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Biomedical subjects

R Pabst

Publications and source records attributed to R Pabst.

At least 235 records · Page 13Linked to original sources

Lymphocyte emigration from lymph nodes by blood in the pig and efferent lymph in the sheep.

Two types of experiment using local labeling of lymph nodes with FITC showed that lymphocytes emigrate from lymph nodes, predominantly in blood in the pig and in efferent lymph in the sheep. In the first type of experiment with the pig, few cells emigrated via the lymph, while the number of labelled cells in the blood increased progressively and the indices in mesenteric blood were always higher than in jugular blood in simultaneously-drawn samples. However, in the sheep, when efferent lymph flowed freely, very low numbers emerged in blood and continuing large numbers of lymphocytes emerged in efferent lymph. In the second type of experiment carried out wholely under anaesthetic on mesenteric lymph nodes in pigs and sheep, and on superficial inguinal lymph nodes in pigs, the lymph node was isolated, the lymph and venous drainage collected and only the arterial supply maintained. Large numbers of FITC+ lymphocytes emigrated via the vein in pigs with either node cannulation (i.e. up to 7% blood lymphocytes were labelled with an emigration rate of approximately 10(8) cells/hr) but in sheep, while lymph contained approximately 30-80% labelled cells and the emigration rate was also approximately 10(8) cells/hr, the mesenteric blood contained very few labelled cells (approximately 0.2%, giving a mean venous emigration rate of 2.7 X 10(6)/hr). Study of the type of lymphocytes emerging from labelled pig lymph nodes and spleen during the phase of major emigration showed that sIg+ B and E rosette-forming T cells, but almost no Null cells, are involved.

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The emigration of lymphocytes from Peyer's patches in sheep.

The magnitude of the output of lymphocytes from Peyer's patches (PP), the morphology of the lymphocytes and their route of exit from PP has been examined in sheep. An extracorporeal perfusion system was used to selectively label a 3-4 m length of the terminal ileum of lambs at 6-10 weeks of age (the mesenteric lymph nodes had previously been excised from most lambs). This part of the intestine contains about 80% of the total PP tissue in a lamb and most of the lymphoid cells in the perfused tissue were within the PP. During a 10 min labeling period, fluorescein isothiocyanate (FITC) was added to the perfusate to label all the cells in the perfused tissues and [3H]thymidine [(3H]dThd) was added at the same time to label only those cells in the S-phase of the cell cycle. The unincorporated label was then washed from the perfused tissues, the normal blood circulation was reestablished and the lamb allowed to recover from anesthesia. It was established that the labeling was restricted to the perfused tissue and therefore that any labeled cells subsequently found elsewhere in the animal must have emigrated from the terminal ileum. During the 24 h after perfusion 1.4 X 10(9)-3.9 X 10(9) lymphoid cells (i.e. FITC labeled) left the perfused tissues via the lymph; 12-19% of these cells were either in the process of dividing or less than 24 h old (i.e. [3H] dThd labeled). The majority of the labeled cells probably came from the PP and most were classified as small lymphocytes although the [3H] dThd labeled population included a high proportion of large lymphocyte and lymphoblasts especially during the early hours after the perfusion. The labeled lymphocytes entered the blood in substantial numbers which increased linearly with time so that by 24 h about 7% of the lymphocytes in the blood were fluorescent. The numbers of newly produced cells began to increase rapidly in the blood only during the 12 h to 24 h period. The number of labeled cells in the blood was reduced by about 95% when the lymph from the perfused tissues was drained from the lamb during the experiment. This result provides clear evidence that the vast majority of all of the cells that leave the PP do so via the lymph and not via the blood.

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The emigration of lymphocytes from palatine tonsils after local labelling.

Lymphocytes in the palatine tonsils of normal young pigs were selectively labelled by minute multiple injections of fluorescein isothiocyanate into the tonsils. One day later the numbers of tonsil-derived lymphocytes were determined in cervical, bronchial and mesenteric lymph nodes, spleen, Peyer's patches, thymus, bone marrow and blood. Although not all tonsillar lymphocytes could be labelled by this technique, a total of approximately 0.6 X 10(9) lymphocytes emigrated from the tonsils. Relatively more lymphocytes were found in lymph nodes than in the spleen and very few in the thymus, Peyer's patches and bone marrow. This organ distribution was different from the results from selectively labelling lymphocytes in lymph nodes, spleen and bone marrow.

Animals↗

Splenic regeneration and blood flow after ligation of the splenic artery or partial splenectomy.

In normal, young miniature piglets, either the major arteries and the veins were ligated or three quarters of the spleen was resected using three different techniques. These techniques served as an animal model for splenic salvage in human subjects. After arterial ligation, no signs of infarction could be seen 1 and 4 weeks later, and after 6 months, normal sized spleens with enlarged vessels in the gastrolienal ligaments were found. When the splenic remnant was left at the main vessels, no obvious growth could be demonstrated after 6 months. The splenic tissue left at the smaller vessels, however, increased in size. Measurements of splenic blood flow in unanesthetized piglets 6 months after operation resulted in greater differences in perfusion per gram among the experimental groups. Not only splenic mass, but also blood flow to the whole splenic tissue seems to be an important factor in the protective function of the spleen.

