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Biomedical subjects

M G Johnston

Publications and source records attributed to M G Johnston.

At least 19 recordsLinked to original sources
PubMed1992

Is endothelium necessary for transmural pressure-induced contractions of bovine truncal lymphatics?

The purpose of this study was to investigate the role of the endothelium in regulating lymphatic contractile activity in bovine mesenteric truncal lymphatics. To test the effects of endothelial denudation on lymphatic pumping, bovine lymphatics were suspended in an organ bath preparation with the vessels cannulated at both inflow and outflow ends. By raising the heights of the Krebs reservoir and the outflow catheters appropriately, a transmural pressure could be applied to the vessels. The relationship between transmural pressure and fluid pumping is expressed as a bell-shaped curve with pumping increasing up to a peak pressure (between 8 and 12 cm H2O), and declining at pressures above this level. We compared pressure/flow curves in endothelium-intact and endothelium-denuded vessels. Chemical methods to remove the endothelium were attempted (collagenase and dispase) but were unsuccessful since they resulted in inhibition of all contractile activity including that induced with KCl. A technique using suture silk (3-O to 6-O) threaded into PE 50 or PE 20 polyethylene catheters with a loop at one end proved to be effective. By passing the catheter with its exposed loop of silk through the lymphatic ducts and imparting a twisting motion, we were able to remove the endothelium and preserve the smooth muscle responses to KCl. Transmission electron microscopy as well as silver staining techniques confirmed that all of the endothelium had been removed leaving the subendothelium and smooth muscle undamaged. We established that removal of the endothelium had no effect on lymphatic pumping. Pumping could not have occurred without the normal function of the one-way valves indicating that the denudation procedures did not damage these elements. Maximum flow rates and the transmural pressures that induced peak flows were similar in the two groups. We conclude that the pumping activity of bovine mesenteric truncal lymphatics in response to transmural pressure changes does not depend on an intact endothelium.

Animals↗
PubMed1992

Role of endothelial cells in regulating hemoglobin-induced changes in lymphatic pumping.

Studies with a sheep isolated duct preparation in vivo demonstrated that the route of administration of hemoglobin was important in demonstrating its inhibitory effect on lymphatic pumping. With autologous oxyhemoglobin administered intravenously (final plasma concentration 5 x 10(-5) M), pumping was not inhibited. However, the addition of oxyhemoglobin (5 x 10(-5) M) into the reservoir (lumen of the duct) resulted in > 95% inhibition of pumping. The extraluminal administration of oxyhemoglobin (10(-5) M) to bovine mesenteric lymphatics in vitro resulted in a 40% inhibition of pumping, whereas the introduction of oxyhemoglobin (10(-5) M) into the lumen of the vessels suppressed pumping 95%. In vessels mechanically denuded of endothelium, intraluminal oxyhemoglobin inhibited pumping 50%. These results suggested that oxyhemoglobin depressed pumping through an effect on both smooth muscle and endothelium. Once pumping was inhibited with oxyhemoglobin administration, stimulation of the duct with elevations in transmural pressure restored pumping activity when endothelial cells were present. However, in the absence of endothelium, pumping decreased with increases in distending pressures. We conclude that oxyhemoglobin has a direct inhibitory effect on lymphatic smooth muscle. The ability of oxyhemoglobin to alter the pressure range over which the lymph pump operates appears to be dependent on an intact endothelium.

Animals↗
PubMed1992

Studies on lymphatic drainage of the peritoneal cavity in sheep.

In the sheep, it is possible to cannulate several of the lymphatics that drain the peritoneal cavity and assess lymphatic drainage of this serous space directly. Indwelling catheters were placed in the caudal mediastinal and thoracic ducts. The right lymph duct could not be cannulated. Lymphatic drainage of the peritoneal cavity based on the movement of 125I-albumin from the cavity into the lymph compartments was not affected by the osmolality of the dialysate but was markedly altered by anesthesia. In addition, lymphatic drainage was assessed from the disappearance of instilled 125I-albumin from the peritoneal cavity and from the appearance of intraperitoneally administered 125I-albumin in the bloodstream and compared with data from the cannulated preparations. Lymph flows derived from tracer movement into the cannulated lymph compartments and from the appearance of tracer in the bloodstream were very similar. However, calculations of lymph flows based on the disappearance of tracer from the peritoneal cavity appeared to overestimate lymphatic drainage.

