Abstract
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.
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.
| Original language | English |
|---|---|
| Pages (from-to) | 450-457 |
| Number of pages | 8 |
| Journal | International Journal of Artificial Organs |
| Volume | 35 |
| Issue number | 6 |
| DOIs | |
| Publication status | Published - Jun 2012 |