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Seraj Ahmed Khan et al, 2018;3(1):099–103.

International Journal of Medicine in Developing Countries

Adenosine deaminase activity in subjects with normal and insufficient liver function

Seraj Ahmed Khan1*, Rajendra Kumar Chaudhari1, Sarita Lamichane1, Madhab Lamsal1

Correspondence to: Seraj Ahmed Khan

*B.P. Koirala Institute of Health Sciences, Dharan, Nepal.

Email: drserajkhan [at] gmail.com

Full list of author information is available at the end of the article.

Received: 30 May 2018 | Accepted: 28 September 2018


ABSTRACT

Background:

Adenosine deaminase (ADA), a nonspecific marker for cellular immunity, is raised in diseases eliciting a cell-mediated immune response. High serum ADA activities were observed in patients with different liver diseases. This study was aimed to determine the association of serum ADA and transaminases levels in liver disease patients and normal control.


Methodology:

In this comparative cross-sectional study, 80 subjects; 40 with liver disease and 40 without liver disease were enrolled. Liver function profile was done by autoanalyzer and ADA activity was measured by Giusti and Galanti’s colorimetric method. Independent t-test and Mann–Whitney U test for the association; Pearson’s correlation and Spearman’s Rho for correlation study. Multiple linear regression model was used for the independent association of the variable with ADA. p < 0.05 set as statistically significant.


Results:

The subjects were divided into Group 1 (normal liver function) and Group 2 (altered liver function) with 40 individuals in each group and mean age was 40.0 ± 16.5 years and 42.4 ± 16.3 years, respectively. Total bilirubin, direct bilirubin, aspartate transaminase (AST), alanine transaminase (ALT), alkaline phosphatase (ALP), and ADA were higher in subjects with liver insufficiency (group 2) and was statistically significant. ADA was significantly correlated with albumin, AST, ALT, and ALP in subjects with normal liver function whereas no such pattern was noticed in patients with liver insufficiency.


Conclusion:

This study demonstrates that the serum ADA activity is predominantly elevated in subjects with altered liver function and highlights its role in diagnosis and monitoring the clinical status of the patients with liver insufficiency.


Keywords:

Adenosine deaminase, aspartate transaminase, alanine transaminase, alkaline phosphatase, liver function test.

Introduction

Adenosine deaminase (ADA) is ubiquitously present in human tissues and its activity is higher in lymphoid tissues, predominantly in T-cells [1]. Its main physiological role is related to lymphocytic proliferation, differentiation, and maturation. Its relationv with cellular immunity causes its increase in disease which triggers immunological response [24]. Patients with liver disorders like acute hepatitis, alcoholic hepatic fibrosis, chronic active hepatitis, liver cirrhosis, and hepatoma have reported high serum ADA activities, sighting its role in immunological disturbance in such diseases [5].

The term “Liver function test” is a frequently used but poorly defined phrase that encompasses the numerous serum chemistries that can be assayed to assess hepatic function or injury. Biochemical markers such as serum bilirubin, alanine transaminase (ALT), aspartate transaminase (AST), ratio of aminotransferases, alkaline phosphatase (ALP), total protein, and albumin are most commonly done in liver function profile test [6]. Liver tests rarely suggest a specific diagnosis; rather, they suggest a general category of liver disease, such as hepatocellular or cholestasis, which then further directs the evaluation. Many tests, such as the aminotransferases or ALP, do not measure liver function at all. Rather, they detect liver cell damage or interference with bile flow.

The exact mechanism involved in the liver damage induced by viral hepatitis is not very elaborate, though both viral and host factors may have a role to contribute in the pathology. Liver lesions could be the result of immune responses or the cytopathic actions of the virus. Cytotoxic T cells and cytokines, produced by both CD4+ (T helper) and cytotoxic T cells, may be responsible for much of the damage that occurs in the livers of infected patients [4]. Similar is the case with non-alcoholic fatty liver disease, where the pathogenesis is yet to be completely understood. However, it involves deposition of excess fat in the liver; followed by increased fatty acid oxidation, oxidative stress, and cytokine production, resulting in progression to steatohepatitis and fibrosis. Various pathogenic mechanisms that play a role include cytokines tumor necrosis factor-α, adiponectin, resistin, interleukins, and transforming growth factor [7].

Liver disease is accompanied by an increased production of free radicals. Hepatocyte membranes are rich in polyunsaturated fatty acid (PUFA), which is susceptible to attack by free radicals. The PUFA of the membranes undergoes peroxidation. Membrane-bound enzymes like ADA get released. Patients in liver disease become slightly immunocompromised. Therefore, the immunological marker may be altered in such patients. Since ADA is a non-specific marker of T-cell activation, there are chances for its alteration. Therefore, this study is undertaken to see an association between serum ADA in normal and altered liver function test profile.

