Clinical Impact of Liver Biopsies in Liver Transplant Recipients

Abstract

Background

Percutaneous liver biopsy is an established diagnostic procedure for the assessment of liver pathologies. Limited data are available on the clinical impact of liver biopsies in liver transplant recipients.

Material/Methods

Liver transplant recipients undergoing liver biopsy between 2000 and 2013 were analyzed. Demographic characteristics and transplantation data were extracted from the transplantation database.

Results

A total of 496 liver biopsies were performed in 312 patients. The main biopsy indications were suspected rejection (66%, 327/496), protocol biopsy (22%, 108/496), and suspected recurrence of the primary disease (7%, 34/496). Histological findings showed acute cellular rejection in 36% (179/496), idiopathic chronic hepatitis in 28% (141/496), and normal histology in 11% (54/496). Liver biopsies in patients with clinically suspected rejection showed histological findings compatible with acute or chronic rejection in 46% (151/327). In 41% (205/496) of the patients, the immunosuppressive therapy was adjusted due to the biopsy result. For alanine-aminotransferase and bilirubin, significant differences were detected between baseline and week 4 and 12 after treatment modification (p<0.05).

Conclusions

Liver biopsies in liver transplant recipients have potential impact on the modification of the immunosuppressive therapy. The correlation between suspected rejection and histological findings is limited; therefore, a liver biopsy is indicated in unclear cases.

Background

Percutaneous liver biopsy is an established diagnostic procedure for the assessment of liver pathologies [1]. In patients after orthotopic liver transplantation (OLT), liver biopsy is used to detect acute or chronic rejection and recurrence of the underlying disease, and to evaluate the degree of fibrosis/stage of liver disease [2,3]. Although potentially life-threatening complications may occur, the reported incidence rate of complications ranges from 1.4% to 9% in recent publications and the procedure is generally well tolerated [4,5].

The selection of patients after OLT for liver biopsy is not standardized and the correlation between clinical suspicion of cellular rejection and histological findings is poor [6,7]. Data on the clinical impact and consequences of liver biopsies in patients after OLT in terms of treatment modification and response are sparse. These considerations prompted us to analyze our cohort of patients after OLT undergoing liver biopsy regarding indication of liver biopsy, treatment regimen, laboratory parameters, modification of treatment, and response to the initiated treatment change (among other factors).

Material and Methods

Liver transplant recipients presenting for liver biopsy to the endoscopic unit of Hannover Medical School between 2000 and 2013 were analyzed retrospectively. Pediatric patients were excluded from the study. In case of repetitive liver biopsies, all presentations were included in the analysis. To adjust for potential bias, further analyses with restriction to the first presentation for liver biopsy were performed. Moreover, the patient cohort was stratified by median time from OLT to liver biopsy and by ≤12 months (early biopsy) or >12 months (late biopsy) of liver biopsy after OLT. Demographic characteristics, transplantation data, immunosuppressive regimens, indication for biopsy, histological findings, treatment modification after liver biopsy, response to therapy, and laboratory parameters were extracted from the endoscopy and transplantation database. An escalation of immunosuppressive therapy was defined as addition of an immunosuppressive agent or dose escalation of the therapy. Dose reduction of the immunosuppressive therapy was defined as reduction of the existing therapy or de-escalation from triple to dual or dual to monotherapy.

Percutaneous liver biopsy was performed as described before after written informed consent was obtained [4]. Twelve or more portal tracts were required for liver biopsy specimens to be considered as adequate. The study was approved by the local institutional Ethics Committee (Ethics Committee of Hannover Medical School).

Statistical analysis

Data are expressed as number/percentages or mean ±SD or median with interquartile range (IQR) in case of non-normal distribution. All data were tested for normality (Shapiro-Wilk test, Kolmogorov-Smirnov test). Non-continuous parameters were analyzed by χ² test or Fisher’s exact test, as appropriate, and continuous parameters were analyzed by Mann-Whitney U test. In case of non-parametric paired data, Friedman test was used to identify differences in repeated measurements (e.g., laboratory values after treatment change). P values <0.05 were considered statistically significant. The software used was the SPSS Statistical Package (version 19.0, SPSS Inc, Chicago, IL).

Results

A total of 496 liver biopsies in 312 patients after OLT fulfilled the inclusion criteria and were retrospectively analyzed. The patients underwent OLT between 1985 and 2010. Demographic characteristics and transplantation data of the study group are presented in Table 1.

Table 1.

