CYTOCHEMICAL OBSERVATIONS ON THE RELATIONSHIP BETWEEN LYSOSOMES AND PHAGOSOMES IN KIDNEY AND LIVER BY COMBINED STAINING FOR ACID PHOSPHATASE AND INTRAVENOUSLY INJECTED HORSERADISH PEROXIDASE

After incubation of formalin-fixed, frozen sections of kidney and liver from peroxidase-treated rats in an azo dye medium for acid phosphatase, and after subsequent incubation of the same sections with benzidine, phagosomes were stained blue and lysosomes were stained red in the same cells. It was observed that newly formed phagosomes were separate from preexisting lysosomes in the tubule cells of the kidney and in the Kupffer cells of the liver at early periods after treatment with peroxidase. At later periods, the color reactions for acid phosphatase and peroxidase occurred in the same granules. The reaction of peroxidase decreased gradually and disappeared from the phago-lysosomes after 2 to 3 days, whereas the reaction for acid phosphatase persisted. In the liver, most of the injected protein was concentrated in large phagosomes located at the periphery of the cells lining the sinusoids. The peribiliary lysosomes showed a relatively weak reaction for peroxidase in the proximity of the portal veins. After pathological changes of permeability, phagosomes and lysosomes lost their normal location and fused, in the interior of many liver cells, to form large vacuoles or spheres. The effects of a reduced load of peroxidase and the effects of the pretreatment with another protein (egg white) on the phago-lysosomes of the kidney were tested. The relationship of the fusion of phagosomes with lysosomes to the size of normal and pathological phago-lysosomes was discussed.     

Colorimetric analysis with N,N-dimethyl-p-phenylenediamine of the uptake of intravenously injected horseradish peroxidase by various tissues of the rat

1. A method is described for the colorimetric determination of peroxidase with N,N-dimethyl-p-phenylenediamine. The amount of red pigment formed by peroxidase is proportional to the concentration of enzyme and to the time of incubation during the first 40 to 90 seconds. The influence of the concentration of enzyme, N,N-dimethyl-p-phenylenediamine, H(2)O(2), the time of incubation, pH, the temperature, and the possible interference by oxidizing and reducing agents of tissues has been tested. 2. The method has been used to follow the uptake of intravenously injected horseradish peroxidase by 18 different tissues of the rat over a period of 30 hours. The highest concentration of the injected tracer enzyme was found in extracts of kidney, liver, bone marrow, thymus, and spleen. Considerable amounts were taken up by pancreas, prostate, epididymis, and small intestine. Lower concentrations were found in extracts of lung, stomach, heart, and skeletal muscle, aorta, skin, and connective tissue. No uptake was observed by brain and peripheral nerve tissue. 3. Tissue homogenates containing high concentrations of the injected peroxidase, in general also showed high or average activity of acid phosphatase. 4. Six hours after intravenous administration, the liver contained 27 per cent, the kidney 12 per cent, and the spleen, 1.4 per cent of the injected dose. 5. Approximately 20 per cent of the injected peroxidase was excreted in the urine during the first 6 hours, and the concentration of peroxidase in blood serum and urine fell exponentially during this time. After 6 hours, only low concentrations were excreted in the urine but low enzyme activity was still detectable after 30 hours. Approximately 6 per cent of the injected dose was excreted in the feces from 6 to 20 hours after administration. 6. After feeding through a stomach tube, low concentrations of peroxidase were found in blood serum and urine. Considerable variations in the extent of absorption from the gastrointestinal tract were observed in individual rats.     

Colorimetric Analysis with N,N-Dimethyl-p-phenylenediamine of the Uptake of Intravenously Injected Horseradish Peroxidase by Various Tissues of the Rat