Anesthetics↗
PubMed1991

Lymphatic removal of dialysate from the peritoneal cavity of anesthetized sheep.

Several investigators have suggested that the lymphatic circulation reduces ultrafiltration in continuous ambulatory peritoneal dialysis (CAPD). The purpose of this study was to assess lymphatic drainage of the peritoneal cavity directly in anesthetized sheep under dialysis conditions. Lymph was collected from the caudal mediastinal lymph node and the thoracic duct, both of which are involved in the lymphatic drainage of the ovine peritoneal cavity, and from the prescapular lymph node, which is not involved in peritoneal lymphatic drainage. Fifty ml/kg volumes of a mildly hypertonic dialysis solution (Dianeal 1.5%) containing 25 microCi 125I-human serum albumin were instilled into the peritoneal cavity, and lymph flows and the appearance of labeled protein in the lymphatic and vascular compartments were monitored for six hours. Following the instillation of dialysis fluid there was a tendency for lymph flow rates from the thoracic duct to increase but these changes were not significant. However, flow rates from the caudal lymphatic demonstrated significant increases, especially in the final three hours of the monitoring period. Only about 8% of the radiolabeled albumin was removed from the peritoneal cavity over six hours (that is, 92% was left in the peritoneal space). Of the albumin removed, approximately 17% of this was drained by abdominal visceral lymphatics into the thoracic duct. About 25% passed through the diaphragm into the caudal mediastinal lymph node and into efferent lymph. Since the efferent lymphatic duct of the caudal mediastinal node empties directly into the thoracic duct, about 42% of all protein removed from the peritoneal cavity of the sheep was ultimately transported to the thoracic duct.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Lymphatic drainage of the peritoneal cavity in sheep.

Lymphatic drainage of the peritoneal cavity has been investigated in anesthetized sheep. Studies involving intraperitoneal administration of a complex of Evans blue dye and bovine serum albumin demonstrated the existence of three anatomically distinct pathways. In the first pathway, dye is removed from the peritoneal cavity by diaphragmatic lymphatics that pass into caudal sternal lymph nodes. Efferent lymphatics from these nodes transport the material to cranial sternal lymph nodes. Efferent cranial sternal lymphatics then convey the material either directly or indirectly, via tracheal lymphatic trunks, to the right lymph duct. In the second pathway, the complex is transported from the peritoneal cavity by diaphragmatic lymphatics that pass into the caudal mediastinal lymph node. Efferent lymphatic ducts from this node transport the material to the thoracic duct. The third pathway appears to involve transport of the dye across the mesothelial lining of the abdominal viscera and removal from the interstitium by afferent visceral lymphatics. Material taken up in this manner is ultimately transported to the thoracic duct by efferent visceral lymphatics. Experiments involving measurements of lymphatic absorption of 125I-labeled human serum albumin from the peritoneal cavity indicated that, over the 6-h period studied, 4.55 +/- 1.20 and 1.43 +/- 0.56% of the injected tracer could be recovered in thoracic duct lymph and caudal mediastinal efferent lymph, respectively, and the sum of these values represented 26% of the recovered radioactivity. On the other hand, 16.95 +/- 6.93% of the injected radioactivity could be found in the blood over the same period.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Production of plasminogen activator and plasminogen activator inhibitor by bovine lymphatic endothelial cells: modulation by TNF-alpha.