Patient characteristics and transplantation data. Demographic data, laboratory values, transplantation data and indications for orthotopic liver transplantation (OLT) from the study cohort (n=312).

Demographics	Patients after OLT (n=312) (number, percentage (IQR))
Gender (Male (M), Female (F))	M204, F108
Age (years)	46 (35–56)
Body mass index (kg/m2)	24.1 (21.4–27.5)
Laboratory values
Bilirubin (<17 μmol/L)	18 (12–31)
ALT (<35 U/L)	83 (35–155)
AST (<32 U/L)	53 (30–115)
GGT (<39 U/L)	141 (60–300)
AP (35–104 U/L)	179 (107–328)
CRP (<8 mg/L)	5 (2–8)
Transplantation data
Duct-to-duct anastomosis	220/312, 71%
Cold ischemia time (min)	572 (475–668)
Complete graft	277/312, 89%
Primary transplantation	253/312, 81%
Indication for liver transplantation
Primary sclerosing cholangitis	60/312, 19%
HBV ±HCC	37/312, 12%
HCV ±HCC	31/312, 10%
Acute liver failure	24/312, 8%
Alcohol-induced liver cirrhosis ±HCC	23/312, 7%
HBV/HDV ± HCC	18/312, 6%
Cryptogenic cirrhosis	17/312, 5%
Primary biliary cirrhosis	15/312, 5%
Autoimmune hepatitis	14/312, 5%
Wilson disease	14/312, 5%
HCC	11/312, 4%
Budd-Chiari syndrome	10/312, 3%
NASH-associated cirrhosis	8/312, 3%
Hemochromatosis	7/312, 2%
Cystic fibrosis	6/312, 2%
Polycystic liver disease	6/312, 2%
Amyloidosis	4/312, 1%
Others	7/312, 2%

ALT – alanine aminotransferase; AST – aspartate aminotransferase; GGT – gamma-glutamyl transferase; AP – alkaline phosphatase; CRP – C-reactive protein; HCC – hepatocellular carcinoma; HBV – hepatitis B virus infection; HCV – hepatitis C virus infection; HDV – hepatitis D virus infection; NASH – non-alcoholic steatohepatitis.

Main indications for OLT were viral hepatitis (86/312, 28%), primary sclerosing cholangitis (60/312, 19%), and acute liver failure (24/312, 8%) (Table 1). The biopsy indications were as follows: suspected rejection (66%, 327/496), protocol biopsy (22%, 108/496), suspected recurrence of the primary disease (7%, 34/496), unclear cholestasis (3%, 16/496), and other reasons (2%, 11/496). The median time interval from transplantation to liver biopsy was 27 months (IQR 8–70 months). Insufficient biopsy material was obtained in 11 (2.2%) of 496 liver biopsies (less than 12 portal tracts).

Histological findings showed acute cellular rejection in 36% (179/496), idiopathic chronic hepatitis in 28% (141/496), normal histology in 11% (54/496), cholestasis/obstructive cholangiopathy in 7% (34/496), and fatty liver disease in 6% (30/496) of the biopsies (Table 2). Liver biopsies in patients with clinically suspected rejection showed histological findings compatible with acute or chronic rejection in 46% (151/327) of the cases.

Table 2.

Histological findings and immunosuppressive regimen in patients after liver transplantation. The biopsies predominantly showed acute cellular rejection and idiopathic chronic hepatitis. In patients with recurrence of the underlying disease histological findings were compatible with hepatits C virus re-infection (n=5), autoimmune hepatitis (n=4) and primary sclerosing cholangitis (n=4). The different immunosuppressive regimens at day of liver biopsy are depicted as percentages and absolute numbers.

Histological findings	%, Number
Acute cellular rejection	36% (179/496)
Idiopathic chronic hepatitis	28% (141/496)
Normal histology	11% (54/496)
Cholestasis/obstructive cholangiopathy	7% (34/496)
Fatty liver	6% (30/496)
Chronic rejection	3% (16/496)
Recurrence of the underlying disease	3% (13/496)
Liver cirrhosis	1% (5/496)
Liver necrosis	1% (4/496)
Others	4% (20/496)
Immunosuppression at day of biopsy	%, Number
Ciclosporine, steroids	23% (113/496)
Ciclosporine, mycophenolate, steroids	22% (107/496)
Ciclosporine, mycophenolate	17% (85/496)
Tacrolimus, mycophenolate, steroids	13% (63/496)
Ciclosporine	9% (46/496)
Tacrolimus	6% (28/496)
Ciclosporine, azathioprin, steroids	3% (14/496)
Mycophenolate, steroids	2% (9/496)
Mycophenolate	2% (9/496)
Ciclosporine, azathioprin	1% (7/496)
Ciclosporine, sirolimus	1% (5/496)
Sirolimus, mycophenolate, steroids	1% (3/496
Sirolimus, steroids	1% (3/496)
Ciclosporine, sirolimus, steroids	0.4% (2/496)
Sirolimus	0.2% (1/496)
Ciclosporine, everolimus, steroids	0.2% (1/496)

