|Year : 2014 | Volume
| Issue : 1 | Page : 13-19
Liver function test profile of Nigerian children with sickle cell anaemia in steady state
Shehu A Akuyam1, Peter O Anaja1, Olufemi G Ogunrinde2, Auwal Abubakar1, Nasir Lawal1, Sa'ad M Ya'uba2, Abdullahi Musa2, Fatima F Abdullah2, Yusuf Garba1, Yakubu Abubakar2, Mustapha N Adebiyi2
1 Department of Chemical Pathology, Ahmadu Bello University Teaching Hospital, Zaria, Nigeria
2 Department of Paediatrics, Ahmadu Bello University Teaching Hospital, Zaria, Nigeria
|Date of Web Publication||7-Apr-2014|
Shehu A Akuyam
Department of Chemical Pathology, Ahmadu Bello University Teaching Hospital, Zaria
Source of Support: Macarthur Foundation,, Conflict of Interest: None
Background and Objective: Several studies across the world have revealed that liver dysfunction is a common finding in children with sickle cell anaemia (SCA). The situation in hospitals in Nigeria is not known. The few studies carried out in Nigeria were on adult subjects. The overall objective of the study was to evaluate the liver function test (LFT) profile of children with SCA, aged 1-14 years, with a view to improve the routine investigation and management of this group of subjects. Materials and Methods: LFT profile of 60 SCA subjects and 60 age- and sex-matched apparently healthy subjects (controls) was evaluated in the present study in the departments of chemical pathology and paediatrics of Ahmadu Bello University Teaching Hospital (ABUTH), Zaria, northern region of Nigeria. Results: The results of serum LFTs of the SCA subjects and age- and sex-matched control subjects showed that the concentrations of serum total bilirubin (TB), conjugated bilirubin (CB), unconjugated bilirubin (UCB), alkaline phosphatase (ALP), and aspartate transaminase/alanine transaminase (AST/ALT) ratio were significantly higher (P < 0.001), while the levels of total protein (TP) were significantly lower (P < 0.001) in SCA subjects compared with the control subjects. Serum levels of ALT, AST and albumin (ALB) among SCA and control subjects were not statistically different (P > 0.05). The results of serum LFTs also showed that there were no significant age differences (P > 0.05) in both SCA subjects and control subjects. There were no correlations between all the LFT analytes and the frequency of crisis (FOC) among SCA subjects (P > 0.05). Conclusion: It could be concluded from the findings of the present study that there are mild LFT abnormalities in children with SCA even in steady state, and that the extent of some of the abnormalities decreases with advancing age of the subjects.
Keywords: Liver dysfunction, liver function test profile, Nigerian children, sickle cell anaemia, sickle cell disease
|How to cite this article:|
Akuyam SA, Anaja PO, Ogunrinde OG, Abubakar A, Lawal N, Ya'uba SM, Musa A, Abdullah FF, Garba Y, Abubakar Y, Adebiyi MN. Liver function test profile of Nigerian children with sickle cell anaemia in steady state. Niger J Basic Clin Sci 2014;11:13-9
|How to cite this URL:|
Akuyam SA, Anaja PO, Ogunrinde OG, Abubakar A, Lawal N, Ya'uba SM, Musa A, Abdullah FF, Garba Y, Abubakar Y, Adebiyi MN. Liver function test profile of Nigerian children with sickle cell anaemia in steady state. Niger J Basic Clin Sci [serial online] 2014 [cited 2019 May 23];11:13-9. Available from: http://www.njbcs.net/text.asp?2014/11/1/13/130157
| Introduction|| |
Sickle cell disease (SCD) is a group of complex genetic disorders with multisystem manifestations, and is an autosomal recessive (inherited) blood disorder characterised by defective (sickle) haemoglobin (HbS) in the red cells when deoxygenated. ,,,, It is a condition resulting from the inheritance of two abnormal allelemorphic genes controlling the formation of the beta (β) chains of haemoglobin.  SCD describes a group of complex, chronic disorders including HbSS, HbSC, HbSβ thalassaemia and other heterozygous conditions.
Sickle cell anaemia (SCA) is the most common and severe form of SCD, and is the condition resulting from the inheritance of two sickle genes. , It results from the substitution of valine for glutamic acid at position 6 of the β-globin gene at chromosome number 11, leading to production of a defective form of haemoglobin, haemoglobin S (HbS). 
