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Molecular Vision 2009Inflammation plays an important role in dry eye syndrome. In this study, inflammatory cytokine expression on the ocular surface in the Botulium toxin B (BTX-B) induced...
PURPOSE
Inflammation plays an important role in dry eye syndrome. In this study, inflammatory cytokine expression on the ocular surface in the Botulium toxin B (BTX-B) induced mouse dry eye model was investigated.
METHODS
CBA/J mice received an injection of saline or 20 milliunits (mU) of BTX-B into the lacrimal gland. Tear production and corneal fluorescein staining were evaluated in all groups before injection and at 3 time points after. The pro-inflammatory cytokines macrophage inhibitory factor (MIF), interleukin-1beta (IL-1 beta), tumor necrosis factor-alpha (TNF- alpha) and interleukin-6 (IL-6) in conjunctival and corneal epithelium were evaluated by real time quantitative PCR and immunohistochemistry.
RESULTS
BTX-B injected mice showed significantly decreased aqueous tear production and increased corneal fluorescein staining at the 1 week and 2 week time points compared with normal control and saline-injected mice. The BTX-B injected mice mRNA expression levels of TNF-alpha and IL-1beta from conjunctival and corneal epithelial cells increased significantly at two early time points comparing with that of normal and saline injected mice, but IL-1beta returned to normal levels at the 4 week time point. Saline injected mice showed no difference in mRNA expression of TNF-alpha, IL-1beta, MIF, and IL-6 on the ocular surface tissue at all time points. Immunohistochemistry confirmed these findings.
CONCLUSIONS
BTX-B induced mouse model showed decreased aqueous tear production, increased corneal fluorescein staining, and TNF-alpha and IL-1beta increased expression on the ocular surface within one month. The patterns seen appeared to mimic those in humans with non-Sjögren's syndrome keratoconjunctivitis sicca (NS-KCS).
Topics: Animals; Botulinum Toxins; Conjunctiva; Cornea; Cytokines; Disease Models, Animal; Dry Eye Syndromes; Female; Fluorescein; Fluorescent Antibody Technique; Gene Expression; Lacrimal Apparatus; Mice; Mice, Inbred CBA; Tears
PubMed: 19190733
DOI: No ID Found -
Frontiers in Neurology 2022This study aimed to construct an animal model of intracranial arterial dolichoectasia (IADE) applying the modified modeling protocol.
BACKGROUND AND PURPOSE
This study aimed to construct an animal model of intracranial arterial dolichoectasia (IADE) applying the modified modeling protocol.
MATERIALS AND METHODS
Twenty five milliunits elastase and inactivated elastase were, respectively, injected into the cerebellomedullary cistern of 60 C57/BL6 mice which were divided into experimental group (EG, = 30) and control group (CG, = 30) by using a computer-based random order generator. The modified modeling protocol clarified these aspects including brain three-dimensional parameters of mouse head fixation, angle of head inclination, fixed position of taper ear, needle holding technique, needle entry depth, prevention of liquid drug back flow, and storage conditions of elastase. And it was observed for the following parts such as mortality, inflammatory factors, craniocerebral arteries scanning, vascular tortuosity index, artery diameter, pathology of the cerebrovascular.
RESULTS
Within differently surveyed stage, the total mortality of mice in EG was 20%. ELISA illustrated that the levels of matrix metalloproteinase-9 (MMP-9) and tumor necrosis factor α (TNF-α) in peripheral blood were increased significantly after modeling. Angiography indicated that 100% of IADE in EG were observed and the diameter and tortuosity index of the basilar artery were significantly increased ( < 0.01). EVG histological processing and staining showed the disrupted internal elastic lamina, the atrophied muscle layer, and the hyalinized connective tissue of the basilar artery with the vascular wall tunica media in EG. Micro-computed tomography reported that the craniocerebral arteries of the mice in EG were outstandingly elongated, tortuous, and dilated.
CONCLUSION
The modified modeling protocol can reduce the mortality, improve the success rate, and provide a stable animal model for IADE.
PubMed: 35518204
DOI: 10.3389/fneur.2022.860541 -
BMC Pregnancy and Childbirth Mar 2023The reproductive hormone oxytocin facilitates labour, birth and postpartum adaptations for women and newborns. Synthetic oxytocin is commonly given to induce or augment...
