In-vivo Uptake Research Articles

Microvascular recruitment is an early insulin effect that regulates skeletal muscle glucose uptake in vivo.

Insulin increases glucose disposal into muscle. In addition, in vivo insulin elicits distinct nitric oxide synthase-dependent vascular responses to increase total skeletal muscle blood flow and to recruit muscle capillaries (by relaxing resistance and terminal arterioles, respectively). In the current study, we compared the temporal sequence of vascular and metabolic responses to a 30-min physiological infusion of insulin (3 mU. min(-1). kg(-1), euglycemic clamp) or saline in rat skeletal muscle in vivo. We used contrast-enhanced ultrasound to continuously quantify microvascular volume. Insulin recruited microvasculature within 5-10 min (P < 0.05), and this preceded both activation of insulin-signaling pathways and increases in glucose disposal in muscle, as well as changes in total leg blood flow. Moreover, l-NAME (N(omega)-nitro-l-arginine-methyl ester), a specific inhibitor of nitric oxide synthase, blocked this early microvascular recruitment (P < 0.05) and at least partially inhibited early increases in muscle glucose uptake (P < 0.05). We conclude that insulin rapidly recruits skeletal muscle capillaries in vivo by a nitric oxide-dependent action, and the increase in capillary recruitment may contribute to the subsequent glucose uptake.

Adipose tissue IL-6 content correlates with resistance to insulin activation of glucose uptake both in vivo and in vitro.

Obesity and type 2 diabetes are associated with insulin resistance, the mechanisms of which remain poorly understood. A significant correlation between circulating IL-6 level and insulin sensitivity has recently been found in humans. Because adipose tissue could be a significant source of IL-6, we analyzed the relationship between the levels of adipose tissue IL-6 and insulin action in vivo, during a hyperinsulinemic normoglycemic clamp, and in vitro by measuring glucose transport in adipocytes from 12 obese subjects with (n = 7) or without (n = 5) diabetes. We observed an inverse correlation between adipose tissue IL-6 content and maximal insulin-responsiveness measured in vivo (P < 0.02) and in vitro (P < 0.02). Conversely, there was no significant correlation between these two later parameters and adipose tissue leptin or tumor necrosis factor-alpha protein contents. Furthermore, we showed, for the first time, the presence of immunoreactive IL-6 receptors in the plasma membrane of human abdominal sc adipocytes. This suggests that locally secreted IL-6 could act on adipocytes by an autocrine/paracrine mechanism. In conclusion, increased IL-6 production by sc adipose cells might participate to the insulin-resistant state observed in human obesity.

In-vitro hydrolysis, permeability, and ocular uptake of prodrugs of N-[4-(benzoylamino)phenylsulfonyl]glycine, a novel aldose reductase inhibitor.

To enhance the ocular uptake of N-[4-(benzoylamino)phenylsulfonyl]glycine (BAPSG), two ester (methyl and isopropyl) prodrugs were synthesized and evaluated for their stability in various buffers (pH 1-9), hydrolysis in rabbit ocular tissues (cornea, conjunctiva, iris-ciliary body, lens, aqueous humor, and vitreous humor), transport across cornea and conjunctiva, and in-vivo uptake following topical administration. Over the pH range of 1-9, the rate constants for degradation ranged from 5.67 to 218.9 x 10(-3) h(-1) for the methyl ester and from 3.14 to 4.45 x 10(-3) h(-1) for the isopropyl ester. At all pH conditions, the isopropyl ester was more stable when compared with the methyl ester. A change in buffer concentration at pH 7.4 did not influence the stability of the prodrugs. The prodrugs were rapidly hydrolysed in the tissue homogenates, with the rate constants for hydrolysis ranging from 1.98 to 7.2x 10(-3) min(-1) for the methyl ester and 3.32 to 6.53 x 10(-3) min(-1) for the isopropyl ester. The in-vitro permeability of the methyl ester was less than the parent drug across cornea and conjunctiva. Isopropyl ester levels were not detectable in the receiver chamber even at the end of the 4-h transport study. Following topical administration of BAPSG and the two prodrugs at a dose of 60 microg/eye, the lowest levels were seen in vitreous humor for parent compound and its methyl ester. In general, the tissue uptake of methyl ester was less than BAPSG. Isopropyl ester levels were below detection limits in all the ocular tissues. Lipophilic ester prodrugs of BAPSG showed good aqueous solution stability in tissue homogenates. However, these prodrugs lacking the free carboxylate anion exhibited reduced in-vitro permeability and in-vivo uptake, suggesting the importance of free carboxylate anion in the delivery of BAPSG.