The different immunosuppressive regimens at day of liver biopsy are illustrated in Table 2. The majority of patients (45%, 222/496) received a dual immunosuppressive therapy, whereas 38% (190/496) were treated with a triple therapy, and only 17% (84/496) received monotherapy. In 41% (205/496) of the patients, the immunosuppressive therapy was adjusted due to the biopsy result.

An escalation of the immunosuppressive therapy was initiated in 31% (63/205) of the cases. In 22% (44/205), a dose reduction of the immunosuppressive therapy was prescribed. Bolus administration of steroids (methylprednisolone 500 mg for 3 days) was applied in 39% (80/205) of the patients. A change of the calcineurin inhibitor therapy (cyclosporin to tacrolimus or tacrolimus to cyclosporin) was documented in 9% (18/205) of the cases.

To analyze surrogate parameters of treatment response, we compared baseline laboratory parameters (at day of liver biopsy) with laboratory values at week 4 and 12. For ALT, a significant difference was detected between the different time points for patients with treatment modification: 128 U/L (IQR 81–229) at baseline, 75 U/L (IQR 44–148) at week 4, and 45 U/L (IQR 26–83) at week 12 (p=0.01) (Figure 1A). For bilirubin, a significant difference between baseline and week 4 and between baseline and week 12 was detected: 22 μmol/L (IQR 13–45) at baseline, 18 μmol/L (IQR 12–46) at week 4, and 16 μmol/L (IQR 10–29) at week 12 (p<0.05) (Figure 1B). For GGT, no difference was detected among the different time points (p>0.05, data not shown). Stratification of the patient cohort according to the time interval from OLT to liver biopsy (≤12 months [n=91] and >12 months [n=114]) showed a significant difference for ALT between the different time points in both groups (p=0.01; ≤12 months: baseline 137 U/L (IQR 79–235), week 4 87 U/L (IQR 44–145), week 12 50 U/L (IQR 29–97); >12 months: baseline 125 U/L (IQR 81–213), week 4 65 U/L (IQR 43–152), week 12 41 U/L (24–70)). The same result was obtained when the patient cohort was stratified by median time from OLT to liver biopsy (group ≤27 months and >27 months) (p<0.05). In contrast, bilirubin was equal between the different time points in both groups (p>0.05). ALT was significantly different between the time points for patients with treatment modification when only the first presentation for liver biopsy was considered (p<0.05).

Figure 1.

(A) Laboratory parameters at different time points after biopsy. Comparison of ALT at day of liver biopsy (baseline) with ALT values at week 4 and 12 for patients with treatment modification (n=205). A significant difference was detected between baseline and week 4 and 12. P-value *** <0.01. (B) Laboratory parameters at different time points after biopsy. Comparison of bilirubin at day of liver biopsy (baseline) with bilirubin values at week 4 and 12 for patients with treatment modification (n=205). A significant difference was detected between baseline and week 4 and 12. P-value *** <0.01; * <0.05.

For patients who did not undergo modification of the immunosuppressive regimen, bilirubin and GGT showed no difference between the time points (p>0.05, data not shown). The same result was obtained when the patient cohort was stratified by median time from OLT to liver biopsy (group ≤27 months and >27 months) and liver biopsy ≤12 months and >12 months after OLT (p>0.05). ALT was 48 U/L (IQR 23–97) at baseline, 42 U/L (IQR 19–90) at week 4, and 33 U/L (IQR 22–72) at week 12 (p<0.05 for the comparison of baseline/week 4 and baseline/week 12). The same result was obtained after stratification of the cohort by median time from OLT to liver biopsy for the comparison of baseline and week 12 (p<0.05). Stratification of the patients by indication of liver biopsy (suspected cellular rejection and protocol biopsy), as well as the number of treatment modifications and laboratory values, are shown in Figure 2A and 2B.

Figure 2.