Maternal and newborn plasma oxytocin levels in response to maternal synthetic oxytocin administration during labour, birth and postpartum - a systematic review with implications for the function of the oxytocinergic system.
BACKGROUND
The reproductive hormone oxytocin facilitates labour, birth and postpartum adaptations for women and newborns. Synthetic oxytocin is commonly given to induce or augment labour and to decrease postpartum bleeding.
AIM
To systematically review studies measuring plasma oxytocin levels in women and newborns following maternal administration of synthetic oxytocin during labour, birth and/or postpartum and to consider possible impacts on endogenous oxytocin and related systems.
METHODS
Systematic searches of PubMed, CINAHL, PsycInfo and Scopus databases followed PRISMA guidelines, including all peer-reviewed studies in languages understood by the authors. Thirty-five publications met inclusion criteria, including 1373 women and 148 newborns. Studies varied substantially in design and methodology, so classical meta-analysis was not possible. Therefore, results were categorized, analysed and summarised in text and tables.
RESULTS
Infusions of synthetic oxytocin increased maternal plasma oxytocin levels dose-dependently; doubling the infusion rate approximately doubled oxytocin levels. Infusions below 10 milliunits per minute (mU/min) did not raise maternal oxytocin above the range observed in physiological labour. At high intrapartum infusion rates (up to 32 mU/min) maternal plasma oxytocin reached 2-3 times physiological levels. Postpartum synthetic oxytocin regimens used comparatively higher doses with shorter duration compared to labour, giving greater but transient maternal oxytocin elevations. Total postpartum dose was comparable to total intrapartum dose following vaginal birth, but post-caesarean dosages were higher. Newborn oxytocin levels were higher in the umbilical artery vs. umbilical vein, and both were higher than maternal plasma levels, implying substantial fetal oxytocin production in labour. Newborn oxytocin levels were not further elevated following maternal intrapartum synthetic oxytocin, suggesting that synthetic oxytocin at clinical doses does not cross from mother to fetus.
CONCLUSIONS
Synthetic oxytocin infusion during labour increased maternal plasma oxytocin levels 2-3-fold at the highest doses and was not associated with neonatal plasma oxytocin elevations. Therefore, direct effects from synthetic oxytocin transfer to maternal brain or fetus are unlikely. However, infusions of synthetic oxytocin in labour change uterine contraction patterns. This may influence uterine blood flow and maternal autonomic nervous system activity, potentially harming the fetus and increasing maternal pain and stress.
Topics: Infant, Newborn; Pregnancy; Female; Humans; Oxytocin; Parturition; Postpartum Period; Labor, Obstetric; Postpartum Hemorrhage
PubMed: 36864410
DOI: 10.1186/s12884-022-05221-w -
Obstetrics and Gynecology Aug 2011To examine the effects and safety of high-dose (compared with low-dose) oxytocin regimen for labor augmentation on perinatal outcomes.
OBJECTIVE
To examine the effects and safety of high-dose (compared with low-dose) oxytocin regimen for labor augmentation on perinatal outcomes.
METHODS
Data from the Consortium on Safe Labor were used. A total of 15,054 women from six hospitals were eligible for the analysis. Women were grouped based on their oxytocin starting dose and incremental dosing of 1, 2, and 4 milliunits/min. Duration of labor and a number of maternal and neonatal outcomes were compared among these three groups stratified by parity. Multivariable logistic regression and generalized linear mixed model were used to adjust for potential confounders.
RESULTS
Oxytocin regimen did not affect the rate of cesarean delivery or other perinatal outcomes. Compared with 1 milliunit/min, the regimens starting with 2 milliunits/min and 4 milliunits/min reduced the duration of first stage by 0.8 hours (95% confidence interval 0.5-1.1) and 1.3 hours (1.0-1.7), respectively, in nulliparous women. No effect was observed on the second stage of labor. Similar patterns were observed in multiparous women. High-dose regimen was associated with a reduced risk of meconium stain, chorioamnionitis, and newborn fever in multiparous women.
CONCLUSION
High-dose oxytocin regimen (starting dose at 4 milliunits/min and increment of 4 millliunits/min) is associated with a shorter duration of first-stage of labor for all parities without increasing the cesarean delivery rate or adversely affecting perinatal outcomes.
LEVEL OF EVIDENCE
II.