Lipoprotein receptors and atherogenic receptor-mediated mechanisms.

The liver plays a decisive role in the regulation of the plasma levels of atherogenic lipoproteins. The primary liver interaction site of chylomicron remnants and VLDL remnants (beta-VLDL) is still unidentified, whereas the subsequent cellular uptake is likely to be mediated in concert by the LDL receptor-related protein and the LDL receptor. The nature of the primary interaction site of remnants (remnant receptor) might be a liver-specific proteoglycan or a liver-specific protein. Atherogenic modified LDL can be recognized by a family of scavenger receptors. A newly identified 95 kDa protein forms the most likely candidate for mediating the in-vivo uptake of oxidized LDL from the circulation and may, therefore, protect the body against the presence of oxidized LDL in the blood compartment.

Regulation of the growth hormone receptor during liver regeneration in the rat.

The GH receptor (GHR) in regenerating rat liver following partial hepatectomy has been characterized in terms of mRNA levels, size of receptor variants, ligand binding, endocytosis and degradation. The GHR mRNA level, quantitated by solution hybridization, was found to be unchanged during the first 3 days of regeneration. The specific binding of 125I-labelled bovine GH to a GHR-enriched low density membrane fraction was, in contrast to the mRNA level, reduced to 10-35% between 12 and 24 h after partial hepatectomy, and remained low until 48 h. Removal of endogenous GH by MgCl2 treatment did not further increase the binding of 125I-labelled bovine GH. Affinity crosslinking experiments revealed reduced labelling of a specific receptor binder at 95 kDa, assuming a 1:1 stoichiometric binding of the ligand, in regenerating liver. Additional evidence for a decrease in the amount of functional GHR was provided by the finding of decreased levels of IGF-I mRNA in the liver remnant, with a nadir at 50% between 24 and 48 h after hepatectomy. GH binding was reduced to a similar extent in hypophysectomized animals subjected to partial hepatectomy, implying that pituitary GH is not involved in the mechanism of GHR down-regulation. The in-vivo uptake of 125I-labelled bovine GH, which is correlated with the number of GHRs available for binding in the plasma membrane, was decreased to 23% in an endosome-enriched membrane fraction from regenerating liver at 24 h. No qualitative differences in the size distribution of endocytosed receptor-ligand complexes in normal and regenerating liver were noted.(ABSTRACT TRUNCATED AT 250 WORDS)

Factors Which Regulate Net Hepatic Glucose Uptake In Vivo

The regulation of net hepatic glucose uptake in vivo occurs by way of the hormonal milieu (insulin and glucagon), the glucose level, and the route of glucose delivery. Hyperglycemia in the absence of changes in pancreatic hormones (i.e., increased insulin and/or decreased glucagon) does not elicit significant glucose uptake by the liver. Net hepatic glucose uptake is augmented in a dose-dependent manner by a rise in insulin and is further stimulated by the presence of a "portal signal." The presence of coordinated changes in insulin, glucagon, and the glucose level in combination with the "portal signal" ensures adequate glucose uptake by the liver in response to a meal.

In-vivo uptake of human growth hormone in male rat liver.

