Imaging of primary human hepatocytes performed with micron‐sized iron oxide particles and clinical magnetic resonance tomography

Abstract

Transplantation of primary human hepatocytes is a promising approach in certain liver diseases. For the visualization of the hepa-tocytes during and following cell application and the ability of a timely response to potential complications, a non-invasive modality for imaging the transplanted cells has to be established. The aim of this study was to label primary human hepatocytes with micron-sized iron oxide particles (MPIOs), enabling the detection of cells by clinical magnetic resonance imaging (MRI). Primary human hepatocytes isolated from 13 different donors were used for the labelling experiments. Following the dose-finding studies, hepatocytes were incubated with 30 particles/cell for 4 hrs in an adhesion culture. Particle incorporation was investigated via light, fluorescence and electron microscopy, and labelled cells were fixed and analysed in an agarose suspension by a 3.0 Tesla MR scanner. The hepatocytes were enzymatically resuspended and analysed during a 5-day reculture period for viability, total protein, enzyme leakage (aspartate aminotransferase [AST], lactate dehydrogenase [LDH]) and metabolic activity (urea, albumin). A mean uptake of 18 particles/cell could be observed, and the primary human hepatocytes were clearly detectable by MR instrumentation. The particle load was not affected by resuspension and showed no alternations during the culture period. Compared to control groups, labelling and resuspension had no adverse effects on the viability, enzyme leakage and metabolic activity of the human hepatocytes. The feasibility of preparing MPIO-labelled primary human hepatocytes detectable by clinical MR equipment was shown in vitro. MPIO-labelled cells could serve for basic research and quality control in the clinical setting of human hepatocyte transplantation.

In Vitro Evaluation of Magnetic Resonance Imaging Contrast Agents for Labeling Human Liver Cells: Implications for Clinical Translation

“…They assumed that released SPIO boosted inflammation locally after their delivery to the damaged tissue [20]. To exclude such unknowables, labeling must be achieved by simple incubation of cells with particles [12]. For this application, MPIO are superior to SPIO because they enable faster labeling, possibly due to their larger size and to negatively charged groups on their surface [36].…”

“…Primary human hepatocytes were isolated using a modified two-step collagenase perfusion technique [12]. In brief, the specimens were flushed under sterile conditions with prewarmed washing buffer, digested by recirculation of digestion buffer with 0.1% collagenase P (Roche Diagnostics GmbH, Mannheim, Germany) and mechanically disrupted in washing buffer supplemented with human albumin.…”

“…The clinically approved dextran-coated SPIO are biodegradable [13], whereas polymer-embedded MPIO are supposedly bio-inert [14]. Several studies have shown that neither type of particle affects the cellular viability or metabolic activity of labeled cells [12,[15][16][17][18], but some recent studies have found adverse effects after administration of SPIO-labeled cells [19][20][21][22]. For MPIO, which have not been investigated to that extent, concerns remain with respect to their large iron core.…”

“…Standardized protocols for labeling human liver cells with both types of particles are available [10][11][12], and we previously used MPIOs in a preclinical study of LCT monitoring (Raschzok et al, unpublished data); however, SPIOs are readily available as clinically approved contrast agents (e.g., Feridex, Bayer HealthCare). Therefore, we compared the efficacy of labeling human hepatocytes by incubation with SPIO or MPIO for visualization under conditions of clinical MRI in vitro.…”

Magnetic targeting of iron-oxide-labeled fluorescent hepatoma cells to the liver

“…This can be achieved either with pathology using recombinant retroviruses reporter genes [11], at the expense of repeated biopsies, or using in vivo imaging and cell labeling, for example, with iron-oxide particles, before transplantation [12]. Intracellular magnetic labeling offers opportunities for both MRI monitoring of cell transplant [13] and control of cell migration by remote magnetic forces.…”