Mutant Hemoglobin Research Articles

Structural basis for the antipolymer activity of Hb [formula omitted] trapped in a tense conformation

The phenotypical severity of sickle cell disease (SCD) can be mitigated by modifying mutant hemoglobin S (Hb S, Hb α2β2s) to contain embryonic ζ globin in place of adult α-globin subunits (Hb ζ2β2s). Crystallographical analyses of liganded Hb ζζ2β2s, though, demonstrate a tense (T-state) quaternary structure that paradoxically predicts its participation in--rather than its exclusion from--pathological deoxyHb S polymers. We resolved this structure-function conundrum by examining the effects of α → ζ exchange on the characteristics of specific amino acids that mediate sickle polymer assembly. Superposition analyses of the βs subunits of T-state deoxyHb α2β2s and T-state CO-liganded Hb ζ2β2s reveal significant displacements of both mutant βsVal6 and conserved β-chain contact residues, predicting weakening of corresponding polymer-stabilizing interactions. Similar comparisons of the α- and ζ-globin subunits implicate four amino acids that are either repositioned or undergo non-conservative substitution, abrogating critical polymer contacts. CO-Hb ζ2βs2 additionally exhibits a unique trimer-of-heterotetramers crystal packing that is sustained by novel intermolecular interactions involving the pathological βsVal6, contrasting sharply with the classical double-stranded packing of deoxyHb S. Finally, the unusually large buried solvent-accessible surface area for CO-Hb ζ2β2s suggests that it does not co-assemble with deoxyHb S in vivo. In sum, the antipolymer activities of Hb ζ2β2s appear to arise from both repositioning and replacement of specific α- and βs-chain residues, favoring an alternate T-state solution structure that is excluded from pathological deoxyHb S polymers. These data account for the antipolymer activity of Hb ζ2β2s, and recommend the utility of SCD therapeutics that capitalize on α-globin exchange strategies.

The Effects of Moringa oleifera Leaves Extracts on Sickle Cell Hemoglobin

Sickle cell disease (SCD) caused by hemoglobin mutant called hemoglobin S or HbS, which originated from the replacement of a hydrophilic amino acid residue-glutamic acid by a hydrophobic moiety valine, at the sixth position of the β-chain of hemoglobin molecule. This loss of charge causes the erythrocytes to sickle under hypoxic conditions. The phytochemical, proximate and amino acid compositions of the leaves extracts of the plant Moringa oleifera were assayed to determine their antisickling effects on sickle cell hemoglobin. The proximate composition of the leaves showed the data:moisture (6.0±1.2mg/100g); crude protein (28.0±1.0mg/100g); carbohydrate (48.10±0.13mg/100g); crude fiber (10.05±0.01mg/100g); ash(9.30±0.01 mg/100g) ; pH 6.25 and crude fat (2.5±0.0 mg/100g). The different extracts including the crude aqueous extracts (CAE) revealed a preponderance of the following phytomedicines:-alkaloids

Mechanistic Insights into Forced Chromatin Looping Mediated Activation of Fetal Globin Gene Expression