1. A method is described for the colorimetric determination of peroxidase with N,N-dimethyl-p-phenylenediamine. The amount of red pigment formed by peroxidase is proportional to the concentration of enzyme and to the time of incubation during the first 40 to 90 seconds. The influence of the concentration of enzyme, N,N-dimethyl-p-phenylenediamine, H₂O₂, the time of incubation, pH, the temperature, and the possible interference by oxidizing and reducing agents of tissues has been tested. 2. The method has been used to follow the uptake of intravenously injected horseradish peroxidase by 18 different tissues of the rat over a period of 30 hours. The highest concentration of the injected tracer enzyme was found in extracts of kidney, liver, bone marrow, thymus, and spleen. Considerable amounts were taken up by pancreas, prostate, epididymis, and small intestine. Lower concentrations were found in extracts of lung, stomach, heart, and skeletal muscle, aorta, skin, and connective tissue. No uptake was observed by brain and peripheral nerve tissue. 3. Tissue homogenates containing high concentrations of the injected peroxidase, in general also showed high or average activity of acid phosphatase. 4. Six hours after intravenous administration, the liver contained 27 per cent, the kidney 12 per cent, and the spleen, 1.4 per cent of the injected dose. 5. Approximately 20 per cent of the injected peroxidase was excreted in the urine during the first 6 hours, and the concentration of peroxidase in blood serum and urine fell exponentially during this time. After 6 hours, only low concentrations were excreted in the urine but low enzyme activity was still detectable after 30 hours. Approximately 6 per cent of the injected dose was excreted in the feces from 6 to 20 hours after administration. 6. After feeding through a stomach tube, low concentrations of peroxidase were found in blood serum and urine. Considerable variations in the extent of absorption from the gastrointestinal tract were observed in individual rats.     

COMPARATIVE ANALYSIS OF THE CONCENTRATION OF INJECTED HORSERADISH PEROXIDASE IN CYTOPLASMIC GRANULES OF THE KIDNEY CORTEX, IN THE BLOOD, URINE, AND LIVER

The concentration of horseradish peroxidase in total particulate fractions from the kidney cortex did not change much during the first few hours after injection, as long as most of the injected protein was not yet cleared from the blood. It decreased at a rate of 6–8% per hr afterwards. The concentration of peroxidase in total particulate fractions increased in proportion to the load (dose) over a wide range, suggesting that a constant fraction of the protein was reabsorbed by micropinocytic vesicles into the tubule cells from the glomerular filtrate. The amount of peroxidase excreted in the urine also increased in proportion to the injected dose. The proportion of peroxidase taken up by the liver, however, decreased several times when the dose was increased. A marked decrease of protein uptake into the kidney cortex and an increase of urinary excretion were observed when rats received a second, equal dose of peroxidase 4 hr after the first injection, and the rate of clearance of peroxidase from the blood was decreased after the second injection. The liver, on the other hand, took up almost twice as much peroxidase after two injections as after one. The uptake of peroxidase by the kidney cortex increased with age. Cytochemical observations on the preferential absorption of peroxidase by certain cell types and segments of the renal tubules in relation to dose are reported.     

Rapid Cytochemical Identification of Phagosomes in Various Tissues of the Rat and their Differentiation from Mitochondria by the Peroxidase Method

1. Granules characterized by their ability to segregate foreign proteins (phagosomes) were identified in the cells of many rat organs after intravenous administration of horseradish peroxidase, by using the conventional test with benzidine for the histochemical detection of peroxidase. The largest numbers of phagosomes were identified in kidney and liver. Considerable numbers were observed cytochemically in pancreas, prostate, epididymis, thymus, spleen, bone marrow, small intestine, heart, pituitary, and mouse mammary carcinoma. 2. The variation in size of the phagosomes ranging from the limit of microscopic visibility up to 5 µ diameter, previously described for kidney, was also observed to occur in many of the other organs. The average size of the phagosomes in different organs was also different, the phagosomes of the liver, for example being on the average smaller than those of the kidney, pancreas, and prostate. 3. In squash preparations of kidney and liver, the phagosomes appeared often in curved rows following the course of the cell membranes of epithelial cells. In several other organs, they appeared aggregated in cells located in the vicinity of blood or lymphatic vessels or capillaries. 4. After injection of peroxidase directly into the brain of a rabbit, a striking concentration of peroxidase was observed in phagosomes of endothelial cells of capillaries and vessels, surrounding the site of injection. It was suggested that this localization may offer an explanation for the so called blood-brain barrier. 5. The cytochemical peroxidase method was applied to smears of isolated fractions of kidney and liver. Only the isolated phagosomes, but not the isolated nuclei, mitochondria, and microsomes, reacted with benzidine after administration of peroxidase. The contamination of conventionally prepared nuclear, mitochondrial, and microsomal fractions of kidney and liver with phagosomes of different sizes was observed. By correlating the cytochemical peroxidase test of smears of isolated fractions with the colorimetric determination of peroxidase, acid phosphatase, and cytochrome oxidase in the same fractions, the differentiation of the phagosomes from mitochondria and other cell granules was facilitated. 6. The marked difference in the osmotic properties of phagosomes and mitochondria, detectable after treatment with 70 per cent alcohol, and the difference in their affinities towards basic fuchsin, made it possible to differentiate the phagosomes from the mitochondria. It was found by this simple procedure that kidney cells of normal rats contain a large number of phagosomes ranging in size from 0.5 to 3 µ, whereas liver cells of normal rats contain relatively few phagosomes of this size but many smaller ones (0.2 to 0.5 µ diameter). These increased in size after treatment of the rats with horseradish peroxidase.     

