human thymus epithelial cells.The electron-microscopic immunocytochemical observations clearly confirm microtopographical differences of epithelial cells not only between the thymic cortex and medulla, but also within the cortex itself. At least four subtypes of epithelial cells could be distinguished: 1) the cortical surface epithelium; 2) the main cortical epithelial cells and thymic nurse cells; 3) the medullary epithelial cells; and 4) the epithelial cells of Hassalls corpuscles.The various epithelial cell types of the thymus display several common features like tonofilaments, desmosomes and some surface antigens as demonstrated by anti-KiM3. In other respects, however, they differ from each other. The cortical subtype of thymic epithelial cells including the thymic nurse cells shows a distinct pattern of surface antigens reacting positively with antibodies against HLA-DR (anti-HLA-DR) and anti-21A62E. Electron-microscopic immunocytochemistry with these antibodies clearly reveals a surface labeling and a narrow contact to cortical thymocytes particularly in the peripheral cortical regions. An alternative staining pattern is realized by antibodies to some antigens associated with other subtypes of thymic epithelial cells. Medullary epithelial cells as well as the cortical surface epithelium react likewise positively with antibodies to special surface antigens (anti-Ep-1), to special epitopes of cytokeratin (anti-IV/82), and to thymic hormones (anti-FTS). The functional significance of distinct microenvironments within the thymus provided by different epithelial cells is discussed in view of the maturation of T-precursor cells.

Ultrastructure of the age-involuted adult human thymus

SummaryAge involuted thymus tissue from a middle aged (33 years) and an old (63 years) man have been examined by electron microscopy and compared with thymus tissue from children. Biopsies had been taken during surgical correction of congenital heart defects.The fine structural architecture of cortex, medulla and connective tissue in the remaining lymphatic islands in the adult thymus investigated was not different to the thymus of children. We were surprised to find vigorous lymphocytopoiesis in the cortical regions and to recognize extended areas of medulla with a cellular composition which obviously provides the same microenvironment for T-cell maturation as the medulla of the non involuted thymus. Our findings are discussed in relation to the increasing arguments that the human thymus serves an immunological function throughout life.

The development of the human spleen

SummarySplenic tissue of human fetuses from the 14th to the 24th week of gestation (menstrual age) were investigated by light- and electron microscopy to describe the development of the red and white pulp in close relationship to the differentiation of the vascular tree. Special interest is focussed on the differentiation of the T-cell- and the B-cell regions and their specific stationary cells.The preliminary stage, here called the “primary vascular reticulum,” lasts up to the 14th gestational week (gw). Numerous erythrocytes, normoblasts and macrophages are seen among a network of mesenchymal cells and argyrophilic fibers. Hematopoiesis, especially erythropoiesis, can be recognized.The characteristic organ structure becomes established during the subsequent transformation stage of the fetal spleen, beginning with the 15th gw. Splenic lobules begin to form during the 15th to 17th gw. They consist of a central artery, surrounded by a sheath of lightly stained stationary cells which resemble myofibroblasts. At the periphery of these lobules the red pulp forms. Initially mobile cells are distributed throughout the reticulum. Soon they begin to accumulate in the venous sinuses, which develop from lacunae among the reticular network and come into contact with the venous system. The endothelial wall of these sinuses remains discontinuous, confirming the theory of the “open” vascularization of the spleen. The development of the larger veins is correlated with the differentiation of the splenic trabeculae.The development of the white pulp is correlated with the stage of lymphoid colonization within the spleen, beginning around the 18th gw. An accumulation of lymphocytes around the central arteries can be recognized during the 19th and 20th gw. These lymphoid cells show morphological and immunohistochemical characteristics of T-precursor cells. Within the now assembling periarterial lymphoid sheath (PALS) a few precursors of interdigitating cells (IDC) are recognizable, giving evidence for the differentiation of the T-cell region.Around the 23rd gw the assemblage of primary follicles is discernible at the periphery of the PALS. Precursors of the follicular dendritic reticulum cell (FDRC), the specific stationary cell of the B-cell region, have been recognized. This observation leads to the conclusion that the small primary follicles represent the beginning formation of the B-cell region.The significance of the vascular system for the differentiation of the specific splenic organization is discussed.

The development of the human lymph node

SummaryLymph nodes of human fetuses from the 11th to the 20th gestational week (g.w.) were investigated by light- and electron microscopy under particular consideration of the development of the T-cell and the B-cell regions and their specific reticulum cells. Lymph node development begins as a mesenchymal condensation, containing capillaries and mesenchymal cells; this primordium bulges into a lymph sac. Within the primordium of the lymph node granulopoiesis and erythropoiesis occur temporarily from the 12th to the 14th g.w. The first lymphoid cells and undifferentiated blast cells are seen in the 12th g.w.; monocytes and macrophages can be found from the 13th g.w. onward.The development of the T-cell regions begins during the 13th g.w., before differentiation into cortex and medulla becomes obvious in the 14th g.w. Near the marginal sinus, cells displaying features of interdigitating reticulum cells (IDC) show similarities to monocytes. The morphological differentiation of the IDC is complete in the 17th g.w. when they are found in the paracortical region. Among the IDC, lymphoid cells with features of thymocytes are arranged in small groups.The first indication of the development of B-cell regions can be recognized in the 14th g.w. when precursors of dendritic reticulum cells (DRC) are seen near the marginal sinus; this area also displays lymphoblasts, immunoblasts, and plasmoblasts. During the 20th g.w. small primary follicles are discernible in the outer cortex; in addition to blast cells they contain small lymphocytes, none of which show features of thymocytes. The morphological development of DRCs is not entirely complete until the 20th g.w.; however, some cells already show a characteristic network of interwoven processes.The probable origin of (i) the IDC from monocytes, and (ii) the DRC and fibroblastic reticulum cells from a common type of mesenchymal precursor cells, as well as their significance for a specific micromilieu in the T-cell and the B-cell regions, are discussed.