SCD presents a major medical problem in certain parts of the world, particularly in tropical Africa, the Caribbean and the Middle East.  The prevalence of SCD is quite variable, but it is estimated that 8% of black people in America and 40% of the population in certain countries of tropical Africa have the sickle cell trait.  In fact, SCD affects millions of people throughout the world. In the United States, it is present in 1 in every 500 African-American births.  The prevalence of SCA in Nigeria is about 20 per 1000 births (2%) as reported by the World Health Organization (WHO) in 2005,  and it was also reported that in Nigeria alone, more than 100,000 children are born annually with SCA. 
SCD is characterised by chronic haemolytic anaemia, red cell sequestration and vaso-occlusive crises arising from widespread vascular occlusion by sickled red blood cells and leading to multiple organ infarctions.  Sickling of red cells in various parts of the body causes acute and chronic ischaemia leading to progressive tissue damage. , The rate of development of various complications is variable, but many subjects develop organ damage. This, in turn, is associated with increased morbidity and mortality. 
The liver is one of the organs involved in the multiorgan failure that occurs in SCD. It was reported that liver and biliary tract dysfunctions are common complications of SCA.  The hepatic dysfunction occurs commonly in children with SCD. ,, Previous reports described jaundice, hepatic infarcts, acute and chronic hepatitis, choledocholithiasis and cirrhosis.  The liver can be affected by a number of complications due to the disease itself and its treatment. , The risk of cholelithiasis, choledocholithiasis and liver failure increases in these subjects due to sickling. In addition, viral hepatitis and other hepatobiliary diseases can also occur. , Various disorders of the hepatobiliary system can occur in subjects with SCA. It was reported that clinical spectrum of SCD ranges from mild liver function test (LFT) abnormalities in asymptomatic subjects to significant hepatic abnormalities with marked hyperbilirubinemia. 
Being a complex multisystem disorder, SCA requires specialised and comprehensive care to achieve an optimal outcome. Timely and appropriate treatment of acute illness in SCA subjects is critical because life-threatening complications develop rapidly. It is essential that every patient with SCA receive comprehensive care that is coordinated through a regular medical check-up in hospital with appropriate expertise. It has been documented that an expert and comprehensive medical care decreases the morbidity and prolongs the life expectancy for individuals with SCA.  In this context, an appropriate treatment requires the active involvement of health care professionals with expertise in the management and treatment of SCA.
Most pathological studies of liver disease in SCA and its variants were performed retrospectively on autopsy specimens, and because of the prominent histological features of intrasinusoidal sickling and Kupffer cell erythrophagocytosis, hepatic dysfunction was attributed to the intrahepatic sickling of erythrocytes in this haemoglobinopathy. Evidence of liver disease in SCD is obtained either from abnormal biochemical tests or postmortem liver biopsy specimen. There is paucity of data on the liver function status of SCA subjects in most of the Nigerian hospitals, including Ahmadu Bello University Teaching Hospital (ABUTH), Zaria, northern region of Nigeria. Most of the studies reported have been carried out elsewhere in the world, and the few studies that have been carried out worldwide were mostly among adults and not among children with SCA.
Early detection of the complications of SCA such as liver and renal diseases is very essential in reducing the morbidity and mortality in SCA subjects. This could be achieved by carrying out various liver and renal functions tests (LFTs and RFTs). Therefore, in view of its clinical importance, there is the need to evaluate the liver function status of Nigerian children with SCA. Therefore, the overall objective of the present study was to evaluate the LFT profile of children with SCA and of age 1-14 years, with a view to improve the routine investigation and management of this disorder.
| Materials and Methods|| |
The study was conducted in the departments of chemical pathology and paediatrics of ABUTH. A total of 120 subjects were consecutively enrolled for the study. These consisted of 60 children with SCA and 60 age- and sex-matched children with HbAA (controls). The control subjects were consecutively enrolled by screening for the presence of SCA and other haemoglobinopathies. This was achieved by determining the genotype (Hb electrophoretic pattern) of the subjects using electrophoretic method. In this case, children with AA genotype were consecutively enrolled. The target population was children aged 1-14 years who were attending the Haemato-oncology Clinic of the Department of Paediatrics of ABUTH.
The 60 SCA subjects aged 1-14 years (mean age 7 ± 3.61 years) were consecutively enrolled for the study. These consisted of 30 males (50%) aged 1-14 years (mean age 6 ± 3.55 years) and 30 females (50%) aged 1-14 years (mean age 8 ± 3.57 years). The subjects were also divided into three different age groups: group I subjects aged <5 years (n = 21, 35.00%), group II subjects aged 5-9 years (n = 20, 33.33%) and group III subjects aged 10-14 years (n = 19, 31.67%). All the subjects were in steady state of health as indicated by the findings from history and physical examination. The SCA subjects had a mean frequency of sickle cell crisis of 4 ± 4.27 crises in a year (range of 0-6 crises in a year). Similarly, 60 children with HbAA who were age- and sex- matched with the cases with SCA also ranged in age from 1 to 14 years (mean age of 7 ± 3.61 years) and were consecutively enrolled for the study as controls. These children were also grouped into three different age groups to tally with their age- and sex-matched SCA counterparts.