Rapid cytochemical identification of phagosomes in various tissues of the rat and their differentiation from mitochondria by the peroxidase method

1. Granules characterized by their ability to segregate foreign proteins (phagosomes) were identified in the cells of many rat organs after intravenous administration of horseradish peroxidase, by using the conventional test with benzidine for the histochemical detection of peroxidase. The largest numbers of phagosomes were identified in kidney and liver. Considerable numbers were observed cytochemically in pancreas, prostate, epididymis, thymus, spleen, bone marrow, small intestine, heart, pituitary, and mouse mammary carcinoma. 2. The variation in size of the phagosomes ranging from the limit of microscopic visibility up to 5 micro diameter, previously described for kidney, was also observed to occur in many of the other organs. The average size of the phagosomes in different organs was also different, the phagosomes of the liver, for example being on the average smaller than those of the kidney, pancreas, and prostate. 3. In squash preparations of kidney and liver, the phagosomes appeared often in curved rows following the course of the cell membranes of epithelial cells. In several other organs, they appeared aggregated in cells located in the vicinity of blood or lymphatic vessels or capillaries. 4. After injection of peroxidase directly into the brain of a rabbit, a striking concentration of peroxidase was observed in phagosomes of endothelial cells of capillaries and vessels, surrounding the site of injection. It was suggested that this localization may offer an explanation for the so called blood-brain barrier. 5. The cytochemical peroxidase method was applied to smears of isolated fractions of kidney and liver. Only the isolated phagosomes, but not the isolated nuclei, mitochondria, and microsomes, reacted with benzidine after administration of peroxidase. The contamination of conventionally prepared nuclear, mitochondrial, and microsomal fractions of kidney and liver with phagosomes of different sizes was observed. By correlating the cytochemical peroxidase test of smears of isolated fractions with the colorimetric determination of peroxidase, acid phosphatase, and cytochrome oxidase in the same fractions, the differentiation of the phagosomes from mitochondria and other cell granules was facilitated. 6. The marked difference in the osmotic properties of phagosomes and mitochondria, detectable after treatment with 70 per cent alcohol, and the difference in their affinities towards basic fuchsin, made it possible to differentiate the phagosomes from the mitochondria. It was found by this simple procedure that kidney cells of normal rats contain a large number of phagosomes ranging in size from 0.5 to 3 micro, whereas liver cells of normal rats contain relatively few phagosomes of this size but many smaller ones (0.2 to 0.5 micro diameter). These increased in size after treatment of the rats with horseradish peroxidase.     

Dynamic pathways of selenium metabolism and excretion in mice under different selenium nutritional statuses

The selenoprotein, cellular glutathione peroxidase (cGPx), has an important role in protecting organisms from oxidative damage through reducing levels of harmful peroxides. The liver and kidney in particular, have important roles in selenium (Se) metabolism and Se is excreted predominantly in urine and feces. In order to characterize the dynamics of these pathways we have measured the time-dependent changes in the quantities of hepatic, renal, urinary, and fecal Se species in mice fed Se-adequate and Se-deficient diets after injection of (82)Se-enriched selenite. Exogenous (82)Se was transformed to cGPx in both the liver and kidney within 1 h after injection and the synthesis of cGPx decreased 1 to 6 h and continued at a constant level from 6 to 72 h after injection. The total amount of Se associated with cGPx in mice fed Se-deficient diets was found to be less than in mice fed Se-adequate diets. This finding indicated that cGPx synthesis was suppressed under Se-deficient conditions and did not recover with selenite injection. Excess Se was associated with selenosugar in liver and transported to the kidney within 1 h after injection, and then excreted in urine and feces within 6 h after injection. Any excess amount of Se was excreted mainly as a selenosugar in urine.     