125I-Labelled human GH (hGH) was injected i.v. to male rats and its subcellular distribution in the hepatocyte was examined using fractionation techniques. Uptake into liver homogenates was maximal by 15 min after injection and represented 24% of the injected radioactivity; it was markedly inhibited by coinjection of native hGH. 125I-Labelled hGH taken up by the liver underwent a time-dependent translocation process. The peak of specific labelling of plasma membranes occurred at 3 min whereas later on the radioactivity was concentrated in low-density structures present in Golgi-endosome fractions. To characterize the ligand-associated structures better, endosome-enriched fractions were prepared from a microsomal fraction by isopycnic centrifugation in a sucrose gradient and a Nycodenz gradient. The radioactivity was in one peak with a median density of 1.096 g/cm3 in the Nycodenz gradient fractions. The peak of radioactivity was distinct from that of galactosyltransferase activity which appeared at a median density of 1.114 g/cm3. The labelled material eluted from the various subcellular fractions appeared as intact hGH. Upon in-vivo interaction with male rat hepatocytes, 125I-Labelled hGH was internalized with a sequential association with plasma membranes and endocytic structures distinct from Golgi elements.

Spiramycin uptake by alveolar macrophages.

The in-vitro and in-vivo uptake of spiramycin by human and animal alveolar macrophages was studied. In-vitro penetration was studied in guinea pig and human alveolar macrophages incubated in medium 199 at 37 degrees C containing spiramycin at various concentrations. Results were expressed as the cellular/extracellular concentration ratio (C/E). The in-vivo study was performed in patients receiving 500 or 1000 mg spiramycin every 8 h as a 1-h infusion on day 1. A single infusion was given on day 2, 2 h before serum and bronchoalveolar lavage (BAL) sampling. Spiramycin was assayed by HPLC, and by a microbiological assay. In guinea pig alveolar macrophages, the C/E ratio of spiramycin after 60 min at 37 degrees C was 20.3 +/- 6.5 when the concentration was 10 mg/l. In human alveolar macrophages, the C/E ratio was 21.3 +/- 8.7 at 5 mg/l spiramycin and 23.8 +/- 8.7 at 50 mg/l. The accumulated spiramycin was slowly released when the cells (guinea pig alveolar macrophages) were washed and re-incubated in antibiotic free medium. Spiramycin was able to penetrate the alveolar space. In BAL supernatant, spiramycin levels were about 24-fold the serum level (n = 6 patients), when the BAL/serum glucose ratios were used as the dilution estimate. Alveolar macrophage levels ranged from 17 to 210 mg/l (n = 6 patients receiving 500 mg spiramycin infusion). These results are consistent with the in-vitro data.

The Accumulation Mechanism of Cationic Mitomycin C-dextran Conjugates in the Liver: In-vivo Cellular Localization and In-vitro Interaction with Hepatocytes

To elucidate the mechanism of the accumulation of mitomycin C-dextran conjugate (MMC-D) in the liver, in-vivo cellular uptake and in-vitro cellular interaction of MMC-D have been studied. Localization of cationic and anionic MMC-D (MMC-Dcat. and MMC-Dan.) in different liver cell types following i.v. administration was examined in rats and the significant contribution of parenchymal cells demonstrated. In-vitro cellular interaction was determined by measuring the drug concentration in the medium after incubation with rat isolated hepatocytes. MMC-Dcat. was highly adsorbed on the surface of hepatocytes at pH 7.2, while the interaction between MMC-Dan. and hepatocytes was negligible. The percentage association of MMC-Dcat. with hepatocytes remained almost constant during the course of incubation and no significant difference was observed between the incubation at 4 and 37 degrees C. The adsorption phenomenon was shown to conform to Langmuir's adsorption isotherm. The amount of MMC-Dcat. associated with hepatocytes increased as the molecular weight of the dextran chain increased. These results showed that MMC-Dcat. was adsorbed on the surface of hepatocytes by an electrostatic force and this binding was responsible for its remarkable accumulation in the liver in-vivo. Thus some physicochemical properties of the MMC-D conjugates are thought to play an important role in the disposition characteristics of the conjugates.