Topics: Adult; Female; Humans; Infant, Newborn; Infant, Newborn, Diseases; Labor Onset; Labor, Induced; Obstetric Labor Complications; Oxytocics; Oxytocin; Parity; Pregnancy; Pregnancy Outcome; Young Adult
PubMed: 21775839
DOI: 10.1097/AOG.0b013e3182220192 -
The Journal of Biological Chemistry Apr 1997The effects of increased plasma free fatty acids (FFA) on insulin-dependent whole body glucose disposal, skeletal muscle glycolysis, glycogen synthesis, pyruvate versus...
The effects of increased plasma free fatty acids (FFA) on insulin-dependent whole body glucose disposal, skeletal muscle glycolysis, glycogen synthesis, pyruvate versus FFA/ketone oxidation, and glucose 6-phosphate (Glu-6-P) were investigated in the awake rat. A control group (glycerol-infused) and high plasma FFA group (Liposyn-infused) were clamped at euglycemia (approximately 6 mM)-hyperinsulinemia (10 milliunits/kg/min) throughout the experiment (180-240 min). In the initial experiment, 13C NMR was used to observe [1-13C]glucose incorporation into [1-13C]glycogen in the rat hindlimb for glycogen synthesis calculations and into [3-13C]lactate and [3-13C]alanine for glycolytic flux calculations. These experiments were followed by 31P NMR measurements of Glu-6-P changes under identical conditions of the initial experiment. Plasma FFA concentrations were 2.25 +/- 0.36 and 0.20 +/- 0.03 mM in the high plasma FFA and control groups respectively (p < 0.0005). Glucose infusion rates (Ginf) decreased significantly in the Liposyn-infused rats (29.5 +/- 0.7 and 27.2 +/- 1.2 mg/kg/min for control and high plasma FFA group, respectively, at 15 min to 30.7 +/- 2.3 and 17.7 +/- 1.3 mg/kg/min, respectively, at the end of the experiment, p < 0.002). Glycogen synthesis rates were 163 +/- 32 and 104 +/- 17 nmol/g/min, and glycolytic rates were 57.9 +/- 8.0 and 19. 5 +/- 3.6 nmol/g/min (p < 0.002) in the control and high plasma FFA groups, respectively. The relative flux of pyruvate versus free fatty acids and ketones entering the tricarboxylic acid cycle was greater in the control (57 +/- 9%) versus high plasma FFA group (25 +/- 4%) (p < 0.005) as assessed by [4-13C]glutamate/[3-13C]lactate steady state isotopic enrichment measurements. Finally, Glu-6-P concentrations increased by 29.8 +/- 7.0 and 52.8 +/- 12.3% (p < 0. 05) in the control and high plasma FFA groups, respectively, above their basal concentrations by 180 min. In conclusion, we have demonstrated the ability to use in vivo NMR to elucidate the metabolic fate of glucose within skeletal muscle of an awake rat during a euglycemic-hyperinsulinemic clamp and increased levels of plasma FFA. These data suggest that increased concentrations of plasma FFA inhibit insulin-stimulated muscle glucose metabolism in the rat through inhibition of glycolysis.
Topics: Alanine; Animals; Carbon Isotopes; Fatty Acids, Nonesterified; Glucose; Glucose Clamp Technique; Glucose-6-Phosphate; Glycogen; Glycolysis; Hyperinsulinism; Infusions, Intravenous; Insulin; Ketones; Kinetics; Lactates; Magnetic Resonance Spectroscopy; Models, Biological; Muscle, Skeletal; Phosphorus; Pyruvates; Rats; Rats, Sprague-Dawley; Wakefulness
PubMed: 9099689
DOI: 10.1074/jbc.272.16.10464 -
Hypertension (Dallas, Tex. : 1979) Dec 2009Mechanisms of formation and growth of intracranial aneurysms are poorly understood. To investigate the pathophysiology of intracranial aneurysms, an animal model of...
