Absorption of beta-casomorphins from autoperfused lamb and piglet small intestine.

beta-Casomorphins (beta-CMs) derived from milk beta-casein may exert various opiate activities in milk-fed infants. To assess the physiological significance of beta-CMs as a source of circulating opioids in infants, we measured absorption rates of several beta-CMs under near-physiological conditions using in situ autoperfused lamb intestine. The naturally occurring beta-CMs, beta-CM-7 and beta-CM-4-amide, were absorbed readily into blood with no transfer into lymph. Uptake peaked within several minutes of the luminal infusion of peptide but then declined sharply and stopped within a further 10-15 min. The recovery in blood, intestinal contents, and tissue at the end of the 30-min experiment was less than 1% of the infused dose. The low recovery was due to rapid proteolysis based on in vitro studies that demonstrated half-lives of less than 5 min in lamb blood, luminal contents, and lymph. The synthetic dipeptidyl peptidase IV-resistant analogue beta-[D-Ala2]CM- 4-amide was stable during incubation in blood, lymph, or luminal contents and was absorbed into blood at rates that were maximal within several minutes and remained steady for the 30-min period. We conclude that although natural beta-CMs are transferred across the lamb small intestine, rapid degradation within the intestinal lumen, gut epithelium, and blood would prevent entry into the circulation under normal conditions. Val-beta-CM-7, a putative stable precursor, had similar stability and kinetics of absorption to beta-CM-7, results that exclude Val-beta-CM-7 as a stable precursor for delivery of beta-CMs to the circulation. Essentially identical results to those in lambs were obtained in 7-day-old piglets.

A comparison of absorption of glycerol tristearate and glycerol trioleate by rat small intestine.

Generally, fats rich in saturated fatty acids raise serum cholesterol, whereas fats rich in polyunsaturated fatty acids lower it. There appear to be exceptions; e.g., stearic acid (18:0)-rich fats have little or no effect on serum cholesterol concentrations. This apparent lack of cholesterolemic effect of stearic acid-rich fat could be because intestinal absorption of fat is poor or subsequent plasma and/or tissue metabolism of fat is different. To investigate mechanisms involved, we compared intestinal digestion, uptake, and lymphatic transport of glycerol tristearate (TS) and glycerol trioleate (TO, 18:1). Two groups of rats bearing intestinal lymph fistulas were used. TO rats were fed intraduodenally for 8 h at a constant rate a lipid emulsion of 25 mumols/h of TO (labeled with glycerol tri[9,10 (n)-3H]oleate), 7.8 mumols of egg phosphatidylcholine, and 57 mumols of sodium taurocholate in 3 ml of phosphate-buffered saline. TS rats were fed the same lipid emulsion except that TS replaced TO and the emulsion was labeled with glyceryl [1,3-14C]tristearate. The lymph triglyceride and radioactivity were determined. After infusion, the luminal and mucosal radioactive lipid content was analyzed. The results showed that there was significantly less lipid transported in the lymph of TS rats compared with TO rats. The results also showed a significant decrease in the absorption of TS as compared with TO. This was due in part to poor lipolysis. In addition, the lipid absorbed by the intestine of the TS rats was transported into lymph less efficiently than in TO rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Temocillin: lymph penetration and protein binding.

Temocillin, a novel betalactam antibiotic, was administered in doses of 1,2 and 4 g i.v. to 12 healthy subjects and the plasma concentrations of free and protein bound temocillin assayed and protein binding parameters were calculated. In a second study 2 g of temocillin was administered i.v. to 12 healthy subjects and samples of lymph were collected and assayed for total temocillin. Using the protein binding parameters so obtained the corresponding free temocillin in lymph was calculated. The clinical significance of the lymph penetration is discussed.

Pressure within the thoracic duct modulates lymph composition.

The amount of lymph received by the thoracic duct depends on each contributing organ's ability to produce interstitial fluid and generate a pressure differential moving lymph into the central lymphatic circulation. It has been reported that varying the pressure within the thoracic duct could alter each organ's contribution to thoracic duct flow. The thoracic duct above the diaphragm was cannulated to obtain lymph from the liver, gut, and lower body. Pressure within the thoracic duct was elevated serially by increasing the lymphatic cannula outflow height. This caused lymph protein concentration to increase while chyle concentration (measured by absorbance) decreased. The data demonstrate that as thoracic duct pressure increases, the percentage contribution of gut lymph flow (as represented by chyle concentration) decreases while the contribution of lymph originating within the liver (as indicated by higher protein concentration) increases. We conclude that pressure variation within the central lymphatic system affects the amount of lymph or edema fluid leaving any given organ.