~(125)Ⅰ—蜂毒肽在小鼠体内分布、吸收、排泄的研究

以氯胺T为氧化剂参照Hunter和Greenwood的多肽类化合物的碘化标记法,制备了~(125)I—标记的蜂毒肽在小鼠体内分布、吸收、排泄的研究,实验证明小鼠肌注~(125)I—蜂毒肽后,吸收很快,主要分布部位为肾、肺、心、肝、小肠、关节、脾与肌肉,脑组织中含量很少,肌肉注射后5分钟血液中含量可达70％,~(125)—I蜂毒肽主要经肾排泄,肌注后30分钟肾脏浓集最高,而尿液中以1.5小时为最高,而粪便中排泄少。     

Influence of high vitamin E dosages on retinol and carotinoid concentration in body tissues and eggs of laying hens

The aim of the study was to contribute to the discussion of overdosing vitamin E in laying hens. A total of 45 laying hens, divided into 5 groups were fed diets supplemented with either 0; 100; 1000; 10 000 or 20 000 mg dl‐α‐tocopheryl acetate/kg diet over a period of 10 weeks. Concentrations of vitamins A and E were measured in plasma, various tissues and egg yolk. Furthermore egg yolk colour and some carotinoids were measured in egg yolks. None of the vitamin E doses significantly influenced performance of the hens. As expected, vitamin E concentration in plasma, all tissue samples and egg yolk was significantly increased with increasing tocopherol content in the diet. The egg yolk showed the highest vitamin E concentration, followed by liver and muscles. Feeding 1000 mg α‐tocopheryl acetate per kg diet resulted in an increase of vitamin A concentration in the liver. Very high doses (10 000 and 20 000 mg/kg diet) significantly decreased retinol concentration in the liver and egg yolk, as well as carotinoid concentration in the egg yolk. The lower carotinoid concentration in egg yolk resulted in a decreased intensity of egg yolk colour. A prooxidative and/or competitive effect of very high doses of vitamin E with other fat soluble substances has been discussed.     

Distribution of cadmium in selected organs of mice: effects of cadmium on organ contents of retinoids and beta-carotene

Cadmium was administered to 32 adult ICR mice i.p. in two single doses (0.25 and 0.5 mg CdCl2, per kg of b.w.). After 48 hours concentrations of cadmium in kidneys, liver, spleen, muscle (m. quadriceps femoris), ovaries and testes and the concentration of retinyl palmitate, retinol and beta-carotene in kidney, liver and testes were determined. Significantly higher cadmium concentration was found in liver, kidney and ovary in both experimental groups in comparison with the control group (p<0.001). In muscle, spleen and testis the cadmium level was higher, however not significantly. No significant differences in the concentration of retinyl palmitate, retinol and alpha-carotene in liver were found. Concentration of alpha-carotene in kidney and testis was significantly decreased in both groups administered with cadmium (p<0.001). Concentration of retinyl palmitate was significantly lower in testis in the group with higher cadmium level (p<0.001) and the concentration of retinol significantly decreased in kidney and testis of mice after an administration of 0.5 mg CdCl2/kg b.w.     

Extrahpetic distribution of sulfobromophthalein.

Plasma clearance of sulfobromophthalein (BSP) is widely used as a measure of hepatic function. Its validity depends upon its exclusive elimination from the body via bile. For example, in the present study, when BSP was administered intravenously (i.v.) to rats at four different doses (18.75, 37.5, 75, and 150 mg/kg), less than 0.5% of each dose was excreted into the urine and between 70 and 85% was excreted into the bile within 6 h after administration. It has been assumed that the distribution of BSP is limited to the blood and liver witith very little appearing in other tissues. When we measured the amount of BSP in the plasma, liver, and the bile 10 min after the i.v. administration of either a high (150 mg/kg) or a low (18.75 mg/kg) dose of BSP, only 60% of the dose was accounted for. The concentration of BSP and 12-I-labelled albumin (RISA) was measured in various tissue samples 10 min after administration of 17.5 or 150 mg of BSP or RISA per kilogram. More BSP was found in all tissues than was contained in the plasma entrapped therein. Thus, the distribution of BSP is not limited to the liver and plasma. During excretion BSP leaves other tissue (kidney, spleen, lung, etc.) and is ultimately excreted into the bile.     