Children with SCA of age between 1 and 14 years, belonging to either sex, and presenting to the Paediatric Haemato-oncology Clinic of ABUTH, and who were confirmed to have laboratory diagnosis of HbSS and were in a steady state were included in the study. Age- and sex-matched children who had HbAA haemoglobin electrophoretic patterns were consecutively enrolled as controls. These were selected from the population of children presenting to the Paediatric Out-patient Department (POPD) of ABUTH with mild ailments and not in any clinical condition that can affect the results of LFT, and with no history of recent or frequent blood transfusion.
All children who were tested positive for HB s Ag and HIV/AIDS and those with documented conditions that can affect the results of LFTs, as well as those with history of recent or frequent blood transfusion were excluded from the study. The children whose parents/guardians declined to give consent for inclusion were also excluded from the study.
Informed consent for inclusion into the study was obtained from the parents/guardians of the selected children using a standard informed consent format. Ethical approval was obtained from the Ethical Committee of the Faculty of Medicine of the Ahmadu Bello University/Ahmadu Bello University Teaching Hospital (ABU/ABUTH), Zaria, in accordance with the Declaration of Helsinki. The sample size for the study was determined from a standard formula for the calculation of minimum sample size. ,
At the clinic, those children who satisfied the study inclusion criteria were consecutively selected. A full medical history was obtained from the parents/guardians of the selected children. This was followed by a detailed physical examination and collection of blood samples. The main findings were documented in the study proforma. For each child with SCA who was consecutively enrolled, an age- and sex-matched control (child with HbAA) was consecutively selected.
Blood specimens were collected from both the SCA and control subjects, and taken into plain tubes by venipuncture in children with prominent veins and by femoral tap in children whose veins were tiny, particularly in those under 5 years of age. The blood specimens in the plain tubes were centrifuged and the sera were carefully drawn into sample bottles and then analysed for LFT immediately.
Serum bilirubin (SB) concentrations were estimated using Van den Bergh diazo reaction method of Malloy and Evelyn.  Serum alanine and aspartate transaminase (ALT and AST) activities were estimated using the colorimetric method of Reitman and Frankel,  while serum alkaline phosphatase (ALP) activity was estimated using the method of King and Armstrong.  Serum albumin (ALB) concentrations were estimated using the method of Doumas et al.,  whereas serum total protein (TP) concentrations were estimated by Biuret method (Kingley.  De Ritis ratio (AST/ALT ratio) was determined by AST activities divided by ALT activities, as calculated by De Ritis et al. 
The data obtained were analysed using Microsoft Office Excel 2007. The results of LFT obtained from children with SCA were compared with those of controls using the paired two-tailed Student's t-test for matched samples. The LFT results obtained from children with SCA, as well as controls in different age groups were compared using one-way analysis of variance (ANOVA). Similarly, the frequency of crisis (FOC) in different age groups of children with SCA was compared using the paired two-tailed Student's t-test for matched samples. In each case where there was significant difference, a post-hoc analysis was carried out using paired two-tailed Student's t-test. This is in order to know specifically where the difference is. Correlations between FOC and each of the LFT analytes in children with SCA were found using Pearson's linear correlation analysis. A P value equal to or less than 0.05 (P ≤ 0.05) was considered as statistically significant.
| Results|| |
The results of serum LFTs among children with SCA and age- and sex- matched controls are shown in [Table 1]. The table shows that the concentrations of serum total bilirubin (TB), conjugated bilirubin (CB), unconjugated bilirubin (UCB) and ALP, and AST/ALT ratio were significantly higher (P < 0.001), while the TP concentrations were significantly lower (P < 0.001) in children with SCA compared with the controls. There was no significant difference in the serum levels of ALT, AST and ALB between children with SCA and controls (P > 0.05).
|Table 1: Liver function test profile (mean±SEM) among children with SCA and controls|
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[Table 2] and [Table 3] show the results of serum LFTs according to different age groups in children with SCA and controls, respectively. The results show that the levels of all the LFTs in different groups of children with SCA were not statistically significant (P > 0.05, ANOVA). Activities of serum ALP in different groups of control subjects were significantly different (P < 0.05, ANOVA). A post-hoc analysis using paired Student's t-test showed that the levels of ALP among the control subjects aged <5 years were significantly higher than those of age 6-9 years (P < 0.05), but were significantly lower than in controls aged 10-14 years (P < 0.05).