[Status of lymphatic vessels, lymph flow and lymph toxicity in patients with obliterating arterial diseases of the lower extremities]. / Sostoianie limfaticheskikh sosudov,limfotoka i toksichnost' limfy u bol'nykh s obliteriruiushchimi zabolevaniiami arterii nizhnikh konechnostei.

The condition of the lymph vessels, the lymph flow, and the toxicity of peripheral lymph were studied in 46 patients with obliterating diseases of the arteries of the lower limbs. The results of the study showed the severity of the destructive changes in the peripheral lymphatic system to be directly dependent on the severity of the arterial ischemia of the limb. Block of the lymphatic system, deceleration of the lymph flow, and changes in the lymph vessels themselves lead to deterioration of the tissue drainage function and intensification of regional endotoxicosis. In view of which correction of the peripheral lymphatic system should be included in the complex of measures for the treatment of patients with obliterating diseases of the lower limb arteries.

Surface-active phospholipid in muscle lymph and its lubricating and adhesive properties.

In order to investigate the role of phospholipid in facilitating lymph flow, "deep-thigh" lymph was collected from ten anesthetized dogs and analyzed for phospholipids using thin-layer chromatography. The lymph was found to be surface active at liquid and at solid surfaces at which it deposited a hydrophobic monolayer in vitro. Extracted phospholipid was found to be an effective release agent as a monolayer, reducing the force of adhesion of 5% protein solutions by 76% according to a standard test for tacky glues. The same monolayers were effective lubricants, reducing friction by 96%; while mixtures of the same phospholipids from synthetic sources gave similar results for release and lubrication. Surfaces in contact with extracellular fluid or lymph in vivo were found to be hydrophobic with a drop of saline on semitendinosus muscle fibers displaying a contact angle of 40.2 degrees +/- 7.2 degrees. The results are considered compatible with the hypothesis that surface-active phospholipid facilitates the flow of lymph; while it could also provide boundary lubrication for sliding of connective tissue in locomotion and for any relative movement of motor units in muscle contraction and fatigue.

[Fibronectin from the lymph of healthy persons]. / Fibronektin limfy zdorovykh liudei.

Content of fibronectin was studied in lymph and blood plasma of 24 healthy 23-57 years old men. Concentration of fibronectin in lymph and blood plasma was 44.2 +/- 21.7 mg/ml and 354 +/- 88 mg/ml, respectively. Interrelations were not found between content of fibronectin in lymph and blood plasma as well as between the total content of protein and fibronectin in lymph. Fibronectin in lymph was shown to occur in its intact form using immunoblotting procedure involving monoclonal antibodies to fibronectin. Simultaneously with whole molecules of fibronectin its fragments appear to be present in lymph.

Recovery of fish oil-derived fatty acids in lymph of thoracic duct-cannulated Wistar rats.

The absorption of equivalent doses of eicosapentaenoic and docosahexaenoic acids was compared in rats when administered as the ethyl ester concentrate, ethyl ester concentrate plus olive oil, free fatty acid or triacylglycerol (menhaden oil). Lymph was collected from a thoracic duct cannula for 24 hr after dosing via an indwelling duodenal catheter. After 24 hr, the absorption of eicosapentaenoic acid was greater for the free fatty acid and menhaden oil than for the ethyl ester form, but docosahexaenoic acid absorption was comparable for all forms. Other rats had greater plasma levels of eicosapentaenoic and docosahexaenoic acids 5 hr after oral gavage dosing with menhaden oil than did rats dosed with the ethyl ester form.

Contractility of human leg lymphatics during exercise before and after indomethacin.

Subcutaneous lymphatics in the lower leg were catheterized in the retrograde direction in 6 healthy male subjects. The catheter was connected to a pressure transducer, and pressure was measured during three stages of exercise including standing, tip-toeing, and running in place. Before the third stage, indomethacin (50mg) was given i.v. Rhythmic pressure waves were registered in each subject. During the second stage, when the subjects were "warmed up," the frequency (min-1) was 2.4 +/- 0.5 (mean +/- SEM). The corresponding values during tip-toeing and running were 5.8 +/- 0.7 (p less than 0.05) and 5.4 +/- 0.5 (p less than 0.05), respectively. The amplitudes (mean values between 3.2-4.7mmHg while standing) were not consistently altered during tip-toeing or running in any of the three stages. During standing there was a negative correlation between frequency and amplitude. No such correlation was found during tip-toeing or running, or after injection of indomethacin. Indomethacin did not significantly alter any of the measured parameters, but in two subjects the frequencies and amplitudes were decreased (about 50%) during standing, tip-toeing, and running.