Josamycin pulmonary penetration determined by broncho-alveolar lavage in man.

Macrolide antibiotics are concentrated by phagocytic cells in vitro. We studied the in-vivo uptake of josamycin by alveolar macrophages recovered by broncho alveolar lavage performed in patients 3 h after ingestion of 1 g of the drug. Simultaneous determination of the antibiotic levels was performed on the BAL supernatant and the serum and the results compared to those obtained by incubating alveolar macrophages and blood polymorphonuclears at 2 and 8 mg/l. Josamycin concentration was measured using a high pressure liquid chromatography method. Results show that intracellular josamycin levels in vivo are similar to those observed in vitro. Accumulation of the drug also occurs in BAL fluid (reaching about a 100-fold the serum concentration). Pulmonary and serum levels are significantly correlated.

Ontogeny of the androgen receptor in rat ventral prostate during sexual development.

Concentrations of cytosolic androgen receptor, DNA and soluble protein, contents of DHT, and in-vivo uptake of 3H-DHT were measured in rat ventral prostates at 5-day intervals during sexual development. Regarding prostate weight two phases of growth were noted being separated by a period of stagnation from Day 40 to 45. Cytosolic androgen receptor, particle-bound DHT, and uptake of 3H-DHT into the 100,000-g sediment showed a clear pattern: a maximum in the prepubertal animal at age Day 20, a minimum at age Day 30 (4 days after the early pubertal rise of LH, testosterone, and DHT) followed by a second maximum on Day 55 (2 days before the beginning of fertility), and a second minimum in the young mature animal on Day 70. An intermediate peak seen at age Day 37 was not significant. Neither the time-dependent profile of the cytosolic androgen receptor nor the contents and in vivo uptake of DHT were correlated to concentrations of circulating gonadotrophins, growth hormone, and sex-steroids measured during puberty in the same strain of animals. Therefore, the regulating mechanism remains unclear.

Recombinant human insulin-like growth factor: testing the somatomedin hypothesis in hypophysectomized rats

The in-vivo biological activity of recombinant methionyl insulin-like growth factor I (met-IGF-I) was demonstrated in hypophysectomized rats by following blood glucose after an i.v. bolus injection of met-IGF-I; a dose-dependent decrease in blood sugar was seen. Membrane transport was studied using the non-metabolizable amino acid alpha-aminoisobutyric acid; stimulation was obtained with the highest dose used (90 micrograms/rat). To test the original somatomedin hypothesis, growth studies were performed in hypophysectomized rats. Two or three doses of met-IGF-I were given with three different administration regimes (i.v. or s.c. infusion, or s.c. injections twice daily) for 6 or 8 days. Little growth-promoting activity was observed, with a significant effect on body weight gain obtained only when met-IGF-I was given continuously at the highest dose used (180 micrograms/day). No effect was seen on the in-vivo uptake of radioactive sulphate into cartilage. Epiphyseal cartilage width increased slightly at the highest dose of met-IGF-I, but only when the hormone was given by infusion. When 180 micrograms met-IGF-I/day were given by injections, a significant effect on longitudinal bone growth was obtained (90 micron above control). The levels of IGF in the serum were not measurably increased after s.c. administration of met-IGF-I, whereas after i.v. infusion, significantly raised levels were obtained at the higher dose rates (3.0 +/- 0.3 and 2.8 +/- 0.1 units/ml). Growth hormone was much more effective than met-IGF-I even at 50-fold lower doses. Priming the animals with 10 mu. bovine GH/day followed by combined infusions of GH and met-IGF-I did not reveal any potentiating effects of met-IGF-I in the presence of GH. We conclude that met-IGF-I is a relatively poor growth-promoting agent when given systemically, and that somatomedins are more likely to act as local growth factors rather than as circulating mediators of the growth-promoting effects of GH.

Lysosomes in skeletal muscle following denervation. Time course of horseradish peroxidase uptake and increase of lysosomal enzymes.