The β-globin enhancer, called locus control region (LCR), is required for high level expression of all b-type globin genes. The LCR is in physical proximity with the genes it controls, with contacts shifting from embryonic (ε) to fetal (γ) and finally to adult (δ and β) globin gene promoters during development. In prior studies we showed that forced chromatin contacts between the LCR and the β-globin promoter led to transcriptional activation, suggesting that LCR-promoter looping causally underlies β-globin transcription (Deng et al. Cell 2012). In these studies, the transcription co-factor Ldb1 was tethered to the β-globin promoter using artificial zinc finger (ZF) DNA binding proteins, to trigger the promoter-LCR interaction. We subsequently showed that tethering Ldb1 to the promoters of developmentally silenced embryonic or fetal globin genes reactivated their expression in adult erythroblasts in a manner dependent on looped contacts with the LCR (Deng et al. Cell 2014). This work established a novel strategy to raise fetal globin expression in patients with sickle cell anemia.To examine mechanistically the effects of chromatin looping on gene expression we performed single molecule RNA FISH experiments to precisely measure transcription output at individual alleles before and after enforced LCR-γ-globin looping. The experiments were carried out in primary human erythroblasts, which produce elevated levels of γ-globin when exposed to culture conditions. Preliminary data suggest that the majority of transcripts emerge from the β-globin gene with a smaller fraction of transcripts coming from the γ-globin gene as expected. Among cells producing any type of globin primary transcripts, a significant fraction (25%-35%) of cells co-express γ- and β-globin. Importantly, γ- and β-globin are frequently transcribed from the same allele. Forced juxtaposition of the LCR and the γ-globin promoter increases the number of alleles expressing only γ-globin while reducing the number of alleles expressing only the β-globin gene. This result is consistent with the γ- and β-globin genes competing for LCR activity, and emphasizes the usefulness of this approach in the context of sickle cell anemia in which not only elevated levels of fetal hemoglobin but also reduction of the mutant hemoglobin are desirable. Surprisingly, however, the proportion of alleles co-expressing γ-globin and β-globin remains largely constant. We are testing whether co-expression from the same allele is LCR independent. Finally, our studies suggest that LCR-promoter contacts increase the probability of transcription of a given allele.We will also present work addressing the critical question as to how alteration of chromatin architecture overcomes the action of transcriptional repressive complexes, such as Bcl11a, which normally maintain embryonic and fetal globin genes in a repressed state throughout adulthood.In sum, our studies produce a deeper understanding of the interplay of chromatin architecture and gene expression in a system that holds great potential for therapeutic application in patients with hemoglobinopathies. DisclosuresNo relevant conflicts of interest to declare.

How people in Benin assess a couple's risk of having a baby with sickle cell disease.

Sickle cell disease (SCD) is a genetic disease resulting from the inheritance from both parents of a mutant hemoglobin gene. Its occurrence can, at best, be prevented, and its daily life consequences can, at least, be limited. As the mutant gene is recessive, it should be useful for people living in countries where SCD is endemic to know their own genetic status and that of their actual or potential partner in order to assess the risk of having a baby with SCD. The present study aimed at examining how a convenience sample of 128 lay people and nine physicians in Benin judge the likelihood that a newborn will suffer from SCD as a function of the genetic status of the baby's mother and father. As expected, several qualitatively different clusters of participants were found. A minority (29%) made judgments that were largely consistent with the correct rule for determining the likelihood of disease. A larger group (37%) expressed, however, the view that (a) to know a child's likelihood of suffering from SCD, information is needed about the genetic statuses of both parents and (b) this likelihood depends additively on these genetic statuses. Finally, another group (34%) thought that, if one parent is suffering from SCD or is a carrier of a sickle gene, the likelihood that the child will have SCD is high, irrespective of the other parent's status. Thus, even among a relatively well-educated group of people in Benin, only a minority used the correct judgment rule when assessing the risk of SCD. Work needs to be done to educate the population regarding the proper way to combine information.

Vasculopathy and pulmonary hypertension in sickle cell disease.

Sickle cell disease (SCD) is an autosomal recessive disorder in the gene encoding the β-chain of hemoglobin. Deoxygenation causes the mutant hemoglobin S to polymerize, resulting in rigid, adherent red blood cells that are entrapped in the microcirculation and hemolyze. Cardinal features include severe painful crises and episodic acute lung injury, called acute chest syndrome. This population, with age, develops chronic organ injury, such as chronic kidney disease and pulmonary hypertension. A major risk factor for developing chronic organ injury is hemolytic anemia, which releases red blood cell contents into the circulation. Cell free plasma hemoglobin, heme, and arginase 1 disrupt endothelial function, drive oxidative and inflammatory stress, and have recently been referred to as erythrocyte damage-associated molecular pattern molecules (eDAMPs). Studies suggest that in addition to effects of cell free plasma hemoglobin on scavenging nitric oxide (NO) and generating reactive oxygen species (ROS), heme released from plasma hemoglobin can bind to the toll-like receptor 4 to activate the innate immune system. Persistent intravascular hemolysis over decades leads to chronic vasculopathy, with ∼10% of patients developing pulmonary hypertension. Progressive obstruction of small pulmonary arterioles, increase in pulmonary vascular resistance, decreased cardiac output, and eventual right heart failure causes death in many patients with this complication. This review provides an overview of the pathobiology of hemolysis-mediated endothelial dysfunction and eDAMPs and a summary of our present understanding of diagnosis and management of pulmonary hypertension in sickle cell disease, including a review of recent American Thoracic Society (ATS) consensus guidelines for risk stratification and management.