Lung vascular permeability changes in lambs with hyaline membrane disease.

To investigate the mechanism of pulmonary edema in hyaline membrane disease (HMD), lymph from the efferent duct of the mediastinal lymph node was collected in premature lambs before and after delivery by cesarean section. Mean lymph flow in 7 lambs with histologically verified HMD increased progressively over 4 h after delivery to 3 times the fetal value, while lymph flow in 7 lambs without HMD increased to 3.5 times at 1 h and decreased thereafter. At 4 h after birth, lung lymph flow was significantly higher in lambs with HMD than in lambs without HMD (0.70 +/- 0.15 (SEM) vs 0.43 +/- 0.07 ml.h-1.kg-1). Lymph/plasma concentration ratio for small endogenous protein fractions (effective molecular radius, 3.6 and 3.8 nm) was significantly higher in lambs with HMD than in lambs without HMD at 2-4 h. Postmortem extravascular lung water was significantly higher in lambs with HMD (6.1 +/- 0.5 vs 4.3 +/- 0.3 ml/g dry lung weight). It is concluded that lung water is high in lambs with HMD, which appears to be a result both of delayed absorption of fetal lung liquid and increased permeability of the pulmonary exchange vessels.

The concentration of apolipoprotein A-II in human peripheral lymph.

The concentration of apolipoprotein A-I (apoA-I) and of apolipoprotein A-II (apoA-II) was determined immunoelectrophoretically in lymph and plasma of six subjects. The concentration in lymph of apoA-I was 20.3 +/- 3.1 mg/dl, that of apoA-II was 4.6 +/- 0.5 mg/dl. The ratio of the concentration in lymph over that in plasma (CL/CP) for apoA-I was 0.14 +/- 0.1, that for apoA-II 0.14 +/- 0.01. Lymph and plasma samples from two subjects were fractionated by exclusion chromatography and the concentration of both apolipoproteins in resulting fractions was determined immunoelectrophoretically. ApoA-I of lymph eluted with fractions spanning a wider range of particle sizes than plasma apoA-I, while lymph apoA-II eluted predominantly with fractions that contained particles corresponding in size to plasma apoA-II-containing particles. It appears that the largest and smallest lymph HDL represent subspecies of Lp(A-I without A-II). These findings are discussed in the context of their possible bearing on initial stages of reverse transport of cholesterol.

A rapid method for staining large chylomicrons.

In this report, we present a rapid method for producing high-quality micrographs suitable for determining the size distributions of particles in concentrated samples of postprandial chylomicrons and chylomicron remnants. The procedure consists of mixing particles with osmium tetroxide in water to stabilize the lipids of the particles. These fixed and positively stained particles are then negatively stained with phosphotungstate in the presence of dilute sucrose. This dual staining procedure prevents the fusion and clustering of chylomicrons during processing for electron microscopy and is effective with particles of different lipid compositions. In addition, this procedure is simple and rapid, adding only one mixing step and 5 min to the preparation time required for conventional negative stains.

Fibronectin fragments in lung lymph after thrombin-induced lung vascular injury.

Fibronectin is an adhesive glycoprotein found in plasma and lymph as well as between lung endothelial cells and their collagenous basement membranes. Fibronectin is highly sensitive to proteolytic cleavage. We determined if fragments of fibronectin appear in lung lymph in association with increased lung protein clearance after thrombin-induced intravascular coagulation. Thrombin was infused intravenously, (80 units/kg for 30 minutes) into sheep (n = 8) surgically prepared with chronic lung lymph fistulas. Plasma and lymph fibronectin was assayed by electroimmunoassay. Fibronectin fragments were detected using Western blot analysis. After thrombin infusion, lymph flow increased 650% above baseline within 1-2 hours in association with a 35% decline in lymph-to-plasma total protein concentration ratio. This was followed by a second phase (3.5-6 hours) of normalized lymph-to-plasma ratios coupled with sustained elevation of lymph flow. Lung protein clearance remained elevated (p less than 0.10) for 5.5 hours. Plasma fibronectin levels declined slightly over 1-5 hours (zero time = 597 +/- 64 micrograms/ml; 1.5 hours = 478 +/- 59 micrograms/ml) and then increased significantly (p less than 0.05) over 24-48 hours (760 +/- 85 micrograms/ml). The amount of low molecular weight fibronectin fragments in lung lymph increased over the 1.5-6 hours post-thrombin and then declined over 12-48 hours. Thus after thrombin infusion, fragments of fibronectin were usually detected in increased amounts of lung lymph in association with an elevation of lung protein clearance.