|Table 2: Liver function test profile (mean±SEM) among children with SCA according to age group|
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|Table 3: Liver function test profile (mean±SEM) among controls according to age group|
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The results of FOC in different groups of children with SCA are shown in [Table 4]. The results show an FOC for the total group of approximately 3 times per year. The results show that there were significant differences between the different groups (P < 0.05, ANOVA). Post-hoc analysis using paired Student's t-test revealed that the values of FOC were significantly higher in the group with subjects <5 years than in the group with subjects older than 5 years (P < 0.05).
|Table 4: Frequency of sickle cell crisis (mean±SEM) among children with SCA according to age group|
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There were no significant correlations between FOC and each of the values of serum TB, CB, UCB, AL, AS, AL, TP, ALB and AST/ALT among children with SCA (r = 0.024, P > 0.05; r = 0.092, P > 0.05; r = 0.003, P > 0.05; r = 0.094, P > 0.05; r = 0.136, P > 0.05; r = 0.088, P > 0.05; r = 0.019, P > 0.05; r = 0.163, P > 0.05; r = 0.057, P > 0.05 respectively).
| Discussion|| |
In the present study we evaluated the LFT among children with SCA in Zaria, northern region of Nigeria. The findings of increased serum TB, CB, UCB and ALP and reduced TP concentrations in children with SCA observed in the present study demonstrate that some hepatic functions were deranged in SCA.
The finding of significantly higher activities of ALP in SCA patients could be due to cholestasis, as it is known that ALP is a marker of cholestasis. It could also be due to long-standing vaso-occlusion which might have affected the bone. Again, the finding of significantly higher activities of ALP in control individuals of age between 10 and 14 years than in younger ones could be attributed to the increased production of this enzyme during the adolescence/pubertal stage. The pattern of ALP in this study is consistent with the literature report showing that ALP activities are usually high in the newborns and young children up to about 5 years of age, and the levels reduce after this age before rising sharply at the adolescent age due to pubertal growth spurt.  The absence of correlation between the frequency of sickle cell crisis and all the LFT components as observed in the present study suggests that the occurrence of the crisis in children with SCA does not influence the LFT. The finding of significant reduction of FOC among SCA subjects aged 10-14 years demonstrates that the FOC decreases with advancing age in SCA.
The finding of mild abnormalities in LFT, as indicated by slight to moderate elevation of the LFT components in the present study, agrees with the previous reports. ,,,,, Kotila et al.  reported the occurrence of jaundice, and minimal increase in liver size and the activities of ALT, AST and ALP among adult SCA subjects in steady state of health in Ibadan, Nigeria. Some other studies conducted by Johnson  and Traina et al.  in the western world suggest variable degrees of LFT abnormalities. The finding of minor derangement in LFTs is consistent with the report of Maher and Mansour  who observed that the clinical spectrum of SCD ranges from mild LFT abnormalities in asymptomatic subjects to significant hepatic abnormalities with marked hyperbilirubinemia. It has been estimated by Diggs  that 10% of children with SCA admitted to hospital developed hepatic crisis. However, many of the studies reported that the abnormalities in LFTs tend to be more severe during vaso-occlusive episodes,  fever and leucocytosis,  and these were not observed in our patients at the time of sampling.
It was reported that the incidence of liver dysfunction in children with SCA is common, being a component of the multiorgan failure that occurs in this disorder.  However, the pathophysiology of the liver disease in SCA is not certain because of its complexities. The clinical manifestations of the different causes of liver failure are also similar and interrelated, thus making the pathophysiology complex. Moreover, it has been observed that enlargement of the liver does not connote disease and a normal-sized liver may be diseased. ,[ 33] Therefore, abnormalities in the LFT reported in the present study could not be ascertained to a particular factor, rather they could be due to a combination of factors.
