PLASMA CLEARANCE OF 57COBALTLABELLED VITAMIN B12 BOUND IN VITRO AND IN VIVO TO TRANSCOBALAMIN I AND II

SummaryPharmacokinetics of intravenously injected 125I-labeled urokinase (125I-UK) of a molecular weight of 33,000 daltons in normal rabbits and patients with various diseases were investigated. The plasma clearance of 125I-UK in rabbits was described by a biexponential curve within six hours with a half-life of 8 minutes, 2.3 hours, respectively. The radioactivity in the liver and kidneys 15 minutes after iv injection with 125I-UK was 9.6% and 14.0% of the radioactivity injected, respectively. Approximately 80% of the total radioactive material injected was excreted in the urine in 18 hours. No increase in activator activity in the urine was observed after a large amount of UK injection. Activity uptake of 125I-UK by experimentally induced arterial thrombus was little. Lysis of the stasis thrombus was produced by injecting 7.5 × 104 IU of UK in only one out of 8 rabbits. In vitro contact experiment revealed that transfer of 125I-UK to plasma clot is slow (24 hours for 10% of 125I-UK by plasma clot). In 4 patients plasma clearance of 125I-UK was essentially similar to that in rabbits. From the results obtained optimal dosage regimen of UK administration for complete thrombolysis in vivo was discussed.

Download Full-text

Catalytic activity and stereoselectivity of engineered phosphotriesterases towards structurally different nerve agents in vitro

Archives of Toxicology ◽

10.1007/s00204-021-03094-0 ◽

2021 ◽

Author(s):

Anja Köhler ◽

Benjamin Escher ◽

Laura Job ◽

Marianne Koller ◽

Horst Thiermann ◽

...

Keyword(s):

Plasma Clearance ◽

Nerve Agents ◽

Inhibition Assay ◽

Ache Inhibition ◽

Catalytic Activities ◽

Chiral Lc ◽

Hydrolysis Of ◽

Medical Countermeasures

AbstractHighly toxic organophosphorus nerve agents, especially the extremely stable and persistent V-type agents such as VX, still pose a threat to the human population and require effective medical countermeasures. Engineered mutants of the Brevundimonas diminuta phosphotriesterase (BdPTE) exhibit enhanced catalytic activities and have demonstrated detoxification in animal models, however, substrate specificity and fast plasma clearance limit their medical applicability. To allow better assessment of their substrate profiles, we have thoroughly investigated the catalytic efficacies of five BdPTE mutants with 17 different nerve agents using an AChE inhibition assay. In addition, we studied one BdPTE version that was fused with structurally disordered PAS polypeptides to enable delayed plasma clearance and one bispecific BdPTE with broadened substrate spectrum composed of two functionally distinct subunits connected by a PAS linker. Measured kcat/KM values were as high as 6.5 and 1.5 × 108 M−1 min−1 with G- and V-agents, respectively. Furthermore, the stereoselective degradation of VX enantiomers by the PASylated BdPTE-4 and the bispecific BdPTE-7 were investigated by chiral LC–MS/MS, resulting in a several fold faster hydrolysis of the more toxic P(−) VX stereoisomer compared to P(+) VX. In conclusion, the newly developed enzymes BdPTE-4 and BdPTE-7 have shown high catalytic efficacy towards structurally different nerve agents and stereoselectivity towards the toxic P(−) VX enantiomer in vitro and offer promise for use as bioscavengers in vivo.

Download Full-text

Abstract 230: Lipoprotein X Causes Renal Disease in LCAT Deficiency

Arteriosclerosis Thrombosis and Vascular Biology ◽

10.1161/atvb.36.suppl_1.230 ◽

2016 ◽

Vol 36 (suppl_1) ◽

Author(s):

Edward B Neufeld ◽

Alice Ossoli ◽

Seth G Thacker ◽

Boris Vaisman ◽

Milton Pryor ◽

...

