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CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
54
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION SUK SEO, MD NATALIE TOROK, MD Division of Gastroenterology and Hepatology, University of California Davis Medical Center, 4150 V Street, Suite 3500, Sacramento, CA, 95817, USA
HISTORICAL NOTES CRYOPRECIPITATION AUTOANTIGEN AUTOANTIBODY METHODS OF DETECTION PREVALENCE/PROGNOSIS TREATMENT TAKE-HOME MESSAGES REFERENCES
ABSTRACT Cryoglobulins are a mixture of immunoglobulins and complement components that precipitate at temperature lower than 37 C. Cryoglobulinaemia (CG) means the presence of cryoglobulins in a patient’s serum, but it is also referring to an inflammatory syndrome that generally involves small to medium vessel vasculitis due to cryoglobulin containing immune complexes. Prior to the identification of hepatitis C virus (HCV) in 1989, CG was largely termed ‘essential’ in patients who do not have associated lymphoproliferative disease or autoimmune disease. It is now recognized that up to 90% of patients with clinically evident CG have chronic HCV infection. The role of HCV in pathogenesis of CG and the treatment options are discussed here.
HISTORICAL NOTES In 1974, Brouet et al. [1] devised the current system of classification of the types of cryoglobulinaemia based upon its immunochemical composition and reported frequency of each type (Table 54.1). Autoantibodies, 2/e Copyright © 2007, Elsevier, B.V. All rights reserved.
437
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CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
TABLE 54.1 Brouet Classification for Cryoglobulinaemia Type I II III
Prevalence (%) 25 25 50
Immunoglobulins Monoclonal IgM, Monoclonal IgG Monoclonal IgM Polyclonal IgM
Type I cryoglobulinaemia is most often seen in patients with underlying lymphoproliferative disorders such as multiple myeloma, Waldenstrom’s macroglobulinaemia, or chronic lymphocytic leukemia. This type classically produces signs related to hyperviscosity. Although Raynaud’s syndrome and transient cold-induced digital ischemia are more common, serious thrombotic events may also occur involving cardiovascular, renal or neurological systems. Most cases of Type II CG are associated with chronic hepatitis C virus (HCV) infection. In such patients, antiHCV antibody and HCV RNA are found in high concentration of cryoprecipitate as well as the vessel walls of affected organ. In Type III, both the IgG and the rheumatoid factor (RF) IgM are polyclonal. Although this type is sometimes seen in chronic autoimmune disorders as well as hematological malignancies, the majority of the Type III CG patients also have chronic HCV infection.
CRYOPRECIPITATION The crucial point in the cryoprecipitating process is the production of IgM-RF molecules. In the presence of IgG molecules with anticore activity and IgM-RF activity, the HCV core protein becomes insoluble at cold temperatures. If IgM-RF/HCV core protein is mixed with non-specific IgG, cold-dependant insolubility does not occur, which suggests that binding of IgG with anticore activity causes molecular conformation changes resulting in increased precipitability [2]. Complement binding to immune complexes also has a role in cryoprecipitation: C1q protein binding activity is increased in the cryoprecipitates. This may trigger the modulation of the T-cell immune response via the globular domain of the C1q receptor (C1qR). In addition, since C1qR is widely expressed in endothelial cells, HCV core protein containing immune complexes could bind to the vascular surface contributing to the development of vasculitis (Figure 54.1) [3, 4].
AUTOANTIGEN Because cryglobulins consist of autoantibodies that take on antigenic properties during the process of forming immune complexes, the distinction of autoantigen and autoantibody becomes arbitrary. In Type I CG, monoclonal IgG or less commonly IgM, produced by dominant B-cell lines with underlying lymphoproliferative disease, can serve as both autoantigen and autoantibody by self-aggregation via Fc-Fc interaction. IgM Fab fragments can also precipitate by themselves. Classical formation of cryoglobulin in Type II or Type III CG involves interaction between Fab fragments of IgM with RF activity and Fc or Fab segment of IgGs.
439
AUTOANTIBODY
HCV core protein IgG
IgM
Clq
Clq receptor
Endothelial cells FIGURE 54.1 Proposed model of the cryoprecipitating immune complex in HCV-related cryoglobulinaemia. From Sansonno et al., Lancet Infect Disease, 2005 [2].