Mechanisms of formation and growth of intracranial aneurysms are poorly understood. To investigate the pathophysiology of intracranial aneurysms, an animal model of intracranial aneurysm yielding a high incidence of large aneurysm formation within a short incubation period is needed. We combined two well-known clinical factors associated with human intracranial aneurysms, hypertension and the degeneration of elastic lamina, to induce intracranial aneurysm formation in mice. Roles of matrix metalloproteinases (MMPs) in this model were investigated using doxycycline, a broad-spectrum MMP inhibitor, and MMP knockout mice. Hypertension was induced by continuous infusion of angiotensin II for 2 weeks. The disruption of elastic lamina was achieved by a single stereotaxic injection of elastase into the cerebrospinal fluid at the right basal cistern. A total of 77% of the mice that received 35 milliunits of elastase and 1000 ng/kg per minute of angiotensin II developed intracranial aneurysms in 2 weeks. There were dose-dependent effects of elastase and angiotensin II on the incidence of aneurysms. Histologically, intracranial aneurysms observed in this model closely resembled human intracranial aneurysms. Doxycycline, a broad-spectrum MMP inhibitor, reduced the incidence of aneurysm to 10%. MMP-9 knockout mice, but not MMP-2 knockout mice, had reduced the incidence of intracranial aneurysms. In summary, a stereotaxic injection of elastase into the basal cistern in hypertensive mice resulted in intracranial aneurysms that closely resembled human intracranial aneurysms. The intracranial aneurysm formation in this model appeared to depend on MMP activation.
Topics: Angiotensin II; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Doxycycline; Enzyme Inhibitors; Hypertension; Injections; Intracranial Aneurysm; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred C57BL; Mice, Knockout; Pancreatic Elastase; Subarachnoid Space; Vasoconstrictor Agents
PubMed: 19884566
DOI: 10.1161/HYPERTENSIONAHA.109.138297 -
Proceedings of the National Academy of... May 1977The location of the somatostatin-containing D-cells of the pancreatic islets between the A- and B-cells suggests that their function might be to inhibit insulin and/or...
The location of the somatostatin-containing D-cells of the pancreatic islets between the A- and B-cells suggests that their function might be to inhibit insulin and/or glucagon secretion by these neighboring cells. To determine if insulin and/or glucagon, in concentrations that might be present in the extracellular space surrounding the D-cells, stimulate immunoreactive somatostatin (IRS) release, we perfused 10 microng of glucagon or 10 milliunits of insulin per ml in 11 isolated dog pancreases, for 40 min in seven experiments and for 100 min in four experiments. In eight of the nine experiments in which glucagon was perfused, a prompt and significant rise in mean IRS release, ranging from 71 to 128% above the control level, was observed. In the eight experiments in which insulin was perfused. IRS did not increase during the first 40 min; in the two 100-min insulin experiments, it did rise during the final 50 min, however. To determine the effect of an A- and B-cell secretogogue on IRS release, we perfused 20 mM arginine for 60 min in six experiments. In all, IRS rose within 3 min and reached a level 71-465% above the control, remaining significantly elevated throughout the perfusion, while glucagon and insulin rose to peak levels at 2 min and then declined somewhat despite continuing arginine perfusion. The results indicate that perfusion of the normal dog pancreas with high doses of glucagon or arginine is accompanied by a prompt increase in IRS release and are compatible with a local feedback circuit involving A- and D-cells. Insulin appears not to augment IRS release, at least not promptly, but IRS stimulated by local endogenous glucagon could inhibit the B-cell response to locally secreted glucagon and thereby influence the composition of the insulin/glucagon secretion mixture.
Topics: Animals; Arginine; Dogs; Glucagon; Insulin; Islets of Langerhans; Male; Secretory Rate; Somatostatin
PubMed: 325567
DOI: 10.1073/pnas.74.5.2140 -
ACS Chemical Biology Feb 2020The enzymatic transamination of ketones into ()-amines represents an important route for accessing a range of pharmaceuticals or building blocks. Although many...
The enzymatic transamination of ketones into ()-amines represents an important route for accessing a range of pharmaceuticals or building blocks. Although many publications have dealt with enzyme discovery, protein engineering, and the application of ()-selective amine transaminases [()-ATA] in biocatalysis, little is known about the actual role and how these enzymes have evolved from the ubiquitous α-amino acid transaminases (α-AATs). Here, we show the successful introduction of an ()-transaminase activity in an α-amino acid aminotransferase with one to six amino acid substitutions in the enzyme's active site. Bioinformatic analysis combined with computational redesign of the d-amino acid aminotransferase (DATA) led to the identification of a sextuple variant having a specific activity of 326 milliunits mg in the conversion of ()-phenylethylamine and pyruvate to acetophenone and d-alanine. This value is similar to those of natural ()-ATAs, which typically are in the range of 250 milliunits mg. These results demonstrate that ()-ATAs can evolve from α-AAT as shown here for the DATA scaffold.
