In vivo detection of gene expression in liver by 31P nuclear magnetic resonance spectroscopy employing creatine kinase as a marker gene.
In vivo assessment of gene expression is desirable to obtain information on the extent and duration of transduction of tissue after gene delivery. We have developed an in vivo, potentially noninvasive, method for detecting virally mediated gene transfer to the liver. The method employs an adenoviral vector carrying the gene for the brain isozyme of murine creatine kinase (CK-B), an ATP-buffering enzyme expressed mainly in muscle and brain but absent from liver, kidney, and pancreas. Gene expression was monitored by (31)P magnetic resonance spectroscopy (MRS) using the product of the CK enzymatic reaction, phosphocreatine, as an indicator of transfection. The vector was administered into nude mice by tail vein injection, and exogenous creatine was administered in the drinking water and by i.p. injection of 2% creatine solution before (31)P MRS examination, which was performed on surgically exposed livers. A phosphocreatine resonance was detected in livers of mice injected with the vector and was absent from livers of control animals. CK expression was confirmed in the injected animals by Western blot analysis, enzymatic assays, and immunofluorescence measurements. We conclude that the syngeneic enzyme CK can be used as a marker gene for in vivo monitoring of gene expression after virally mediated gene transfer to the liver.
Auricchio
A
A
Department of Molecular and Cellular Engineering, Institute for Human Gene Therapy, University of Pennsylvania, Philadelphia, PA 19104, USA.
Zhou
R
R
Wilson
J M
JM
Glickson
J D
JD
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000201:enzymology; D002457:Cell Extracts; D003401:Creatine / Q000378:metabolism / Q000494:pharmacology; D003402:Creatine Kinase / Q000235:genetics / Q000378:metabolism*; D005260:Female; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D018014:Gene Transfer Techniques; D017930:Genes, Reporter / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D008819:Mice, Nude; D009928:Organ Specificity; D010725:Phosphocreatine / Q000032:analysis / Q000378:metabolism; D014161:Transduction, Genetic
D002457:Cell Extracts; D007527:Isoenzymes; D010725:Phosphocreatine; D003402:Creatine Kinase; D003401:Creatine