Clinical heterogeneity of sickle cell disease in Indian subcontinent

INTRODUCTION

Sickle cell disease (Includes Sickle Cell Anemia) is one of the prevalent hemoglobinopathies in India and is present in both Tribal and non-tribal populations in different degrees spreading from north eastern states to central India states down to southern states of India around Nilgiri hills encompassing Kerala, Tamil Nadu, and Karnataka states.^[1-6] In the central Indian distribution stretching from western states of Gujarat and Maharashtra across Jharkhand, Chhattisgarh, Madhya Pradesh, up to eastern borders occupied by Odisha and Andhra Pradesh, are home to sickle cell disease with carrier frequency between 1% and 40% of the groups studied.^[1,4,6]

With this wide distribution of this monogenic disease, a great deal of clinical heterogeneity in their presentation and complications is not only seen across different states where the disease is common, the heterogeneity is also visible in the same population groups within limited geographical distribution. Thus making the disease extremely interesting for various expert groups, i.e., Clinicians from various specialties, anthropologists, population geneticists, molecular biologists, statisticians, Blood Transfusion Experts, Public health workers, and government of the day who tries to manage, understand, and control this disease. In this paper, an attempt will be made to understand the heterogeneity of clinical presentation of this monogenic disorder by analyzing relevant papers published from this country and around the world over the years.

SICKLE CELL DISEASE

Sickle cell disease is one of the very important symptomatic hemoglobinopathies in the world as well as in India. This disease is basically a monogenic disorder involving a single nucleotide mutation of beta globin gene leading to the replacement of an electronegative amino acid glutamine with an electroneutral amino acid valine at the position 6 of beta globin gene. This small change leads to a profound alteration in red cell biology causing it to adopt a sickled shape when exposed to low oxygen tension in the circulation. These sickled red cells quickly lose water becomes dehydrated and express strong adhesive molecules on its surface leading to red cell aggregation, its adhesion to endothelial cells and other formed elements of blood with recruitment of neutrophils and platelets, leading to vaso occlusion and hemolysis. Several cycles of this sickling process may lead to hemolysis, development of permanently sickled cells, clogging, and infarction of the spleen in some cases. The basic pathology of sickle cell disorders stems from anemia, vaso-occlusion, vaso-occlusion, and hemolysis in different degrees and in different parts of the circulation in different ages, leading to its diverse symptoms [Flow Diagram 1]. It is not easily understood that variability of clinical presentation of this disease among one particular ethnic group and in different parts of the country is contributed by molecular diversity of the beta globin gene itself related to production of different amount of sickling inhibitory fetal hemoglobin (HbF), molecular changes in alpha globin gene i which may contribute to amount of hemoglobin contained within a given red cells altering its intrinsic viscosity. Innumerable trans acting genetic elements across the genome finally determine the sickling status of the red cells as well as different components of vascular, cytokine/endocrine, and immune systems in the background where this hemolysis and vasoocclusion occur.^[7-14] Not to speak about environmental parameters such as nutrition and infection, surrounding temperature and humidity as well as the working condition of the individual where the disease finally expresses.

Close

Flow Diagram 1:

Pathobiology of SCD.

Export to PPT

The homozygous state of sickle cell gene causes sickle cell anemia and compound heterozygosity with other beta globin genes or interacting alpha globin gene decide some of the complex phenotypes of sickle cell gene.^[7,12-19] Double eloquent mutation on the same beta globin gene may produce severe disease with death in utero or at birth and trait status of this condition can cause sickling even in a heterozygous state as in Hb S- Oman or Hemoglobin S- Antilles. Eleven or more such hemoglobin variants have been reported in the international literature.^[20] These hemoglobinopathies are not generally found in India. One should be on the lookout for such double mutant on the same B^s gene if an apparent sickle cell trait show vaso occlussive changes.

CLINICAL PRESENTATION OF SICKLE CELL DISEASE

Everywhere in the world clinical presentation is very variable in SCD, though it is a monogenic disease. The reason for this has been described above. However, the clinical presentation of this hemoglobinopathy varies with different age groups and the underlying reason for some of the presentation also varies in different age groups. Tables 1 and 2 show different clinical presentations of Sickle Cell Disease.

