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J L Haar

Publications and source records attributed to J L Haar.

At least 37 records · Page 2Linked to original sources

The liver hemopoietic environment: II. Peroxidase reactive mouse fetal liver hemopoietic cells.

Through the combined use of peroxidase cytochemistry and examination at the ultrastructural level, the present study has identified liver hemopoietic foci containing three forms of erythropoietic cells, two forms of myelopoietic cells, and a population of peroxidase nonreactive cells within the extravascular compartments of mouse fetal liver. The nonreactive cells were 10 micron in diameter, displayed no peroxidase activity and were designated type I cells. This cell had an irregular nucleus, small profiles of rough endoplasmic reticulum (RER), a considerable population of monoribosomes and a few polyribosomes. The incidence of this cell type decreased significantly from 50% at 12 days gestation to approximately 10% of the hemopoietic cells at 17 days gestation. Type I cells could not be classified into a hemopoietic lineage and may represent undifferentiated hemopoietic stem cells. Three forms of erythropoietic cells, designated types II, III, and IV, were identified. These cells had a diffuse cytoplasmic peroxidase reaction, no peroxidase positive membrane-bound organelles, and were approximately 7 micron in diameter. They corresponded to the more classically defined proerythroblast, polychromatophilic erythroblast, and nucleated normoblast, respectively. Types II and III had moderate cytoplasmic reactions, whereas type III, in addition, had a slight nuclear reaction. Type IV cells had a very dense cytoplasmic reaction but no nuclear reaction. Of the myelopoietic cells detected, one form had a slightly reactive Golgi and a few reactive granules. The other form possessed a clearly positive nuclear envelope (NE), RER, Golgi, and a population of reactive granules. The phagocytic sinusoidal lining cells (Kupffer cells) were peroxidase negative in contrast to similar cells in the rat. A population of peroxidase-positive granules was detected in fetal liver developing hepatocytes at 17 days gestation and increased in number with age. The morphology and organization of these various cell types in the liver hemopoietic environment are discussed.

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Migration of chicken yolk sac cells to bursa of fabricius supernatants.

Investigators have suggested that a bursal chemotactic factor exists that selectively attracts hemopoietic stem cells. This theory was investigated by determining the ability of embryonic (10- through 14-day) bursal supernatants to cause migration of age-matched yolk sac cells using blind well chambers. Supernatants of 10, 11, 13 and 14-day bursae caused specific migration of yolk sac cells whereas 12-day bursal supernatants inhibited migration of 12-day yolk sac cells. Ten-day migrating cells consisted of pro- and basophilic erythroblasts and hemopoietic stem cells which formed erythroid colonies in a chorioallantoic membrane-colony forming unit (CAM-CFU) assay.

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Selective migration of 'null' cells towards a thymus factor in vitro.

Mouse fetal liver, nude adult bone-marrow and normal adult bone-marrow cells are depleted of theta+, Ig+ cells by the use of in vitro migration through a porous membrane towards a thymus supernatant attractant. The attracted cells are free in suspension, viable and can be obtained in large numbers through the use of multiple migration wells for in vivo testing.

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The role of the nuclear protein matrix during development of rabbit granulocytes.

A proteinaceous nuclear substructure (nuclear protein matrix or nuclear pore complex-lamina) has been described in a number of cells and may be a universal feature of cell nuclei. We have investigated the nuclear protein matrix (NPM) in the rabbit blood granulocyte and its precursor cells to determine (A) whether the NPM composition is similar to other cells that have been studied and (B) to determine whether the dramatic morphological changes that the granulocyte nucleus undergoes during cell maturation are related to changes in the composition or structure of the NPM. NPM preparations from rabbit granulocytes were similar but not identical to those found in HeLa cells or rat liver nuclei. The NPM structure of more mature cells retained more DNA during the isolation procedures than did immature cell NPMs, and the DNA was less accessible to DNAse in the mature cell nuclei. Scanning and transmission electron microscopy revealed a continuous outer covering of the NPM preparation and a lattice-like internal structure. Recognizable nuclear form persisted although the preparations represented less than 20% of the original nuclear protein. NPM preparations from mature cells were similar in overall dimensions and internal structure to NPM from immature cells, suggesting reexpansion during isolation of a previously compacted NPM structure in segmented neutrophils. NPM proteins are synthesized primarily in early stages of cell development. The NPM appears to play a major, but passive, structural role in the nuclear changes observed during maturation of granulocytes.

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Applications of light scatter to separation of stem cells.