Distribution and breakdown of labeled coenzyme Q10 in rat

Radioactive coenzyme Q(10) ([(3)H]CoQ) was synthesized in a way that the metabolites produced retained the radioactivity. Administration of the lipid to rats intraperitoneally resulted in an efficient uptake into the circulation, with high concentrations found in spleen, liver, and white blood cells; lower concentrations in adrenals, ovaries, thymus, and heart; and practically no uptake in kidney, muscle, and brain. In liver homogenate most [(3)H]CoQ appeared in the organelles, but it was also present in the cytosol and transport vesicles. Mitochondria, purified on a metrizamide gradient, had a very low concentration of [(3)H]CoQ, which was mainly present in the lysosomes. All organs that took up the labeled lipid also contained water-soluble metabolites. The majority of metabolites excreted through the kidney and appeared in the urine. Some metabolites were also present in the feces, which further contained nonmetabolized [(3)H]CoQ, excreted through the bile. The major metabolites were purified from the urine, and the mass spectrometric fragmentation showed that these compounds, containing the ring with a short side chain, are phosphorylated. Thus, the results demonstrate that CoQ is metabolized in all tissues, the metabolites are phosphorylated in the cells, transported in the blood to the kidney, and excreted into the urine.     

Rate of concentration and digestion of radioactive growth hormone preparations injected in rats,as measured by the amount and nature of radioactivity in the tissues

Summary The distribution of radioactivity in the tissues of the rat has been established after the administration of radioactive bovine growth hormone preparations.Bovine growth hormone was used either transformed in to a¹⁴C-guanidinated derivative, which was fully active, of labeled with less than 1 mole per mole of¹²⁵I.The tissue radioactivity distribution curves obtained belong to two different categories: in kidney, liver and spleen there is an early concentration which attains a maximum in 15 minutes after the injection of the hormone, and rapidly declines. In heart, skeletal muscle, pancreas, intestine, bone and fat, the radioactivity increases gradually and a steady-state is reached after 30 to 60 minutes.Kidney is the organ where the highest concentration of radioactivity occurs. However, muscle accumulates more than 60% of the initial doses after 2 hours. Very little radioactivity appears in the urine, in this period.Similar results have been obtained with pharmacological or physiological doses of the labeled hormones.Blood plasma does not degrade the injected hormone but kidney, liver and muscle rapidly produce radioactive fragments soluble in 10% trichloro-acetic acid.Dedicated to ProfessorLuis F. Leloir on the occasion of his 70^th birthday.     

Quantification of trimesic acid in liver, spleen and urine by high-performance liquid chromatography coupled to a photodiode-array detection

The quantification of trimesic acid, a constitutive organic linker from the biodegradable porous iron(III) trimesate MIL-100(Fe) (MIL stands for Materials from Institut Lavoisier), has been performed in different biological complex media (liver, spleen and urine) using a liquid-liquid extraction procedure. A recovery exceeding 92 wt% was achieved from rat tissues and urine spiked with trimesic acid. After extraction, the determination of the trimesic acid concentration was realised by using a simple and accurate high-performance liquid chromatography (HPLC) method using photodiode-array detection (PDA) and aminosalicylic acid, as internal standard. Linearity of this method was kept from 0.01 to 100mg of trimesic acid per liter of urine and from 0.05 to 5.00 wt% of trimesic acid per tissue weight. The limit of detection of the method was 0.01 μg per injection. This method was finally applied to analyze and quantify the amount of trimesic acid in rat urine and tissue samples at the different stages of degradation of MIL-100(Fe).     

The metabolism of selenomethionine, Se-methylselenocysteine, their selenonium derivatives, and trimethylselenonium in the rat