We have investigated whether lymphatic endothelial cells in culture produce plasminogen activators (PAs) and their inhibitors (PAIs) and if these activities can be modulated by the inflammatory cytokine Tumor Necrosis Factor alpha (TNF-alpha). Examination by reverse fibrin autography of the conditioned medium from these cells revealed a PAI of Mr 50 kDa. Also evident by fibrin autography were two species of PAs, of Mr 110 kDa and Mr 60 kDa. The 110 kDa protein co-migrated with the PA-PAI complexes and the 60 kDa protein co-migrated with tissue Plasminogen Activator (tPA). Functional and immunological assays indicated the human TNF-alpha increased the type 1 plasminogen activator inhibitor (PAI-1) in a time dependent manner. Treatment of the cells with recombinant human TNF-alpha for 24 hours resulted in a 3 to 7 fold increase in the amount of PAI released into the conditioned media. Immunoblot analysis identified the PAI in the TNF-alpha treated cell conditioned media, as PAI-1. Deposition of PAI-1 in the extracellular matrix then became apparent. TNF-alpha increased 4 fold the amount of tPA-PAI-1 complexes (Mr 110 kDa) detected in the conditioned media. Free tPA (Mr 60 kDa) decreased to 1/5 of control. Net fibrinolytic activity, as determined by a chromogenic substrate assay, decreased after TNF-alpha treatment. No urokinase type Plasminogen Activator (uPA) activity was detected in control or treated cells. This fibrinolytic activity may be important in maintaining free fluid movement in the interstitium and lymphatic vessels and in inflammatory states this potential may be decreased by the increase in PAI-1.

Animals↗
PubMed1990

Overview of proposed air emission standards and guidelines for municipal waste combustors.

The EPA proposed regulations for municipal waste combustors (MWCs) on December 20, 1989. The regulations include (1) performance standards under Section 111(b) of the Clean Air Act (CAA) for new, modified, or reconstructed MWCs and (2) draft emission guidelines and compliance schedules for the states to use to develop control requirements from existing MWCs under Section 111(d). This paper will outline the proposed air emission standards and guidelines, as well as the basis for the prescribed emission limits. The schedule for the remainder of the regulations development will also be discussed.

Air Pollution↗
PubMed1990

Modulation of lymphatic pumping by lymph-borne factors after endotoxin administration in sheep.

The purpose of this study was to test the hypothesis that endotoxin administration to sheep results in host-derived lymph-borne factors that modulate lymphatic pumping activity. To achieve this, two sheep were used for each experiment. In the test animal, a segment of intestinal lymphatic was isolated from all lymph input and provided with lymph from a reservoir. Pumping activity was initiated with a fixed transmural pressure applied to the test vessel, and the only input to this duct was provided by lymph from an indwelling catheter in a second donor sheep. The intravenous administration of endotoxin to the donor animals (33 micrograms/kg) generally resulted in increased pumping in the test vessels over the 1st h, but this was followed by reductions in pumping until flow stopped in all preparations. In control experiments (no endotoxin administered) pumping was unaffected. Further investigation revealed that these activities were relatively unstable and, in the case of the inhibitory material, appeared to act by decreasing the sensitivity of the vessel to changes in transmural pressure, because flow could be reestablished in the test vessels by elevating transmural pressures above the level originally chosen for the experiment. Endotoxin itself had no direct effect on sheep lymphatics in vivo or on bovine lymphatic vessels in vitro. However, the appearance of erythrocyte hemolysate (erythrolysate) in lymph was regularly observed after endotoxin infusion, and we demonstrated that erythrolysate (diluted to contain 10(-5) M hemoglobin) was a potent inhibitor of lymphatic pumping in vivo and in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Lymphatic pumping in response to changes in transmural pressure is modulated by erythrolysate/hemoglobin.

