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 Table of Contents  
ORIGINAL ARTICLE
Year : 2018  |  Volume : 43  |  Issue : 3  |  Page : 115-118

Effect of homocysteine and folic acid on vaso-occlusive crisis in children with sickle cell disease


1 Department of Clinical Biochemistry, Liver Institute, Menoufia University, Menoufia, Egypt
2 Department of Pediatrics, Faculty of Medicine, Cairo University, Cairo, Egypt
3 Department of Molecular Diagnostics and Therapeutics, Genetic Engineering and Biotechnology Research Institute, University of Sadat City, Sadat, Egypt

Date of Submission17-Jan-2018
Date of Acceptance12-Feb-2018
Date of Web Publication3-Dec-2018

Correspondence Address:
Ahmed M Badr
Pediatrics Department, Faculty of Medicine, Cairo University, 12613
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/ejh.ejh_1_18

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  Abstract 


Introduction Vaso-occlusion is a determinant for most manifestations of sickle cell anemia (SCA). Elevated concentration of homocysteine contributes to thrombosis, a frequent event in SCA. Folic acid deficiency leads to increase in plasma homocysteine. The aim of study was to test whether children with SCA have elevated serum homocysteine with diminished folate level and to determine whether hyperhomocysteinemia has a correlation with the frequency of vaso-occlusive crisis.
Patients and methods A case–control study was carried over a period of 1 year (January to December 2014) in the hematology clinic, Abo El-Reesh Hospital, Cairo University. A total of 50 patients with SCA were included together with 30 age-matched and sex-matched healthy children recruited from Menoufia Hospital. Venous blood samples were obtained from both groups to measure serum homocysteine and folic acid levels.
Results The mean±SD of age of the patients and controls were 6.20±2.55 and 6.03±2.64 years, respectively. Homocysteine level was significantly higher in the patients group compared with control group, with a mean±SD of 44.68±9.096 and 18.81±3.76 µmol/l, respectively (P>0.01). Folic acid level was lower in the patients group than control group, with 12.02±2.76 and 14.68±2.99 ng/ml, respectively (P<0.05). Significant inverse correlation was found between homocysteine and folic acid (correlation coefficient −0.337 and P=0.017). A strong positive correlation between homocysteine level and the frequency of vaso-occlusive crisis was found (P=0.04).
Conclusion Patients with sickle cell disease have high serum homocysteine with low folate levels. This hyperhomocysteinemia is positively correlated with the frequency of vaso-occlusive crisis.

Keywords: folic acid, homocysteine, sickle cell disease, vaso-occlusive crisis


How to cite this article:
Raouf AA, Hamdy MM, Badr AM, Shalaan O, Sakr M, Rahman AR. Effect of homocysteine and folic acid on vaso-occlusive crisis in children with sickle cell disease. Egypt J Haematol 2018;43:115-8

How to cite this URL:
Raouf AA, Hamdy MM, Badr AM, Shalaan O, Sakr M, Rahman AR. Effect of homocysteine and folic acid on vaso-occlusive crisis in children with sickle cell disease. Egypt J Haematol [serial online] 2018 [cited 2018 Dec 16];43:115-8. Available from: http://www.ehj.eg.net/text.asp?2018/43/3/115/246772




  Introduction Top


Sickle cell anemia (SCA) is a genetic disorder caused by homozygosity for a single β-globin gene mutation (β6GAG→GTG), in which valine replaces glutamic acid at the sixth codon of β-globin chain.

Despite this fact, the clinical course of patients with SCA is extremely variable, with the severity of manifestations ranging from asymptomatic to a very severe course [1],[2]. The phenotypic variability may be explained by some genetic factors; those related to globin genes have been recognized [2]. There is evidence that SCA and other chronic hemolytic anemia are characterized by a hypercoagulable state with increase of thrombin and fibrin generation as well as platelet activation with an increased risk for thromboembolic complications [3].

