T-cell clones derived by CD3 stimulation from hepatitis C virus explanted liver tissue are not representative of dominant clones present in vivo.
Liver tissue from hepatitis C virus (HCV)-related end-stage disease contains T-cell infiltrates. The goal of this study is to determine whether CD4 T-cell clones established in vitro using an antigen-independent technique from explanted liver tissue (n = 3) are representative of dominant clones present in vivo. T-cell receptor (TCR) use by intrahepatic CD4 T cells was assessed by spectratype analysis. Clones were established from single CD4 T cells by culturing in vitro with anti-CD3 and interleukin-2 (n > 25 per patient). TCR genes expressed by each clone were identified by sequencing. When identical clones were isolated, the original spectratype was analyzed further to determine whether the clone was a dominant T-cell expansion in vivo. Evidence for clonal expansions was found in all patients by spectratyping. T cells expressing the same TCRBV genes used for spectratyping were cloned in vitro. Duplicate clones expressing the same TCR genes were observed in 2 patients. Confirmation that clones established in vitro matched those present in vivo was obtained for 2 clones. Many dominant clones identified by spectratyping did not proliferate in vitro. Although spectratyping suggested the widespread accumulation of clonal expansions in HCV-related end-stage liver disease, clones established in vitro using anti-CD3 were poorly representative of dominant clones present in vivo. Although cloning with anti-CD3 has the advantage of generating T-cell clones not biased a priori toward a specific antigen, modified cloning strategies will need to be developed to expand those clones that appear dominant in end-stage organs.
Davey
M P
MP
Department of Veterans Affairs Medical Center, Oregon Health Sciences University, 3710 SW U.S. Veterans Hospital Rd., Bldg. 101, Rm. 502, Portland, OR 97201, USA. daveym@ohsu.edu
Rosen
H R
HR
Woody
C N
CN
Haley
D P
DP
Kurkinen
J
J
Lewinsohn
D M
DM
D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D017252:CD3 Complex / Q000276:immunology*; D002999:Clone Cells; D003584:Cytological Techniques; D005260:Female; D016174:Hepacivirus / Q000302:isolation & purification*; D006801:Humans; D066298:In Vitro Techniques; D008099:Liver / Q000473:pathology* / Q000821:virology*; D016031:Liver Transplantation; D008297:Male; D008875:Middle Aged; D013601:T-Lymphocytes / Q000187:drug effects* / Q000473:pathology*
D000911:Antibodies, Monoclonal; D017252:CD3 Complex