AUTOANTIBODY Chronic HCV infection occurs due to the host’s inability to mount effective virusspecific T-cell response and viral factors that allow immune escape via mutations at the antigenic site. In this setting, non-organ specific autoantibodies are frequently found (antinuclear antibodies with titer >1:40 in 21%, anti-smooth muscle antibody with titer >1:40 in 21% of patients, and anti-liver kidney microsomal antibodies in 5%) [5]. Unregulated production of other non-neutralizing antibodies in the setting of uncontrolled HCV infection may contribute to the development of immune complexes. Many disease states linked with CG share common characteristics in that that they are associated with induction of B-cell hyperactivation. In cases associated with HCV, the B-cell hyperactivation and production of autoantibodies may be initiated as the HCV envelope protein E2 binds to CD81 [6]. CD81 is a viral receptor present on B lymphocytes. The E2-CD81 interaction induces activation and expansion of peripheral CD5+ cells [7]. The CD5+ B cells are considered the major source of the IgM-RF, and they are also prone to other autoantibody production. With persistent stimulation, a dominant monoclonal IgM may emerge. Most of the monoclonal IgM-RF is of an unusual idiotype (Wa), and the B cell clones producing monoclonal IgM show evidence of ongoing somatic hypermutation [6]. When postulating a model for development of CG it is important to remember a caveat that CG is also seen in HCV negative patients or patients who have achieved complete HCV clearance. In addition, not all patients with HCV infection produce cryoglobulins. Furthermore, clinical recovery from CG is observed in patients with persistent viral replication. Rarely, CG is also seen in some patients with hepatitis B virus, Epstein-Barr virus and human immunodeficiency virus, and human T-cell lymphotropic virus-1 infection. In such cases, how cryoglobulins are produced and which antigen triggers this process, is still largely speculative.
440
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
METHODS OF DETECTION The serum concentration of cryoglobulins can be measured semiquantitatively. Blood is collected at a fasting state because lipids may interfere with the test. After it is placed in a warm tube and centrifuged, the serum is cooled to 4 C, and then observed for precipitation over a 48–72 hour period. The tube is centrifuged in the cold and the percent precipitate as a function of 1 ml serum volume (called the cryocrit) can be measured. Positive cryocrit corroborates the diagnosis; however, there is no correlation between the cryocrit value and the severity of end organ involvement. The type of the abnormal protein (mono- or polyclonality of IgM or IgG) can be determined by immunofixation electrophoresis, enzyme-linked immunosorbent assay (ELISA) or other specific immunological assays. The HCV core antigen may be detected using ELISA. Measurement of immunologic parameters such as IgM, IgG, and C3 and C4 fractions, RF activity, C1q- and C1q-binding activity can be monitored by chromatography and nephelometry. In patients where the diagnosis is in doubt, direct immunohistochemistry on biopsies (purpuric skin, nerve, renal biopsy) can often confirm the cryoglobulinaemia diagnosis.
PREVALENCE/PROGNOSIS With clinical manifestation of CG occurring in 10–15% of patients with HCV, it is now recognized that CG is the most common extrahepatic complication in patients with HCV. In those patients diagnosed with mixed CG, 40–98% have HCV [8]. Infection with hepatitis B is implicated in less than 5% of patients with CG. The prevalence of CG is increased with prolonged duration of liver disease and with the presence of cirrhosis [9]. The overall prevalence of CG is not known, but it is probably underestimated. In addition, there is a high rate of false negative serologic HCV tests in CG. Testing for HCV-RNA by PCR could increase diagnostic accuracy. Finding persistently elevated cryocrit (>1% for 3–6 months) also corroborates the diagnosis. Mortality and morbidity in patients with cryoglobulinaemia often depend on co-morbidities, but patients with renal involvement and lymphoproliferative disorder have worse prognosis overall. A meta-analysis of 19 studies published between 1994 and 2001 identified that CG was associated with increased risk of cirrhosis and higher overall mortality [9].