Red blood cells and lysate products (erythrolysate) are observed consistently in lymph draining acute and chronic inflammatory reactions and from tissues subjected to trauma or surgical procedures. Using hemoglobin as a marker for erythrolysate, we have measured hemoglobin in lymph up to the 10(-6) M range in a number of pathophysiological states. Data demonstrate that erythrolysate alters the pumping characteristics of lymphatic vessels. To test the effects of erythrolysate on lymphatic pumping, bovine lymphatics were suspended in an organ bath preparation with the vessels cannulated at both inflow and outflow ends. By raising the heights of the Krebs reservoir and the outflow catheters appropriately, a transmural pressure that stimulated pumping activity could be applied to the vessels. With a fixed transmural pressure of 6 cm H2O applied to the ducts, sheep erythrolysate depressed pumping activity between 40% and 100%, with dilutions containing between 10(-8) and 10(-5) M hemoglobin. Although the active principle in the red blood cells has not been characterized, evidence from precipitation purification experiments suggests that hemoglobin is an important component. Once suppressed, pumping could be restored in many but not all vessels (often to control levels) by elevating the distending pressure above 6 cm H2O. The relation between transmural pressure and fluid pumping is expressed as a bell-shaped curve, with pumping increasing up to a peak pressure (usually 8 cm H2O) and declining at pressures above this level. By comparing pressure/flow curves, we were able to ascertain that hemoglobin shifted the lymphatic function curve to the right and, on average, reduced the maximum pumping capability of the vessels. We speculate that the presence of erythrolysate/hemoglobin in lymph may modulate the ability of lymphatic vessels to drain liquid and protein from the tissue spaces.

Animals↗
PubMed1990

Quantitation of human melanoma, carcinoma and sarcoma tumor cell adhesion to lymphatic endothelium.

We have used an in vitro adhesion assay to study the interaction of tumor cells with lymphatic endothelium, a dynamic event that leads to tumor metastasis in vivo. 3H-thymidine-labeled human tumor cells from: one primary Ewing sarcoma, two established melanoma cell lines, two colon and two breast carcinomas (one established line and one primary culture of each) were added to 24-well culture dishes containing confluent monolayers of bovine lymphatic endothelium. Radioactivity associated with either the cells in suspension or the attached cells was assessed and compared at frequent intervals up to 360 minutes. Generally, tumor cell attachment increased as a function of time reaching a plateau between 180 and 360 minutes. the modular media system described here facilitates the primary and secondary culture (or co-culture) of a variety of normal and transformed cells. Primary cultures with a rounded morphology (one breast and one colon carcinoma) showed the lowest preferential attachment for lymphatic endothelium. All established cell lines and the primary Ewing sarcoma cell line displayed a more fibroblastic morphology and achieved the highest adhesion profiles. There was a correlation between the malignancy and attachment potential for the melanoma and breast carcinoma cell lines. Collectively, these data show that established tumor cell lines with fibroblastic-like morphology exhibit more rapid adhesion than primary tumor cell cultures with more rounded morphologies. While this property may reflect in vitro selection and/or adaptation, it does correlate with the metastatic propensity for some human tumor cells.

Adenocarcinoma↗
PubMed1989

Suppression of fluid pumping in isolated bovine mesenteric lymphatics by interleukin-1: interaction with prostaglandin E2.

