N. B. Everett

Distribution of Blood (Fe59) and Plasma (I131) Volumes of Rats Determined by Liquid Nitrogen Freezing

A standard method is described which gives reproducible blood volume values for rats and their various organs and tissues.

RECIRCULATION OF LYMPHOCYTES

Abstract : The long-lived small lymphocyte population of the rat includes approximately 90 per cent of the cells which appear in thoracic duct lymph. About two-thirds of the small lymphocytes in blood are long-lived. These long-lived cells are produced at a rate proportional to body growth and recirculate from blood to lymph. Althouohtthough a major percentage of the small lymphocytes in lymph nodes and a smaller percentage of those in spleen are of the long-lived variety, the thymus may be their primary source of origin. The small lymphocytes in the bone marrow, approximately 95 per cent of those in the thymus and a major percentage of those in the spleen are short lived (five days or less). Evidence is presented which suggests that the long-lived lymphocyte plays an active role in immune response mechanisms. (Author)

Lymphopoiesis in the thymus and other tissues: functional implications.

Publisher Summary This chapter reviews the evidence relative to the life span and turnover times of the lymphocyte populations in the various organs with special emphasis on the thymus, and relates these life span studies with the many advances that have come from studies of lymphocyte fate and function. The formation of long-lived small lymphocytes is controlled by the thymus and may be mediated by a humoral factor produced by the epithelial cells of the thymus. Thus neonatal thymectomy prevents the formation of the long-lived population and thymectomy of the young adult prevents new cells from being added to the already existing pool. The long-lived population is highly radiosensitive and the thymic epithelium is required to mediate the regeneration of a new population in heavily irradiated animals. Animals made deficient in long-lived small lymphocytes by such methods as chronic lymphatic drainage, neonatal thymectomy, or irradiation, show a deficiency in their ability to reject a homograft or heterograft and in other immunological reactions. This immunological incompetence may be restored by injecting long-lived small lymphocytes from an isogenic strain.

Localization of newly synthesized proteins in regenerating newt limbs as determined by radioautographic localization of injected methionine-S35

Abstract Sites of amino acid incorporation within the regenerating forelimb of adult Triturus viridescens were determined by the radioautographic localization of injected methionine-S 35 . The limb regenerates studied ranged in age from 24 hours to 27 days. During the first few days after amputation the epidermis only produces an intense radioautographic reaction. The intensity of the reaction of the mesodermal tissues increases by the fifth day, when the periosteum of the transected humerus and the terminal portions of the cut muscles and nerves display a concentration of radioactivity. Fragments of sarcoplasm detached from the damaged muscles produce a strong reaction, suggesting that they are not moribund fragments. Those osteocytes apparently being released from the degenerating bone are highly radioactive. The intensity of the radioautographic reaction is generally even throughout the blastema during the growth phase. The radioautographs suggest that during the growth phase the peripheral nerve may exert an influence on protein synthesis within the undifferentiated blastemal cell. In later stages the more intense reduction of emulsion is produced by those cells undergoing differentation. The observations thus suggest that increased incorporation of amino acid is characteristic of the regenerate generally, and comprises a significant feature of dedifferentiating as well as differentiating cells.

MALL LYMPHOCYTE POPULATIONS IN THE MOUSE BONE MARROW

Autoradiography and scintillation counting have been used for evaluation of lymphocyte turnover and life span in the bone marrow, peripheral blood and thoracic duct lymph of BALB/C mice. It was shown that the bone marrow contained two populations of small lymphocytes. One population was labelled 100% after 3–4 days of intensive injections of 3H‐thymidine and constituted about 75% of the lymphocytes. The remaining 25% of the lymphocytes turned over at a much slower rate comparable to the rate of increase in labelled small lymphocytes of the thoracic duct. More than 10% of the small lymphocytes of the bone marrow were found to be unlabelled after 10 days of intensive injections of 3H‐thymidine. Nine weeks after giving 3H‐thymidine for 30 consecutive days, 8·6% of the small lymphocytes in the bone marrow remained labelled. The mean grain counts of cells in this population were comparable to those of thoracic duct lymphocytes at corresponding times. About 90% of the peripheral blood lymphocytes were found to have a slow turnover and a long life span.