TREATMENT Because HCV has a clear biological role in pathogenesis of CG in most patients, one must consider eradicating HCV with antiviral therapy. Interferon-based treatment reduces viral replication rate (even without complete or permanent elimination), inhibits lymphocyte proliferation and immunoglobulin synthesis, and improves immune complex clearance by enhancing macrophage activity. In terms of clinical response, the effectiveness of interferon-alpha is comparable to that observed in the management of HCV without CG. In a recent randomized controlled study with eighteen patients affected by mixed CG, PEG-interferon alfa-2b in combination
441
REFERENCES
with ribavirin showed efficacy and safety for initial treatment of HCV-associated CG [10]. Cryoglobulin levels usually decrease and clinical symptoms of CG improve as the viral load decreases with interferon based treatment. Interferon-based therapy, however, should be considered with a caveat in that its direct immunomodulating effect can induce de novo autoimmune diseases during treatment. In addition, patients with clinically evident peripheral neuropathy, nephropathy, or skin ulcers, IFN based therapy may aggravate these manifestations. In such patients, first-line treatment usually is immunosuppression with steroids, with or without cyclophosphamide depending on the overall clinical severity. Since steroids can increase the HCV RNA levels and serum cryoglobulins, this drug should be administered in low doses and tapered quickly. In the most severe cases, removal of circulating immune complexes by plasma exchange may be useful, particularly in active cryoglobulinemic nephropathy. The latest development against the abnormal B-cell clone driven by HCV is the anti-CD 20 monoclonal antibody, rituximab. This agent has previously been shown activity in B-cell lymphomas and autoimmune disorders. Recent studies showed that treatment with rituximab resulted in a significant and rapid improvement of clinical signs and end organ function, and a decline of RF activity and cryocrit in most patients with CG, including patients resistant to IFN. However, because rituximab decreases anti-HCV antibody titers and increases viremia, long term effect of rituximab on the liver disease is not known [11].
TAKE-HOME MESSAGES • B-cell clonal expansion is hallmark of all types of CG. • HCV is an important etiological factor in most CG, and HCV directly induces clonal expansion of IgM-RF-synthesizing B-cells. • HCV core protein seems to be an important factor for formation of CG, engagement of CG onto the vascular endothelium, and propagation of B-cell clonal expansion. • Treatment for HCV-associated CG should be tailored to individual patients. • New modalities such as pegylated interferon and rituximab may further improve treatment responses for clinically significant CG.
REFERENCES 1. Brouet J, Clauvel J, Danon F. Biologic and clinical significance of cryoglobulins. A report of 86 cases. Am J Med 1974; 57(5): 775–88. 2. Sansonno D, Lauletta G, Nisi L. Non-enveloped HCV core protein as constitutive antigen of coldprecipitable immune complexes in type II mixed cryoglobulinaemia. Clin Exp Immunol 2003; 133: 275–82. 3. Sansonno D, Dammacco F. Hepatitis C virus, cryoglobulinemia, and vasculitis: Immune complex relations. Lancet Infect Dis 2005; 5: 227–36. 4. Yao Z, Nguyen D, Hiotellis A, Hahn Y. Hepatitis C virus core protein inhibits human T lymphocyte responses by a complement-dependent regulatory pathway. J Immunol 2001; 167: 5264–72. 5. Berenger M, Wright T. Viral Hepatitis, Sleisenger & Fordtran’s Gastrointestinal and Liver Disease, 7th edn. University of Texas Southwestern, Dallas: W.B. Saunders Co, 2002; 2: 1309. 6. Machida K, Cheng K, Pavio N, Sung V, Lai M. Hepatitis C virus E2-CD81 interaction induces hypermutation of the immunoglobulin gene in B cells. J Virol 2005 Jul; 79(13): 8079–89. 7. Curry M, Golden-Mason L, Doherty D. Expansion of innate CD5 pos B cells expressing high levels of CD81 in hepatitis C virus infected liver. J Hepatol 2003; 38: 642–50.
442
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
8. Ali A, Zein N. Hepatitis C infection: A systemic disease with extrahepatic manifestations. Cleve Clin J Med 2005; 72(11): 1005–8, 1010–14. 9. Kayali Z, Buckwold V, Zimmerman B, Schidt W. Cryoglobulinemia, and cirrhosis: A meta-analysis. Hepatology 2002; 36: 9978–85. 10. Mazzaro C, Zorat F, Caizzi M. Treatment with peg-interferon alfa-2b and ribavirin of hepatitis C virus-associated mixed cryoglobulinemia: A pilot study. J Hepatol 2005; 42: 632–8. 11. Sansonno D, Valli De Re, Lauletta G. Monoclonal antibody treatment of mixed cryoglobulinemia resistant to interferon-alpha with an anti-CD20. Blood 2003; 101: 3818–26.