In addition to local physiological forces, the modulation of lymphatic pumping by chemical mediators may play an important role in the regulation of extravascular water in inflammation and shock. Since Interleukin-1 (IL-1) appears to be of major importance in the host's response to infection by mediating many inflammatory events, we thought it important to determine if this cytokine could affect the lymphatic circulation and in particular to ask whether IL-1 was capable of altering lymphatic pumping in response to changes in transmural pressure. Bovine lymphatic segments (6 to 8 cm in length) were cannulated at both ends and suspended in an organ bath preparation. The vessels were provided with Krebs solution from a reservoir. With no net driving pressure, a transmural pressure applied to the ducts elicited contractile activity and fluid pumping with increases in pumping up to 8 cm H2O and reductions in flow above this level of distension. Human recombinant IL-1 alpha (10(-7) to 10(-9) M) administered into the lumina of the vessels depressed pumping activity approximately 5-30% at transmural pressures between 2 and 16 cm H2O. With limited supplies, we could only assess the effects of human recombinant IL-1 beta at 10(-8) M. However, it was more potent than IL-1 alpha, inhibiting pumping at all transmural pressures with maximum suppression in the range of 70% at peak flows. The ability of IL-1 to induce prostaglandin synthesis may be one of its most important biologic functions. It is likely therefore that IL-1 and PGE2 are closely linked and are probably present together in inflammatory lesions. With this in mind, we investigated the effects of PGE2 alone and in combination with IL-1 alpha. PGE2 by itself reduced pumping at concentrations between 10(-6) and 10(-9) M. When mixed with IL-1 alpha (both agents at a final concentration of 10(-9) M), the mixture had a marked inhibitory effect on flow, reducing pumping 50 to 70% at peak flows. The effect of the mixture of IL-1 alpha and PGE2 compared with the two agents administered separately was greater than the predicted additive effect at transmural pressures above 6 cm H2O. At lower transmural pressures, however, the level of real inhibition was less than the predicted additive effect.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

Modulation of fluid pumping in isolated bovine mesenteric lymphatics by a thromboxane/endoperoxide analogue.

A thromboxane/endoperoxide analogue (compound U46619) is known to stimulate phasic and tonic contractions in quiescent bovine lymphatic vessels and enhance contractile activity in spontaneously active vessels. In order to determine how these effects relate to changes in fluid propulsion by the lymphatics, we have assessed the effects of U46619 on the ability of isolated bovine mesenteric lymphatics to pump fluid in vitro. Bovine lymphatic segments (up to 8 cm in length with a minimum of 4 valves) were cannulated at both ends and fluid input provided from a reservoir. Flow through the vessels was regulated by intraluminal pressures. On average, changes in transmural pressures up to 8 cm H2O resulted in enhanced pumping; pressures above this level depressed flow. The dominant effect of U46619 (added to the reservoir) was to depress pumping; 10(-7) and 10(-9)M decreased flow at all transmural pressures tested; 10(-8)M had a dual effect, slightly inhibiting flow at low transmural pressures and enhancing flow at higher pressures. These results suggest that thromboxane may stimulate or inhibit lymphatic pumping depending on the concentration of the agent and the transmural pressure applied to the vessel. These effects may relate to its ability to induce variable changes in luminal diameter and frequency and force of contractions.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1988

Coagulation of sheep intestinal and prefemoral lymph.

We have determined the most suitable method for the automated analysis of the clotting parameters in sheep intestinal and prefemoral lymph as defined by the Activated Partial Thromboplastin Times (APTT; measure of intrinsic coagulation pathway) and the Prothrombin Times (PT; measure of extrinsic coagulation pathway). As opposed to optical density systems, the use of a Fibro-System Fibrometer was found to provide the most consistent assessment of coagulation with the endpoint being the time to fibrin strand formation. We measured APTT in sheep intestinal and prefemoral lymph of 59.78 +/- 7.69 seconds and 51.03 +/- 10.49 seconds respectively. These values were more prolonged than those obtained from sheep blood plasma but only in the case of intestinal lymph were the differences significant (p less than 0.025). Human blood APTT values were significantly less than both sheep blood (p less than 0.05) and sheep intestinal (p less than 0.001) and prefemoral lymph (p less than 0.01). PT values were found to be 21.56 +/- 1.14 seconds in intestinal and 22.00 +/- 1.88 seconds in prefemoral lymph. These values were also significantly greater than those obtained from sheep blood (both p less than 0.001). Human blood PTs were significantly less than both sheep blood (p less than 0.001) and intestinal and prefemoral lymph (both p less than 0.001). Measurement of APTT and PT values in intestinal lymph and PT determinations in prefemoral lymph were not affected by storage in the refrigerator or freezer. There was some indication that APTT values in prefemoral samples were susceptible to storage artifacts; however, the differences in coagulation times were not significant.

Animals↗
PubMed1987

Decreased lymphatic pumping after intravenous endotoxin administration in sheep.