Keyword(s):

Renal Disease ◽

Plasma Clearance ◽

Mesangial Cells ◽

Chemical Properties ◽

Low Hdl ◽

Lcat Deficiency ◽

Physical And Chemical ◽

Dependent Pathway

Familial lecithin:cholesterol acyltransferase (LCAT) deficiency (FLD) is characterized by low HDL, accumulation of an abnormal cholesterol-rich multilamellar particle called lipoprotein-X (LpX) in plasma, and renal disease. The aim of our study was to determine if LpX is nephrotoxic and to gain insight into the pathogenesis of FLD renal disease. We administered a synthetic LpX, nearly identical to endogenous LpX in its physical, and chemical properties, to wild-type and Lcat -/- mice. Our in vitro and in vivo studies demonstrated an apoA-I and LCAT-dependent pathway for LpX conversion to HDL-like particles, which likely mediates normal plasma clearance of LpX. Plasma clearance of exogenous LpX was markedly delayed in Lcat -/- mice, which have low HDL but only minimal amounts of endogenous LpX and do not spontaneously develop renal disease. Chronically administered exogenous LpX deposited in all renal glomerular cellular and matrical compartments of Lcat -/- mice, and induced proteinuria and nephrotoxic gene changes, as well as all of the hallmarks of FLD renal disease as assessed by histological, TEM, and SEM analyses. Extensive in vivo EM studies revealed LpX uptake by macropinocytosis into mouse glomerular endothelial cells, podocytes, and mesangial cells and delivery to lysosomes, where it was degraded. Endocytosed LpX appeared to be degraded by both human podocyte and mesangial cell lysosomal PLA 2 and induced podocyte secretion of pro-inflammatory IL-6 in vitro and renal Cxl10 expression in Lcat -/- mice. In conclusion, LpX is a nephrotoxic particle that in the absence of LCAT induces all of the histological and functional hallmarks of FLD and hence may serve as a biomarker for monitoring recombinant LCAT therapy. In addition, our studies suggest that LpX-induced loss of endothelial barrier function and release of cytokines by renal glomerular cells likely plays a role in the initiation and progression of FLD nephrosis.

Download Full-text

Protamine enhances the proliferative activity of hepatocyte growth factor in rats

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1998.274.1.g21 ◽

1998 ◽

Vol 274 (1) ◽

pp. G21-G28 ◽

Cited By ~ 1

Author(s):

Ke-Xin Liu ◽

Yukio Kato ◽

Tai-Ichi Kaku ◽

Kunio Matsumoto ◽

Toshikazu Nakamura ◽

...

Keyword(s):

Growth Factor ◽

Hepatocyte Growth Factor ◽

Proliferative Activity ◽

Plasma Clearance ◽

Mitogenic Response ◽

Enhancing Effect ◽

A Cell ◽

Hepatocyte Growth

The effect of protamine on the proliferative activity of hepatocyte growth factor (HGF) was examined in α-naphthyl isothiocyanate-intoxicated rats. Protamine preinjection increased the hepatocyte labeling index induced by HGF four- to fivefold. A similar effect was also observed in partially hepatectomized rats. Because a cell surface heparin-like substance can bind to HGF and protamine has an affinity for heparin, protamine may affect HGF pharmacokinetics. In fact, protamine injection caused a transient increase in plasma HGF concentrations after administration of HGF and, in vitro, protamine eluted HGF prebound to heparin-Sepharose. Protamine also reduced the plasma clearance of HGF and increased 2.5-fold the exposure of hepatocytes to HGF in vivo. The enhancing effect of protamine on the mitogenic response of hepatocytes to HGF was also observed in vitro (∼2-fold after protamine pretreatment compared with HGF alone), suggesting that the enhancing effect of protamine on HGF-induced liver regeneration results from dual effects exerted by protamine 1) lowering the overall elimination of HGF and 2) directly stimulating hepatocyte mitosis induced by HGF.