The formation of dimethylselenide (respiratory) and trimethylselenonium (urinary) metabolites from [75Se]selenomethionine, [75Se]methylselenomethionineselenonium, [75Se]methylselenocysteine, [75Se]dimethylselenocysteineselenonium, and [75Se]trimethylselenonium was determined using single sc doses of 2 or 0.064 mg Se/kg in male and female rats. The 75Se content of liver, kidney, pancreas, testis, spleen, blood, heart, brain, and skeletal muscle was determined at 0.5 and 24 h. Respiratory 75Se after 24 h was greatest from Se-dimethylselenocysteineselenonium (38 and 17% for the high and low doses, respectively). Respiratory 75Se was about 8% for the high dose of Se-methylselenocysteine and was less for all other compounds. Total 75Se excretion in the urine was highest from rats given trimethylselenonium (about 90%, both doses) and was lowest from rats given selenomethionine (4%, low dose). Urine samples were chromatographed on SP-Sephadex cation-exchange columns and 75Se was eluted with ammonium formate; trimethylselenonium was precipitated with ammonium Reineckete solution and trimethylsulfonium carrier. Urinary trimethylselenonium excretion was greatest from rats given trimethylselenonium, but rats given Se-dimethylselenocysteineselenonium (low dose) excreted 35-45% of the dose as trimethylselenonium ion. The lowest quantity of trimethylselenonium was excreted by rats given the low dose of selenomethionine (0-3%). Pancreas, kidney, and liver showed the highest uptake (% of dose/g) of the selenium compounds. Trimethylselenonium was highly concentrated by the kidney and also showed high myocardial uptake (heart/blood ratio = 5) 0.5 h after injection; the selective uptake of trimethylselenonium in heart was not observed for the other selenonium compounds.     

The Metabolism of Salidroside to Its Aglycone p-Tyrosol in Rats following the Administration of Salidroside

Salidroside is one of the major phenolic glycosides in Rhodiola, which has been reported to possess various biological activities. In the present study the in vivo deglycosylation metabolism of salidroside was investigated and its aglycone p-tyrosol but not the original salidroside was identified as the main form in rat tissues following the administration of salidroside. After the i.v. administration of salidroside at a dose of 50 mg/kg in rats, salidroside was quantified only in the liver, kidney and heart tissues. The highest level of p-tyrosol was detected in the heart, followed by the spleen, kidney, liver and lungs, in order. Salidroside was detected only in the liver, in contrast, p-tyrosol was detectable in most tissues except the brain, and the kidney tissues contained a significant amount of p-tyrosol compared to the other tissues after the i.g. administration of 100 mg/kg salidroside. The excretion behaviour revealed that the administrated salidroside mainly eliminated in the form of salidroside but not its aglycone metabolite p-tyrosol through urine. After i.v. and i.g. administration in rats, 64.00% and 23.80% of the total dose was excreted through urine in the form of salidroside, respectively. In addition, 0.19% and 2.25% of the dose was excreted in the form of p-tyrosol through urine after i.v. and i.g. administration, respectively. The faecal salidroside and p-tyrosol concentrations were 0.3% and 1.48% of the total dose after i.v. administration, respectively. After the i.g. administration of salidroside, trace salidroside and p-tyrosol were quantified in faeces within 72 h. In addition, the biliary excretion levels of salidroside after i.v. and i.g. administration were 2.86% and 0.02% of the dose, respectively. The obtained results show that salidroside was extensively metabolised to its aglycone p-tyrosol and distributed to various organs and the orginal salidroside was cleared rapidly through urine following the administration of salidroside.     

Metabolism of bufotenine-2′-14C in human volunteers

The major ultimate metabolites of bufotenine-2′-¹⁴C in two volunteers have been identified and quantitated in the urine after intravenous administration of doses of 0.2 mg and 1 mg (10 μc/mg). Both subjects excreted over 90% of the ¹⁴C within 12 hours. Most of this was in the form of 5-hydroxyindoleacetic acid (68% and 74%). Only small amounts of bufotenine itself were excreted (1% and 6%). The remaining radioactive materials in the urine were not identified.     

Disposition and hepatoprotection by phosphatidyl choline liposomes in mouse liver