The in-vivo uptake of exogenously applied horseradish peroxidase and the activities of the lysosomal enzymes acid phosphatase and cathepsin D were studied histochemically and/or biochemically in innervated and 2-14 day-denervated tibialis anterior muscles of the mouse. The biochemically determined uptake of horseradish peroxidase showed a large increase already 4 days after denervation. The activities of the lysosomal enzymes increased in a more gradual fashion, and only cathepsin D showed an increase in activity when expressed as total activity per muscle. Histochemically horseradish peroxidase was found to be localized in muscle fibres in characteristic spindle-shaped segments after denervation. The main increase in the number of such segments per transverse section of the muscle occurred between 3 and 6 days after denervation. In serial sections these segments frequently showed positive staining also for acid phosphatase. It is concluded that exogenously applied horseradish peroxidase is taken up into the lysosomal system, which after denervation becomes organized into characteristic spindle-shaped segments in the muscle fibres. The endocytic activity of muscle fibres increases early after denervation. This is followed by a more gradual increase in activity of lysosomal enzymes and finally by an organization of the lysosomal system into characteristic spindle-shaped segments. The results are compatible with the working hypothesis that increased endocytosis may initiate lysosomal activation in denervated skeletal muscle.

In-vivo uptake and metabolism of testosterone by the skin and other tissues of the rat.

The uptake and metabolism of radioactively labelled testosterone was investigated in vivo at various time intervals in the skin, plasma and other tissues of adult rats castrated 24 h earlier. After the addition of marker steroids the components of the tissue extracts were separated by partition and development in several chromatographic systems; steroid identification was confirmed by derivative formation and recrystallization to constant 3H:14C ratio. The skin resembled the classical androgen target organs in the uptake and retention of both total and unconjugated radioactivity, but the actual composition of the radioactivity present in the skin was markedly different from that in the accessory glands. Little testosterone was recovered from the ventral prostates and 5α-dihydrotestosterone (5α-DHT), often considered the active intracellular androgenic steroid, constantly occupied the largest proportion of the extracted radioactivity. In the skin samples, however, testosterone was always the major unconjugated steroid detected and 4-androstene-3,17-dione, 5α-DHT, 5α-androstane-3α,17β-diol, androsterone and 5α-androstane-3β,17β-diol were only identified in much smaller quantities, even after 5 h. Although the majority of the plasma radioactivity was conjugated or very polar by 20 min, all the steroids found in the skin were present, albeit in different proportions; testosterone was always predominant and 5α-androstane-3β,17β-diol was also detected in larger amounts than the very low levels of the other steroids. The results suggest that 5α-DHT is not as important in the skin as it is in the prostate of the rat and that other androgenic steroids, most notably testosterone itself, may be involved in the mechanism of androgen action in the skin.

Effects of growth hormone, prolactin and thyroxine on body wegith, somatomedin-like activity and in-vivo sulphation of cartilage in hypopituitary dwarf mice.

Hypopituitary dwarf mice were found to have reduced levels of serum somatomedin-like activity compared with normal mice of the Snell strain. Treatment with bovine growth hormone for 3 and 7 days resulted in growth without significantly increased levels of serum somatomedin-like activity, as detected by in-vitro uptake of 35SO4(2-) into normal rat cartilage; only after treatment for 14 days was somatomedin activity significantly raised. However, treatment for 2 days with bovine growth hormone, bovine prolactin or thyroxine resulted in a dose-dependent increase in in-vivo uptake of 35SO4(2-) into dwarf mouse costal cartilage; growth hormone and thyroxine did not act synergistically. Ten days of treatment with growth hormone promoted a dose-dependent increase in both growth (increased weight gain and tail length) and in-vivo of 35SO4(2-). Increase in tail length was correlated with uptake of 35S04(2-). Thus, in-vivo uptake of 35SO4(2-) into dwarf mouse costal cartilage provides a sensitive method for detecting a dose-related effect of growth hormone.