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION SUK SEO, MD NATALIE TOROK, MD Division of Gastroenterology and Hepatology, University of California Davis Medical Center, 4150 V Street, Suite 3500, Sacramento, CA, 95817, USA
HISTORICAL NOTES CRYOPRECIPITATION AUTOANTIGEN AUTOANTIBODY METHODS OF DETECTION PREVALENCE/PROGNOSIS TREATMENT TAKE-HOME MESSAGES REFERENCES
ABSTRACT Cryoglobulins are a mixture of immunoglobulins and complement components that precipitate at temperature lower than 37 C. Cryoglobulinaemia (CG) means the presence of cryoglobulins in a patient’s serum, but it is also referring to an inflammatory syndrome that generally involves small to medium vessel vasculitis due to cryoglobulin containing immune complexes. Prior to the identification of hepatitis C virus (HCV) in 1989, CG was largely termed ‘essential’ in patients who do not have associated lymphoproliferative disease or autoimmune disease. It is now recognized that up to 90% of patients with clinically evident CG have chronic HCV infection. The role of HCV in pathogenesis of CG and the treatment options are discussed here.
HISTORICAL NOTES In 1974, Brouet et al. [1] devised the current system of classification of the types of cryoglobulinaemia based upon its immunochemical composition and reported frequency of each type (Table 54.1). Autoantibodies, 2/e Copyright © 2007, Elsevier, B.V. All rights reserved.
437
438
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
TABLE 54.1 Brouet Classification for Cryoglobulinaemia Type I II III
Prevalence (%) 25 25 50
Immunoglobulins Monoclonal IgM, Monoclonal IgG Monoclonal IgM Polyclonal IgM
Type I cryoglobulinaemia is most often seen in patients with underlying lymphoproliferative disorders such as multiple myeloma, Waldenstrom’s macroglobulinaemia, or chronic lymphocytic leukemia. This type classically produces signs related to hyperviscosity. Although Raynaud’s syndrome and transient cold-induced digital ischemia are more common, serious thrombotic events may also occur involving cardiovascular, renal or neurological systems. Most cases of Type II CG are associated with chronic hepatitis C virus (HCV) infection. In such patients, antiHCV antibody and HCV RNA are found in high concentration of cryoprecipitate as well as the vessel walls of affected organ. In Type III, both the IgG and the rheumatoid factor (RF) IgM are polyclonal. Although this type is sometimes seen in chronic autoimmune disorders as well as hematological malignancies, the majority of the Type III CG patients also have chronic HCV infection.
CRYOPRECIPITATION The crucial point in the cryoprecipitating process is the production of IgM-RF molecules. In the presence of IgG molecules with anticore activity and IgM-RF activity, the HCV core protein becomes insoluble at cold temperatures. If IgM-RF/HCV core protein is mixed with non-specific IgG, cold-dependant insolubility does not occur, which suggests that binding of IgG with anticore activity causes molecular conformation changes resulting in increased precipitability [2]. Complement binding to immune complexes also has a role in cryoprecipitation: C1q protein binding activity is increased in the cryoprecipitates. This may trigger the modulation of the T-cell immune response via the globular domain of the C1q receptor (C1qR). In addition, since C1qR is widely expressed in endothelial cells, HCV core protein containing immune complexes could bind to the vascular surface contributing to the development of vasculitis (Figure 54.1) [3, 4].
AUTOANTIGEN Because cryglobulins consist of autoantibodies that take on antigenic properties during the process of forming immune complexes, the distinction of autoantigen and autoantibody becomes arbitrary. In Type I CG, monoclonal IgG or less commonly IgM, produced by dominant B-cell lines with underlying lymphoproliferative disease, can serve as both autoantigen and autoantibody by self-aggregation via Fc-Fc interaction. IgM Fab fragments can also precipitate by themselves. Classical formation of cryoglobulin in Type II or Type III CG involves interaction between Fab fragments of IgM with RF activity and Fc or Fab segment of IgGs.
439
AUTOANTIBODY
HCV core protein IgG
IgM
Clq
Clq receptor
Endothelial cells FIGURE 54.1 Proposed model of the cryoprecipitating immune complex in HCV-related cryoglobulinaemia. From Sansonno et al., Lancet Infect Disease, 2005 [2].
