Prognostic Potential of RDW in Discriminating Hemoglobinopathies among Patients reporting to Aga Khan Hospital, Kisumu – Egyptian Journal of Medical Human Genetics

The red cell distribution width was significantly (p < 0.0001) elevated in the overall hemoglobinopathy cases with an overall median of 20.7 (IQR = 8.3) meaning it could be used to differentiate diseased (hemoglobinopathy) from non-diseased (hemoglobinopathy-free) individuals, thus serving as a potential biomarker of hemoglobinopathies in Western Kenya. However, when hemoglobinopathies were broken into their respective phenotypes, some phenotypes had statistically significant (p < 0.0001) elevated RDW, while others did not have statistical significance. Therefore, the present study obtained RDW cutoff value of 19.9 [14.5, IQR = 2.7; 95% CI 9.1–19.9, p < 0.0001], meaning that 95% of the normal population [32] have RDW ≤ 19.9; thus, above that could be indicating a marked anisocytosis suggesting a hemoglobinopathy that needs confirmation using the gold standard tests. This cutoff value was developed from the normal controls who had RDW median of 14.5 (IQR = 2.7), which is higher compared to 11.3 documented by a previous study [33]. This explains the issue of addressing establishment of local reference values as recommended by previous studies, that clinical laboratories need to obtain their own normal ranges [34]. Variation of hematological normal ranges has been reported in many parts of Africa due to several factors that include sex, geographical location, race, altitude and diet [34, 35]. In a study done in Nakuru County, similarly, documented reduced hematological ranges with the author citing genetic, ethnic and demographic variations which she recommends regions to develop their reference ranges for better patient management [36]. In another study done in Kenya medical research institute in Kisumu, found out that use of locally established reference ranges resulted in fewer participants classified as having abnormal hematological or biochemical values compared to use of USA-derived reference intervals [37]. In a similar population-based cohort study done in Kericho, Kenya, in a potential Phase I/II HIV vaccine trial site, necessitated establishment of region-specific clinical reference ranges for trial enrollment and participant monitoring. The authors documented that use of (USA) reference range which defines anemia in African-American men and women as hemoglobin of 12.9 g/dL and 11.5 g/dL, respectively, would result into the entire Kericho population classified as anemic, leading to erroneous and unnecessary treatments [38]. To this effect, the present study developed a RDW standard cutoff value of 19.9 at 95% confidence interval calculated from the control median of 14.5 + (IQR = 2.7 × 2) as required by normal distribution curves [32].

RDW was higher in males with a median of 17.1 (IQR = 8.0) than females who had a median of 15.5 (IQR = 6.7) but did not have statistical significance (p = 0.317) between the two sexes as shown in Table 3, similar to respective control groups (p = 0.089). Similar findings are reported by Qurtom et al. that RDW showed no statistically significant difference in the mean RDW for males and females [26], consistent with results reported by Hoffmann et al. [22]. The red cell distribution width (RDW) decreased significantly as the study subjects aged (p < 0.05), starting with ≤ 5-year-olds who has the highest median, while those ≥ 13 years have the lowest RDW median (Table 3). However, the RDW for participants aged 6–12 years, like those for age ≥ 13 years, did not differ significantly (p = 0.347), irrespective of gender, from those for the respective control groups (Table4). This implies that same normal reference RDW values could apply to persons aged ≥ 6 years, but children aged < 6 years should have separate values. The reason behind high RDW in children ≤ 5 years may be due to a vigorous activity of bone marrow releasing immature RBC (reticulocytes) that are usually larger than normal red blood cells bringing about this variation of erythrocytes in children.

Red cell distribution normal range for children ≤ 5 years does not exist currently and globally despite studies showing similar findings of children < 2 years having a higher significant RDW mean as compared to the adult population [26]. It has also been noted that RDW becomes abnormal early before Hb or even MCV; thus, it is an excellent biomarker in children ≤ 5 years with anemia-related disorders [26]. The mean RDW of 13.2 ± 0.9 was reported by Qurtom et al. in individuals < 2 years old which varied slightly with our findings (control groups) of 15.95 (IQR = 4.2) of children ≤ 5 years and 14.2 (IQR = 1.7) of individuals aged 6–12 years, which could be due to variation of study designs or sociocultural epigenetic differences that affect phenotypic gene expression of biological characteristics in humans based on geographical locations [8, 28]. Another study reported a strong association between RDW and age; however, the age sets were majorly < 18 years and those > 18 years, where RDW increased with age which could be due to exposure of environmental factors or infections to these individuals as they aged [22].

The overall case group had a RDW median of 20.7 (IQR = 8.3), significantly (p < 0.0001) elevated compared to control group 14.5 (IQR = 2.7) indicating a marked anisocytosis in hemoglobinopathies. Hb SS had a RDW median of 25.4 (IQR = 5.5), the highest (p < 0.0001) among hemoglobinopathies listed by the present study with similar findings being recorded by Webster and Castro whose Hb SS had a mean of 22.4 ± 4 [39]. Hb SS + β thalassemia, was the second highest (p < 0.0001) with a RDW median of 23.3 (IQR = 7.9) while Hb SS + HbF recorded a median of 20.9 (IQR = 5.5), the lowest among Hb SS phenotypes. Similar order of decreasing magnitude of RDW was reported in anemic patients, with the highest value being seen in sickle cell anemia, sickle cell + β thalassemia, sickle cell trait, β-thalassemia and iron deficiency anemia [33]. RDW was the lowest in Hb SS + HbF due to the stability of Hb F that is known to reduce red cell anisocytosis since it does not enter HbS-polymerization phase, thus regulating clinical and hematological features of SCD [40]. In a prospective cohort study done in Kilifi area in Kenya, noted reduced morbidity and mortality in children < 5 years with Hb SS + HbF demonstrating the stability of hemoglobin F in resisting hemolysis [7]. In a study done in Kisumu County to differentiate malaria in SCD using hematological parameters, documented RDW of 14.9 (3.3) in HbSS phenotypes contrasts our finding of 25.4 (IQR = 5.45) in Hb SS, 20.85 (IQR = 5.47) in HbSS + HbF and 23.3(IQR = 7.9) in HbSS + β-thalassemia. This reduced RDW in that study could be associated with sample size error, analysis error or equipment used in the study since anisocytosis is inevitable in Hb SS phenotypes based on previous studies [12, 26, 33, 39].

Pure HbAS recorded a RDW median of 16.4 (IQR = 6.5) which was the lowest among hemoglobin disorders investigated by the present study. Though significantly elevated (p < 0.0001), RDW would generate many false positive results if used as a screening tool for SCT since its median was within the standard cutoff value of 19.9. Similar findings are documented by studies showing SCT patients having normal RDW, which makes it difficult to use this parameter to distinguish diseased (SCT) from the healthy population [41].

HbAS + HbF and HbAS + β thalassemia had medians of 24.15 (IQR = 7.43) and 20.9 (IQR = 10.5), respectively, which were high compared to the control group but, when subjected under statistical test, did not have statistical significance (p = 0.449, p = 0.791, respectively), suggesting that small sample size could have resulted into an abnormally high RDW. Similarly, there was no statistical significance (p = 1.0) between RDW of pure β-thalassemia compared to the control group with similar information being documented by previous studies that RDW cannot be used to discriminate β-thalassemia from β-thalassemia-free population [26].