Small unilamellar liposomes with an average diameter of 80 nm were prepared from phosphatidyl choline of various sources using the dialysis method with cholate as a detergent. When 14C-labeled soybean liposomes were intravenously injected into male NMRI mice, up to 10% of the total label was found in the liver lipid. The uptake was dose-dependent and reached an apparent saturation 4 h after injection. The liver maintained a constant radioactivity corresponding to 1.9 +/- 0.13 mg phospholipid/g liver until ten hours after injection of 850 mg labeled phosphatidyl choline/kg body wt. Little radioactivity was taken up by the spleen. Analogous doses of liposomes prepared from egg yolk phosphatidyl choline led to a radioactivity corresponding to 1.3 +/- 0.4 mg lipid/g liver 4 h after injection. Liposomes with a similar size were prepared from hydrated, i.e., saturated phosphatidyl choline. After intravenous administration of these liposomes, an amount of 5.3 +/- 0.5 mg labeled lipid was found per g liver after 4 h. In contrast to unsaturated liposomes, 5.8 +/- 0.8 mg lipid per gram spleen was trapped by the spleen. The pharmacodynamic effect of these different liposomes was studied in benzo[a]pyrene-pretreated mice intoxicated with 400 mg/kg paracetamol. Animals which received paracetamol exhibited serum alanine aminotransferase activities of 4220 +/- 1140 units/l after 4 h and exhaled 120 +/- 19 nmol ethane kg-1 h-1. When pretreated with 850 mg soybean phosphatidyl choline/kg body wt. (i.v.) 2 h prior to paracetamol, the increase in serum transaminase activity was reduced to 117 +/- 104 units/l and ethane exhalation amounted to 18 +/- 8 nmol kg-1 h-1. In contrast, similar pretreatment with egg yolk phosphatidyl choline or hydrated phosphatidyl choline failed to protect against paracetamol-induced hepatotoxicity. The different pharmacodynamic effects of the two phosphatidyl cholines of plant or animal origin cannot be explained on the basis of their different pharmacokinetics. In the case of soybean phosphatidyl choline liposomes, the amount of radioactive lipid found in the liver correlated with the hepatoprotective potency.     

OCCURRENCE OF PHAGOSOMES AND PHAGO-LYSOSOMES IN DIFFERENT SEGMENTS OF THE NEPHRON IN RELATION TO THE REABSORPTION, TRANSPORT, DIGESTION, AND EXTRUSION OF INTRAVENOUSLY INJECTED HORSERADISH PEROXIDASE

The size, number, and location of lysosomes, phagosomes, and phago-lysosomes in different segments of the proximal and distal tubules, in the collecting tubules, and in invading macrophages of the kidneys of rats were compared by staining lysosomes (acid phosphatase) red, and phagosomes (injected horseradish peroxidase) blue in separate sections, and by staining phago-lysosomes purple by successive application of the reactions for the two enzymes in the same sections. It was concluded from these observations that the absorption of the foreign protein from the lumen and its gradual digestion in large phago-lysosomes took place mainly in the cells of the proximal convoluted tubules of the outer cortex. Several segments of the proximal convoluted tubules were distinguished on the basis of differences in the size and location of the phago-lysosomes and the amounts of peroxidase ingested. The distal tubules showed, in addition to moderate numbers of phago-lysosomes, many small phagosomes in the apical and basal zones of the cells. Moderate numbers of phagosomes and phago-lysosomes were observed in the cells of the collecting tubules. Macrophages showing very large phago-lysosomes were seen in the peritubular capillaries of the medulla, after injection of peroxidase. When high doses of peroxidase were administered, enlarged phago-lysosomes, parts of which seemed to be extruded into the lumen, were formed in the terminal segments of the proximal convoluted tubules.     

ENHANCEMENT OF BRAIN THIAMINE DIPHOSPHATASE ACTIVITY OF RATS BY INJECTION OF CHOLINERGIC DRUGS

Abstract— The effects of cholinergic drugs on thiamine diphosphatase (TDPase) in rat brain, liver and kidney were studied to clarify the role of the enzyme in the central nervous system.
Brain TDPase activity was markedly increased by intraperitoneal injection of a sub-lethal dose of physostigmine, ambenonium or pentetrazol. These drugs also increased the activity in the kidney, but not liver. Strychnine, atropine, and scopolamine did not affect the activity of brain TDPase, but decreased the enzyme activity of liver and kidney. Physostigmine also increased the activity of brain thiamine monophosphatase.
Brain TDPase activity reacheda maximum 30 minafterphysostigmine injection (1.0mg/kg). However, inhibition of brain acetylcholinesterase activity was greatest 45 min after physostigmine injection. The TDPase and AChE activities had both returned to normal values 60 min after the injection. The durations of these changes of TDPase and AChE activity corresponded to the duration of the tremor induced by physostigmine. The contents of total and phosphorylated thiamines in the brain but not in the liver or kidney were significantly reduced by physostigmine.
The relationship between ACh and activation of TDPase activity by cholinesterase inhibitors is discussed.