Homocysteine is sulfur-containing amino acid and a normal intermediate in methionine metabolism. In many individuals with inborn errors of homocysteine metabolism, kidney and liver diseases, or nutritional deficiencies, homocysteine levels can rise beyond normal levels and lead to adverse health outcomes. Elevated plasma total homocysteine is an independent risk factor for both cardiovascular and noncardiovascular-related mortality [4],[5].

The aim of this study was to test whether children with sickle cell disease (SCD) have elevated concentration of serum homocysteine with diminished levels of folate and to determine if there is any correlation between hyperhomocysteinemia and the frequency of vaso-occlusive crisis.


  Patients and methods Top


A case–control study was carried over a period of 1 year from first of January 2104 to the end of December 2014. A total of 50 patients (SCA and sickle cell thalassemia) were recruited from the Hematology Center in Abo El-Reesh Hospital along with 30 age-matched and sex-matched healthy controls taken from Menoufia Hospital.

History of renal, cardiac, or hepatic disease was considered as exclusion criteria. All the patients were subjected to the following after signing an informed consent by one of the parents or the patient himself: full history taking with special emphasis on age, sex, residence, frequency of occurrence of vaso-occlusive crisis, severity and site of pain, and adherence to folic acid therapy. The severity of vaso-occlusive crisis was determined according to the pain scale (1–10) plus whether the patient needed hospital, emergency, or unscheduled ambulatory care for pain in the previous day. Routine physical examination was done for all patients. Overall, 3 ml of venous blood sample was obtained from both groups and centrifuged. The serum was taken to the Medical Research Unit in the Genetic Engineering Institute/University of Sadat City, Menoufia, for measurement of serum homocysteine and folic acid level using high-performance liquid chromatography (Shimadzu, Kyoto, Japan), which consisted of a system controller model SCL-10 AVP, a degasser model DGU-12A, two liquid delivery pumps model LC-8AVP, UV-visible detector model SPD-10AVP, and injector model SIL-10A, equipped with 20 µl sample loop.

Biochemical assessment

Total homocysteine level was determined after reverse-phase high-performance liquid chromatography by using isocratic elution and fluorimetric detection. Plasma folate concentrations were determined by a microbial assay with the use of a 96-well plate and manganese supplementation, as described previously [6].

Statistical analysis

Data were analyzed using SPSS Windows statistical package version 18 (SPSS Inc., Chicago, Illinois, USA). Numerical data were expressed as mean, SD, and range. Qualitative data were expressed as frequency and percentage. Pearson’s correlation analysis was done between serum homocysteine level and folic acid, as well as between homocysteine level and the frequency of vaso-occlusive crises. The r value ranges from −1.0 to 1.0 inclusive and reflects the extent of a linear relationship between two data sets. P value was estimated, and a value less than 0.05 indicates statistical significance.


  Results Top


In this study, a total of 50 patients and 30 healthy controls were studied. The mean and SD of age (years) of the patients and controls was 6.20±2.55 and 6.03±2.64 years, respectively (P=0.781). Almost two-thirds of the studied cases were males.

The patients group includes 18 (36%) cases with SCA and 32 (64%) cases with sickle thalassemia.

Patients with SCD (SS) included 12 cases with frequent vaso-occlusive crisis, whereas patients with sickle thalassemia (S/B) included 11 cases only. χ2-test revealed a significant association between sickle genotype and frequency of vaso-occlusive crisis (χ2=4.836 and P=0.04) ([Table 2]).
Table 2 χ2-Test between vaso-occlusive crisis and sickle genotype in patients with sickle cell disease

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Homocysteine level was higher in the patients group compared with control group, with mean and SD of 44.68±9.096 and 18.81±3.76 µmol/l, respectively ([Table 1]). When applying the Student t-test, the result is statistically significant, with a P value less than 0.01.
Table 1 Clinical and laboratory measurements of study group and homocysteine correlations in patients with sickle cell disease

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Folic acid level was lower in the patients group compared with control group, with mean±SD of 12.02±2.76 and 14.68±2.99 ng/ml, respectively, When applying the Student t-test, the result is statistically significant, with a P value of 0.002.