In-vivo uptake of topically applied fluoride by human dental enamel

In-vivo F determinations were carried out using a non-destructive physical technique based on prompt activation by proton bombardment. 74 sound incisors were submitted to 4 min topical applications of commercial gels and solutions containing fluoride. The F uptake was measured for many hours, days and weeks following fluoride application. The largest amounts of fluoride were acquired by the enamel 30 min after an amine fluoride solution application (4460 parts/10 6). Two acidulated phosphate fluoride (APF) gels gave a mean enrichment of 1774 and 3277 parts/10 6 respectively, whereas amine fluoride (AmF) and monofluorophosphate gels were less efficient (228 and 105 parts/10 6 of enrichment respectively). The superiority of AmF solution treatment was observed for many weeks after fluoride application; 6 weeks were needed to leach all the fluoride acquired by the enamel whereas with APF gels, the initial F level was recovered 2 weeks after the topical application.

Mechanisms involved in the regulation of steroid receptor levels

The turnover of steroid receptors comprises: synthesis in the cytoplasm, translocation into the cell nucleus and degradation at a still unknown site. From studies on the oestradiol/receptor system, the following conclusions can be drawn. 1. The in-vivo uptake of oestradiol by target cell nuclei is receptordependent. Steroid and receptor are translocated from the cytoplasm in a 1:1 ratio. A recycling of receptor is undetectable after pulse administration of oestradiol. Receptor replenishment in the cytoplasm is accomplished by synthesis. 2. The nuclear uptake of receptor-in contrast-proceeds also in the absence of oestradiol. Both forms of receptor, monomer and “activated” dimer are present in oestrogen-free nuclei. 3. Oestradiol enhances the “nucleotropy” and the turnover rate of receptor. 4. Oestrogenicity and antioestrogenicity are apparently linked to effects exerted on the “nucleotropy” of receptor and its ability to interact with the relevant nuclear structures.

Response of human enamel to topical application of ammonium fluoride

In-vitro uptake of F by intact enamel from 0.62 M solutions of NH 4F was greater than that from NaF at low pH, but similar at nearly neutral or alkaline pH. Fluoride contents in enamel biopsies taken 2 weeks after a 3 min topical treatment with the acid solutions also were higher for the NH 4F treatment. The in-vitro and in-vivo uptake from both solutions was enhanced by pretreating the enamel for one minute with 0.05 M phosphoric acid. Examination by the electron microscope of the F-exposed enamel before and after washing revealed the deposit formed from NH 4F was retained more tenaciously than that formed from NaF. X-ray diffraction patterns showed formation of CaF 2 when ground enamel was treated with either of the two acid fluoride solutions.

THE IN VIVO UPTAKE AND METABOLISM OF TRITIATED PROGESTERONE BY THE VAGINA OF THE OVARIECTOMIZED MOUSE

Journal Article THE IN VIVO UPTAKE AND METABOLISM OF TRITIATED PROGESTERONE BY THE VAGINA OF THE OVARIECTOMIZED MOUSE Get access Janet Stokes, Janet Stokes Imperial Cancer Research Fund, Lincoln's Inn Fields, London W.C.2. England Search for other works by this author on: Oxford Academic Google Scholar Len Martin Len Martin Imperial Cancer Research Fund, Lincoln's Inn Fields, London W.C.2. England Search for other works by this author on: Oxford Academic Google Scholar Acta Endocrinologica (Norway), Volume 56, Issue 1_Supplement, Dec 1967, Page S147, https://doi.org/10.1530/acta.0.056S147 Published: 01 December 1967

Effect of Chronic Lead Intoxication on in-vivo I131 Uptake by the Rat Thyroid

SummaryIn vivo depression of I131 uptake by the rat thyroid, and the conversion ratio has been produced by chronic lead intoxication. Female rats are more susceptible than male rats.The authors would like to thank Dr. William J. Darby for his encouragement and suggestions.The opinions are those of the authors and do not necessarily reflect the opinion of the U.S. Navy.