Insight into the allosteric mechanism of Scapharca dimeric hemoglobin.

Allosteric regulation is an essential function of many proteins that control a variety of different processes such as catalysis, signal transduction, and gene regulation. Structural rearrangements have historically been considered the main means of communication between different parts of a protein. Recent studies have highlighted the importance, however, of changes in protein flexibility as an effective way to mediate allosteric communication across a protein. Scapharca dimeric hemoglobin (HbI) is the simplest possible allosteric system, with cooperative ligand binding between two identical subunits. Thermodynamic equilibrium studies of the binding of oxygen to HbI have shown that cooperativity is an entropically driven effect. The change in entropy of the system observed upon ligand binding may arise from changes in the protein, the ligand, or the water of the system. The goal of this study is to determine the contribution of the change in entropy of the protein backbone to HbI cooperative binding. Molecular dynamics simulations and nuclear magnetic resonance relaxation techniques have revealed that the fast internal motions of HbI contribute to the cooperative binding to carbon monoxide in two ways: (1) by contributing favorably to the free energy of the system and (2) by participating in the cooperative mechanism at the HbI subunit interface. The internal dynamics of the weakly cooperative HbI mutant, F97Y, were also investigated with the same methods. The changes in backbone NH dynamics observed for F97Y HbI upon ligand binding are not as large as for the wild type, in agreement with the reduced cooperativity observed for this mutant. The results of this study indicate that interface flexibility and backbone conformational entropy of HbI participate in and are important for the cooperative mechanism of carbon monoxide binding.

Forming A Blood Tie

Last month, the health care giant Baxter agreed to buy AesRx, a virtual biotech firm developing a drug for sickle cell disease. If successful, the drug, Aes-103, could be the first dedicated treatment for the more than 85,000 people in the U.S. with sickle cell. But despite a sizable market and significant need, the drug might just as easily have disappeared into obscurity. Developing drugs for sickle cell has proven daunting, and investors were not interested in Aes-103. The compound was kept alive in large part because of a National Institutes of Health program called Therapeutics for Rare & Neglected Diseases (TRND). It was started in 2009 to forge partnerships with companies or institutions in need of help with drug development projects that are high risk but potentially offer high therapeutic reward. Sickle cell is the result of a mutation in hemoglobin, the component of red blood cells tasked with ...

Detection of sickle cell hemoglobin in Haiti by genotyping and hemoglobin solubility tests.

Sickle cell disease is a growing global health concern because infants born with the disorder in developing countries are now surviving longer with little access to diagnostic and management options. In Haiti, the current state of sickle cell disease/trait in the population is unclear. To inform future screening efforts in Haiti, we assayed sickle hemoglobin mutations using traditional hemoglobin solubility tests (HST) and add-on techniques, which incorporated spectrophotometry and insoluble hemoglobin separation. We also generated genotype data as a metric for HST performance. We found 19 of 202 individuals screened with HST were positive for sickle hemoglobin, five of whom did not carry the HbS allele. We show that spectrophotometry and insoluble hemoglobin separation add-on techniques could resolve false positives associated with the traditional HST approach, with some limitations. We also discuss the incorporation of insoluble hemoglobin separation observation with HST in suboptimal screening settings like Haiti.