India is a vast country and sickle cell disease arises or inherited with different genetic backgrounds. There are broadly four areas in the country where the disease is prevalent^[1-6] and shows some clinical variation. Areas 1 and 2: The broad area, i.e., central band of India stretching from states of Gujarat and Maharashtra in the west, spreading over Jharkhand, Chhattisgarh, and Madhya Pradesh in central India and in the east covering the states of Odisha and Andhra Pradesh. Maximum number of carriers and patients live in these areas distributed over many tribal and some cast groups.^[1,4,6] From this broad area, clinical heterogeneity of substantial magnitude is seen between patients in Gujarat and those from central Indian areas, i.e., Nagpur of Maharashtra, Chhattisgarh, etc., so patients in central India and Nagpur are more severely affected than those from state of Gujarat.^[4,6] 3: Third area is contributed by tea garden areas of Bengal and Assam, the population was originally from tribal communities of MP/Chhattisgarh and Odisha. They were transported generations ago as indentured laborers by the British rulers to work in the tea garden areas.^[2,21] However, there is very little mixing of this population with the native tribe or caste population. 4: This population group is in south India, i.e., in the states of Kerala and Tamil Nadu, affecting the tribal populations of the Nilgiri hills area and in the Waynad district of Kerala.^[3,22,23] 55% of this population were said to have milder disease though painful crisis is not uncommon.

Clinical presentation: Certain clinical presentations like priapism, pedal ulcer, hyposplenism^[24] in early infancy, and dactylitis are rare all over in Indian sickle cell disease patients. However, pedal ulceration has been reported from Delhi hospital in 8% of the patients attending the hospital while none of the HbS beta thal patients had pedal ulceration. Details of the origin of these patients are not known. This is a very small selected group.^[25]

Splenic sequestration is also rare (5% newborns). The proportion of patients in central India and in the Nagpur area having painful crisis is very high so also bilateral necrosis of hip joints and strokes in the young. In Gujarat areas and in south India, these complications are milder and shift to higher age groups.

Transfusion requirements are high in Nagpur and Central as well as in parts of Jharkhand and Chhattisgarh areas compared to tea garden workers and south Indian sickle cell patients. No large-scale data are available on the development of allo antibodies in sickle disease patients across in India but varies from 3% to 11%.^[26] There are widely different criteria used in this country to transfuse SCD patients.

Transfusion-transmitted hepatitis is a relatively common occurrence and is related not only to transfusion practice but also widespread practice of tattooing using common needle. Splenomegaly is present in a large number of SCD patients in India often exceeding 50% of the patient studied.^[5,6,17] In one of the studies, a large number of them turned out to be double heterozygote for HbS and the Beta thalassemia gene.^[17]

Ambient weather is very variable so also malaria intensity in different parts of India. The painful crisis in central India which has both extremes of hot and cold weather in different seasons the painful crisis increase during summer months in contrast in Gujarat and in the North eastern states of Assam and in Kerala state, this tendency increases during hot and humid rainy season.

Pulmonary syndromes, i.e., acute, chronic, and recurrent acute chest syndrome is also seen in higher proportion of patients in central India states.

Chronic pain is also very prevalent across the country but shows higher proportion of affected patients in Nagpur and Central India.^[12,23,27] though this is a universal symptom in the disease across the length and breadth of the country.

Bhat et al., in systematic study spanning over 24 weeks on a large number of patients across India in a multicentric study showed at least one episode of pain in 87% of patients.^[27]

However, opiate dependence in this disease is not widely reported and only occasional case report is available in English literature.

Cholelithiasis is common across the country with the disease and by the time an SCD patient reaches 30 years of age almost a third of such patients have gallstones. Increased serum bilirubin is also a very a common occurrence in this disease across the country and fluctuates with other associated complications.

Degree of Jaundice in these patients at stable state has been shown to be linked to UGTA1 enzyme gene polymorphism through the long TA repeat^[28] and hemoxygenase enzyme gene polymorphism. Hyperbilirubinemia in many studies has been linked to higher TA repeats in UGTA1 gene and recently, high levels of HbF in SCD have been described from central India with long TA repeat of Hem Oxygenase 1 gene.^[29] Gallstones were found by random screening in 10% adult patients with sickle cell disease.^[30]

In summary, it may be said that we do not have extensive data comparing the clinical presentation of a well-maintained cohort of sickle cell disease patients across various corners of the country; however, the general impression is that the disease is relatively less severe than its African counterparts. The severity regarding painful crisis, vaso-occlusive crises, and hemolytic crisis is generally more in central Indian patients. Splenomegaly during adulthood is common and so is jaundice. Strokes and bilateral necrosis of the hip joint are quite common in central India reaching >10% by 10 years of age. Same complication in Gujarat is seen about 5 years later.As transfusion practice is variable across the country, distribution of SCD patients with this infection is likely to be variable and is sparsely available in the literature. Chest syndrome in the young as well as in the elderly. Splenic sequestration is uncommon but seen.