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An animal model for aseptic necrosis after intramuscular injections.

Ten drugs or vaccines commonly given to patients by intramuscular injection were injected into the femoral artery of normal young anaesthetized pigs, in order to establish an animal model for macroscopically identifiable aseptic tissue necrosis (Nicolau syndrome). Despite the wide range of constituents and chemical groupings in the drugs which had caused Nicolau syndrome in patients, when injected into the pigs a typical pattern of reactions could be observed for many of them, as follows: the leg contracted rapidly, the skin area supplied by this artery initially became pale and then bluish-red with an irregular reticular appearance before finally tissue necrosis developed. These reactions are comparable to the symptoms of Nicolau syndrome in man. However, no reactions were seen when drugs or vaccines which have not been known to cause aseptic necrosis in man, e.g. tetanus toxoid, influenza vaccine or triamcinolon, were injected i.a.

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Cell renewal of glomerular cell types in normal rats. An autoradiographic analysis.

Normal adult Sprague-Dawley rats received either a single or repetitive injection of the DNA precursor 3H-thymidine (3H-TdR). For autoradiography semi-thin sections were prepared 2 hr to 14 days after labeling. The majority of labeled cells noted in glomerular tufts were endothelial cells. Mesangial cells had a lower production rate. Podocytes revealed no evidence of proliferation. Bowman's capsule cells showed a higher labeling index than tuft cells at all times. Neither the urinary nor the vascular pole was found to be a proliferative zone for Bowman's capsule cells. The flash and repetitive labeling experiments demonstrated a constant rate of cell renewal of about 1% per day, resulting in a long life span for endothelial and mesangial cells as well as Bowman's capsule cells. These data provide a basis for cell kinetic studies in models of glomerular diseases.

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In situ labelling of bone marrow lymphocytes with fluorescein isothiocyanate for lymphocyte migration studies in pigs.

In normal young pigs, the femoral artery and vein were cannulated and after occluding other vessels to one hind leg they were connected to an extracorporeal perfusion system. Fluorescein isothiocyanate (FITC) was added to the perfusate to selectively label cells in the bone marrow. Large numbers (approximately 0.9 X 10(9] of labelled lymphocytes left the bone marrow of one leg within 1 d and migrated via the blood to the bone marrow in other bones, lymph nodes, spleen, Peyer's patches and even into the thymus. On average 1.7% of the lymphocytes in the blood and about 1% within the spleen were labelled. Peyer's patches and the thymus showed very low indices. Thus the bone marrow is an integral part of the migratory route of lymphocytes. Selective labelling of bone marrow cells in their normal microenvironment with FITC is a suitable method for studies of cell migration from the bone marrow.

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Regeneration and function of autologous splenic grafts in pigs.

The growth kinetics, perfusion and immune response of autotransplanted splenic tissue were studied in pigs. Splenic fragments were transplanted subfascially or in the greater omentum and regenerated to small splenules with a normal histologic structure 6 months later. The grafts showed a normal function of the white pulp after antigenic stimulation, and of the red pulp as demonstrated scintigraphically. The blood flow was reduced. The growth kinetic was less rapid than in rodents and resulted in a relatively lower mass of splenic tissue, but these results are closer to findings in human splenosis.

Animals↗

Subsets of blood, spleen and recirculating lymphocytes in man.

Lymphocyte subpopulations were characterized in human blood and spleens. In addition the spleens were perfused by a closed extracorporeal perfusion system under almost physiological conditions. Lymphocytes released from the spleen during perfusion were taken to be representative of recirculating lymphocytes. B lymphocytes were classified by their surface immunoglobulin, T lymphocytes and T lymphocyte subsets by cytochemistry, sheep red blood cell rosette formation and in some experiments by monoclonal antibodies. In the blood 71 +/- 4.3% of the lymphocytes were rosette forming cells and 23.3 +/- 3.8% B lymphocytes. In the spleen 49.8 +/- 3.6% were T and 53.3 +/- 2.1% were B lymphocytes. In three spleens the mean number of OKT3+ lymphocytes were 27.6 +/- 7.0% OKT4+ 8.6 +/- 1.4% and OKT8+ 13.7 +/- 2.2%. The ratio of T helper to T suppressor lymphocytes was 0.67 for the spleen and 1.7 for the blood. The lymphocytes released from the perfused spleen showed a similar distribution pattern of surface markers to that of the splenic subpopulations.

Aged↗

Classification of newly formed and migrating splenic lymphocytes.

Newly formed lymphoid cells in the spleens of normal young pigs were selectively labelled with 3H-TdR, using an extracorporeal perfusion system. One day later splenic and blood lymphocytes were separated by rosetting with sheep red blood cells for classical T cells and by a direct anti-globulin rosetting reaction for B cells. The proportion of newly formed lymphocytes in these subsets was determined by autoradiography. In this study we show that while both B and T lymphocytes are produced, B cells show a much higher incidence of newly formed cells. Labelled populations of spleen derived T and B lymphocytes were also found in the blood, but the B cells showed a lower labelling incidence than in the spleen.