Measurement and Radioautographic Localization of Albumin in Rat Tissues After Intravenous Administration

Radioautography and conventional counter methods have been used to determine the relative amount and localization of I131 albumin in a number of tissues at 3, 30, and 60 min. after intravenous injection. Certain tissues showed essentially no change in albumin content with time, others contained extravascular albumin which increased with time, and the liver and adrenal evidenced a decreasing level of albumin with time. The transcapillary passage of albumin and its accumulation in extravascular compartments are correlated with tissue structure.

MIGRATION OF SMALL LYMPHOCYTES IN ADULT MICE DEMONSTRATED BY PARABIOSIS

Parabiotic BALB/C mice were used to study the traffic of small lymphocytes in immunological mature but unchallenged mice.

RADIOAUTOGRAPHIC STUDIES OF THE STEM CELL IN THE THYMUS OF THE IRRADIATED RAT

Radioautographic evidence is presented which characterizes the marrow derived stem cell which promotes thymic recovery following irradiation in the rat. These immigrant cells are similar in morphology to blood monocytes and have been called monocytoid, meaning monocyte‐like in appearance. The typical cell had abundant pale staining cytoplasm and a nucleus with many invaginations and folds and a fine chromatin structure. There was no prominent nucleolus. The majority of these cells entered the thymus of the irradiated rat via the blood vessels into the septa and made their way through the connective tissue to the outer cortex. Three distinct morphological cell types appeared to be derived from the immigrant cells. These were fibrocyte‐like cells which were located within the septa, macrophages located mainly within the medulla and septa, and large blast cells within the cortex, which proliferated giving rise to large thymocytes. The blast cells were characterized as having abundant moderately basophilic (and pyroninophilic) cytoplasm with a distinct cytoplasmic boundary, a large nucleus which still had invaginations and folds, a loose chromatin structure and one or more very prominent nucleoli. They were located in groups primarily within the outer cortex and often adjacent to blood vessels. They were found to be highly susceptible to damage in smear preparations. In contrast, their progeny, the large thymocytes were not highly susceptible to damage in smear preparations but teased out as large round cells with a highly basophilic rim of cytoplasm. The large thymocytes were precursors to medium and small cells.

Radioautographic Studies of Reticular and Lymphoid Cells in Germinal Centers of Lymph Nodes

Earlier studies from our laboratory have shown that there are two populations of small lymphocytes in the rat with respect to rate and site of formation and circulating life span (Caffrey et al., 1962). One of these has a rapid turnover rate with a circulating life span of about 5 days. The other is produced at a slow and rather constant rate which is roughly proportional to the growth rate of young adult animals. It is the latter population which has a long circulating life span and recirculates from blood to lymph. It is believed that at least some of these may have a life span commensurate with that of the animal since heavily labeled small lymphocytes have been recovered for as long as one year after labeling with Thymidine-3H. Subsequent studies have shown the percentages of the two populations in blood, thoracic duct lymph, and in each of the major lymphoid organs. It was determined that approximately 75% of the small lymphocytes of the mesenteric lymph node were of the long-lived variety and approximately 25% were short-lived (Everett et al., 1964 a). From attempts to assess the respective functions of the two varieties of small lymphocytes, evidence was obtained which suggested that the long-lived lymphocyte has an active role in immune response mechanisms. For example, these cells evidenced enlargement, both nuclear and cytoplasmic, in animals treated with pertussis vaccine (Everett et al., 1964 a). Additionally, as an indirect response to total body irradiation (Everett et al, 1964 b), many long-lived cells enlarged and showed a pronounced increase in the rate of protein synthesis (Everett et al., 1965).

The distribution and excretion of S35 sodium sulfate in the albino rat.

Summary Bile fistula rats given S35 sodium sulfate intravenously excreted 89% of the injected sulfur within 24 hr. Seventy-five per cent of the sulfur was recovered in the urine, 10% in the bile, and 4% in the feces. Determinations for sulfur were made on segments of the gastrointestinal tract at intervals after injections. The results reveal that in the bile fistula animals sulfur enters the upper third of the small intestine and moves through the intestines to be eliminated in the feces. The suggestion is made that sulfate passes directly from the blood or tissue fluid into the upper small intestine.