The occurrence of liver disease in SCA may be due to a variety of causes such as obstruction of sinusoids by sickle cells with subsequent hepatic infarction during vaso-occlusive episodes, red cell sequestration, cholelithiasis and cardiac failure. Other multiple factors that may contribute to the aetiology of the liver disease in SCD include ischaemia, transfusion-related viral hepatitis, iron overload (haemosiderosis) and gall stones. Coiner et al.  suggested that the most common causes are those related to repeated blood transfusion, such as haemosiderosis and viral hepatitis, which lead to chronic liver disease. These authors reported haemosiderosis and erythrophagocytosis in the liver biopsies of all sickle cell subjects with chronically elevated LFTs in their study. This was supported by Johnson et al.,  Traina et al.  and Mills.  However, the SCA patients selected for the present study were screened for the presence of hepatitis and there was no history of repeated blood transfusion, and therefore, these are unlikely to be the causes of abnormal LFTs in these patients. Presence of haemosiderosis and cardiac failure was not established in the SCA patients under study, and therefore, their role in the causation of the observed abnormal LFTs could not be ascertained. Therefore, the most likely causes of altered LFTs in the SCA patients in our study could be haemolysis, red cell sequestration and hyperbilirubinemia. The presence of hyperbilirubinemia might have caused bilirubin toxicity and inability for the liver to metabolise. The excess bilirubin could precipitate in the liver to form gall stones which then damage the liver cells.
It was suggested by Benerjee et al.  that the hepatic complications of the SCDs, including SCA, can be separated into the following categories: (1) disorders related to haemolysis, (2) the problem related to anaemia and subsequent transfusion management, (3) the consequences of sickling and vaso-occlusion and (4) defects unrelated to SCD. Maher and Mansour  have suggested a multifactorial aetiology for liver disease in SCD subjects, including factors associated with chronic haemolytic anaemia (cholelithiasis), multiple transfusions (viral hepatitis and iron overload), vascular damage and the sickling process itself. However, data points to the importance of vascular changes and the significant participation of the sickling process in most of subjects, as reported by Charlotte et al. 
The elevation of SB concentrations as found in the children with SCA in the present study could be due to haemolysis which is ongoing even in the absence of crisis. High levels of ALP found in the subjects of the present study may be because of either cholestasis or bone lesion being one of the common complications of SCA. Acute intrahepatic cholestasis may be a consequence of widespread sickling within the sinusoid or extreme haemolysis with resultant hyperbilirubinemia, which is often accompanied by elevated ALP and variable levels of ALT and AST. This could be supported by the study of Kotila et al.  in which it was reported that 74% of the sickle cell subjects showed elevated serum ALP levels, but no significant correlation was found between this and the liver size. Thus, it suggests that liver pathology may not be solely accountable for the elevation of this enzyme.
A distinct clinical presentation of sickle cell intrahepatic cholestasis, and a syndrome characterised by progressive cholestasis in the absence of cirrhosis had been reported in a small number of cases of SCA.  These cases are characterised by right upper quadrant pain, extreme elevation of bilirubin, striking elevation of ALP and variable elevation of transaminases, as well as histological features including intracanalicular cholestasis, sinusoidal dilatation, Kupffer cell hyperplasia and erythrophagocytosis. The findings of marked elevation of SB and ALP and slight elevation of serum transaminases reported in our study could be strongly suggested to be due to cholestasis, which is consistent with the previous reports as explained above.
A previous study has identified bone ALP as the principal enzyme fraction that increases during sickle cell crises, and it also appeared that there is a correlation between the severity of crises, serum ALP activities and isoenzyme patterns.  However, it was suggested that these abnormalities could also be detected even when the subjects are asymptomatic,  and this is consistent with the finding of our study of elevated ALP despite the fact that the SCA children selected were in a steady state of health and not in crisis.
| Conclusion|| |
It could be concluded from the findings of the present study that there are mild LFT abnormalities in children with SCA even in steady state, and that the extent of some of the abnormalities decreases with advancing age of the subjects. It could be recommended from the findings of the present study that a routine evaluation of liver function status be considered in the management of children with SCA in Nigerian hospitals and elsewhere. This could assist in the early detection of liver dysfunction in this group of subjects and, hence, reduce morbidity and mortality from SCA. Evaluation of activities of ALP isoenzymes is recommended to ascertain the origin of its elevation among SCA subjects.
| Acknowledgements|| |
The authors acknowledge and appreciate the financial support provided by Macarthur Foundation for conducting this study. They are also grateful to Drs M. A. Bugaje, F. D. Akeredolu, I. E. Ezeukwu, K. Hamzat and S. O. Ige of Paediatrics Department, as well as Matrons E. F. Idowu, M. N. Maigari, M. N. Iyoke, A. J. Raji and J. D. Haruna of POPD Clinic and Mr. C. R. Gabriel, Mr. D. Musa and Mal H. Yusuf of the Institute of Maternal and Child Health of ABUTH, Zaria for their assistance during the course of sampling processes.
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[Table 1], [Table 2], [Table 3], [Table 4]