Hb KODAIRA II: A HIGH OXYGEN AFFINITY VARIANT WITH A NOVEL MUTATION IN THEβ-GLOBIN GENE AND PHENOTYPIC IDENTITY TO Hb KODAIRA

We report a case of α⁺-thalassemia (α⁺-thal) trait caused by a novel frameshift mutation in exon 2 of the α2-globin gene, specifically a deletion of a single nucleotide at amino acid codon 81 [HBA2 c.244delT]. The mutation results in a premature termination of translation at codon 83.     

Molecular Variations Linked to the Grouping of β- and α-Globin Genes in Neonatal Patients with Sickle Cell Disease in the State of Pernambuco,Brazil

Various factors have been described as phenotypic modulators of sickle cell disease, such as levels of fetal hemoglobin (Hb F), presence of α-thalassemia (thal), and haplotypes of the β-globin genes. In order to characterize and determine the frequency of the β^S and β^C mutations and the prevalence of ?α^3.7-thal, 74 patients with sickle cell disease detected during neonatal screening in the State of Pernambuco, Brazil, were studied. The haplotypes of the β gene and ?α^3.7-thal were determined using polymerase chain reaction (PCR), and specific restriction endonucleases were used to establish the polymorphic sites of the haplotypes. The results showed the high frequency of the Central African Republic (CAR) or Bantu haplotype in the State of Pernambuco, Brazil. The low frequency of the Benin haplotype recorded in this study, in comparison with other states in northeast Brazil, suggests the diversity of origins of Afro-Brazilians in this region.     

Novel Mutations Responsible for α-Thalassemia in Iranian Families

α-Thalassemia (α-thal) is usually caused by deletions on the α-globin gene cluster and the role of point mutations is less well investigated. In the present study, a total of 1048 individuals with hypochromic microcytic anemia, who did not present the most common α-thal deletions, were referred for α-globin gene DNA sequencing. The nucleotide changes were studied and a total of five new mutations was identified, of which three were located on the α2 gene [codon7 (Lys→Stop), codon 34 (Leu→Pro) and codon 83 (Leu→Arg)] and two on the α1 gene [IVS-I-116 (A>G) and codon 44 (+C)]. These novel mutations not only explain new findings by molecular analysis of the α-globin gene but also have clinical importance due to their changes in α-globin production in means of decreased hemoglobin (Hb) related values. Moreover, considerations of its role in combination with other mutations, and the possibility of causing Hb H (β4) are yet to be studied.     

Characterization of Clinical and Laboratory Profiles of the Deletional α2-Globin Gene Polyadenylation Signal Sequence (AATAAA > AATA– –) in an Indian Population

Abstract

α-Thalassemia (α-thal) is characterized by large deletions involving the variable regions of α2 and/or α1 genes. Nondeletional mutations and polyadenylation (polyA) signal sequence motif mutations are less common. In this retrospective study, we describe a fragment length analysis-based polymerase chain reaction (PCR) assay for screening the T^Indian (AATAAA?>?AATA–?–; HBA2: c.*93_*94delAA) polyA signal deletion along with its clinical and laboratory presentation in 21 patients. Most of the patients were diagnosed in early adulthood with a clinical presentation ranging from asymptomatic in the heterozygous state to severe Hb H disease with a prominent hemolytic component in the homozygous state. On genetic analysis, 14 patients were found to be homozygotes, five were compound heterozygotes and two were heterozygotes. Thus, the T^Indian polyA signal deletion is common in the Indian population and should be screened for in patients with nondeletional α-thal mutations.     

Decline of Renal Function Is Associated with Proteinuria and Systolic Blood Pressure in the Morning in Diabetic Nephropathy

The aim of this study was to investigate a significance of increased proteinuria in the morning and the effects of antihypertensive treatment on proteinuria and arterial blood pressure in the progression of chronic renal insufficiency in type 2 diabetic patients with hypertension and nephropathy. In three 24-hr urine samples and blood pressure monitoring, separated into a night-and daytime and spot urine in the morning, variation in protein-creatinine ratio (g/g) and blood pressure were assessed in 24 (58 ± 3 years old; M/F: 17/7) diabetic patients with hypertension and nephropathy. Furthermore, the effects of antihypertensive therapy of combinations of angiotensin converting enzyme (ACE) inhibitor, calcium antagonists, diuretics, and α1 blocker were evaluated in 3 years. Home blood pressure measurement was carried out every month and 24-hr urine was collected every 2 months. The baseline urine excretion of protein-creatinine ratio and blood pressure were (1.22 ± 0.13 g/g creatinine: 154/96 ± 6/5 mmHg) in daytime and (1.39 ± 0.13: 168/88 ± 15/7) in the morning. At the end of the study, significant associations among a decline of 24-hr creatinine clearance and both of the urine excretion of protein-creatinine ratio (r = 0.47, p < .01) and the levels of systolic blood pressure (r = 0.46, p < .01) and between the levels of systolic blood pressure and the urine excretion of protein-creatinine ratio in the morning (r = 0.57, p < .001) were demonstrated. However, there were no significant associations among other variables. Analysis of patients who had systolic blood pressure in the morning less than 140 mmHg revealed that 65% of these patients received doxazosin-averaged doses of 4.8 ± 1.5 mg daily. The levels of both blood pressure and proteinuria-creatinine ratio in the morning mainly associate with progression of renal function in diabetic patients with hypertension and nephropathy.     