Lung function and six-minute walk test performance in individuals with sickle cell disease

BackgroundSickle Cell Disease (SCD), which is characterized by a mutation in the gene encoding beta hemoglobin, causes bodily dysfunctions such as impaired pulmonary function and reduced functional capacity. ObjectiveTo assess changes in pulmonary function and functional capacity in patients with SCD and to identify the relationships between these variables.MethodWe evaluated sociodemographic, anthropometric, lung function (spirometry), respiratory (manovacuometer), peripheral muscle strength (Handgrip strength - HS) and functional capacity (i.e., the six-minute walk test) parameters in 21 individuals with SCD (average age of 29±6 years). Shapiro-Wilk, paired Student's, Wilcoxon, Pearson and Spearman correlation tests were used for statistical analyses, and the significance threshold was set at p<0.05. ResultsA total of 47.6% of the study subjects exhibited an altered ventilation pattern, 42.8% had a restrictive ventilatory pattern (RVP) and 4.8% exhibited a mixed ventilatory pattern (MVP). The observed maximal inspiratory pressure (MIP) values were below the predicted values for women (64 cmH2O), and the maximum expiratory pressure (MEP) values, HS values and distance walked during the 6MWT were below the predicted values for both men (103 cmH2O, 39 Kgf and 447 m, respectively) and women (64 cmH2O; 27 Kgf; 405 m, respectively). Positive correlations were observed between maximum voluntary ventilation (MVV) and MEP (r=0.4; p=0.046); MVV and BMI (r=0.6; p=0.003); and between HS and MIP (r=0.7; p=0.001), MEP (r=0.6; p=0.002), MVV (r=0.5; p=0.015), distance walked in the 6MWT (r=0.4; p=0.038) and BMI (r=0.6; p=0.004). ConclusionsSCD promoted changes in lung function and functional capacity, including RVPs and a reduction in the distance walked in the 6MWT when compared to the predictions. In addition, significant correlations between the variables were observed.

Proteomic analysis reveals novel proteins associated with progression and differentiation of colorectal carcinoma.

The objective of this study is to characterize differential proteomic expression among well-differentiation and poor-differentiation colorectal carcinoma tissues and normal mucous epithelium. The study is based on quantitative 2-dimensional gel electrophoresis and analyzed by PDquest. Excluding redundancies due to proteolysis and posttranslational modified isoforms of over 600 protein spots, 11 proteins were revealed as regulated with statistical variance being within the 95th confidence level and were identified by peptide mass fingerprinting in matrix assisted laser desorption/ionization time-of-flight mass spectrometry. Progression-associated proteins belong to the functional complexes of tumorigenesis, proliferation, differentiation, metabolism, and the regulation of major histocompatibility complex processing and other functions. Partial but significant overlap was revealed with previous proteomics and transcriptomics studies in CRC. Among various differentiation stage of CRC tissues, we identified calreticulin precursor, MHC class I antigen (human leukocyte antigen A ), glutathione S-transferase pi1, keratin 8, heat shock protein 27, tubulin beta chain, triosephosphate, fatty acid-binding protein, hemoglobin (deoxy) mutant with val b 1 replaced by met (HBB), and zinc finger protein 312 (FEZF2). Their functional networks were analyzed by Ingenuity systems Ingenuity Pathways Analysis and revealed the potential roles as novel biomarkers for progression in various differentiation stages of CRC.

1P094 α鎖あるいはβ鎖の近位ヒスチジンがグリシンに置換された変異ヘモグロビンの機能と構造(02. ヘム蛋白質,ポスター,第52回日本生物物理学会年会(2014年度))

1P094 α鎖あるいはβ鎖の近位ヒスチジンがグリシンに置換された変異ヘモグロビンの機能と構造(02. ヘム蛋白質,ポスター,第52回日本生物物理学会年会(2014年度))