AUTOANTIBODY Chronic HCV infection occurs due to the host’s inability to mount effective virusspecific T-cell response and viral factors that allow immune escape via mutations at the antigenic site. In this setting, non-organ specific autoantibodies are frequently found (antinuclear antibodies with titer >1:40 in 21%, anti-smooth muscle antibody with titer >1:40 in 21% of patients, and anti-liver kidney microsomal antibodies in 5%) [5]. Unregulated production of other non-neutralizing antibodies in the setting of uncontrolled HCV infection may contribute to the development of immune complexes. Many disease states linked with CG share common characteristics in that that they are associated with induction of B-cell hyperactivation. In cases associated with HCV, the B-cell hyperactivation and production of autoantibodies may be initiated as the HCV envelope protein E2 binds to CD81 [6]. CD81 is a viral receptor present on B lymphocytes. The E2-CD81 interaction induces activation and expansion of peripheral CD5+ cells [7]. The CD5+ B cells are considered the major source of the IgM-RF, and they are also prone to other autoantibody production. With persistent stimulation, a dominant monoclonal IgM may emerge. Most of the monoclonal IgM-RF is of an unusual idiotype (Wa), and the B cell clones producing monoclonal IgM show evidence of ongoing somatic hypermutation [6]. When postulating a model for development of CG it is important to remember a caveat that CG is also seen in HCV negative patients or patients who have achieved complete HCV clearance. In addition, not all patients with HCV infection produce cryoglobulins. Furthermore, clinical recovery from CG is observed in patients with persistent viral replication. Rarely, CG is also seen in some patients with hepatitis B virus, Epstein-Barr virus and human immunodeficiency virus, and human T-cell lymphotropic virus-1 infection. In such cases, how cryoglobulins are produced and which antigen triggers this process, is still largely speculative.
440
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
METHODS OF DETECTION The serum concentration of cryoglobulins can be measured semiquantitatively. Blood is collected at a fasting state because lipids may interfere with the test. After it is placed in a warm tube and centrifuged, the serum is cooled to 4 C, and then observed for precipitation over a 48–72 hour period. The tube is centrifuged in the cold and the percent precipitate as a function of 1 ml serum volume (called the cryocrit) can be measured. Positive cryocrit corroborates the diagnosis; however, there is no correlation between the cryocrit value and the severity of end organ involvement. The type of the abnormal protein (mono- or polyclonality of IgM or IgG) can be determined by immunofixation electrophoresis, enzyme-linked immunosorbent assay (ELISA) or other specific immunological assays. The HCV core antigen may be detected using ELISA. Measurement of immunologic parameters such as IgM, IgG, and C3 and C4 fractions, RF activity, C1q- and C1q-binding activity can be monitored by chromatography and nephelometry. In patients where the diagnosis is in doubt, direct immunohistochemistry on biopsies (purpuric skin, nerve, renal biopsy) can often confirm the cryoglobulinaemia diagnosis.
PREVALENCE/PROGNOSIS With clinical manifestation of CG occurring in 10–15% of patients with HCV, it is now recognized that CG is the most common extrahepatic complication in patients with HCV. In those patients diagnosed with mixed CG, 40–98% have HCV [8]. Infection with hepatitis B is implicated in less than 5% of patients with CG. The prevalence of CG is increased with prolonged duration of liver disease and with the presence of cirrhosis [9]. The overall prevalence of CG is not known, but it is probably underestimated. In addition, there is a high rate of false negative serologic HCV tests in CG. Testing for HCV-RNA by PCR could increase diagnostic accuracy. Finding persistently elevated cryocrit (>1% for 3–6 months) also corroborates the diagnosis. Mortality and morbidity in patients with cryoglobulinaemia often depend on co-morbidities, but patients with renal involvement and lymphoproliferative disorder have worse prognosis overall. A meta-analysis of 19 studies published between 1994 and 2001 identified that CG was associated with increased risk of cirrhosis and higher overall mortality [9].
