Many mediators activate eosinophils via transduction pathways involving the enzyme phosphatidylinositol 3-kinase. The initial investigation of wortmannin, a specific inhibitor of PI3-kinase, was of its effect on human and guinea pig eosinophil superoxide (O(2)(-)) release and degranulation in vitro. Subsequently, the effect on allergen- and Sephadex-induced bronchial inflammation and airway hyperresponsiveness (AHR) in vivo in guinea pigs was investigated. Wortmannin potently inhibited complement C5a-induced O(2)(-) generation and eosinophil peroxidase (EPO) release from human eosinophils, with 50% inhibition produced by a 1-10 nM concentration. Both aerosol allergen challenge of sensitized guinea pigs and intravenous injection of Sephadex beads in normal guinea pigs caused, in 24 h, significant eosinophilia and increased EPO activity in bronchoalveolar lavage fluid (BALF) and AHR to intravenous acetylcholine and histamine. In the allergic model, intranasal pretreatment with wortmannin had no effect on BALF eosinophilia, but dose dependently inhibited BALF EPO activity. At 1 mg/kg, the drug abolished the AHR to histamine, but not acetylcholine. In the Sephadex model, the drug significantly inhibited all three parameters (eosinophilia, increased EPO activity, and AHR to both spasmogens). These results show that wortmannin is a potent inhibitor of human eosinophil degranulation and that when administered intranasally can prevent AHR in allergen-challenged guinea pigs, probably by inhibiting eosinophil degranulation, but not their accumulation in BALF. This may be relevant to the possible clinical utility of wortmannin in conditions involving eosinophilic inflammation and AHR.

Objective: This study aimed to document the transition of hemoglobin (Hb) F levels from early childhood to adulthood in Kuwaiti sickle cell disease patients, investigating its relationship to sex, Hb genotype and coexistence of α-thalassemia trait. Subjects and Methods: The following parameters were extracted from the patients’ records: age, sex, Hb, mean corpuscular volume, mean corpuscular Hb, red blood cell count, Hb F, Hb S, Hb A₂ and α-globin genotype. Hb quantitation was performed with cation exchange HPLC, while α-globin genotype was determined by PCR. Results: Records were available for 149 patients, made up of 94 SS and 55 Sβ⁰thal; 83 males and 66 females, aged 3 months to 60 years (mean 10.5 ± 1.8). The mean Hb F level in the whole population was 21.5 ± 8.1% and was not significantly different between males and females, and SS or Sβ⁰thal. When the age groups were analyzed, the Hb F level was highest (28.9 ± 10.9%) in those below 5 years. Indeed, patients ≤2 years had a mean level of 31.9 ± 13.0%. There was no significant difference in the Hb F levels in SS patients with or without coexistent α-thal trait. Conclusions: Kuwaiti sickle cell disease patients below 5 years of age have close to 30% Hb F and this is probably a major reason why they usually do not present before this age, unlike patients elsewhere who present within the first year of life.

Hemoglobin genotype and HBB haplotype are established genetic factors that modify the clinical phenotype in sickle cell disease (SCD). Current methods of establishing these two factors are cumbersome and/or prone to errors. The throughput capability of next generation sequencing (NGS) makes it ideal for simultaneous interrogation of the many genes of interest in SCD. This study was designed to confirm the diagnosis in patients with HbSS and Sβ-thalassemia, identify any ß-thal mutations and simultaneously determine the ßS HBB haplotype. Illumina Ampliseq custom DNA panel was used to genotype the DNA samples. Haplotyping was based on the alleles on five haplotype-specific SNPs. The patients studied included 159 HbSS patients and 68 Sβ-thal patients, previously diagnosed using high performance liquid chromatography (HPLC). There was considerable discordance between HPLC and NGS results, giving a false +ve rate of 20.5% with a sensitivity of 79% for the identification of Sβthal. Arab/India haplotype was found in 81.5% of βS chromosomes, while the two most common, of the 13 β-thal mutations detected, were IVS-1 del25 and IVS-II-1 (G>A). NGS is very versatile and can be deployed to simultaneously screen multiple gene loci for modifying polymorphisms, to afford personalized, evidence-based counselling and early intervention.

Patients with sickle cell disease (SCD) in Kuwait have elevated HbF levels ranging from ~10–44%; however, the modulating factors are unclear. We investigated the association of single nucleotide polymorphisms (SNPs) at BCL11A, HBS1L-MYB and HBB with HbF levels in 237 Kuwaiti SCD patients, divided into 3 subgroups according to their HbF levels. Illumina Ampliseq custom DNA panel was used for genotyping and confirmed by arrayed primer extension or Sanger sequencing. In the BCL11A locus, the CC genotype of rs7606173 [χ2 = 16.5] and (GG) of rs10195871 [χ2 = 15.0] were associated with Hb-F1 and HbF-2 subgroups, unlike rs1427404-T [χ2 = 17.3], which showed the highest association across the three subgroups. HBS1L-MYB locus revealed 2 previously-described SNPs (rs66650371 [χ2 = 9.5] and rs35795442 [χ2 = 9.2]) and 2 previously-unreported SNPs, (rs13220662 [χ2 = 6.2] and rs1406811 [χ2 = 6.7]) that were associated with the HbF-3 subgroup, making this the key locus elevating HbF to the highest levels. HBB cluster variants were associated with lower levels of HbF (β = −1.1). We report four previously-unpublished variants showing significant association with HbF. Each of the three quantitative trait loci affects HbF levels differently; unique SNPs, especially in HBS1L-MYB, elevate HbF to the highest levels.

Background Systemic lupus erythematosus (SLE) is an autoimmune inflammatory disease which involves the loss of self-tolerance with hyperactivation of autoreactive T- and B-cells. Protein tyrosine phosphatase non-receptor type 22 (PTPN22) encodes for lymphoid specific phosphatase (LYP) which is a key negative regulator of T lymphocyte activation. The aim of this study was to investigate the association between PTPN22 gene functional variant R620W and systemic lupus erythematosus (SLE) by comparing its prevalence in Kuwaiti SLE patients and controls. Methods The study included 134 SLE patients and 214 controls from Kuwait. The genotypes of PTPN22 gene functional variant R620W were determined by PCR-RFLP and confirmed by DNA sequence analysis in both SLE patients and the controls. Results A relatively high prevalence of the variant 620 W (T-allele) of the PTPN22 gene was detected in the SLE patients from Kuwait. 35.7% of the SLE patients had at least one variant allele (T-allele) compared to 15.9% in the controls. A statistically significant difference was detected in the frequency of variant genotypes, TT and CT between SLE patients and the controls ( p < 0.0001). No association was detected between the PTPN22 gene variant and the Raynaud’s phenomenon, renal involvement and severity of the SLE. Conclusions The frequency of PTPN22 gene functional variant R620W reported in this study is amongst the highest compared to other world populations. A high prevalence of this variant in SLE patients in comparison to the healthy controls suggests its significant contribution in conferring susceptibility to SLE together with other factors.