Analysis of sickle hemoglobin

Background: Sickle Hemoglobin is the most common pathological hemoglobin mutation worldwide which forms sickle shape or elongated forms on deoxygenation. According to malaria hypothesis, there is resistance against malaria by the heterozygous carrier state. There is spread of Hb S allele from the highly malarial regions to most other regions worldwide. The study was conducted as Sickle Hemoglobin has been frequently seen in our institute. Materials and Methods: This study was conducted in the department of pathology, Maharajgunj medical campus, Tribhuwan University Teaching hospital, from January 2011 to January 2013. All the relevant data were obtained from the archives of the department. Hemoglobin electrophoresis was performed by Cellulose Acetate Electrophoresis at alkaline pH Method. Sickling test was done using sodium dithionite. Results: A total of 35 cases were diagnosed as sickle cell disorder. The male: female ratio was 2.5:1with the commonest age group 11-20 years (42.8%). Sixty percent showed hemoglobin between 6-8 gm/ dl. Tharu, Chaudhary and Tharu, Rana (91.3%) were the commonest ethnic group with both sickle cell anemia and trait. Lama, Neupane and Baral suffered only from trait. Mean corpuscular volume ranged from 82 to 94 fL. On peripheral blood smear reversible and irreversible sickle cells were present in 88.56 % of cases and 11.42% were devoid of sickle cells. Sickling test was positive in all cases. On cellulose acetate electrophoresis 22 (62.85%) cases had Hb S, Hb F and Hb A2 or HbS and Hb A2 where as 13(37.1%) showed Hb S, Hb A and Hb A2 variants. Conclusion: The commonest ethnic group of Tharus(91.3%), Chaudhary and Rana are from Terai region of Nepal which is a malarial zone. However 8.4% of cases were seen in other ethnic group who are from non-endemic region. DOI: http://dx.doi.org/10.3126/jpn.v3i6.8989 Journal of Pathology of Nepal (2013) Vol. 3, 437-440

Autoxidation and Oxygen Binding Properties of Recombinant Hemoglobins with Substitutions at the αVal-62 or βVal-67 Position of the Distal Heme Pocket

The E11 valine in the distal heme pocket of either the α- or β-subunit of human adult hemoglobin (Hb A) was replaced by leucine, isoleucine, or phenylalanine. Recombinant proteins were expressed in Escherichia coli and purified for structural and functional studies. (1)H NMR spectra were obtained for the CO and deoxy forms of Hb A and the mutants. The mutations did not disturb the α1β2 interface in either form, whereas the H-bond between αHis-103 and βGln-131 in the α1β1 interfaces of the deoxy α-subunit mutants was weakened. Localized structural changes in the mutated heme pocket were detected for the CO form of recombinant Hb (rHb) (αV62F), rHb (βV67I), and rHb (βV67F) compared with Hb A. In the deoxy form the proximal histidyl residue in the β-subunit of rHb (βV67F) has been altered. Furthermore, the interactions between the porphyrin ring and heme pocket residues have been perturbed in rHb (αV62I), rHb (αV62F), and rHb (βV67F). Functionally, the oxygen binding affinity (P50), cooperativity (n50), and the alkaline Bohr Effect of the three α-subunit mutants and rHb (βV67L) are similar to those of Hb A. rHb (βV67I) and rHb (βV67F) exhibit low and high oxygen affinity, respectively. rHb (βV67F) has P50 values lower that those reported for rHb (αL29F), a B10 mutant studied previously in our laboratory (Wiltrout, M. E., Giovannelli, J. L., Simplaceanu, V., Lukin, J. A., Ho, N. T., and Ho, C. (2005) Biochemistry 44, 7207-7217). These E11 mutations do not slow down the autoxidation and azide-induced oxidation rates of the recombinant proteins. Results from this study provide new insights into the roles of E11 mutants in the structure-function relationship in hemoglobin.

Hb Papanui [α99(G8)Lys→Arg; HBA2: c.299A>G]: A Novel Silent Substitution Interfering in Hb A1c Determination

We report a novel hemoglobin (Hb) mutation [Hb Papanui [α99(G8)Lys→Arg; HBA2: c.299A>G] that was identified in an individual with an inappropriately low Hb A1c value of 2.8% when using an ion exchange method. Although occurring at a well conserved site, the conservative substitution was not associated with any adverse clinical features.

Crystallographic structure determination of B10 mutants of Vitreoscilla hemoglobin: role of Tyr29 (B10) in the structure of the ligand-binding site.