TREATMENT Because HCV has a clear biological role in pathogenesis of CG in most patients, one must consider eradicating HCV with antiviral therapy. Interferon-based treatment reduces viral replication rate (even without complete or permanent elimination), inhibits lymphocyte proliferation and immunoglobulin synthesis, and improves immune complex clearance by enhancing macrophage activity. In terms of clinical response, the effectiveness of interferon-alpha is comparable to that observed in the management of HCV without CG. In a recent randomized controlled study with eighteen patients affected by mixed CG, PEG-interferon alfa-2b in combination
441
REFERENCES
with ribavirin showed efficacy and safety for initial treatment of HCV-associated CG [10]. Cryoglobulin levels usually decrease and clinical symptoms of CG improve as the viral load decreases with interferon based treatment. Interferon-based therapy, however, should be considered with a caveat in that its direct immunomodulating effect can induce de novo autoimmune diseases during treatment. In addition, patients with clinically evident peripheral neuropathy, nephropathy, or skin ulcers, IFN based therapy may aggravate these manifestations. In such patients, first-line treatment usually is immunosuppression with steroids, with or without cyclophosphamide depending on the overall clinical severity. Since steroids can increase the HCV RNA levels and serum cryoglobulins, this drug should be administered in low doses and tapered quickly. In the most severe cases, removal of circulating immune complexes by plasma exchange may be useful, particularly in active cryoglobulinemic nephropathy. The latest development against the abnormal B-cell clone driven by HCV is the anti-CD 20 monoclonal antibody, rituximab. This agent has previously been shown activity in B-cell lymphomas and autoimmune disorders. Recent studies showed that treatment with rituximab resulted in a significant and rapid improvement of clinical signs and end organ function, and a decline of RF activity and cryocrit in most patients with CG, including patients resistant to IFN. However, because rituximab decreases anti-HCV antibody titers and increases viremia, long term effect of rituximab on the liver disease is not known [11].
TAKE-HOME MESSAGES • B-cell clonal expansion is hallmark of all types of CG. • HCV is an important etiological factor in most CG, and HCV directly induces clonal expansion of IgM-RF-synthesizing B-cells. • HCV core protein seems to be an important factor for formation of CG, engagement of CG onto the vascular endothelium, and propagation of B-cell clonal expansion. • Treatment for HCV-associated CG should be tailored to individual patients. • New modalities such as pegylated interferon and rituximab may further improve treatment responses for clinically significant CG.
REFERENCES 1. Brouet J, Clauvel J, Danon F. Biologic and clinical significance of cryoglobulins. A report of 86 cases. Am J Med 1974; 57(5): 775–88. 2. Sansonno D, Lauletta G, Nisi L. Non-enveloped HCV core protein as constitutive antigen of coldprecipitable immune complexes in type II mixed cryoglobulinaemia. Clin Exp Immunol 2003; 133: 275–82. 3. Sansonno D, Dammacco F. Hepatitis C virus, cryoglobulinemia, and vasculitis: Immune complex relations. Lancet Infect Dis 2005; 5: 227–36. 4. Yao Z, Nguyen D, Hiotellis A, Hahn Y. Hepatitis C virus core protein inhibits human T lymphocyte responses by a complement-dependent regulatory pathway. J Immunol 2001; 167: 5264–72. 5. Berenger M, Wright T. Viral Hepatitis, Sleisenger & Fordtran’s Gastrointestinal and Liver Disease, 7th edn. University of Texas Southwestern, Dallas: W.B. Saunders Co, 2002; 2: 1309. 6. Machida K, Cheng K, Pavio N, Sung V, Lai M. Hepatitis C virus E2-CD81 interaction induces hypermutation of the immunoglobulin gene in B cells. J Virol 2005 Jul; 79(13): 8079–89. 7. Curry M, Golden-Mason L, Doherty D. Expansion of innate CD5 pos B cells expressing high levels of CD81 in hepatitis C virus infected liver. J Hepatol 2003; 38: 642–50.
442
CRYOGLOBULINS AND CRYOGLOBULINS SECONDARY TO HEPATITIS C VIRUS INFECTION
8. Ali A, Zein N. Hepatitis C infection: A systemic disease with extrahepatic manifestations. Cleve Clin J Med 2005; 72(11): 1005–8, 1010–14. 9. Kayali Z, Buckwold V, Zimmerman B, Schidt W. Cryoglobulinemia, and cirrhosis: A meta-analysis. Hepatology 2002; 36: 9978–85. 10. Mazzaro C, Zorat F, Caizzi M. Treatment with peg-interferon alfa-2b and ribavirin of hepatitis C virus-associated mixed cryoglobulinemia: A pilot study. J Hepatol 2005; 42: 632–8. 11. Sansonno D, Valli De Re, Lauletta G. Monoclonal antibody treatment of mixed cryoglobulinemia resistant to interferon-alpha with an anti-CD20. Blood 2003; 101: 3818–26.