Pearson correlation shows a negative significant correlation between homocysteine level and folic acid level (r=−0.1, P=0.04) as shown in [Figure 1].
Figure 1 Homocysteine and folic acid level correlation.

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A strong positive correlation was found between homocysteine level and the frequency of vaso-occlusive crises (r=0.9, P<0.05).


  Discussion Top


In the present study, patients with homozygous SCA have a higher frequency of vaso-occlusive crises than patients with sickle–β+thalassemia genotype. This observation was consistent with the results of other previous studies [7],[8].

In the current study, homocysteine level was higher whereas folic acid level is lower in the patients than the control. There is a significant negative correlation between homocysteine level and folic acid level [9].

The elevated homocysteine level was consistent with the results of other previous studies [10],[11].

In a recent study conducted on 20 children from Yemen, the authors reported significantly elevated homocysteine level in the studied patients [10]. This concordance in results with our study may be explained by sharing some environmental factors. SCD has wide variability of severity, and environmental factors, including climate, air quality, socioeconomic status, exercise, and infection, can be important factors in determining the severity of SCD [12].

Hyperhomocysteinemia affects ∼20% of adults with SCA despite routine folate supplementation as reported by Dhar et al. [13].

On the contrary, some pediatric studies found no significant difference in homocysteine level compared with controls [14],[15].

Concerning serum folate level, Lowenthal et al. [16] recorded higher homocysteine and folate level in patients with SCD than control. Patients with SCD may have higher nutritional requirement for folic acid than the general population.

In our study, significant positive correlation was found between homocysteine level and the frequency of vaso-occlusive crisis, which indicates that hyperhomocysteinemia status contributes toward initiation of vaso-occlusive crisis through occlusion of small blood vessels. This hyperhomocysteinemia may be attributed to pyridoxine (vitamin B6) deficiency. Homocysteine is an intermediate compound of methionine degradation pathway and is normally remethylated to methionine. This methionine-sparing reaction is catalyzed by the enzyme methionine synthase, which requires 5-methyltetrahydrofolate (metabolite of folic acid) as a methyl donor and methylcobalamin (metabolite of vitamin B12) as a cofactor [17]. In addition, homocysteine is trans-sulferated to cystathionine. This pathway requires vitamin B6 [18] and deficiency of this vitamin will contribute to high serum homocysteine level through disturbance in the metabolic pathway of homocysteine [19].


  Conclusion Top


Children with SCA and sickle thalassemia have high serum level of homocysteine and low level of folic acid compared with normal population. There is significant positive correlation between homocysteine level and the frequency of vaso-occlusive crisis.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Steinberg MH, Adewoye AH. Modifier genes and sickle cell anemia. Curr Opin Hematol 2006; 13:131–136.  Back to cited text no. 1
    
2.
Adams GT, Snieder H, McKie VC, Clair B, Brambilla D, Adams RJ et al. Genetic risk factors for cerebrovascular disease in children with sickle cell disease: design of a case-control association study and genome wide screen. BMC Med Genet 2003; 4:6.  Back to cited text no. 2
    
3.
Ataga KI, Orringer EP. Hypercoagulability in sickle cell disease: a curious paradox. Am J Med 2003; 115:721–728.  Back to cited text no. 3
    
4.
Vollset SE, Refsum H, Tverdal A, Nygård O, Nordrehaug JE, Tell GS, Ueland PM. Plasma total homocysteine and cardiovascular and noncardiovascular mortality: the Hordaland Homocysteine Study. Am J Clin Nutr 2001; 74:130–136.  Back to cited text no. 4
    
