Prediction of hepatic plasma clearance in vivo from gel-entrapped rat and human hepatocytes

2013 ◽  
Vol 91 (2) ◽  
pp. 178-186 ◽  
Author(s):  
Jian Yin ◽  
Hongxia Qiu ◽  
Jing Dai ◽  
Yanhua Lu ◽  
Rong Zhao ◽  
...  
Keyword(s):  
Plasma Clearance ◽  
Rat Hepatocytes ◽  
Drug Clearance ◽  
Human Hepatocytes ◽  
Drug Binding ◽  
Parallel Tube ◽  
Predicted Values ◽  
The Mathematical Model

This paper aimed to evaluate the applicability of gel-entrapped rat and human hepatocytes in the prediction of hepatic plasma clearance (CLh,plasma) in vivo. The in vitro intrinsic clearances (CLint,in vitro) for the selected compounds were determined from the substrate disappearance rate, and further used to predict CLh,plasma using 3 classical mathematical models (well-stirred, parallel-tube, and dispersion) and disregarding drug binding. As a result, the predicted values from gel-entrapped rat hepatocytes were mostly within 2 SE of the literature data with a high correlation coefficient (R2) of 0.88–0.91. The predicted data with human hepatocytes also fitted well with the clinical data, indicating a high accuracy in prediction of in-vivo clearance. With respect to the mathematical model for predicting CLh,plasma, the parallel-tube and dispersion models produced a better prediction than the well-stirred model, and we suggest using the parallel-tube model because it is less complex mathematically. In conclusion, gel-entrapped hepatocytes predicted the drug clearance well and seemed to be a useful tool in the process of drug discovery.

scholarly journals Novel testing strategy for prediction of rat biliary excretion of intravenously administered estradiol-17β glucuronide

2020 ◽  
Author(s):  
Annelies Noorlander ◽  
Eric Fabian ◽  
Bennard van Ravenzwaay ◽  
Ivonne M. C. M. Rietjens
Keyword(s):  
Biliary Excretion ◽  
Rat Hepatocytes ◽  
Scaling Factor ◽  
Blood Levels ◽  
Testing Strategy ◽  
Predicted Values ◽  
Definition Of ◽  
Estradiol 17Β

Abstract The aim of the present study was to develop a generic rat physiologically based kinetic (PBK) model that includes a novel testing strategy where active biliary excretion is incorporated using estradiol-17β glucuronide (E217βG) as the model substance. A major challenge was the definition of the scaling factor for the in vitro to in vivo conversion of the PBK-model parameter Vmax. In vitro values for the Vmax and Km for transport of E217βG were found in the literature in four different studies based on experiments with primary rat hepatocytes. The required scaling factor was defined based on fitting the PBK model-based predicted values to reported experimental data on E217βG blood levels and cumulative biliary E217βG excretion. This resulted in a scaling factor of 129 mg protein/g liver. With this scaling factor the PBK model predicted the in vivo data for blood and cumulative biliary E217βG levels with on average of less than 1.8-fold deviation. The study provides a proof of principle on how biliary excretion can be included in a generic PBK model using primary hepatocytes to define the kinetic parameters that describe the biliary excretion.

Hepatocyte Labeling with 99mTc-GSA: A Potential Non-Invasive Technique for Tracking Cell Transplantation

2012 ◽  
Vol 35 (6) ◽  
pp. 450-457 ◽  
Author(s):  
Manil Chouhan ◽  
Juliana Puppi ◽  
Estela Solanas ◽  
Ragai R. Mitry ◽  
Anil Dhawan ◽  
...  
Keyword(s):  
Rat Hepatocytes ◽  
Fold Increase ◽  
Cell Types ◽  
Human Hepatocytes ◽  
Hepatocyte Transplantation ◽  
Invasive Technique ◽  
Promising Alternative ◽  
And Function