Serum amyloid A and cholesterol: a pivotal role on inflammation

Abstract

Oxidative stress and the formation of cytotoxic aggregates of the presynaptic protein α-synuclein (AS) are two important events associated with the pathogenesis of Parkinson's disease (PD) and several other neurodegenerative diseases. In this context, extensive efforts have been made to elucidate the molecular basis of the cytotoxic synergy between oxidative stress and AS aggregation. In this study, we demonstrate that the exposure of AS to oxidative stress induced by UV radiation (AS_UV) blocks the protein fibrillation, leading to the formation of highly toxic fibril-incompetent oligomers. In addition, AS_UV exhibited stronger anti-fibrillogenic properties than H₂O₂-treated AS, inhibiting the fibrillation of unmodified AS at notably low concentrations. Mass spectrometry indicated that Met5 oxidation to Met-sulfoxide was the only modification promoted by UV exposure, which is reinforced by NMR data indicating that Met5 is the only residue whose amide resonance completely disappeared from the ¹H-¹⁵N HSQC spectrum after UV exposure. This result is supported by previous data that indicate that C-terminal Met residues (Met116 and Met127) and N-terminal Met1 are less susceptible to oxidation than Met5 because of the residual structure of the disordered AS monomer. Overall, our findings suggest that specific oxidation of Met5 might be sufficient to promote the formation of highly neurotoxic oligomers of AS.     

In Silico analysis of perturbed steroidogenesis and gonad growth in fathead minnows (P. promelas) exposed to 17α-ethynylestradiol

Abstract

The multi-factorial nature of adverse reproductive effects mediated by endocrine disrupting compounds (or EDCs) makes understanding the mechanistic basis of reproductive dysfunction a highly pertinent area of research. As a consequence, a main motivator for continued research is to integrate ‘multi-leveled’ complexity (i.e., from genes to phenotype) using mathematical methods capable of encapsulating properties of physiological relevance. In this study, an in silico stoichiometric model of piscine steroidogenesis was augmented with a ‘biomass’ reaction associating the underlying stoichiometry of steroidogenesis with a reaction representative of gonad growth. The ability of the in silico model to predict perturbed steroidogenesis and subsequent effects on gonad growth was tested by exposing reproductively active male and female fathead minnows (Pimephales promelas) to 88?ng/L of the synthetic estrogen, 17α-ethynylestradiol (EE2). The in silico model was parameterized (or constrained) with experimentally quantified concentrations of selected steroid hormones (using mass spectrometry) and fold changes in gene expression (using RT-qPCR) for selected steroidogenic enzyme genes, in gonads of male and female fish. Once constrained, the optimization framework of flux balance analysis (FBA) was used to calculate an optimal flux through the biomass reaction (analogous to gonad growth) and associated steroidogenic flux distributions required to generate biomass. FBA successfully predicted effects of EE2 exposure on fathead minnow gonad growth (%gonadosomatic index or %GSI) and perturbed production of steroid hormones. Specifically, FBA accurately predicted no effects of exposure on male %GSI and a significant reduction for female %GSI. Furthermore, in silico simulations accurately identified disrupted reaction fluxes catalyzing productions of androgens (in male fish) and progestogens (in female fish), an observation which agreed with in vivo experimentation. The analyses presented is the first-ever to successfully associate underlying flux properties of the steroidogenic network with gonad growth in fish, an approach which can incorporate in silico predictions with toxicological risk assessments.     

Leukotriene B4 12-hydroxydehydrogenase/15-ketoprostaglandinΔ13-reductase (LTB4 12-HD/PGR) responsible for the reduction of a double-bond of the α,β-unsaturated ketone of an aryl propionic acid non-steroidal anti-inflammatory agent CS-670

CS-670 is a non-steroidal anti-inflammatory agent with an α,β-unsaturated ketone structure. It exerts its pharmacological activity after being transformed to the active metabolite (2S,1′R,2′S)-trans-alcohol. Two consecutive reductions are needed for the formation of the active metabolite, reduction of the double-bond of the α,β-unsaturated ketone moiety, followed by reduction of the resulting saturated ketone. The objective of the current study was to identify the enzyme responsible for reduction of the double-bond. An enzyme purified from rat liver cytosol as a single band on sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was analysed by a Mascot database search of nano-LC tandem mass spectrometry (MS/MS) data and the enzyme was identified as 2-alkenal reductase (EC 1.3.1.74), which is known as an β-nicotinamide adenine dinucleotide phosphate (NADPH)-dependent alkenal/one oxidoreductase and has a role for leukotriene B₄ 12-hydroxydehydrogenase/15-ketoprostaglandinΔ13-reductase (LTB₄ 12-HD/PGR). The identification was confirmed by cloning LTB₄ 12-HD/PGR cDNA from rat liver, expressing it in Escherichia coli, and characterizing the properties of the enzyme. The identity was further supported by the subcellular localization in cytosol, a cofactor requirement for NADPH, substrate specificity, and substantial inhibition by 15-ketoPGF_2α, benzylideneacetophenone, indomethacin, and quercitrin. In addition to catalysing the biological reduction of eicosanoids, including prostaglandins, leukotrienes, and lipoxins, LTB₄ 12-HD/PGR was also determined to function as a xenobiotic-metabolizing enzyme.