Cardiovascular manifestations of sickle cell disease as a result of widespread endothelial disease become evident in the form of pulmonary hypertension, cardiac failure, and various types of arrhythmias, which appear late in the natural history of SCD and have not been studied extensively in the Indian context with regard to its heterogeneity. Cardiac dysfunction was studied in a cohort of patients with SCD in south Gujarat aged 18–45 years. 51% had diastolic dysfunction (improper filling is diastole 31% and inadequate relaxation in 20% of 37 patients studied. None of them had any systolic dysfunction by ECHO cardiography.^[31]

A group of fifty patients with sickle cell disease under 18 years of age were studied in Mumbai with tricuspid jet velocity for pulmonary arterial hypertension almost 40% had PAH and 7% had moderate to severe disease.^[32] Pulmonary fibrosis and chronic respiratory insufficiency are also a late manifestations of the disease and have not been looked extensively in this disease.

Chronic renal failure and hematuria related to papillary necrosis have been recorded in central India. Chronic kidney disease was more common in sickle cell anemia compared to sickle beta thalassemia (at 10% versus <1% by the age of 16). Glomerular hyperfiltration is one of the hallmarks of the condition. And was detected in 11% of sickle cell anemia patients and in similar proportion of sickle beta thalassemia patients.^[33]

Infections are common in sickle cell disease patients. In early childhood, death due to septicemia has been recorded; however, in contrast to elsewhere in the world, the organism identified in most of these patients was not pneumococcus but gram-negative bacilli.^[17,34] With the passing of age, splenic insufficiency supervenes despite splenomegaly in large groups of Indian patients and unusual organism like Salmonella Typhi are found in osteomyelitis cases, as also many other organisms associate themselves with various forms of sepsis.^[35,36] In a fairly large investigation on fever with sickle cell disease in children below 12 years, 18% had evidence of bacterial infection in the form of bacteremia in 20%, urinary infection 15%, meningitis 13%, and osteomyelitis 4% approximately. Majority of the infection was due to a gram-negative organism and chest infection was the most common.

Tendency to develop stroke was studied by transcranial Doppler velocity in Chhattisgarh SCD patient and at 11% patients younger than 12 had predisposing Doppler signal.^[37]

BIOLOGICAL AND ENVIRONMENTAL BASIS OF THE CLINICAL HETEROGENEITY OF SICKLE CELL DISEASE

As described in Table 1, major symptomatology of sickle cell disease emerges from two basic mechanisms, i.e., vaso-occlusion and hemolysis. They are interdependent in a complex way but some of the clinical manifestations are predominantly related to one or both the mechanisms. Critical vaso-occlusion leads to ischemia, infarction, organ insufficiency, pain, and chronic inflammatory response which indirectly increases plasma viscosity and makes endothelium a more adherent procoagulant carpet. Vaso-occlusion tends to occur in organs where blood moves slowly and also through sinusoidal networks like bone, spleen and in places where increased viscosity (intrinsic related to red cell hemoglobin content, degree of hemoglobin polymerization, dehydration, change in shape and adhesiveness of red cells, and extrinsic in terms of dehydration and increased sum total of cellular viscosity and plasma viscosity.

Blood is a non-Newtonian fluid hence its viscosity tends to increase disproportionately in smaller blood vessels due to changes in shear force and plasma skimming in the capillaries. In the red cell hemoglobin content, its polymerization and hydration status of the cell are the major determinant of viscosity of red cells as well as whole blood. HbF is a potent inhibitor of HbS polymerization, it influences the sickling behavior of red cells.

At birth HbF levels are very high and also hemoglobin levels and SCD patients are at birth may be asymptomatic but within few weeks HBF levels fall disproportionate to Hb levels and symptoms start. The rate of fall of HBF is not uniform in all patients. In HbS Beta thalassemia, this fall is slower. Co-inheritance of several other common hemoglobinopathies in India has different effects on sickling behavior of the red cells. Hb E does not influence this function greatly, but Hb D may aggravate some of the vasoocclussive feature of the disease.^[15,16]

Deletional alpha thalassemia is a common and largely a silent condition in India. There are two different types of deletional alpha thalassemia seen in this country, e.g., −4.2 deletion with higher amount of hemoglobin and −3.7 deletion associated with slightly lower hemoglobin level. Co-inheritance of these two types of deletion singly or in tandem produces varying degree of microcytosis and low intracellular Hb levels leading to lowering of viscosity and improvement of sickling phenomenon, though viscosity may slightly increase on the balance in India it improves many symptoms of Sickle Cell Disease.^[9-11] Prevalence of inheritance of alpha thalassemia gene reaches more than 90%,^[6,10-12] in some tribal populations and is variable across the country. Overall prevalence of this condition in India is around 14%. Alpha hemoglobin variant of the hemoglobin gene that produces abnormal alpha globin like Hb- J meerut, Hb Constant spring, etc., is rare variants and its impact on defining the severity of sickle cell disease in India is minimal.