Background: Hepatocyte transplantation is a promising alternative to orthotopic liver transplantation, however, the fate of transplanted hepatocytes is not well defined. 99mTc-galactosyl-serum albumin (99mTc-GSA) is a clinical scintigraphic agent which is specifically taken up by the hepatocyte asialoglycoprotein receptor (ASGPR). Aims: To investigate labeling of fresh and cryopreserved human hepatocytes and fresh rat hepatocytes in vitro using 99mTc-GSA Methods: Human and rat hepatocytes were isolated from liver tissue by collagenase perfusion. The ASGPR were characterized using immunohistochemistry and RT-PCR. Hepatocytes were incubated with 99mTc-GSA in suspension at 4°C and 37°C. Cell viability and function was determined using cell mitochondrial dehydrogenase (MTS) and sulphorhodamine B (SRB) assays. Results: Fresh and cryopreserved human hepatocytes expressed the ASGPR. Incubation of hepatocytes in suspension with 99mTc-GSA reduced the viability of hepatocytes, but this was similar to unlabeled control cells. Greater loss of viability was seen on incubation at 37°C compared to 4°C, but there was a significantly greater uptake of 99mTc-GSA at the physiological temperature (6.6 ± SE 0.6-fold increase, p<0.05) consistent with ASGPR-mediated endocytosis. MTS and SRB assays were not significantly affected by labeling with 99mTc-GSA in all three cell types. A mean of 18.5% of the radioactivity was released over 120 min when 99mTc-GSA - labeled hepatocytes were shaken in vitro at 37°C. Conclusions: Human and rat hepatocytes can be labeled with 99mTc-GSA, which may have potential application for in vivo imaging after hepatocyte transplantation.

Studies on Metabolism of Urokinase and Mechanism of Thrombolysis by Urokinase

1979 ◽  
Vol 42 (03) ◽  
pp. 885-894 ◽  
Author(s):  
Tatsuo Ueno ◽  
Norio Kobayashi ◽  
Tadashi Maekawa
Keyword(s):  
Molecular Weight ◽  
Plasma Clearance ◽  
Radioactive Material ◽  
Optimal Dosage ◽  
Plasma Clot ◽  
Optimal Dosage Regimen ◽  
Arterial Thrombus ◽  
Contact Experiment

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.

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

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.

scholarly journals Alcohol-and-HIV-Induced Lysosomal Dysfunction Regulates Extracellular Vesicles Secretion in Vitro and in Liver-Humanized Mice

Biology ◽  
2021 ◽  
Vol 10 (1) ◽  
pp. 29
Author(s):  
Raghubendra Singh Dagur ◽  
Moses New-Aaron ◽  
Murali Ganesan ◽  
Weimin Wang ◽  
Svetlana Romanova ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Extracellular Vesicles ◽  
Treated Group ◽  
Human Hepatocytes ◽  
In Vivo Studies ◽  
People Living With Hiv ◽  
Humanized Mouse Model ◽  
And Function

Background: Alcohol abuse is common in people living with HIV-1 and dramaticallyenhances the severity of HIV-induced liver damage by inducing oxidative stress and lysosomaldysfunction in the liver cells. We hypothesize that the increased release of extracellular vesicles(EVs) in hepatocytes and liver humanized mouse model is linked to lysosome dysfunction. Methods:The study was performed on primary human hepatocytes and human hepatoma RLWXP-GFP (Huh7.5 cells stably transfected with CYP2E1 and XPack-GFP) cells and validated on ethanol-fed liverhumanizedfumarylacetoacetate hydrolase (Fah)-/-, Rag2-/-, common cytokine receptor gamma chainknockout (FRG-KO) mice. Cells and mice were infected with HIV-1ADA virus. Results: We observedan increase in the secretion of EVs associated with a decrease in lysosomal activity and expressionof lysosomal-associated membrane protein 1. Next-generation RNA sequencing of primary humanhepatocytes revealed 63 differentially expressed genes, with 13 downregulated and 50 upregulatedgenes in the alcohol–HIV-treated group. Upstream regulator analysis of differentially expressedgenes through Ingenuity Pathway Analysis identified transcriptional regulators affecting downstreamgenes associated with increased oxidative stress, lysosomal associated disease, and function andEVs biogenesis. Our in vitro findings were corroborated by in vivo studies on human hepatocytetransplantedhumanized mice, indicating that intensive EVs’ generation by human hepatocytes andtheir secretion to serum was associated with increased oxidative stress and reduction in lysosomalactivities triggered by HIV infection and ethanol diet. Conclusion: HIV-and-ethanol-metabolisminducedEVs release is tightly controlled by lysosome status in hepatocytes and participates in thedevelopment of double-insult-induced liver injury.

