No evidence for abnormal immune activation in peripheral blood T cells in patients with hepatitis C virus (HCV) infection with or without cryoglobulinaemia. Multivirc Group.

The aim of this study was to investigate the peripheral blood lymphocyte (PBL) phenotypes and T cell repertoire in patients with HCV infection, with or without mixed cryoglobulinaemia (MC). The patients were: Group 1, 23 patients with HCV infection and MC; Group 2, 14 patients with HCV infection but without MC; Group 3, 10 patients with symptomatic essential MC. Twenty healthy blood donors were used as controls. Blood lymphocyte counts were determined, and flow cytometry was used to measure proportions of B cells (CD19+), natural killer (NK) cells (CD16+CD56+), T cells (CD3+), CD4+ T cell subsets (memory CD4+CD45RO+; naive CD4+CD45RO-; Th0/Th2CD4+CD7-; activated CD4+CD25+), and CD8+ T cell subsets (immunoregulatory CD8+CD57+; cytotoxic CD8+S6F1+, activated CD8+CD25+). Bias in the usage of T cell receptor (TCR) Vbeta chains was studied in a subgroup of 10 representative patients of Group 1 using a polymerase chain reaction (PCR) analysis of the Vbeta segments with a series of 20 oligonucleotides specific for the Vbeta families. The three groups were comparable for blood lymphocyte counts, and we observed no abnormal repartition of the following PBL subsets: T cells (CD3+), CD4+ and CD8+ subpopulations, B cells (CD19+), and the NK cells (CD16+56+). In none of the groups could we observe lymphocyte ex vivo activation as assessed by the normal expression of the activation cell markers: CD25 on CD4+ or CD8+ T cells, or CD5 on B cells. The repartition of naive and memory (CD45RO-/RO+) CD4+ T cells was normal and we did not observe any amplification of the CD4+CD7- T cell subset differentiated in vivo in Th0/Th2 cells. There was no significant amplification of cytotoxic (SF6+) and immunoregulatory (CD57+) CD8+ T cells in HCV patients with or without MC. Finally, the usage of Vbeta families in the TCR repertoire was normal in the patients tested. In patients with chronic HCV infection, with or without MC, we did not find any significant expansion or abnormal activation of T, B and NK cell subsets, dysbalance of the naive/memory subsets, or expansion of the Th0/Th2 subpopulation. These findings differ from the profound immune alterations that are observed in other chronic infections such as HIV or Epstein-Barr virus. Although this study was restricted to the peripheral blood, it suggests that in chronic HCV infection, MC is not the consequence of a chronic activation or dysregulation of the peripheral blood immune cells.

Mixed cryoglobulinemia and hepatitis C virus.

BACKGROUND: Mixed cryoglobulinemia (MC) is frequently associated with clinical and biological evidence of liver disease and has recently been reported in cases of hepatitis C virus (HCV) infection. The aim of this study was to assess prospectively in a large series of MC patients: (1) the prevalence of HCV markers (anti-HCV antibodies and HCV RNA in serum and cryoprecipitate); (2) the main clinical, biologic and liver histologic features in patients with or without HCV infection. PATIENTS: One hundred fifteen consecutive unselected MC patients were studied: 45% had well-defined underlying diseases ("nonessential" MC). Fifty-five percent with no cause of MC were considered to have "essential" MC and were subjected to in-depth examination. METHODS: Patients were considered to have MC if two successive determinations of their serum cryoglobulin level were above 0.05 g/L. Anti-HCV antibodies (Ab) were detected in all patients by second-generation tests (ELISA, RIBA). We also looked for HCV RNA sequences amplified by polymerase chain reaction (PCR) in the sera and cryoprecipitates of 39 patients; HBs antigen, anti-HBs Ab and anti-HBc Ab in all patients; and HBV DNA in 20 sera and 17 cryoprecipitates. Quantitative HCV Ab and RNA studies were performed on whole serum, cryoprecipitates, and supernatants. Clinical features were recorded retrospectively for each patient. Liver biopsies from 23 anti-HCV Ab-positive and 7 anti-HCV Ab-negative patients were examined histologically, with qualitative and quantitative analysis. RESULTS: Anti-HCV Ab were found in 47/115 (41%) patients by ELISA and RIBA: 33/63 (52%) essential MC and 14/52 (27%) nonessential MC. Among the 63 essential MC patients, the 33 anti-HCV Ab-positive (Group 1) were compared to the 30 anti-HCV Ab-negative patients (Group 2). Group 1 patients had more cutaneous involvement (Raynaud's phenomenon, purpura, livedo, distal ulcers, or gangrenous changes) (17 versus 5: p = 0.004), higher alanine aminotransferase levels (110 +/- 22 versus 41 +/- 10 IU; p < 0.005), higher serum cryoglobulin levels (0.35 +/- 0.07 versus 0.12 +/- 0.04 g/L; p = 0.01), lower CH50 (28 +/- 3 versus 44 +/- 2 CH50/mL; p = 0.0001) and lower C4 levels (0.20 +/- 0.02 versus 0.29 +/- 0.03 g/L; p < 0.04). The prevalence of HBV serum markers was low in both groups, and HBV DNA was never detected in any of the sera and cryoprecipitates tested. HCV RNA sequences were detected in 10/16 (63%) sera and 12/16 (75%) cryoprecipitates from Group 1 patients, whereas they were not in the sera or cryoprecipitates from 23 Group 2 patients. Using quantitative PCR, HCV RNA in cryoprecipitates was concentrated 20 to 100 times despite the absence of significant anti-HCV Ab concentration in these samples. Histologic examination of liver biopsies revealed a spectrum of lesions ranging from chronic active hepatitis to cirrhosis, but Knodell's score did not differ between the groups. CONCLUSION: (1) About 50% of the essential MC patients had anti-HCV Ab, and these patients had more severe cryoglobulinemia-associated clinical and biological signs; (2) HCV RNA sequences were found in the large majority of sera and cryoprecipitates from patients with essential MC and anti-HCV Ab and were more concentrated in cryoprecipitates than in supernatants. These results suggest a role for HCV in the pathogenesis of MC and indicate that many cases of essential MC may be secondary to HCV infection and thus nonessential.