The biochemical tests involved in the evaluation of liver function are frequently nonspecific since they reflect the basic pathological processes common to many conditions; therefore they provide limited diagnostic information. Hence, tests reflecting liver cell damage, particularly the measurement of the activities of certain enzymes that are altered in conditions with the immunological response can give a clearer picture of the etiopathogenesis of the liver disease. Therefore, ADA measurement in serum or plasma could add up the diagnostic value in liver diseases [8].

Subjects and Methods

This hospital-based comparative cross-sectional study was conducted in the clinical lab of the department of Biochemistry, B.P. Koirala Institute of Health Sciences. A total of 80 subjects were enrolled in the study, out of which 40 had liver insufficiency and 40 had a normal liver function (based on the liver function test). The subjects were briefly interviewed to fill up the preformed questionnaire sheet. Informed and written consent were taken from the participants before enrolling them in the study.

About 5 ml of blood sample was collected through Vacutainer from antecubital vein following the standard protocol. Sample was collected in two aliquots, one was sent to the Clinical lab for the assay of liver function test profile and the other aliquot was sent to the Special lab for the assay of ADA. The serum was separated by centrifuging the blood samples at 3,000 rpm for 10 minutes. Biochemical assay was done after the separation.

The ADA activity was measured at 37°C according to the method of Giusti and Galanti (1986) based on the Berthelot reaction, i.e., the formation of colored indophenol complexes from ammonia liberated from adenosine, quantified spectrophotometrically. One unit of ADA is defined as the amount of enzyme required to release 1 μmol minute−1 of ammonia from adenosine at standard assay conditions.

Total bilirubin and direct bilirubin were estimated by Diazo method, alanine aminotransferase, aspartate aminotransferase, and alkaline phosphate by Enzymatic International Federation of Clinical Chemistry (IFCC) method, total protein by colorimetric biuret method, and albumin by colorimetric Bromocresol Green (BCG) method. All the parameters were analyzed in autoanalyzer (Cobas 311, Germany).

After the liver function profile was done, the subjects were categorized into two groups: Group 1 with normal liver function test (LFT) and Group 2 with altered LFT based on their lab report.

Data were entered in Microsoft Excel and analyzed using Statistical Package of Social science (SPSS) version 16. All data were expressed in terms of figure, percentage, mean, and standard deviation. Descriptive statistics were used to express demographic data. Bivariate analysis between groups was done using Chi-square test for categorical data. For parametric variables, independent t-test for continuous data and the Pearson correlation coefficient was used to assess the relation between quantitative variables. For non-parametric variables, the Mann–Whitney U test and Spearman correlation were used. Multiple linear regression model was used to see the independent association of the parameters on ADA. A p value < 0.05 was considered statistically significant.

Results

There were 40 subjects each in Group 1 and Group 2 with a mean age of 40.0 ± 16.5 years and 42.4 ± 16.3 years, respectively. There were 17 males and 23 females in Group 1 as compared to 23 males and 17 females in Group 2. Basic and clinical parameters of the participants were depicted in Table 1. There was a significant increase in total bilirubin, direct bilirubin, AST, ALT, ALP, and ADA in Group 2 as compared to Group 1 and was statistically significant. In correlation analysis, it was found that albumin, AST, ALT, and ALP were significantly correlated with ADA in Group 1, while in Group 2 only direct bilirubin was positively correlated with ADA and was statistically significant with a p-value of 0.030 (Table 2).

Multiple linear regression model was constructed to see the independent association of different variables with ADA in both groups. The model explained 62% variance in the dependent variable and fit the data quite well in Group 1; while AST, ALP, total protein, and albumin were independently associated with ADA and were statistically significant (Table 3).

Table 1. Basic and clinical parameters in the study participants.

Variables Group 1 (n = 40) (normal liver function) Group 2 (n = 40) (insufficient liver function) p value
Age (Years) 40.0 ± 16.5 42.4 ± 16.3 0.510*
Gender Male (n) Female (n) 17 23 23 17
Total Bilirubin (mg/dl) 0.40 (0.2–1.4) 0.85(0.1–16.3) <0.001**
Direct Bilirubin (mg/dl) 0.20(0.1–0.8) 0.4(0.1–5.0) <0.001**
Total protein (g/dl) 7.9 ± 0.9 7.8 ± 1.5 0.742*
Albumin (g/dl) 4.5 ± 0.9 4.1 ± 1.1 0.083*
AST (IU/l) 24.5 (11–64) 86.5 (20–792) <0.001**
ALT (IU/l) 22.0 (8–61) 71.5 (11–543) <0.001**
ALP (IU/l) 101.5 (41–333) 123.0 (65–542) <0.001**
ADA (IU/l) 35.7 ± 16.3 50.8 ± 22.0 0.001*

*Independent t-test; **Mann–Whitney U test; p < 0.05 considered as statistically significant.