Interrelated effects of dietary fiber and fat on lymphatic cholesterol and triglyceride absorption in rats.

Lymph cannulated rats were administered intragastrically a test emulsion containing 25 mg of [14C]cholesterol, 50 mg of either guar gum, cellulose or chitosan, and 200 mg of either safflower, high-oleic safflower or palm oil, and the absorption of labeled cholesterol and fatty acids was measured. The type of both dietary fiber (P less than 0.001) and fat (P less than 0.05) significantly influenced cholesterol absorption. A significant interaction of fiber and fat on cholesterol absorption (P less than 0.05) was also observed. Chitosan effectively lowered cholesterol absorption more than did guar gum or cellulose, and this effect was more significant when given with safflower or high-oleic safflower oil than with palm oil. When guar gum was the source of dietary fiber, dietary fats did not modify cholesterol absorption. Dietary fiber also significantly affected triglyceride absorption (P less than 0.05). Absorption tended to be low in the chitosan, high in the cellulose and intermediate in the guar gum group. Absorption of safflower and high-oleic safflower oils tended to be higher than that of palm oil when cellulose or guar gum was fed. Guar gum, as compared with the other fibers, altered the absorption pattern of both cholesterol and triglyceride. The results showed that the type of dietary fat significantly influenced the effect that dietary fiber exerted on lipid absorption.

Pulmonary deposition and clearance of aerosolized alpha-1-proteinase inhibitor administered to dogs and to sheep.

Augmentation of lung antiprotease levels may be an important therapeutic intervention in the prevention of pulmonary emphysema. We have administered aerosols of plasma-derived human alpha 1 proteinase inhibitor (A1PI) to the lungs of dogs and sheep to investigate (a) delivery of the protein to the distal air spaces of the lung; (b) maintenance of functional activity of the protein; and (c) flux of the protein across the components of the alveolar-capillary membrane. A1PI (26.4 mg/kg body weight) was administered as an aerosol to anesthetized animals; sheep were prepared for the chronic collection of lung lymph. Immunoperoxidase staining of lung tissue obtained 2 h after administration of A1PI demonstrated the presence of human A1PI on the surface of alveoli and distal bronchioles. Bronchoalveolar lavage fluid recovered at intervals after A1PI administration demonstrated time-dependent elevations of human A1PI levels with augmentation of lavage fluid antielastase activity in proportion to the content of human A1PI. Using radiolabeled A1PI as a tracer, we found that 32% of the aerosol was retained in the animals' lungs. Measurements of the rate of loss of A1PI from the lung and of the rate of appearance of human A1PI in plasma resulted in a calculated permeability of the alveolar-capillary membrane to A1PI of 3.49-6.39 X 10(-10) cm/s. Experiments using instrumented sheep allowed independent calculation of endothelial permeability to A1PI of 122-236 X 10(-10) cm/s and calculation of epithelial permeability of 4.70-4.81 X 10(-10) cm/s. Modeling of aerosol delivery of A1PI to humans using the results of these studies predicts that the ratio of plasma/alveolar levels of delivered A1PI will be 0.024, and that aerosolization of 175 mg A1PI/d will result in an A1PI alveolar fluid level of 1.0 mg/ml. Aerosol administration of A1PI may provide an efficient method of augmenting alveolar antiprotease levels.

Alternative pathways for drainage of cerebrospinal fluid in the canine brain.

Although the brain has no formal lymphatic system, a substantial quantity of cerebrospinal fluid (CSF) has nonetheless been shown to drain via cervical lymphatics. To pursue further the issue of alternative drainage pathways for CSF, we infused a solution of Ringer's lactate (RL) into the cisterna magna of the dog brain and monitored both the flow and concentration of total protein of cervical lymph. This maneuver promoted a nearly three-fold rise in intracranial pressure and was accompanied by a rise in cervical lymph flow and fall in its protein content. In addition, a profuse nasal discharge (11.4 ml/hr) developed with a moderately high protein content of the rhinorrhea fluid (1.8 g/dl), along with similar appearance times of Evans blue dye (instilled in the cisterna magna) in both cervical lymph and the rhinorrhea fluid (48-70 minutes after infusion). These findings suggest alternative drainage pathways for CSF besides the arachnoid villi (Pacchionian bodies) including connections with lymphatics in the neck and along the olfactory nerve, and around the cribiform plate to the nasal submucosa, and with proptosis, perhaps also through the aqueous humor-canal of Schlemm and nasolacrimal duct.

