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Inclusions in red blood cells containing Hb S or Hb C
Insights
Red cell inclusions in sickle cell disorders are linked to denatured hemoglobin S. Acid elution (AE) is more accurate than dark field microscopy (DFM) for visualizing these inclusions, correlating better with denatured hemoglobin levels.
Area of Science:
- Hematology
- Red Blood Cell Disorders
- Hemoglobinopathies
Background:
- Hemoglobinopathies like sickle cell disease (Hb SS) and Hb C disorders are characterized by abnormal hemoglobin variants.
- Red blood cell inclusions are a pathological feature in these disorders, but their accurate detection and characterization are crucial.
Purpose of the Study:
- To demonstrate and characterize red cell inclusions in individuals with Hb S or Hb C disorders.
- To compare the efficacy of three distinct methods for detecting and quantifying these inclusions.
- To investigate the correlation between red cell inclusions, denatured hemoglobin, and clinical severity, including splenic function.
Main Methods:
- Examination of unstained blood smears using dark field microscopy (DFM).
- Examination of blood smears after acid elution and staining (AE).
- Measurement of membrane-associated denatured hemoglobin (MADH) in red cell ghosts.
Main Results:
- Highest percentages of red cells with inclusions and MADH were observed in severe hemoglobin disorders (e.g., Hb SS, Hb SOArab).
- Results from AE and MADH measurements showed better correlation with clinical severity and splenic function compared to DFM.
- AE demonstrated higher accuracy in visualizing inclusions, correlating better with MADH levels than DFM.
Conclusions:
- Inclusions in sickling disorders are likely caused by denatured hemoglobin S.
- Acid elution (AE) is a more accurate method for visualizing red cell inclusions in hemoglobinopathies than DFM.
- The presence and quantity of inclusions correlate with disease severity and splenic status.
Abstract:
To demonstrate and characterize red cell inclusions in 101 persons with Hb S or Hb C disorders three methods were used: (1) examination of unstained blood smears by dark field microscopy (DFM), (2) examination of blood smears after acid elution and staining (AE), and (3) measurement of membrane-associated denatured haemoglobin (MADH) in ghosts. The control group had inclusions in less than 5% of red cells by DFM and AE and the mean percentage of MADH per total cellular Hb was 0.030+/-0.016%. The highest percentages of red cells with inclusions and of MADH were present in clinically severe haemoglobin disorders, e.g. homozygous sickle cell disease (Hb SS) with less than 10% Hb F and Hb SOArab, with successively lower percentages in moderate to severe disorders, e.g. Hb SS-alpha thalassaemia, Hb-S-beta0 thalassaemia, Hb SC disease, and Hb SS with more than 10% Hb F, indicating agreement in results by three methods. In asymptomatic or mild disorders, e.g. Hb-S-beta+ thalassaemia, Hb CC, Hb AC and Hb AS, the results by AE and measurements of MADH were the same or similar to those in controls, while those by DFM were different. Of 56 patients with Hb SS or Hb SC, the group with functional asplenia had higher percentages of MADH and of red cells with inclusions than those with functioning spleens. Our study suggests that inclusions in sickling disorders may be due to denatured Hb S, with AE being the more accurate method for visualizing these inclusions, as results by this method correlate better with the amount of MADH than those by DFM.