Topics: Bacillus subtilis; Escherichia coli; Escherichia coli Proteins; Mutagenesis, Site-Directed; Mutation; Phenethylamines; Protein Binding; Stereoisomerism; Substrate Specificity; Transaminases
PubMed: 31990173
DOI: 10.1021/acschembio.9b00888 -
The Journal of Biological Chemistry Mar 1985Treatment of isolated rat adipocytes with adrenocorticotropin (ACTH) caused a 1.5-fold increase in phospholipid methyltransferase activity within 5 min. This effect of...
Treatment of isolated rat adipocytes with adrenocorticotropin (ACTH) caused a 1.5-fold increase in phospholipid methyltransferase activity within 5 min. This effect of ACTH was concentration-dependent with maximal activation at 2 milliunits/ml ACTH, and was reproduced by dibutyryl cyclic AMP. ACTH (2 milliunits/ml) caused an increase in the Vmax value of phospholipid methyltransferase without changing the Km for S-adenosyl-L-methionine. Insulin caused a concentration-dependent inhibition of both control and ACTH-stimulated phospholipid methyltransferase. Half-maximal inhibition by insulin was demonstrated with 5 microunits/ml insulin in control cells and with 25 microunits/ml insulin in ACTH-stimulated cells. The rapid and sensitive activation of adipocyte phospholipid methyltransferase by ACTH and inhibition by insulin are consistent with a role for this pathway in the hormonal response of the adipocyte.
Topics: Adipose Tissue; Adrenocorticotropic Hormone; Animals; Bucladesine; Chromatography, Thin Layer; Dose-Response Relationship, Drug; Insulin; Male; Methylation; Methyltransferases; Phosphatidyl-N-Methylethanolamine N-Methyltransferase; Phosphatidylcholines; Phosphatidylethanolamine N-Methyltransferase; Phosphatidylethanolamines; Phospholipids; Rats; Rats, Inbred Strains
PubMed: 2982871
DOI: No ID Found -
The Journal of Biological Chemistry May 1985Morphine 6-dehydrogenase, which catalyzes the dehydrogenation of morphine to morphinone, has been purified about 440-fold from the soluble fraction of guinea pig liver...
Morphine 6-dehydrogenase, which catalyzes the dehydrogenation of morphine to morphinone, has been purified about 440-fold from the soluble fraction of guinea pig liver with a yield of 38%. The purified enzyme was a homogeneous protein on polyacrylamide gel disc electrophoresis and isoelectric focusing. The molecular weight and isoelectric point of the enzyme were 29,000 and 7.6, respectively. The enzyme utilizes both NAD and NADP as a cofactor, and the Km values were 0.12 mM for NAD and 0.42 mM for NADP. The Vmax values for morphine were 588 milliunits/mg of protein (with NAD) and 1600 milliunits/mg of protein (with NADP). The Km values for morphine were 0.12 mM (with NAD) and 0.49 mM (with NADP). The enzyme also exhibited activity for morphine-related compounds: nalorphine, normorphine, codeine, and ethylmorphine; however, 7,8-saturated congeners such as dihydromorphine and dihydrocodeine were poor substrates. The enzyme was inactivated by removal of 2-mercaptoethanol from the enzyme solution. The inactivated enzyme was rapidly recovered by the addition of 2-mercaptoethanol. Phenylarsine oxide and CdCl2 (dithiol modifiers) inhibited competitively toward cofactor binding and noncompetitively toward morphine binding. These results suggest that the enzyme possesses the essential thiol groups, probably vicinal dithiol, at or near the cofactor-binding site. Using the partially purified enzyme, 8-(2-hydroxyethylthio)dihydromorphinone was isolated as the product and identified by UV, mass, and NMR spectra. It was confirmed that morphinone proposed as the dehydrogenation product was nonenzymatically and covalently bound to 2-mercaptoethanol. Accordingly, the isolated morphinone-2-mercaptoethanol conjugate must be formed by two steps: enzymatic production of morphinone from morphine and then nonenzymatic binding of 2-mercaptoethanol to morphinone.
Topics: Alcohol Oxidoreductases; Animals; Guinea Pigs; Hydromorphone; Kinetics; Liver; Magnetic Resonance Spectroscopy; Mercaptoethanol; Morphine; NAD; NADP
PubMed: 2580834
DOI: No ID Found