It was noticed that sickle cell anemia in Indian and in Arabian peninsula are generally milder than its African/USA/Jamaican counterparts and this was related to higher levels of HBF in Indian and Arabian patients (more than 6% and up to 20% or more). This was related to the inheritance of HbS gene in the backbone of Arab - Indian haplotype of the beta globin gene. African variety of sickle cell anemia arose on three different Independent occasions on three different beta-globin haplotype background (Bantu, Benin, and Senegal haplotypes. Bantu is associated with the severest manifestation). In India, around 90% of our sickle cell gene is linked to the Arab-Indian haplotype as evidenced by + Xmn1 restriction enzyme cutting site on the beta-globin gene, Other 10% cases has combination of different haplotypes.^[8,8,13]

Being a very small number of cases in India having these aberrant haplotypes, their total impact on HBF production is not known. In Odisha cohort of sickle cell patients, hereditary persistence of HbF was shown to be another contributor to lowering severity of the disease.^[13] Similar changes could be preset in more extensive areas of the country but needs to be studied in the future.

Hemolysis in addition to contributing to anemia, splenomegaly, and Jaundice also releases free hemoglobin, lactate dehydrogenase (LDH), and other chemicals such as ADP and Arginase in the circulation. This free hemoglobin is involved in endothelial damage and vaso constriction by scavenging nitric oxide released from endothelial cells and free radicals generated from the process through methemoglobin, catalase, superoxide dismutase, and peroxidase-based reactions. This cause wide spread vascular damage. ADP stimulates platelet aggregation and release from platelets various ligands that influence neutrophil behavior. Free hemoglobin after binding with plasma haptoglobin is taken up by monocytes and macrophages through CD163 receptor and Heme moiety of hemoglobin is also taken up by macrophage through HCP19 Heme carrier protein 1) depending on haptoglobin phenotype and degree of involvement of macrophage/monocytes polarization of macrophage occurs with pro or anti-inflammatory cytokines. Product of hemolysis and its interaction on the circulatory system are given in Table 3 and combined effects of vasoocclusion and hemolysis cycle are depicted in a flow diagram in the paper.

Red cell enzymopathies like G6PD deficiency which is very common in different population groups in the country, reaching as high as 25% in some population group^[38] however in general, the figure of 6–10% across the country is quoted. This deficiency could contribute to varying association of clinical severity of the disease. As severity of the enzyme deficiency is variable, also the hemolysis and active generation of oxygen and peroxide radicals. African variety of G5 PD deficiency, i.e., A- is less severe and was shown to give varied interaction with sickle cell gene.^[39,40] Oxy, dexyhemoglobin combines with varying affinity with band 3 of red cell membrane so also hemichromes associated with SCD. These hemoglobin derivatives compete with Aldolase, Pyruvate dehydrogenase, and LDH of Embden Meyerhof pathway and alter the availability of these enzymes and NADPH generation. G6PD also can also work in a complementary manner, influencing the availability of these enzymes and NADPH flux and ATP in the red cells,^[41] providing a valid mechanism for aggravating the disease if severe G6PD deficiency is associated with. More work in this area is needed as also prevalence of some polymorphism in different population group of B and 3 protein of red cells which may influence such binding.

Large association studies^[42] involving 19 single-nucleotide variants in BCL11A, HBS1L-MYB, and HBG2 showed significant association with fetal hemoglobin [HbF] and α-thalassemia deletions were significantly associated with increased hemoglobin level and reduced risk of albuminuria, abnormal transcranial Doppler velocity, and stroke.^[42,43]

Stroke susceptibility was found to correlate with genetic polymorphism [ ]. In the interleukin (IL)4R 503, tumor necrosis factor (TNF) (−308), and ADRB2 27 genes were independently associated with susceptibility in the large vessel (LV) stroke subgroup, while variants in the VCAM1 (−1594) and LDLR NcoI genes were associated with small vessel (SV) stroke risk. The combination of TNF (−308) GG homozygosity and the IL4R 503P variant carrier status was associated with a particularly strong predisposition to LV stroke (odds ratio 5.5; 95% confidence interval (2.3–13.1).^[44]