Site-directed mutants of the gene encoding wild-type Vitreoscilla hemoglobin were made that changed Tyr29 (B10) of the wild-type Vitreoscilla hemoglobin (VHb) to either Phe or Ala. The wild-type and the two mutant hemoglobins were expressed in Escherichia coli and purified to homogeneity. The binding of the two mutants to CO was essentially identical to that of wild-type VHb as determined by CO-difference spectra. Circular-dichroism spectra also showed the two mutants to be essentially the same as wild-type VHb regarding overall helicity. All three VHbs were crystallized and their structures were determined at resolutions of 1.7-1.9 Å, which are similar to that of the original wild-type structure determination. The Tyr29Phe mutant has a structure that is essentially indistinguishable from that of the wild type. However, the structure of the Tyr29Ala mutant has significant differences from that of the wild type. In addition, for the Tyr29Ala mutant it was possible to determine the positions of most of the residues in the D region, which was disordered in the originally reported structure of wild-type VHb as well as in the wild-type VHb structure reported here. In the Tyr29Ala mutant, the five-membered ring of proline E8 (Pro54) occupies the space occupied by the aromatic ring of Tyr29 in the wild-type structure. These results are discussed in the context of the proposed role of Tyr29 in the structure of the oxygen-binding pocket.

Abstract TP261: Spontaneous Strokes In Sickle Cell Transgenic Mice Captured Ex Vivo With Magnetic Resonance Imaging

With 11% of children with sickle cell disease (SCD) having strokes, many as early as 1 to 5 years old, the underlying mechanisms are still unclear, and it is still unknown why this subpopulation has greater risk. More will suffer from subclinical silent strokes that cause cognitive rather than physical impairments. Sickling of red blood cells due to a point mutation in hemoglobin induces chronic inflammation and vascular remodeling in SCD. We have previously shown that powerful proteases, cathepsins, are activated in SCD to promote vascular remodeling that could lead to stroke. However, a mouse model of detectable sickle cell stroke is required to study mechanism and intervention strategies. Here, we used Townes sickle transgenic mouse, with murine hemoglobin knocked out and replaced with human, sickle hemoglobin, to test the hypothesis that sickle transgenic mice have MRI detectable strokes and increased cathepsin proteolytic activity. Three mice homozygous for sickle mutation (SS) and 6 heterozygous littermate controls (AS) were perfused at 8 weeks. Brains were isolated, embedded in gadolinium-agarose, and scanned using a Bruker 9.4 Tesla magnet with a T2-weighted protocol (TE = 13.4 ms. TR= 10,000, 512 x 512 x 70). Of the 3 SS mice, 2 had abnormalities suggestive of stroke (Figure 1, red arrows), with none noted in the AS controls. Separately, immunohistochemistry with anti-cathepsin K antibody indicated a gene dosing effect of sickle allele on cathepsin K expression (SS&gt;AS&gt;AA), verified in aorta homogenates by multiplex cathepsin zymography assays (n=3, p&lt;.05). To conclude, we have identified strokes in sickle transgenic mice with T2 images. These mouse stroke scans, without occlusive or chemical intervention, will be an improved model to test spontaneous stroke interventions specific to sickle cell disease including targeting upregulated cathepsins.

Screen detection of serrated polyps by stool DNA multi-target testing (sDNA-MT): Comparison against fecal immunochemical occult blood testing (FIT).

341 Background: Precursors to 1/3 of colorectal cancer (CRC), serrated polyps have historically been underdetected by screening due to their inconspicuous, nonhemorrhagic, and proximal nature. New generation sDNA-MT show high sensitivity for both CRC and advanced adenomas (Gastroenterology 2012;142:248), but screen detection of serrated polyps by this approach requires further validation. Aim: To assess and compare noninvasive detection of sessile serrated polyps &gt;1cm (SSP) by sDNA-MT and a commercial quantitative FIT. Methods: In a blinded prospective study, a single stool sample used for both tests was collected from 456 asymptomatic adults prior to screening or surveillance colonoscopy (criterion standard). All 29 patients with SSP were included as cases and all 232 with no neoplastic findings as controls. Buffered stool samples were processed and frozen on receipt; Exact Sciences performed sDNA-MT in batches using optimized analytical methods (Gastroenterology 2012;142:S770). The sDNA-MT panel targets methylated BMP3 (mBMP3) and NDRG4, mutant KRAS, β-actin, and hemoglobin. FIT (Polymedco OC-FIT Check) was performed in separate lab &lt;2 days post defecation and evaluated at cutoffs of 50 (FIT-50) and 100 ng/ml (FIT-100). Results: Age/sex distributions were similar in cases and controls: 40% vs 45% &gt; 65 years and 51% vs 59% women, p=NS respectively. SSP median size was 1.2cm (range 1-3cm), 93% were proximal, and 64% had synchronous diminutive polyps. Among sDNA-MT markers, mBMP3 proved highly discriminant for SSP detection (AUC=0.87, p&lt;0.00001); other DNA markers were less informative and provided no incremental sensitivity. Hemoglobin alone showed no discrimination (AUC=0.50, p=NS). At matched specificities, SSP detection by stool mBMP3 was significantly greater than by either FIT-50 or FIT-100 (Table). Conclusions: From a screening and surveillance setting, SSP can be detected noninvasively by stool assay of exfoliated DNA markers, especially mBMP3. FIT appears to have no value in SSP detection. [Table: see text]

