Inflamed glomeruli-specific gene activation that uses recombinant adenovirus with the Cre/loxP system

J Am Soc Nephrol. 2001 Nov;12(11):2330-2337. doi: 10.1681/ASN.V12112330.

Abstract

The authors previously reported that bone marrow-derived CD11b(+)CD18(+) cells could be used as a vehicle to deliver foreign genes into inflamed glomeruli and that this vehicle cell (v-cell) could retard the progression of nephritis by delivering anti-inflammatory molecules. As a next step, the authors tried to establish a switching system by which v-cells are activated only at the inflamed glomeruli. A recombinant adenovirus (Ad) that expressed Cre recombinase under the control of the interleukin-1 beta (IL-1 beta) promoter (AxIL-1pr/Cre) was constructed and transfected into v-cells. After confirming that AxIL-1pr/Cre expresses Cre by lipopolysaccharide (LPS) treatment, AxIL-1pr/Cre was infected together with another Ad bearing a switching reporter unit in which the LacZ gene is activated under the control of the CAG promoter by the Cremediated excisional deletion of interposed stuffer DNA. Only a negligible number of double-infected (Cre/loxPCAG) cells expressed LacZ. This number, however, was significantly increased by LPS, which suggests that LPS-induced Cre effectively deletes the stuffer DNA, which allows for a complete CAG promoter. DBA/2j mice were then transplanted with Cre/loxPCAG cells via a tail vein and treated with anti-glomerular basement membrane (GBM) serum. To trace the transplanted cells, marker v-cells, infected with AxCANLacZ to constitutively express the LacZ gene, were also used. Although transplanted cells expressing LacZ collected in the spleen independent of anti-GBM treatment, they did not express the LacZ gene in the mice transplanted with Cre/loxPCAG cells. On the other hand, transplanted cells were recruited in the glomeruli and expressed the LacZ gene upon anti-GBM treatment. These results suggested that only the v-cells recruited in the glomeruli could be switched on and activate foreign genes.

MeSH terms

  • Adenoviridae / genetics
  • Animals
  • Base Sequence / genetics
  • Basement Membrane / immunology
  • Bone Marrow Cells / drug effects
  • Bone Marrow Cells / physiology
  • Bone Marrow Transplantation
  • Cells, Cultured
  • Female
  • Gene Expression / drug effects
  • Gene Expression Regulation*
  • Gene Transfer Techniques*
  • Genes, Switch
  • Glomerulonephritis / genetics*
  • Immune Sera / pharmacology
  • Integrases / genetics
  • Kidney Glomerulus / immunology
  • Lac Operon
  • Lipopolysaccharides / pharmacology
  • Mice
  • Mice, Inbred DBA
  • Promoter Regions, Genetic / physiology
  • Recombination, Genetic
  • Transcriptional Activation
  • Transfection
  • Viral Proteins / genetics

Substances

  • Immune Sera
  • Lipopolysaccharides
  • Viral Proteins
  • Cre recombinase
  • Integrases