(A) Stratification of the patients for indication of liver biopsy. The flowchart shows the total number of patients undergoing liver biopsy for suspected cellular rejection and the number of treatment modifications. For patients without treatment modification, only limited data were available (week 4, n=49; week 12, n=97). (B) Stratification of the patients for indication of liver biopsy. The flowchart shows the total number of patients undergoing liver biopsy in the protocol biopsy program and the number of treatment modifications. For patients without treatment modification, only limited data were available (week 4, n=34; week 12, n=56).

Discussion

Data on the clinical impact and consequences of liver biopsies, especially for liver transplant recipients, are sparse. In the worsening organ shortage, it is becoming increasingly important to preserve long-term graft function and to minimize adverse effects of the immunosuppressive medication [8]. To fulfil these obligations, it is necessary to detect acute or chronic rejection to adapt the immunosuppressive therapy and to identify patients who may benefit from a reduction of therapy.

In our cohort, the immunosuppressive therapy was adjusted in 41% of the cases after liver biopsy. The high number of treatment modifications underlines the potential impact of liver biopsies in transplant recipients for the adjustment of the immunosuppressive therapy at our institution. Mells et al. reported that the histological findings of protocol liver biopsy led to a documented change of immunosuppression in 32% of patients [9]. The authors concluded that liver biopsies in transplant recipients provide important histological information about graft function that is not available from non-invasive tests and that liver biopsy allows safe modification of immunosuppressive therapy [9].

In our cohort, the immunosuppressive therapy was mainly changed in patients who underwent event-driven biopsies (suspected cellular rejection) compared to patients from the protocol biopsy program (52% vs. 14%). This finding is plausible because event-driven biopsies are performed in patients who display elevated liver function tests (LFTs), which increases the chance of pathological findings in liver biopsies. The lower number of treatment modifications in the protocol biopsy program compared to other publications may be explained by the retrospective character of the studies, because no general algorithm of treatment modification is applied or it may vary between institutions. Nevertheless, our data show the utility of protocol biopsies in liver transplant recipients for the adjustment of the immunosuppressive regimen.

Due to the retrospective nature of our study, it may be difficult to measure treatment success. However, we wanted to analyze LFTs as surrogate parameters of liver inflammation and damage at different time points after liver biopsy (baseline, week 4, and 12). Interestingly, we found that ALT and bilirubin gradually decreased over time after modification of therapy. Unfortunately, we cannot distinguish clearly between treatment success due to modification of therapy and natural history of the liver disease in our study. Moreover, it is difficult to compare patients with and without treatment modification, as the latter group displayed almost normal LFTs at baseline. However, the comparison of the aforementioned groups (suspected cellular rejection) showed that at baseline, 81% of the patients who underwent treatment modification had ALT values greater than 70 U/L. At week 12, this number decreased to 25%. In contrast, 41% of the patients had ALT values greater than 70 U/L in the group without treatment modification at baseline. After 12 weeks, this number remained nearly unchanged, at 34%. These observations indicate that the initiated treatment change led to an improvement of LFT and therefore can be interpreted as treatment success. Moreover, exclusion of repetitive liver biopsies and stratification of the patient cohort by median time from OLT to liver biopsy and/or ≤12 months or >12 months after OLT showed similar results, demonstrating the robustness of the analysis. In contrast, patients without treatment modification displayed constantly elevated LFTs, which may deteriorate graft function over time.

In addition, we found that clinically suspected allograft rejection and histological findings correlate poorly (46%). In a different study, we showed that only 1.4% of percutaneous liver biopsies caused complications [4]. Given the improved safety of the procedure [4,5,7], liver biopsy should be recommended in case of unclear elevation of LFTs in a transplant recipient or for minimization of immunosuppressive therapy.

Non-invasive tests like transient elastography are increasingly used to assess liver fibrosis and portal hypertension [10–12]. Recently, several reports have shown the potential value of transient elastography in patients after liver transplantation [13,14]. In an exploratory prospective study, liver stiffness was associated with severity of acute cellular rejection in patients after liver transplantation [15]. This study indicates a potential use of transient elastography in the management of post-transplant complications. However, a reliable discrimination of other liver pathologies, such as cholestatic liver disease or fatty liver disease leading to increased liver stiffness, is not possible; therefore, non-invasive tests may deliver additional information for the treatment of post-transplant complications, but cannot replace histological examination. Moreover, liver biopsy may deliver important information in patients with coexisting disorders such as steatosis, hemochromatosis, or overlap syndromes [16–18].

Conclusions

Liver biopsies in liver transplant recipients have potential impact on the modification of the immunosuppressive therapy. The correlation between suspected rejection and histological findings is limited. Consequently, liver biopsy is required in unclear cases or liver function disorders.