C4d as a rejection marker in liver transplantation a valuable tool for differential diagnosis in hep

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C4d as a rejection marker in liver
transplantation - a valuable tool for
differential diagnosis in hepatitis-C patients
M.Schmeding1, A.Dankof 2, V.Krenn 2,
U.Neumann1, P.Neuhaus1 1 Department of General
and Transplantation Surgery, Charité University
Hospital Berlin, Germany 2 Department of
Pathology, Charité University Hospital Berlin
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Background

• In renal transplantation humoral components play
  a greater role in rejection mechanisms compared
  to liver transplants. This can be measured by C4d
  expression.
• It has recently been shown that B-cell activation
  is involved in rejection after OLT
• (Moeller et al. Virchows Archives 2005)
• (Dankof, Schmeding et al. Virchows Archives, in
  press)
• Discrimination of recurrent hepatitis-C and acute
  rejection in biopsy specimen is not reliable

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Aim of the study

• Are B-cell related mechanisms regularly involved
  in acute rejection after liver transplantation?
• May C4d then serve as a marker for differential
  diagnosis in distinguishing rejection and
  HCV-reinfection in hepatitis-C patients?

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Patients and Methods I

• Retrospective C4d staining of 97 liver biopsies
  from LTX patients
• Biopsies as protocol biopsies ½, 1 or 3 years
  after LTX or for acute rejection /
  HCV-reinfection
• Immunosuppression based on CNI- and steroids
• 3 groups 34 rejection cases
• 34 hepatitis-C recurrence cases
• 29 controls

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Patients and Methods II

• C4d staining by immunohistochemistry and
  immunofluorescence of parafinized liver tissue
• Evaluation by two independent and blinded
  pathologists (A.D., V.K.)
• Classification of C4d expression as focal,
  moderate or diffuse only moderate and
  diffuse cases were regarded as positive

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C4d staining (immunohistology)
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C4d staining (immunofluorescence)
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Results I

• Group I (AR) C4d pos. staining in 23/34 (67,7)
• Group II (RE-HCV) C4d pos. in 4/34 (11,8),
  (3 of 4 pos. patients treated with
  interferone)
• Group III (control) C4d pos. in 2/29 (6,9)
• Significantly increased C4d expression in
  acute rejection compared to recurrent HCV-cases
  (p lt 0.001, RR 15.68) and controls (p lt 0.001,
  RR 28.5).

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C4d expression ()
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Results II

• Among 34 rejection patients 9 were HCV-positive.
  
• In 6 of these 9 C4d confirmed AR.
• C4d negativity and retrospective clinical
  evaluation identified false diagnosis of
  rejection in 4/4 HCV-patients.
• 2 patients with histologically diagnosed early
  HCV-recurrence received a second biopsy after
  clinical deterioration then showing AR.
• Both cases tested positive for C4d in initial
  biopsies.

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Discussion I

• Humoral response mechanisms can be detected in
  the majority of acute cellular rejections after
  liver transplantation (67) marked by C4d
  expression in periportal tracts of liver
  specimen.
• This is supported by presence of plasma cells,
  macrophages and complement factors in AR liver
  tissue.
• (Krukemeyer et al. Transpl. 2004 78)
• C4d staining may serve to distinguish between
  acute rejection and hepatitis-C recurrence.

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Discussion II

• 3 of 4 patients with C4d positivity in the
  HCV-recurrence group had previously received
  interferone therapy.
• This might suggest activation of the complement
  system by interferone treatment and interference
  with C4d based diagnosis.
• (Baid et al., Am. J. of Transpl. 2003 3
  74-78)
• The reasons for C4d positivity among the
  remaining HCV patient and the 2 control
  individuals (cryptogenic cirrhosis and cystic LD)
  remain unclear.

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Conclusion

• For the first time a significant involvement of
  the activated classical complement pathway and
  the presence of C4d could be demonstrated in
  acute cellular rejection after liver
  transplantation.
• Especially in HCV patients with the difficult
  task of distinguishing between acute rejection
  and acute hepatitis-C recurrence C4d may be of
  great and rather specific value for differential
  diagnosis.
• This appears not to be valid for patients
  undergoing interferone treatment.