Stealth doxorubicin conjugated bimetallic selenium/silver nanoparticles for targeted cervical cancer therapy

2021 ◽  
Vol 12 (4) ◽  
pp. 045006
Author(s):  
Thoko Malinga ◽  
Tukayi Kudanga ◽  
Londiwe Simphiwe Mbatha
Keyword(s):  
Cervical Cancer ◽  
Folic Acid ◽  
Colloidal Stability ◽  
Controlled Drug Release ◽  
Delivery Systems ◽  
Drug Binding ◽  
Cervical Cancer Cells ◽  
Diphenyl Tetrazolium Bromide

Abstract Bimetallic nanosized delivery systems are attracting a lot of research interest as alternatives to monometallic delivery systems. This study evaluated the ability of bimetallic selenium silver chitosan pegylated folic acid targeted nanoparticles (SeAgChPEGFA NPs) to deliver doxorubicin (DOX) in cervical cancer cells. Comparison studies using monometallic selenium chitosan pegylated folic acid (SeChPEGFA NPs) targeted NPs and free DOX were also conducted. The prepared NPs and their drug nanocomplexes were characterised morphologically and physico-chemically. Drug binding and releasing studies were conducted under a simulated environment in vitro. The cytotoxicity and apoptosis studies were studied using the 3-[(4, 5-dimethylthiazol-2-yl)−2, 5-diphenyl tetrazolium bromide] (MTT) assay and the dual dye staining. The findings revealed that the bimetallic SeAgChPEGFA NPs displayed better colloidal stability, superior physico-chemical qualities, and higher binding abilities in comparison with monometallic SeChPEGFA NPs. In addition, the SeAgChPEGFA NPs showed the pH-triggered controlled drug release and cell-specific cytotoxicity. These findings suggest that the bimetallic NPs are superior delivery systems when compared to their monometallic NPs and free drug counterparts, thus, setting a platform for further in vivo examination.

scholarly journals Comparative metabolism of the antiviral dimer 3'-azido-3'-deoxythymidine-P-2',3'-dideoxyinosine and the monomers zidovudine and didanosine by rat, monkey, and human hepatocytes.

1997 ◽  
Vol 41 (11) ◽  
pp. 2502-2510 ◽  
Author(s):  
X R Pan-Zhou ◽  
E Cretton-Scott ◽  
X J Zhou ◽  
M Y Xie ◽  
R Rahmani ◽  
...  
Keyword(s):  
High Performance ◽  
Rat Hepatocytes ◽  
Human Hepatocytes ◽  
Metabolic Fate ◽  
Extracellular Medium ◽  
Isolated Rat Hepatocytes ◽  
Cell Extracts ◽  
Comparative Metabolism ◽  
Isolated Rat

AZT-P-ddI is an antiviral heterodimer composed of one molecule of 3'-azido-3'-deoxythymidine (AZT) and one molecule of 2',3'-dideoxyinosine (ddI) linked through their 5' positions by a phosphate bond. The metabolic fate of the dimer was studied with isolated rat, monkey, and human hepatocytes and was compared with that of its component monomers AZT and ddI. Upon incubation of double-labeled [14C]AZT-P-[3H]ddI in freshly isolated rat hepatocytes in suspension at a final concentration of 10 microM, the dimer was taken up intact by cells and then rapidly cleaved to AZT, AZT monophosphate, ddI, and ddI monophosphate. AZT and ddI so formed were then subject to their respective catabolisms. High-performance liquid chromatography analyses of the extracellular medium and cell extracts revealed the presence of unchanged dimer, AZT, 3'-azido-3'-deoxy-5'-beta-D-glucopyranosylthymidine (GAZT), 3'-amino-3'-deoxythymidine (AMT), ddI, and a previously unrecognized derivative of the dideoxyribose moiety of ddI, designated ddI-M. Trace extracellular but substantial intracellular levels of the glucuronide derivative of AMT (3'-amino-3'-deoxy-5'-beta-D-glucopyranosylthymidine [GAMT]) were also detected. Moreover, the extent of the formation of AMT, GAZT, and ddI-M from the dimer was markedly lower than that with AZT and ddI alone by the hepatocytes. With hepatocytes in primary culture obtained from rat, monkey, and human, large interspecies variations in the metabolism of AZT-P-ddI were observed. While GAZT and ddI-M, metabolites of AZT and ddI, respectively, as well as AZT 5'-monophosphate (MP) and ddI-MP were detected in the extracellular media of all species, AMT and GAMT were produced only by rat and monkey hepatocytes. No such metabolites were formed by human hepatocytes. The metabolic fate of the dimer by human hepatocytes was consistent with in vivo data recently obtained from human immunodeficiency virus-infected patients.

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