scholarly journals The Release of a Soluble Glycosylated Protein from Glycogen by Recombinant Lysosomal α-Glucosidase (rhGAA) In Vitro and Its Presence in Serum In Vivo

Biomolecules ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 1613
Author(s):  
Allen K. Murray
Keyword(s):  
Pompe Disease ◽  
Incubation Medium ◽  
Core Material ◽  
Protein Core ◽  
In Vitro Incubation

In studies on the degradation of glycogen by rhGAA, a glycosylated protein core material was found which consists of about 5–6% of the total starting glycogen. There was an additional 25% of the glycogen unaccounted for based on glucose released. After incubation of glycogen with rhGAA until no more glucose was released, no other carbohydrate was detected on HPAEC-PAD. Several oligosaccharides are then detectable if the medium is first boiled in 0.1 N HCl or incubated with trypsin. It is present in serum either in an HCl extract or in a trypsin digest. The characteristics of the in vivo serum material are identical to the material in the in vitro incubation medium. One oligosaccharide cannot be further degraded by rhGAA, from the incubation medium as well as from serum co-elute on HPAEC-PAD. Several masked oligosaccharides in serum contain m-inositol, e-inositol, and sorbitol as the major carbohydrates. The presence of this glycosylated protein in serum is a fraction of glycogen that is degraded outside the lysosome and the cell. The glycosylated protein in the serum is not present in the serum of Pompe mice not on ERT, but it is present in the serum of Pompe disease patients who are on ERT, so it is a biomarker of GAA degradation of lysosomal glycogen.

IN VITRO UPTAKE OF TRITIATED OESTRADIOL BY THE ANTERIOR HYPOTHALAMUS AND HYPOPHYSIS OF THE RAT

1970 ◽  
Vol 64 (4) ◽  
pp. 687-695 ◽  
Author(s):  
Junzo Kato
Keyword(s):  
Incubation Medium ◽  
Posterior Hypothalamus ◽  
Anterior Hypothalamus ◽  
Ovariectomized Rats ◽  
Free Medium ◽  
Cortex Cerebri ◽  
Anterior Hypophysis ◽  
In Vitro Uptake

ABSTRACT The anterior, middle, and posterior hypothalamus, the cortex cerebri, the anterior hypophysis as well as the diaphragm of adult ovariectomized rats were incubated in vitro with tritiated 17β-oestradiol. The uptake of tritiated oestradiol was differentially distributed intracerebrally with higher accumulation in the anterior hypothalamus and the hypophysis. Lowering the temperature of the incubation medium caused a reduction in the uptake of radioactivity by the anterior hypothalamus as compared to that found in other brain tissues. Tritiated oestradiol taken up in vitro by the anterior hypothalamus and the hypophysis tended to be retained after further incubation in a steroid-free medium. The addition of non-radioactive 17β-oestradiol to the medium inhibited the uptake of tritiated oestradiol by these tissues. Moreover, pretreatment with non-radioactive 17β-oestradiol in vivo prevented the preferential accumulation of tritiated oestradiol in vitro in the anterior hypothalamus and the hypophysis. These results indicate that oestradiol is preferentially taken up in vitro by the anterior hypothalamus and the hypophysis of the rat.

Synthesis of taurocholate by rat fetal liver in organ culture: effects of cortisol in vitro.

1979 ◽  
Vol 237 (2) ◽  
pp. E177 ◽  
Author(s):  
T O Graham ◽  
D H Van Thiel ◽  
J M Little ◽  
R Lester
Keyword(s):  
Organ Culture ◽  
Dose Response ◽  
Incubation Medium ◽  
Response Pattern ◽  
Fetal Liver ◽  
Dry Weight ◽  
Organ Cultures ◽  
In Vitro Incubation ◽  
Culture Effects