Hb Plasencia [α125(H8)Leu→Arg (α2)] is a Frequent Cause of α+-Thalassemia in the Portuguese Population

Hb Plasencia is a thalassemic hemoglobin (Hb) mutation caused by a leucine to arginine replacement at residue 125 of the α2-globin chain (HBA2:c.377T>G). This variant was first described in the heterozygous state in association with a very mild α-thalassemic phenotype in three members of a Spanish family from Plasencia, Western Spain. Reviewing the molecular characterization of 308 Portuguese individual suspected of having α-thalassemia (α-thal) we found Hb Plasencia to be the second most frequent mutation after the –α3.7 deletion.

Sickle Cell Therapy and the Kinetics of Polymerization in the Presence of Ligands

Sickle cell disease results from a mutation of normal human hemoglobin that renders it capable of polymerization once oxygen is delivered. The process of deoxygenation involves quaternary as well as tertiary changes in Hb, and both changes appear to be involved in permitting HbS to assemble into the stiff polymers that distort cells and lead to the occlusion of the microvasculature. Kinetics are central to the disease, because if polymerization is slow enough to occur in the veinous return, the polymerization can be reversed upon reoxygention with minimal if any consequences. We have recently completed a detailed study of the polymerization kinetics of HbS in the presence of ligands. We have found that the kinetics are consistent with equilibrium measurements that show singly ligated HbS molecules will polymerize with only about 0.35 the probability of a deoxy HbS molecule. Given knowledge of these highly concentration sensitive rates, and a distribution of intracellular concentrations, we can calculate the likelihood of sickling at various points in the microcirculation. Because of the finite rate of oxygen delivery, we find sickling is most likely at the exit of the capillaries, which is where obstruction has been observed with intravital microscopy. When this is combined with the results for mixtures of fetal hemoglobin (HbF) or HbA, it becomes possible to determine therapeutic targets for the reduction of rates of sickling.

Search for antisickling agents from plants

The sickle cell disease is fatal in nature. Thousands of children are dying off due to this health problem throughout the globe. Due to the rapid development of diagnosis and clinical managements such patients are living up to a respectable age. But as there is no permanent cure the patients are suffering from bone and joint pain, jaundice, hepato-splenomegaly, chronic infections etc. The main physiological complicacy is due to the polymerization of sickle hemoglobin (HbS), (sickling process) inside the red blood cell (RBC) of these patients during deoxygenating state. The change of RBC from spherical to sickle shape is due to the polymerization of mutant hemoglobin (HbS) inside the RBC and membrane distortion during anoxic condition. The mechanism and the process of sickling are very complex and multifactor in nature. To get rid from such complicacies it is necessary to suitably and accurately stop the sickling of RBC of the patients. The potential anti-sickling agents either from natural sources and/or synthetic molecules may be helpful for reducing the clinical morbidity of the patients. A lot of natural compounds from plant extracts have been tried by several workers in recent past. Most of the studies are based on in vitro red cell sickling studies and their mode of action has not been properly understood. Although, few studies have been in vivo in nature pertaining to transgenic sickle animal model, there is paucity of data on the human studies. The result of such studies although has shown some degree of success, a promising anti-sickling agent is yet to be established.