A novel approach to investigation of the pathogenesis of active minimal-change nephrotic syndrome using subtracted cDNA library screening.
Clinical and experimental observations suggest that minimal-change nephrotic syndrome (MCNS) results from T cell dysfunction, via unknown mechanisms. For the identification of genes that are potentially involved in MCNS, a subtractive cDNA library was constructed from cDNA from T cell-enriched peripheral blood mononuclear cells obtained from the same patient during relapse versus remission ("relapse minus remission"). This library was screened by differential hybridization with forward ("relapse minus remission") and reverse ("remission minus relapse") subtractive cDNAs probes, as well as unsubtracted probes from relapse and remission, and irrelevant nephrotic syndrome (membranous nephropathy). A total of 84 transcripts were isolated, of which 12 matched proteins of unknown function and 30 were unknown clones. Among the 42 known transcripts, at least 18 are closely involved in the T cell receptor-mediated complex signaling cascade, including genes encoding components of the T cell receptor and proteins associated with the cytoskeletal scaffold, as well as transcription factors. In particular, it was demonstrated that the expression levels of Fyb/Slap, L-plastin, and grancalcin were increased during relapse, suggesting that the integration of proximal signaling after T cell engagement involves the preferential recruitment of these cytoskeleton-associated proteins in MCNS. Because very low levels of interleukin-12 receptor beta2 mRNA were detected in relapse samples, the interleukin-12 signaling pathway might be defective, suggesting that, in MCNS, T cell activation evolves toward a T helper 2 phenotype. Therefore, the combination of subtractive cloning and differential screening constitutes an efficient approach to the identification of genes that are likely to be involved in the pathophysiologic processes of MCNS.
Sahali
Djillali
D
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.|*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Pawlak
André
A
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Valanciuté
Asta
A
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Grimbert
Philippe
P
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Lang
Philippe
P
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.|*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Remy
Philippe
P
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Bensman
Albert
A
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.|*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
Guellën
Georges
G
*INSERM Unit 99, Upress B JE 2129, and Adult Nephrology Service, Henri Mondor Hospital, University of Paris XII, Créteil, France, and Pediatric Nephrology Service, Armand Trousseau Hospital AP-HP, Paris, France.
D000328:Adult; D018076:DNA, Complementary / Q000032:analysis*; D015723:Gene Library; D005821:Genetic Techniques; D006801:Humans; D008297:Male; D009404:Nephrotic Syndrome / Q000235:genetics* / Q000503:physiopathology; D012008:Recurrence; D013601:T-Lymphocytes / Q000473:pathology
D018076:DNA, Complementary