Taurocholate production by fetal hepatic organ cultures was measured by radioimmunoassay. Taurocholate production was maximal on day 1 of in vitro incubation, but was demonstrable in organ cultures maintained for periods up to 15 days. Explants obtained from fetuses of 18 gestational days of age produced only 82 pmol taurocholate per milligram dry weight of tissue during the first 24 h of incubation. Explants obtained from fetuses 21 gestational days of age produced 1,043 pmol taurocholate per milligram dry weight. The presence of cortisol (2.0 X 10(-6) M) in the incubation medium increased synthesis of taurocholate by rat fetal liver in which total taurocholate rose 50-fold above control after 120 h of incubation. In increasing concentrations from 2.0 X 10(-9) M to 2.0 X 10(-7) M, cortisol produced an incremental rise in taurocholate. However, additional increases in cortisol dose failed to provide further stimulation, and taurocholate production was inhibited by cortisol concentrations of 2.0 X 10(-5) M. The results provide further validation for the technique of fetal hepatic organ culture. They demonstrate that taurocholate synthesis is increasing rapidly during the final stages of gestation and show that cortisol augments taurocholate synthesis in a dose-response pattern.

EFFECTS OF HYDROXYSTEROID DEHYDROGENASE INHIBITORS ON IN-VITRO AND IN-VIVO STEROIDOGENESIS IN THE OVINE ADRENAL GLAND

1982 ◽  
Vol 92 (2) ◽  
pp. 205-212 ◽  
Author(s):  
P. SINGH-ASA ◽  
G. JENKIN ◽  
G. D. THORBURN
Keyword(s):  
Adrenal Gland ◽  
Incubation Medium ◽  
Bovine Serum ◽  
Hydroxysteroid Dehydrogenase ◽  
Inhibitory Effect ◽  
Bovine Serum Albu ◽  
Stimulatory Action ◽  
Adrenal Steroidogenesis

The effectiveness of trilostane and azastene as inhibitors of adrenal steroidogenesis was compared by in-vitro and in-vivo methods. A radioimmunoassay was developed for the measurement of cortisol in ovine plasma, incubation medium and tissue extract using a specific antiserum raised against cortisol 21-acetate,3-carboxymethyloxime : bovine serum albu Trilostane (20 μmol/l) decreased cortisol synthesis and release both in unstimulated and in ACTH-stimulated adrenal tissues in vitro. The same concentration of azastene had a lesser effect on unstimulated adrenals and was completely ineffective in blocking the stimulatory action of ACTH. In vivo, trilostane suppressed adrenal steroidogenesis in pregnant and cyclic ewes but the suppression in pregnant ewes was over a longer period, and after lower doses. It is concluded that trilostane had an inhibitory effect on ovine adrenal steroidogenesis both in vitro and in vivo.

THE STIMULATION OF INSULIN RELEASE BY ESSENTIAL AMINO ACIDS FROM RABBIT PANCREAS IN VITRO

1970 ◽  
Vol 47 (3) ◽  
pp. 347-356 ◽  
Author(s):  
R. D. G. MILNER
Keyword(s):  
Amino Acids ◽  
Insulin Release ◽  
Incubation Medium ◽  
Statistical Significance ◽  
Adenosine Monophosphate ◽  
Essential Amino Acids ◽  
Isoleucine Leucine ◽  
Rabbit Pancreas

SUMMARY Pieces of rabbit pancreas were incubated in vitro in an incubation medium containing no glucose or 1·5 mg. glucose/ml. In each of these conditions the effect on insulin release of each of the essential amino acids at 5 mm concentration was studied. Leucine was the only essential amino acid that stimulated insulin release to a level which reached statistical significance in an incubation medium containing no glucose. In medium containing 1·5 mg. glucose/ml., arginine, isoleucine, leucine and lysine stimulated insulin release and phenylalanine inhibited insulin release. Glucagon, theophylline or dibutyryl cyclic adenosine monophosphate stimulated insulin release significantly in the presence of leucine but not in the presence of arginine. Arginine stimulated insulin release in the presence of leucine. The results of these experiments characterize further the difference in the mechanism of action of leucine and arginine on the pancreatic β-cell and indicate possible explanations for results obtained in other species in vivo.