DISCUSSION

India is a huge country of continental dimension harboring populations with diverse ancestry and lineage frequently modified by invasion settlement in addition in different era of large scale human destruction due to calamities and diseases have created population bottlenecks and subsequent expansion of small amount of remaining population. Moreover, insulation of our many tribal populations in relatively unaccessible parts of jungle and mountainous areas coupled with endogamy created unique sets of population groups sometimes defined by their tribal names. Infection like malaria, environmental conditions, i.e., humidity, temperature, and occupation, way of life (moving barefoot in jungle or mountainous area may initiate pedal ulcers which in SCD may refuse to heal), nutrition and nutritional deficiency, access to medical care all may determine the clinical presentation of Sickle cell disease in our country. Detailed studies of clinical presentation in each of these locations where the disease is common are not available. Only a snapshot of the varied clinical presentation of the disease is available and presented here.

Though the clinical feature of the disease is a composite of hemolysis and vaso occlusion; these two features are contributed by coexistence of multiple genetic factors all of which have not yet deciphered. It is also not quite understandable why the disease becomes milder after the age of 30 and different age groups present with different sets of problems, e.g., Young strokes are mostly vaso-occlussive but stroke in later years are mostly hemorrhagic? Vascular obstruction in sickle cell disease could involve both large and SV. Moa-moa formation as a cause of stroke has been reported more often from Indian subcontinent. The reason or genetic background for such causation is not known.

Asymptomatic elderly patients with sickle cell disease remain an enigma and remain to be explained. HBF and alpha thalassemia are two important determinants of the severity and presentation of the disease across the world in many studies. Beta hemoglobin gene haplotype in India does not explain the variation in symptoms as 90% of SCD patients HbS gene is on milder Arab- Indian haplotype. With same Arab Indian haplotype, HBF could be very variable, i.e., from 10% to 30%. The other addition of HBF levels is contributed by many quantitative trait loci (QTL) spread across the human genome including on that of X chromosome. Each of these QTL contributes a small amount of HBF positively and out of these BCL11 gene has been found to be important.^[7,42,45] Manipulation of this gene is now a basis of common gene therapy protocol for sickle cell disease and beta thalassemia.

Although it is said that high HBF reduces HbS polymerization making sickle cell disease (anemia) milder needs some modification. Only high level of HbF does not ensure mildness of the disease, but this HBF needs to be distributed in red cells in a way that at least 20% of the red cells should each have more than 10 pg of HBF or more. This is determined by F cells, i.e., HBf containing cell by HBF ratio.^[14]

Jaundice is an important manifestation of sickle cell anemia and its degree in Indian population particularly the hemolytic variety is contributed by a TA repeat polymorphism of UGTA! Gene and Heme Oxygenase gene. Many of our sickle cell disease patients are in effect sickle beta thalassemia patients who have more prominent splenomegaly and relatively milder presentations. These patients have not been identified in many series of SCD patients while describing their clinical features. Very little data is available in Indian literature on sickle retinopathy^[46] and diverse types of hepatopathy are seen in this disease. The two deletional alpha thal mutations, i.e., −4.2 and −3.7 in alpha gene is common but mostly in the trans form. Hence, HbH disease and Hb B Barts are rare in India. −4.2 deletion produces relatively less lowering of Hb in red cells, i.e., slightly more MCH than the other variety whether clinical severity of these two different mutations in homozygous state is different is not known. A major gap in our understanding of SCD in India is later development of various organs in sufficiency and their degree (Lung, Heart, and Kidneys) in different populations. However, a meta-analysis and case–control study from Chhattisgarh and Nagpur area on lung function test in sickle cell disease in younger patients aged between 11 and 17 years showed abnormal lung function and low nitric Oxide production in a large number of patients. Only 11% of the patients had normal lung function test and obstructive, restrictive, and mixed pattern was almost equally distributed.^[38,47,48] Severely affected patients constitute 15% of the patients studied.

In general, it is said that sickle cell trait is a benign condition but complications such as thrombosis and renal insufficiency with sickle cell trait have been reported and need more intense epidemiologic attention considering their huge numbers. Modulation of sickle cell disease associated with hemopoietic nutrient deficiency^[49] and trace elements like Zn deficiency is also another open area for study.^[50-54]

CONCLUSION

A few good review of this disease on Indian patients and more extensively distributed patients across the world will give our readers a broad view of this condition with protean clinical manifestations.