Table 2. Correlation of the variables with ADA in both groups.

Variables Group 1 Group 2
Coefficient p value Coefficient p value
Age −0.007 0.963 −0.104 0.523*
Total Bilirubin (mg/dl) −0.055 0.735 0.299 0.061**
Direct Bilirubin (mg/dl) 0.058 0.721 0.343 0.030**
Total Protein (g/dl) 0.082 0.616 0.133 0.413*
Albumin (g/dl) −0.347 0.028* 0.002 0.989*
AST (IU/l) 0.594 0.000* 0.045 0.783**
ALT (IU/l) 0.400 0.011* −0.184 0.255**
ALP (IU/l) 0.325 0.040* 0.211 0.191**

*Pearson’s correlation; **Spearman’s Rho; p < 0.05 is statistically significant.

AST and ALT levels were categorized into higher and normal level based on their values. AST >40 IU/l was in higher and <40 IU/l in the normal group. Similarly, ALT >55 IU/l was in higher and <55 IU/l in the normal group. A significant difference in ADA value in the groups with higher AST level (p = 0.001) was observed whereas no such association was noticed in ALT category (p = 0.290) (Table 4).

Discussion

The study in the recent past regarding biomarker of liver function had been limited only to its elevation and without evaluation the mechanisms that cause its increase. This positive correlation between serum ADA activity and liver disease in the current study was an important finding. Hence, evaluation of ADA activity in the serum of patients with liver disease can be considered a useful tool for enhancing the diagnostic modality and monitoring the clinical status of chronic liver diseases. Increased serum ADA activities have been observed in infectious diseases caused by microorganisms infected mainly by the macrophages, like tuberculosis, leprosy, visceral, and cutaneous leishmaniasis, brucellosis, typhoid fever, and human immunodeficiency virus infection [913].

Many previous studies have demonstrated an increased ADA activity in liver diseases, such as chronic active hepatitis and liver cirrhosis [1416]. In this study, a significantly higher ADA activity in subjects with liver insufficiency as compared to normal subjects (p < 0.001) was reported. As all types of liver diseases were included as one category so the association of ADA in different types of liver diseases could not be established because of the very small sample size. Some studies have even reported no significant difference between liver diseases like Hepatitis B virus (HBV) and Hepatitis C virus (HCV)infected patients [4], which is in accordance with the present study. Torgutalp et al. [17], in his study reported that serum ADA levels >20 IU/l were found in 63.5% of autoimmune hepatitis patients; and in healthy controls, any increase was not observed beyond the normal level (>20 IU/l). Many recent studies also have come up highlighting the potential diagnostic role of ADA as a marker in evaluating the liver diseases [18,19]. The elevated serum ADA activity in patients with hepatitis may reflect the phagocytic activity of macrophages and proliferation of lymphocytes and may provide useful additional diagnostic information on the pathogenesis of hepatitis.

Table 3. Multiple linear regression for the association of ADA with independent variables.

Variables Coefficient β 95% confidence interval p value
Lower bound Upper bound
Age −0.182 −0.476 0.070 0.140
Total Bilirubin 0.121 −21.50 34.93 0.631
Direct Bilirubin −0.199 −88.64 38.28 0.425
AST 0.486 0.148 1.17 0.013*
ALT −0.083 −0.535 0.334 0.642
ALP 0.260 −0.001 0.154 0.054
TP 0.578 4.589 19.29 0.002*
Albumin −0.639 −22.45 −3.934 0.007*

* p < 0.05 is statistically significant.

Table 4. Serum ADA level according to high and normal AST and ALT level.

Variables ADA( U/l) p value
AST
High Level (n = 41)
Normal Level (n = 39)

51.1 ± 23.9
34.9 ± 15.1
0.001*
ALT
High Level (n = 34)
Normal Level (n = 46)

46.2 ± 21.1
41.0 ± 21.9
0.290

*Independent t-test. p < 0.5 is statistically significant.

Some researchers noticed a stepwise increase in serum ADA levels with increasing severity of liver cirrhosis. The probability of ADA being greater than the mean was approximately 2.5 times higher (2.48, CI 95%: 1.36–4.52) in patients with liver cirrhosis due to HCV infection than in those patients with cirrhosis of a different etiology [15]. The study done by Sánchez et al., also supported the findings of the current study. They observed the highest serum ADA level in hepatic cirrhosis and viral hepatitis as compared with the control [16]. Inflammatory liver diseases display higher levels of serum ADA activity compared with non-inflammatory ones. The most drastic increase in activity is found in acute virus-induced hepatitis, in active liver cirrhosis, extremely high levels in some liver tumors [20].