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Proliferation and emigration of newly formed lymphocytes from pig spleens during an immune response.

Normal young pigs were immunized intravenously with sheep red blood cells (SRBC). At various times after a second SRBC injection the spleens were connected to an extracorporeal perfusion system, and proliferating lymphoid cells in the spleens were selectively labelled with tritiated thymidine. One day later the relative and absolute numbers of spleen-derived lymphocytes were determined by autoradiography in the following organs: various parts of the spleen, mesenteric and cervical lymph nodes, thymus, bone marrow, Peyer's patches, tonsils, intestine, lung, liver and blood. From 1 to 7 days after the second SRBC injection, the spleens produced increasing numbers of lymphocytes, and labelled cells were found especially in the blood and bone marrow. The newly formed splenic lymphocytes migrated preferentially to T- but also to B-cell areas in lymph nodes, Peyer's patches and tonsils. In all organs outside the spleen nearly all labelled spleen-derived lymphocytes were small lymphocytes. However, the bone marrow contained a high proportion of labelled immature and mature plasma cells. The spleen produced large numbers of lymphocytes during the secondary immune response, many of which migrated to different organs probably as memory cells, while others were found in the bone marrow as effector cells from the immune response.

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Replacement of the ureter by an ileal loop. Quantitative aspects of long-term morphological alterations in minipigs.

In minipigs 1 ureter was replaced by a loop of the terminal ileum and the contralateral kidney removed. After 2.5 to 3 years the morphology of the replaced ureter was compared with the normal ileum. Independent of the 3 different operative techniques used, in about half of the pigs there was loss or flattening of the villi. In the other pigs the morphometrically determined number of intraepithelial lymphocytes, the cell density in the lamina propria and the length of the villi did not differ significantly compared with the normal ileum. There was no increase in goblet cells in the crypts. The transitional epithelium covered only a short distance at the anastomotic junctions. Peyer's patches of normal age-related size were found in the replaced ureter. Despite the long-term contact with urine instead of gut contents, in many pigs a normal amount of lymphocytes remained in the "ileal ureter".

Animals↗

Organ distribution and fate of newly formed splenic lymphocytes in the pig.

The production of lymphoid cells in the pig spleen was studied autoradiographically after selective labeling of the spleen using an extracorporeal perfusion circuit. Tritiated thymidine was added as a DNA precursor. One to 4 days after local labeling of the spleen the relative and absolute number of spleen-derived lymphocytes were determined in the following organs: mesenteric, cervical and inguinal lymph nodes, thymus, bone marrow, Peyer's patches, tonsils, three different parts of the gut, lung, liver, and blood. The labeled lymphocytes which migrated to these organs were all small lymphocytes, except for some large cells in the lamina propria. In the bone marrow, however, a considerable number of the spleen-derived immigrants were transformed into plasma cells. The total number of labeled lymphocytes decreased dramatically from Day 1 to Day 4 after labeling, indicating a high percentage of short-lived cells. Within the spleen, plasma cells had the highest labeling index of about 30% at Day 1 but this dropped to only 1.5% on Day 3. The organ distribution of the splenic emigrants changed from Day 1 to Day 4 with a relative increase in lymphocytes found in lymph nodes and a decrease in the lung and intestinal wall. The newly formed splenic lymphocytes migrated to T-and B-cell areas in lymph nodes, Peyer's patches and tonsils. In the intestinal wall labeled lymphocytes were found in the lamina propria and also as intraepithelial lymphocytes. There was no obvious redistribution between organ compartments with time after labeling of the spleen. The spleen produces large numbers of lymphocytes, which show typical organ distribution and homing to areas in lymphoid and nonlymphoid organs.

Animals↗

The three-dimensional reticular structure of the pig spleen demonstrated by labeling with fluorescein isothiocyanate.

Pig spleens were selectively perfused with a fluorescein isothiocyanate (FITC) containing medium. Histological sections of spleens excised one day later revealed the structure of the reticulum in thick cryostat sections directly in three dimensions. The pattern of the reticulum in silver impregnated sections was comparable to the FITC labelling pattern. This technique preferentially outlines the reticulum of the white pulp and the marginal zone of the spleen.

Animals↗

Innervation of the vas deferens and its importance for vasectomy and vasovasostomy.

The histotopographic localization of the nerves of the vas deferens was studied in dogs. The nerves of the vas are concentrated at two poles. A vasectomy with a nerve-sparing technique and a conventional vasectomy were performed in two groups of dogs. 5 months later all dogs underwent vasovasostomy. The intact nerve supply after vasectomy was controlled by electrophysical measurement. Sperm evaluation and morphological findings are compared and the advantages of a nerve-sparing technique demonstrated. The proliferative capacity of the epithelial layer of the vas deferens has to be considered in operations on the vas.

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