Bone marrow fails to differentiate into liver epithelium during murine development and regeneration.

UNLABELLED: Recent reports have provided conflicting conclusions regarding the role for bone marrow (BM)-derived cells in the regeneration of liver. Our aim was to investigate the potential of BM to contribute to liver epithelium using different BM transplant models designed to explore differentiation during normal liver development and regeneration after toxic injury. BM cells from transgenic green fluorescent protein (GFP) mice were injected into neonatal and adult immunodeficient and neonatal immune-competent mice. Three distinct models of liver injury were employed to test the contribution of marrow to the regeneration of hepatocytes, cholangiocytes, and oval cells in immune-deficient adult animals after neonatal transplant. Immunohistochemistry was combined with flow cytometry (FACS) and reverse transcription (RT)-PCR to increase the sensitivity and specificity of the analyses. Although GFP+ marrow-derived cells were observed in the livers of all transplanted animals, immunohistochemistry failed to demonstrate any marrow derived hepatocytes or cholangiocytes. FACS confirmed that GFP+ marrow-derived cells in the liver maintained expression of CD45, a leukocyte marker. Gene expression studies of GFP+ cells isolated by FACS failed to demonstrate expression of liver specific genes in these marrow-derived cells. CONCLUSION: Through highly sensitive and specific analyses, we were unable to demonstrate any evidence of transdifferentiation of BM-derived cells into epithelial hepatic tissue during the period of rapid growth in the neonatal period. Furthermore, although increased migration of hematopoietic cells to the liver occurred after toxic injury, these cells did not contribute directly to the replacement of hepatocytes, cholangiocytes, or oval cells.

[99mTc]HYNIC-RGD for imaging integrin alphavbeta3 expression.

There has been increasing interest in peptides containing the Arg-Gly-Asp (RGD) sequence for targeting of alpha(v)beta(3) integrins to image angiogenesis. [(18)F]Galacto-RGD has been successfully used for positron emission tomography applications in patients. Here we report on the preclinical characterization of a (99m)Tc-labeled derivative for single-photon emission computed tomography. c(RGDyK) was derivatized with HYNIC at the amino group of the lysine [c(RGDyK(HYNIC)) or HYNIC-RGD]. (99m)Tc labeling was performed using coligands (tricine and EDDA), as well as (99m)Tc(CO)(3)(H(2)O)(3). Radiolabeled peptides were characterized with regard to lipophilicity, protein binding and stability in buffer, serum and tissue homogenates. Integrin receptor activity was determined in internalization assays using alpha(v)beta(3)-receptor-positive M21 and alpha(v)beta(3)-receptor-negative M21L melanoma cells. Biodistribution was evaluated in normal and nude mice bearing M21, M21L and small cell lung tumors. HYNIC-RGD could be labeled at high specific activities using tricine, tricine-trisodium triphenylphosphine 3,3',3''-trisulfonate (TPPTS), tricine-nicotinic acid (NA) or EDDA as coligands. [(99m)Tc]EDDA/HYNIC-RGD, [(99m)Tc]tricine-TPPTS/HYNIC-RGD and [(99m)Tc]tricine-NA/HYNIC-RGD showed protein binding (<5%) considerably lower than [(99m)Tc](CO)(3)/HYNIC-RGD and [(99m)Tc]tricine/HYNIC-RGD. [(99m)Tc]EDDA/HYNIC-RGD revealed high in vitro stability accompanied by low lipophilicity with a log P value of -3.56, comparable to that of [(18)F]Galacto-RGD. In M21 cells for this compound, the highest level of specific and rapid cell uptake (1.25% mg protein(-1)) was determined. In vivo, rapid renal excretion, low blood retention, low liver and muscle uptakes and low intestinal excretion 4 h postinjection were observed. Tumor uptake values were 2.73% ID/g in M21 alpha(v)beta(3)-receptor-positive tumors versus 0.85% ID/g in receptor-negative tumors 1 h postinjection. Small cell lung tumors could be visualized using gamma camera imaging. [(99m)Tc]EDDA/HYNIC-RGD shows encouraging properties to target alpha(v)beta(3) receptors in vivo with high stability and favorable pharmacokinetics. Tumor uptake studies showed specific targeting of alpha(v)beta(3)-receptor-positive tumors with tumor-to-organ ratios comparable to those of [(18)F]Galacto-RGD.