The significant increase of ADA in subjects with higher AST enzymes could be potentially explained by the fact that as liver damage increases it leads to the excessive destruction of hepatocytes and this causes the release of liver enzyme AST and while this happens the membrane bound and intracellular enzyme are leaked into the circulation thereby increases its serum level. The current finding suggests that liver insufficiency increases the serum levels of ADA.

Conclusion

A significant increase in serum ADA activity in liver disease patients as compared with control subjects was found. The strong positive association of ADA with high AST level potentially highlights its diagnosis in liver insufficiency. However, further studies, in subjects with different liver diseases so as to understand the link with etiology, are needed to provide conclusive evidence to support the use of serum ADA levels for monitoring the clinical status and to enhance the diagnostic modality of the patients with liver insufficiency.

Acknowledgment

We would like to acknowledge the Technologists of Clinical Biochemistry Lab of our institute for their help and support in the analysis of the parameters. We would also extend our thanks to all the participants who allowed us to carry out this study by giving their consent.


List of Abbreviations

ADA Adenosine deaminase

ALT Alanine aminotransferase

AST Aspartate aminotransferase

ALP Alkaline phosphatase

HBV Hepatitis B Virus

HCV Hepatitis C Virus

IFCC International federation of Clinical Chemistry

LFT Liver function test

PUFA Polyunsaturated fatty acid


Funding

None.


Declaration of conflicting interests

None.


Consent for publication

Informed consent was obtained from all the participants.


Ethical approval

Ethical clearance was taken by Institutional Review Committee (IRC), B.P. Koirala Institute of Health Sciences, Dharan, Nepal (Code No.: IRC/0717/016).


Author details

Seraj Ahmed Khan1, Rajendra Kumar Chaudhari1, Sarita Lamichane1, Madhab Lamsal1

  1. B.P. Koirala Institute of Health Sciences, Dharan, Nepal

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How to Cite this Article
Pubmed Style

Khan SA, Chaudhari RK, Lamichane S, Lamsal M. Adenosine deaminase activity in subjects with normal and insufficient liver function.. IJMDC. 2019; 3(1): 99-103. doi:10.24911/IJMDC.51-1527660074


Web Style

Khan SA, Chaudhari RK, Lamichane S, Lamsal M. Adenosine deaminase activity in subjects with normal and insufficient liver function.. https://www.ijmdc.com/?mno=300602 [Access: January 28, 2022]. doi:10.24911/IJMDC.51-1527660074


AMA (American Medical Association) Style

Khan SA, Chaudhari RK, Lamichane S, Lamsal M. Adenosine deaminase activity in subjects with normal and insufficient liver function.. IJMDC. 2019; 3(1): 99-103. doi:10.24911/IJMDC.51-1527660074



Vancouver/ICMJE Style

Khan SA, Chaudhari RK, Lamichane S, Lamsal M. Adenosine deaminase activity in subjects with normal and insufficient liver function.. IJMDC. (2019), [cited January 28, 2022]; 3(1): 99-103. doi:10.24911/IJMDC.51-1527660074



Harvard Style

Khan, S. A., Chaudhari, . R. K., Lamichane, . S. & Lamsal, . M. (2019) Adenosine deaminase activity in subjects with normal and insufficient liver function.. IJMDC, 3 (1), 99-103. doi:10.24911/IJMDC.51-1527660074



Turabian Style

Khan, Seraj Ahmed, Rajendra Kumar Chaudhari, Sarita Lamichane, and Madhab Lamsal. 2019. Adenosine deaminase activity in subjects with normal and insufficient liver function.. International Journal of Medicine in Developing Countries, 3 (1), 99-103. doi:10.24911/IJMDC.51-1527660074



Chicago Style

Khan, Seraj Ahmed, Rajendra Kumar Chaudhari, Sarita Lamichane, and Madhab Lamsal. "Adenosine deaminase activity in subjects with normal and insufficient liver function.." International Journal of Medicine in Developing Countries 3 (2019), 99-103. doi:10.24911/IJMDC.51-1527660074



MLA (The Modern Language Association) Style

Khan, Seraj Ahmed, Rajendra Kumar Chaudhari, Sarita Lamichane, and Madhab Lamsal. "Adenosine deaminase activity in subjects with normal and insufficient liver function.." International Journal of Medicine in Developing Countries 3.1 (2019), 99-103. Print. doi:10.24911/IJMDC.51-1527660074



APA (American Psychological Association) Style

Khan, S. A., Chaudhari, . R. K., Lamichane, . S. & Lamsal, . M. (2019) Adenosine deaminase activity in subjects with normal and insufficient liver function.. International Journal of Medicine in Developing Countries, 3 (1), 99-103. doi:10.24911/IJMDC.51-1527660074