The effects of endotoxin on the ability of lymphatic vessels to pump fluid in vivo have been assessed with the use of a sheep model system that permits analysis of lymph pumping in sheep without the complication of variable lymph inputs. This involved the isolation of intestinal lymphatic vessels from all lymph input, with saline or lymph provided from a reservoir. The blood and nerve supplies to the vessel were left intact. With no net driving pressure, but with a transmural pressure applied to the vessel to initiate spontaneous contractions and fluid pumping, the intravenous administration of endotoxin (3.3 micrograms/kg in anesthetized sheep and 33 micrograms/kg in nonanesthetized animals) reduced fluid propulsion in both groups of animals (P less than 0.02 and P less than 0.03, respectively). Comparisons with animals that did not receive endotoxin revealed maximum inhibition greater than 90% in anesthetized and 50% in nonanesthetized sheep. Normal pulsatile lymphatic pressures (produced from lymphatic contractions) were reduced in frequency and amplitude after endotoxin administration. Endotoxin itself had no effect on the vessels when added to the fluid in the reservoir, suggesting that the inhibition of the "lymph pump" was mediated through the interaction of endotoxin with cellular or humoral elements in the host. In addition to suppression of lymphatic contractile activity, the intravenous injection of endotoxin enhanced lymph formation as indicated by the 3- to 10-fold increases in lymph flow rates in the two groups. We conclude that, for a given transmural pressure, the systemic administration of endotoxin reduces lymphatic pumping activity. We speculate that this effect may be important in the pathogenesis of the edema associated with sepsis.

Anesthesia↗
PubMed1987

Increased intrinsic pumping of intestinal lymphatics following hemorrhage in anesthetized sheep.

The return of fluid and protein to the bloodstream by the lymphatic circulation may play an important role in reconstituting intravascular volume following hemorrhage. In this study, we have defined the lymph-flow changes that occur in cannulated mesenteric lymphatics following a 25% blood loss in anesthetized sheep and investigated the effects of hemorrhage on the intrinsic contractile activity (lymph pumping) of these vessels in vivo using a new model system. The removal of 25% of the calculated blood volume resulted in increases in lymph flow over a 6-hour period, with peak changes to 3.5 times the prebleed levels. Systemic arterial pressures dropped to roughly 50% of control values immediately following the bleed and returned to control in 3 hours. To directly assess the effects of hemorrhage on lymphatic pumping, a segment of intestinal lymphatic was isolated from all lymph input and supplied with fluid from a reservoir. While there was no net pressure driving fluid through the duct, a transmural distending pressure was applied to the vessel, which stimulated resting lymphatic contractions and fluid pumping. A 25% blood loss resulted in increased activity of the lymph pump; up to 6 times more fluid was propelled through this "isolated" vessel in vivo than in similar preparations in sheep that were not bled (p less than 0.01). Measurements of fluid pulse pressures in this preparation indicated increased pumping frequency and/or force after hemorrhage compared with prebleed levels. We conclude that lymphatic contractile activity is stimulated after a blood loss independent of changes in lymph formation and speculate that this mechanism may play an important role in the reexpansion of the vascular space.

Animals↗
PubMed1987

The regulation of lymphatic pumping.

An important functional property of lymphatic vessels is their ability to pump fluid. To quantitate this activity in vivo, sheep mesenteric lymphatic segments were isolated from all lymph input and provided with lymph plasma or saline from a reservoir. Lymphatic pumping was controlled by transmural pressure with increases in pressure resulting in elevated fluid pumping followed by reductions in flow at high intraluminal pressures. With lymph input to the vessels denied, but with blood and nerve supply left intact, the pumping activity could be altered with systemic physiological perturbations including a major blood loss and the intravenous infusion of endotoxin. In each case, it was clear that the modulation of lymphatic pumping resulted from the direct effects of the test procedure on the 'lymph pump' and not from effects on vascular parameters or lymph formation.

Animals↗