A case is presented for light scatter as a potentially important discrimination parameter for electronic sorting of hemopoietic stem cells. Examples are presented of application of light scatter to sorting of primitive hemopoietic cells from bone marrow, thymus, and fetal yolk sac, using simple empirical approaches and a cell sorter equipped with a standard fixed-angle detector. Cells sorted in this way were ultrastructurally indistinguishable from stem cells observed in whole tissue sections or unfractionated suspensions and remained viable under culture conditions. Cells were further sorted into proliferative and nonproliferative stages whose morphologies were those expected for primitive and mature cells, respectively. Light scatter was a more specific marker for primitive thymocytes than binding of peanut agglutinin. Cells sorted from fetal yolk sac suspensions were ultrastructurally similar to candidate pluripotential stem cells of bone marrow. Therefore, where sorting rather than positive identification is the primary objective of flow analysis and histochemical correlations can be established after sorting, viable stem cells can be sorted from the simple hemopoietic tissues in high and verifiable purity. Sorting of pure stem cells from marrow will require an additional parameter for which an additional angle of light scatter may suffice.

Agglutination↗

Separation of non-proliferating from proliferating thymic lymphocytes based on light scatter analysis.

Two populations of thymic lymphocytes from eight-week-old mice were separated based on the light scatter intensity they generated while passing through a flow cell analyser. The two populations were subsequently studied morphologically and analysed for the relative amount of DNA present in each cell. Cells having a low light scatter intensity had a high nuclear:cytoplasmic ratio, a heterochromatic nucleus, and a proliferative index (S + G2 + M) of only 23%. Cells with a high light scatter intensity had a low nuclear:cytoplasmic ratio, dispersed chromatin, frequently a uropod with budding microvilli and a proliferative index of 71%. The low light scatter cells resemble small cortical thymocytes or lymphocytes from the thoracic duct described by others. The highest light scatter cells resemble medium lymphocytes shown by others to be proliferative.

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Fine structural differentiation of germ layers in the mouse at the time of mesoderm formation.

The morphology of early postimplantation mouse egg cylinders was studied using light and electron microscopy. Implantation sites at seven, seven and one-half and eight days of gestation were dissected from the myometrium and whole implants, including both decidua and egg cylinders were processed for electron microscopy. Pre-primitive streak egg cylinders were composed of two germ layers, a tall columnar ectoderm and an outer visceral endodermal layer. Ectodermal cells demonstrated large oval nuclei and an organelle sparse cytoplasm except for many free polyribosomes. The visceral endodermal layer was composed of two cell populations. One visceral endodermal cell type observed was tall columnar in shape and appeared absorptive as demonstrated by many microvilli, pinocytotic profiles and lysosomal granules. This population was confined to extraembryonic regions of the egg cylinder. The second visceral endodermal cell type, squamous in shape, evidenced only a few microvilli, pinocytotic profiles and lysosomal granules. This population was confined to the embryonic region of the egg cylinder. Concurrent with the formation of the primitive streak an increased number of cellular junctions and nuclear pores became evident in the ectoderm. Mesodermal cells were large and stellate-shaped exhibiting many filapodia which made contact with adjacent mesodermal elements. Later the cephalic region of the primitive streak proliferated resulting in the migration of wedge-shaped mass of cells, the head process. At the most ventral extremity of the post-primitive streak egg cylinder the cells of the head process became intimately associated with the ectoderm by areas of focal contact and gap junctions.

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Fine structural analysis of the effect of trypan blue on the visceral endoderm of the mouse egg cylinder.

The effects of a maternal injection of trypan blue on primitive streak mouse embryos were studied with electron microscopy. Commercial trypan blue was purified by descending paper chromatography, and pregnant females received an intraperitoneal injection of the collected blue fraction on the evening of the 7th day of gestation. Ultrastructurally, the changes in the visceral endoderm were apparent 10 min after the injection and included an increase in the number of fuzzy-coated vesicles in the apical cytoplasm. By 20 min the apical cytoplasm of the extraembryonic visceral endodermal cells was filled with many fuzzy-coated vesicles and numerous vacuoles of various size and electron density. 30 min after the injection, the extraembryonic visceral endodermal cells were relatively smooth lacking a microvillous border and evidence of endocytic activity was rare. Many embryonic visceral endodermal cells were observed in various stages of degeneration although the underlying embryonic ectoderm appeared unaffected. Morphologically, it appears that trypan blue exerts its first effect by altering the endocytic activity of the visceral endoderm.

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An in vitro morphological study of the mouse visceral yolk sac and possible yolk sac immunocyte precursors.