Chylomicron synthesis in experimental nephrotic syndrome.

Mesenteric lymph was collected for 48 h from rats with aminonucleoside-induced nephrotic syndrome, receiving an intraduodenal infusion of a triacylglycerol emulsion. In nephrosis, the rates of lymph flow and triacylglycerol transport were approx. 2-fold higher, but the transport of total protein and of apoproteins A-I and E was 2- to 3-fold lower than that in control rats, resulting in chylomicrons with a 3-fold approx. elevated triacylglycerol/protein ratio. Supplementation of the triacylglycerol infusate with glucose and amino acids did not increase the protein or apoA-I and apoE transport. Production or transport of B and C apoproteins in nephrotic rats was also reduced, as indicated by tetramethylurea solubility, incorporation of intraduodenally infused [3H]leucine and staining of the chylomicron proteins on SDS-PAGE gels. Apoprotein A-IV was the only chylomicron component into which the leucine incorporation was elevated, but its relative content was not increased on SDS-PAGE gels. Lymph chylomicrons of nephrotic rats were larger in size (1498 +/- 37 vs. 1235 +/- 23 A), consistent with the higher triacylglycerol/protein ratio. The concentration of all lipoprotein classes was markedly elevated in the plasma of nephrotic rats, as was that of the total A-I and E apoproteins. Intravenous injection of 125I-labelled HDL, followed by tracing of the label in lymph chylomicrons, indicated a lower rate of transfer of HDL apoproteins from plasma to lymph in nephrotic rats. We conclude that the intestinal chylomicron formation in nephrosis is characterised by an enhanced triacylglycerol transport without the appropriate apoprotein complement. This is probably due to the limited capacity of enterocytes, in marked contrast to hepatocytes, to respond to the hypoproteinemia of nephrosis with increased production and/or transport of the apoproteins.

Rat intestinal lymph osmolarity during glucose and oleic acid absorption.

The two major purposes of this study were to determine 1) how glucose and oleic acid absorption by the intestinal villi influenced the osmotic composition of lymph as it exited the villus base and 2) what if any changes in lymph osmolarity occurred as the lymph traversed through the bowel wall. The rat jejunum was used in all studies and lymph was collected from individual lymphatics at 0.5-1 nl/min during control states and luminal exposure to 35-550 mg% glucose solutions (isotonic in saline) and 5 and 20 mM oleic acidtaurocholate solutions. Lymph collected from the base of villi during vigorous motility had an osmolarity of 403 +/- 15 mosM at rest and was only increased 30-50 mosM more except during exposure to 550 mg% glucose, where osmolarity increased over 100 mosM. Under comparable conditions, the submucosal lymph osmolarity at rest was 302 +/- 3.5 mosM and increased to 330-350 mosM during exposure to all of the solutions tested. When intestinal motility was virtually stopped, the submucosal lymph osmolarity was isotonic for all solutions tested. These observations indicate that absorption of glucose and oleic acid increased the osmolarity of lymph, leaving the villus only 30-50 mosM unless a glucose concentration of 550 mg% was present. Furthermore, the increased flow of villus lymph during absorption raised the osmolarity of the submucosal lymph when bowel motility assisted the lymph propulsion. This movement of materials from the villus to the submucosa by venular blood and lymph flow provides an opportunity for the villus tissue to influence the composition of the submucosal interstitial environment.

[Lipid metabolism and lymph coagulability in the postresuscitation period]. / Lipidnyi obmen i svertyvaemost' limfy v postreanimatsionnom periode.

The values of lipid metabolism and lymph coagulation in the restoration period after resuscitation were studied in experiments on rabbits. A clear correlation between the values was revealed. Increase of concentration of total lipids and of lipid fractions was accompanied by increase of the lymph coagulation potential, which led to disturbed tissue drainage of lymph and was one of the important causes of the development of postresuscitation complications. The elaboration of pathogenetic therapy for regulation of lipid metabolism and lymph coagulation in the postresuscitation period is necessary.

Sampling of lymph from lymph vessels afferent to the supramammary lymph gland in the cow.

A technique for catheterization of a lymph vessel afferent to the supramammary lymph gland in the cow is described. The operation was made with the cow under general anaesthesia. The lymph vessel was catheterized with a 30 cm long heparinized polyethylene catheter with an external diameter of 2.5 to 3 mm. Generally at least 5 ml of lymph could be collected within a period of 3 to 5 minutes. In 11 out of 15 cows lymph could be collected for 7 days or longer.