Inhibition of matrix metalloproteinase activity by ACE inhibitors prevents left ventricular remodeling in a rat model of heart failure

2007 ◽  
Vol 292 (6) ◽  
pp. H3057-H3064 ◽  
Author(s):  
Gregory L. Brower ◽  
Scott P. Levick ◽  
Joseph S. Janicki
Keyword(s):  
Heart Failure ◽  
Matrix Metalloproteinase ◽  
Ace Inhibitors ◽  
Left Ventricular ◽  
Av Fistula ◽  
Functional Changes ◽  
In Vitro Incubation ◽  
In Vivo Treatment

Angiotensin-converting enzyme (ACE) inhibitors represent the front-line pharmacological treatment of heart failure, which is characterized by left ventricular (LV) dilatation and inappropriate hypertrophy. The mechanism of action of ACE inhibitors is still unclear, but evidence suggests that they may act by influencing matrix metalloproteinase (MMP) activity. This study sought to determine whether ACE inhibitors can directly regulate MMP activity and whether this results in positive structural and functional adaptations to the heart. To this end, MMP-2 activity in LV tissue extracted from rats with an aortocaval (AV) fistula was assessed by in vitro incubation as well as in vivo treatment with captopril, lisinopril, or quinapril. Furthermore, LV size and function were determined in untreated AV fistula rats, AV fistula rats treated with lisinopril (3, 5, and 8 wk), and age-matched sham-operated controls. In vitro incubation with captopril, lisinopril, or quinapril significantly reduced MMP-2 activity, as did in vivo treatment. This occurred without a reduction in the available pool of MMP-2 protein. Long-term in vivo administration of lisinopril also prevented LV dilatation, attenuated myocardial hypertrophy, and prevented changes in myocardial compliance and contractility. The results herein demonstrate that ACE inhibitors prevent MMP-2 activity and, in so doing, represent a mechanism responsible for preventing the negative structural and functional changes that occur in the rat AV fistula model of heart failure.

Early Dissociation of Protein Synthesis and Amylase Secretion Following Hormonal Stimulation of the Pancreas

1974 ◽  
Vol 52 (2) ◽  
pp. 198-205 ◽  
Author(s):  
R. Mongeau ◽  
Y. Couture ◽  
J. Dunnigan ◽  
J. Morisset
Keyword(s):  
Protein Synthesis ◽  
Incubation Medium ◽  
Single Injection ◽  
Pancreatic Enzyme ◽  
Amylase Secretion ◽  
Hormonal Stimulation ◽  
Pancreatic Enzyme Secretion ◽  
Metabolic Conditions

The secretion of the various pancreatic enzymes can be increased by hormonal and cholinergic stimulation. However, it is not yet clear among the different investigators if their synthesis remains constant or can be modified according to different metabolic conditions. The secretion and synthesis of the pancreatic proteins were then studied in parallel to evaluate if secretion triggers synthesis or both phenomenons are controlled by separate mechanisms.The approach for these studies consists mainly in a combination of in vivo and in vitro experiments. The stimulants were injected in vivo and the pancreatic secretions were collected for different periods of time. The animals were then sacrificed and protein synthesis was measured in vitro along with the amylase secreted into the incubation medium. The results show that protein synthesis is decreased during the first 15 min after a single injection or infusion of both cholecystokinin–pancreozymin (CCK–PZ) and secretin. This reduction was associated with an increase in amylase secreted into the incubation medium. However, at 30 min after the hormonal stimulation, protein synthesis is increased while secretion into the incubation medium had returned to control levels. This increase in protein synthesis lasts for at least 1 h. These results strongly suggest that pancreatic enzyme secretion and synthesis are dissociated in the early minutes following hormonal stimulation.