Mouse visceral yolk sac has been organ cultured from 9 days of gestation, a time prior to the thymus being lymphoid, until 12 days of gestation, a time after which the thymus is lymphoid. During the culture period the endodermal epithelial cells survived well, erythropoiesis diminished, endothelial-lined cavities formed in the mesodermal mass, and cells developed which have been classified as large, medium and small immunocyte precursors. The cytoplasm of the immunocyte precursors contains polysomes, spherical mitochondria, a few profiles of rough endoplasmic reticulum, occasional granules and a large Golgi complex. This study offers morphological support for the yolk sac origin of immunocyte precursors in the mouse which may seed the thymus and liver.

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Epithelium and associated lymphocytes of developing human fetal appendix. Electron microscopic study.

The epithelium and lamina propria of the human fetal appendix between 7.5 and 18 weeks of gestation was examined with electron microscopy. At 7.5 weeks the epithelium was composed of stratified columnar cells; scattered mesenchymal cells appeared in the lamina propria. By 13.5 weeks of gestation the epithelium was composed of simple columnar cells with a microvillous surface, goblet cells containing numerous mucinogen granules and argentaffin cells with elongated electron-dense granules. Also at 13.5 weeks of gestation some free lymphocytes were observed within the lamina propria as well as an occasional migrating lymphocyte within the epithelium between the columnar cells. At 18 weeks of gestation the simple columnar epithelial cells were tightly packed, the goblet cells were secretory and lymphocytes again appeared singly within the epithelium and lamina propria. No evidence of groups of lymphocytes or developing primary follicles within the lamina propria was observed in this study.

Appendix↗

A light and electron microscopic investigation of the hepatic parenchyma of the adult newt, Notophthalmus viridescens.

This light and electron microscopic study demonstrates that the liver of the adult newt, Notophthalmus viridescens, consists of a mass of hepatocytes interrupted by blood sinusoids which allow blood to percolate through the parenchyma. The plates separating adjacent sinusoids are usually two or more cells thick and the bile canaliculi lie between 2-6 neighboring cells. Fine structural characteristics of hepatocytes include abundant lipid and glycogen inclusions. Melanophores with developing melanosomes are situated throughout the hepatic parenchyma.

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A light and electron microscopic study of myelopoietic cells in the perihepatic subcapsular region of the liver in the adult aquatic newt, Notophthalmus viridescens.

The liver of the newt, Notophthalmus viridescens, consists of several incompletely separated lobes of parenchymal tissue each of which is covered by a perihepatic subcapsular region (SSR) of the myeloid tissue. This tissue contains neutrophils and eosinophils in various stages of differentiation. As neutrophils developfrom myeloblasts to late neutrophilic myelocytes, two types of granules appear. The primary granules (type of granules formed first) are more electron dense and smaller than the secondary granules (type of granules formed later). The primary granules first appear at the stage designated early neutrophilic myelocyte, and the secondary granules appear at the stage of the maturing neutrophilic myelocyte. The eosinophils present are characterized by much larger granules than those observed in neutrophils. Cells in the PSR which superficially resemble small lymphocytes are primitive stem cells that give rise to neutrophils and eosinophils. The liver PSR is invested by a visceral peritoneum of simple squamous mesothelial cells some of which are ciliated.

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Fine structural observations on hemopoiesis in the chorioallantoic placenta of the marmoset.

Hemopoietic sinuses in the chorionic trabeculae of marmoset placenta were examined electron microscopically at several stages of pregnancy. Tissues were prepared both by standard technique and following injection of exogenous peroxidase into the maternal circulation prior to autopsy, and subsequent cytochemical localization of its reaction product in the fetal placenta. Aside from their dilated size and cellular content, the sinuses resembled the allantoic capillaries of the peripheral umbilical circulation in this species; and in early and midgestational placentas, sinusal lumina, differentiating elements in the erythroid line were present in placentas at all stages. Such cells appeared to arise both by a process of proliferation of undifferentiated progenitors from the sinusal lining and by mitosis of pre-existing erythroid cells. Only the latter process was observed in the mature placenta near term. Granulopoiesis was not observed at any stage; on the other hand, cells in the thromboid line were identified in placentas believed to be near midgestation; and large intrasinusal phagocytes were present in both midgestational and late placentas. In the latter, they were seen to have endocytosed peroxidase, and, by tenuous, dendriform processes, to be related anatomically to large numbers of erythroid cells. Early in gestation, the developing erythroid elements were closely related by areas of contact, both to each other and to the delimiting endothelium of the sinuses.

Allantois↗