Download Full-text

Stimulation in vitro of vitamin B12-dependent methionine synthase by polyamines

Biochemical Journal ◽

10.1042/bj3160661 ◽

1996 ◽

Vol 316 (2) ◽

pp. 661-665 ◽

Cited By ~ 10

Author(s):

Susan H. KENYON ◽

Anna NICOLAOU ◽

Tamara AST ◽

William A. GIBBONS

Keyword(s):

Enzyme Activity ◽

Vitamin B12 ◽

Cell Signalling ◽

Methionine Synthase ◽

Polyamine Metabolism ◽

Physiological Concentration ◽

Methylation Pathway ◽

Important Enzyme

Vitamin B12-dependent methionine synthase is an important enzyme for sulphur amino acid, folate polyamine metabolism, S-adenosylmethionine metabolism and also in the methylation pathway of DNA, RNA, proteins and lipids. Consequently, studies aiming at exploring the control and regulation of methionine synthase are of particular interest. Here we report the modulation of enzyme activity in vitro by polyamines. Although putrescine, cadaverine, spermine and spermidine all stimulated enzyme activity, the last two were the most potent, causing increases in enzyme activity up to 400%. The EC50 for spermine was determined as 8 μM and for spermidine 40 μM. The physiological concentration for spermine has been reported to be 15–19 μM. Spermine was found to increase both the Km and the Vmax with respect to methyltetrahydrofolate for the enzyme. These data support the hypothesis that spermine and spermidine are feedback regulators of methionine synthase both in vivo and in vitro and are consistent with the polyamines' regulating cell signalling pathways.

Download Full-text

Vitamin B12–Loaded Buccoadhesive Films as a Noninvasive Supplement in Vitamin B12 Deficiency: In Vitro Evaluation and In Vivo Comparative Study With Intramuscular Injection

Journal of Pharmaceutical Sciences ◽

10.1016/j.xphs.2017.03.040 ◽

2017 ◽

Vol 106 (7) ◽

pp. 1849-1858 ◽

Cited By ~ 6

Author(s):

Soad A. Mohamad ◽

Hatem A. Sarhan ◽

Hamdy Abdelkader ◽

Heba F. Mansour

Keyword(s):

Comparative Study ◽

Vitamin B12 ◽

Intramuscular Injection ◽

Vitamin B12 Deficiency ◽

In Vitro Evaluation ◽

B12 Deficiency

Download Full-text

Hepatic fibronectin matrix turnover in rats: involvement of the asialoglycoprotein receptor

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1999.277.6.g1189 ◽

1999 ◽

Vol 277 (6) ◽

pp. G1189-G1199 ◽

Cited By ~ 1

Author(s):

Robert F. Rotundo ◽

Peter A. Vincent ◽

Paula J. McKeown-Longo ◽

Frank A. Blumenstock ◽

Thomas M. Saba

Keyword(s):

Intravenous Infusion ◽

Plasma Clearance ◽

Endocytic Pathway ◽

Male Rats ◽

Adult Rats ◽

Adhesive Protein ◽

Terminal Galactose ◽

Exposed Terminal

Fibronectin (Fn) is a major adhesive protein found in the hepatic extracellular matrix (ECM). In adult rats, the in vivo turnover of plasma Fn (pFn) incorporated into the liver ECM is relatively rapid, i.e., <24 h, but the regulation of its turnover has not been defined. We previously reported that cellular Fn (cFn) and enzymatically desialylated plasma Fn (aFn), both of which have a high density of exposed terminal galactose residues, rapidly interact with hepatic asialoglycoprotein receptors (ASGP-R) in association with their plasma clearance after intravenous infusion. With the use of adult male rats (250–350 g) and measurement of the deoxycholate (DOC)-insoluble125I-labeled Fn in the liver, we determined whether the ASGP-R system can also influence the hepatic matrix retention of various forms of Fn. There was a rapid deposition of 125I-pFn,125I-aFn, and125I-cFn into the liver ECM after their intravenous injection. Although125I-pFn was slowly lost from the liver matrix over 24 h, more than 90% of the incorporated125I-aFn and125I-cFn was cleared within 4 h ( P < 0.01). Intravenous infusion of excess nonlabeled asialofetuin to competitively inhibit the hepatic ASGP-R delayed the rapid turnover of both aFn and cFn already incorporated within the ECM of the liver. ECM retention of both125I-aFn and125I-cFn was also less than125I-pFn ( P < 0.01) as determined in vitro using liver slices preloaded in vivo with either tracer form of Fn. The hepatic ASGP-R appears to participate in the turnover of aFn and cFn within the liver ECM, whereas a non-ASGP-R-associated endocytic pathway apparently influences the removal of normal pFn incorporated within the hepatic ECM, unless it becomes locally desialylated.

Download Full-text