ÉTUDE DE L'ACTIVITÉ BIOLOGIQUE IN VITRO DES HORMONES GONADOTROPES

1963 ◽  
Vol 42 (4) ◽  
pp. 509-513 ◽  
Author(s):  
D. Gospodarowicz ◽  
J. Legault-Démare
Keyword(s):  
Incubation Medium ◽  
Cholesterol Synthesis ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Corpora Lutea ◽  
Lactogenic Hormone ◽  
Normal Rat ◽  
Stimulation Of

ABSTRACT Human chorionic gonadotrophin (HCG) and lactogenic hormone (LTH or prolactin) were found practically inactive on the incorporation of 14Cacetate into cholesterol of normal rat corpus luteum in vitro. On the contrary, when added simultaneously to the incubation medium, they increased by 90% the labeling of cholesterol. When pseudopregnancy corpora lutea were used, HCG alone stimulated to the same amount, but no stimulation was observed with LTH alone. These results show that the stimulation of cholesterol synthesis is produced by a synergic action of LTH and HCG, LTH being introduced either in vivo (pseudopregnancy) or in vitro.

CORTISONINAKTIVIERUNG IN DER RATTENLEBER NACH INJEKTION VON BUTAZOLIDIN

1960 ◽  
Vol XXXIV (I) ◽  
pp. 77-83 ◽  
Author(s):  
Ludger Lutzmann ◽  
Wilhelm Dirscherl
Keyword(s):  
Incubation Medium ◽  
Liver Slices ◽  
Distribution Studies

ABSTRACT The breakdown of cortisone by the liver of Butazolidin treated rats as well as of untreated controls was examined. At the same time Butazolidin was determined in serum and liver at various times after its injection together with its distribution between incubation media and liver slices. The results are discussed in connection with former experiments in vitro in which Butazolidin in therapeutical doses was added to the incubation medium. From the distribution studies it appears that, in vivo as well as in vitro, the breakdown of cortisone is affected only at a certain Butazolidin concentration.

EFFECTS OF GONADOTROPHINS AND CYCLIC 3′,5′-AMP ON IN VITRO INCORPORATION OF [3H]URIDINE INTO RNA OF THE PREPUBERTAL RAT OVARY

1973 ◽  
Vol 72 (4) ◽  
pp. 771-785 ◽  
Author(s):  
Jan Jarlstedt ◽  
Lars Nilsson ◽  
Lars Hamberger ◽  
Kurt Ahrén
Keyword(s):  
Cyclic Nucleotide ◽  
Gel Electrophoresis ◽  
Incubation Medium ◽  
Polyacrylamide Gel Electrophoresis ◽  
Labelling Pattern ◽  
Total Rna ◽  
Rat Ovary ◽  
Prepubertal Rats

ABSTRACT In vivo and in vitro effects of FSH and LH on in vitro incorporation of [3H]uridine into RNA of the prepubertal rat ovary have been studied. RNA was fractionated with composite agarose-polyacrylamide gel electrophoresis. When FSH was injected into the prepubertal rats 4 h before incubation of the ovaries, the incorporation of labelled uridine into total RNA was decreased showing relatively more radioactivity concentrated to the RNA fractions lighter than 28S as compared to the controls. These effects were not seen when FSH was added to the incubation medium in vitro. When LH was added in vitro to the isolated ovaries a higher percentage of incorporated radioactivity was concentrated in the RNA fractions heavier than 28S without any change in the incorporation of [3H]uridine into total RNA. LH administered in vivo 30 min before incubation of the ovaries gave the same change in the labelling pattern between the RNA fractions as LH in vitro but in addition showed a decreased incorporation of radioactivity into total RNA. The in vitro effects of cyclic 3′,5′-AMP were also studied. When prepubertal rat ovaries were incubated in 10 mmol/l of this cyclic nucleotide, the incorporation of [3H] uridine into total RNA was decreased without any change in the labelling pattern.

Sign in / Sign up

Export Citation Format

Share Document