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Bostom AG, Silbershatz H, Rosenberg IH, Selhub J, D’Agostino RB, Wolf PA. et al.. Nonfasting plasma total homocysteine levels and all-cause and cardiovascular disease mortality in elderly Framingham men and women. Arch Intern Med 1999; 159:1077–1080.  Back to cited text no. 5
    
6.
Tamura T, Freeberg LE, Cornwell PE. Inhibition by EDTA of growth of Lactobacillus casei in the folate microbiological assay and its reversal by added manganese or iron. Clin Chem 1990; 36:1993.  Back to cited text no. 6
    
7.
Platt OS, Brambilla DJ, Rosse WF. Mortality in sickle cell disease. Life expectancy and risk factors for early death. N Engl J Med 1994; 330:1639.  Back to cited text no. 7
    
8.
Nagel RL, Fabry ME, Pagnier J. Hematologically and genetically distinct forms of sickle cell anemia in Africa. The Senegal type and the Benin type. N Engl J Med 1985; 312:880.  Back to cited text no. 8
    
9.
Houston PE, Rana S, Sekhsaria S, Perlin E, Kim KS, Castro OL. Homocysteine in sickle cell disease: relationship to stroke. Am J Med 1997; 103:192–196.  Back to cited text no. 9
    
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Al Nuzaily M, Ali F. Sickle cell anemia in relation to total homocysteine levels and the role of anticoagulant proteins. Int J Pharm Clin Res 2014; 6:192–195.  Back to cited text no. 10
    
11.
Pandey S, Pandey HR, Mishra RM, Pandey SW, Saxena R. Increased homocysteine level in Indian sickle cell anemia patients. Ind J Clin Biochem 2012; 27:103–104.  Back to cited text no. 11
    
12.
Tewari S, Brousse V, Piel FB, Menzel S, Rees DC. Environmental determinants of severity in sickle cell disease. Haematologica 2015; 100:1108–1116.  Back to cited text no. 12
    
13.
Dhar M, Bellevue R, Brar S, Carmel R. Mild hyperhomocysteinemia in adult patients with sickle cell disease: a common finding unrelated to folate and cobalamin status. Am J Hematol 2004; 76:114–120.  Back to cited text no. 13
    
14.
Balassa VV, Kalinyak KA, Bean JA, Stroop D, Gruppo RA. Hyperhomocysteinemia is associated with low plasma pyridoxine level in children with sickle cell disease. J Pediatric Hematol Oncol 2002; 24:374–379.  Back to cited text no. 14
    
15.
Balassa VV, Kalinyak K. Homocysteine and vitamin co-factor levels in patients with sickle cell disease. Proceeding of 43 annual meeting of the American Society of Hematology, 2002.  Back to cited text no. 15
    
16.
Lowenthal EA, Mayo MS, Cornwell PE, Thornley-Browen D. Homocysteine elevation in sickle cell disease. J Am Coll Nutr 2000; 19:608–612.  Back to cited text no. 16
    
17.
Rezvani I, David SR. Metabolic disease. In: Kliegman RM, Jenson HB, editors. Nelson textbook of pediatrics. 18th ed. Philadelphia: WB Saunders Company; 2008. 85:536.  Back to cited text no. 17
    
18.
Guilland JC, Favier A, Potier de Courcy G, Galan P, Hercberg S. Hyperhomocysteinemia: an independent risk factor or a simple marker of vascular disease? 1. Basic data. J Pathol Biol (Paris) 2003; 51:101–110.  Back to cited text no. 18
    
19.
Rodriguez-Cortes HM, Griener JC, Hyland K, Bottiglieri T, Bennett MJ, Kamen BA, Buchanan GR. Plasma homocysteine levels and folate status in children with sickle cell anemia. J Pediatr Hematol Oncol 1999; 3:219.  Back to cited text no. 19
    


    Figures

  [Figure 1]
 
 
    Tables

  [Table 2], [Table 1]



 

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