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Subcellular spatio-temporal intravital kinetics of aflatoxin B₁ and ochratoxin A in liver and kidney.

ABSTRACT: Local accumulation of xenobiotics in human and animal tissues may cause adverse effects. Large differences in their concentrations may exist between individual cell types, often due to the expression of specific uptake and export carriers. Here we established a two-photon microscopy-based technique for spatio-temporal detection of the distribution of mycotoxins in intact kidneys and livers of anesthetized mice with subcellular resolution. The mycotoxins ochratoxin A (OTA, 10 mg/kg b.w.) and aflatoxin B₁ (AFB₁, 1.5 mg/kg b.w.), which both show blue auto-fluorescence, were analyzed after intravenous bolus injections. Within seconds after administration, OTA was filtered by glomeruli, and enriched in distal tubular epithelial cells (dTEC). A striking feature of AFB₁ toxicokinetics was its very rapid uptake from sinusoidal blood into hepatocytes (t_1/2 ~ 4 min) and excretion into bile canaliculi. Interestingly, AFB₁ was enriched in the nuclei of hepatocytes with zonal differences in clearance. In the cytoplasm of pericentral hepatocytes, the half-life (t_1/2~ 63 min) was much longer compared to periportal hepatocytes of the same lobules (t_1/2 ~ 9 min). In addition, nuclear AFB₁ from periportal hepatocytes cleared faster compared to the pericentral region. These local differences in AFB₁ clearance may be due to the pericentral expression of cytochrome P450 enzymes that activate AFB₁ to protein- and DNA-binding metabolites. In conclusion, the present study shows that large spatio-temporal concentration differences exist within the same tissues and its analysis may provide valuable additional information to conventional toxicokinetic studies.

SUBMITTER: Ghallab A

PROVIDER: S-EPMC8166722 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundDifferentiating infiltrating myeloid cells from resident microglia in neuroinflammatory disease is challenging, because bone marrow-derived inflammatory monocytes infiltrating the inflamed brain adopt a 'microglia-like' phenotype. This precludes the accurate identification of either cell type without genetic manipulation, which is important to understand their temporal contribution to disease and inform effective intervention in its pathogenesis. During West Nile virus (WNV) encephalitis, widespread neuronal infection drives substantial CNS infiltration of inflammatory monocytes, causing severe immunopathology and/or death, but the role of microglia in this remains unclear.MethodsUsing high-parameter cytometry and dimensionality-reduction, we devised a simple, novel gating strategy to identify microglia and infiltrating myeloid cells during WNV-infection. Validating our strategy, we (1) blocked the entry of infiltrating myeloid populations from peripheral blood using monoclonal blocking antibodies, (2) adoptively transferred BM-derived monocytes and tracked their phenotypic changes after infiltration and (3) labelled peripheral leukocytes that infiltrate into the brain with an intravenous dye. We demonstrated that myeloid immigrants populated only the identified macrophage gates, while PLX5622 depletion reduced all 4 subsets defined by the microglial gates.ResultsUsing this gating approach, we identified four consistent microglia subsets in the homeostatic and WNV-infected brain. These were P2RY12hi CD86-, P2RY12hi CD86+ and P2RY12lo CD86- P2RY12lo CD86+. During infection, 2 further populations were identified as 'inflammatory' and 'microglia-like' macrophages, recruited from the bone marrow. Detailed kinetic analysis showed significant increases in the proportions of both P2RY12lo microglia subsets in all anatomical areas, largely at the expense of the P2RY12hi CD86- subset, with the latter undergoing compensatory proliferation, suggesting replenishment of, and differentiation from this subset in response to infection. Microglia altered their morphology early in infection, with all cells adopting temporal and regional disease-specific phenotypes. Late in disease, microglia produced IL-12, downregulated CX3CR1, F4/80 and TMEM119 and underwent apoptosis. Infiltrating macrophages expressed both TMEM119 and P2RY12 de novo, with the microglia-like subset notably exhibiting the highest proportional myeloid population death.ConclusionsOur approach enables detailed kinetic analysis of resident vs infiltrating myeloid cells in a wide range of neuroinflammatory models without non-physiological manipulation. This will more clearly inform potential therapeutic approaches that specifically modulate these cells.

Project description:The liver is the central organ for detoxification of xenobiotics in the body. In pharmacokinetic modeling, hepatic metabolization capacity is typically quantified as hepatic clearance computed as degradation in well-stirred compartments. This is an accurate mechanistic description once a quasi-equilibrium between blood and surrounding tissue is established. However, this model structure cannot be used to simulate spatio-temporal distribution during the first instants after drug injection. In this paper, we introduce a new spatially resolved model to simulate first pass perfusion of compounds within the naive liver. The model is based on vascular structures obtained from computed tomography as well as physiologically based mass transfer descriptions obtained from pharmacokinetic modeling. The physiological architecture of hepatic tissue in our model is governed by both vascular geometry and the composition of the connecting hepatic tissue. In particular, we here consider locally distributed mass flow in liver tissue instead of considering well-stirred compartments. Experimentally, the model structure corresponds to an isolated perfused liver and provides an ideal platform to address first pass effects and questions of hepatic heterogeneity. The model was evaluated for three exemplary compounds covering key aspects of perfusion, distribution and metabolization within the liver. As pathophysiological states we considered the influence of steatosis and carbon tetrachloride-induced liver necrosis on total hepatic distribution and metabolic capacity. Notably, we found that our computational predictions are in qualitative agreement with previously published experimental data. The simulation results provide an unprecedented level of detail in compound concentration profiles during first pass perfusion, both spatio-temporally in liver tissue itself and temporally in the outflowing blood. We expect our model to be the foundation of further spatially resolved models of the liver in the future.

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Subcellular spatio-temporal intravital kinetics of aflatoxin B₁ and ochratoxin A in liver and kidney.

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Subcellular spatio-temporal intravital kinetics of aflatoxin B1 and ochratoxin A in liver and kidney.

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Subcellular spatio-temporal intravital kinetics of aflatoxin B₁ and ochratoxin A in liver and kidney.