IL-35 and IL-37 are negatively correlated with high IgE production among children with asthma in Saudi Arabia.

BACKGROUND: Asthma is one of the most prevalent inflammatory disorders among children in Saudi Arabia. OBJECTIVE: This study aimed to determine the correlation between the serum levels of vitamin D, immunoglobulin E (IgE), and cytokine (interferon-gamma (IFN-γ), interleukin (IL)-1ß, IL-4, IL-6, IL-10, IL-13, IL-35, and IL-37) in relation to the severity of disease in patients with asthma. METHODS: This case-control study was carried out at King Abdullah Specialist Children's Hospital, Saudi Arabia, and included 48 patients with asthma and 47 matched controls, aged 6-14 years. A validated questionnaire was administered to the participants, after which each patient with asthma underwent pulmonary function tests. The serum levels of vitamin D, IgE, IFN-γ, IL-1ß, IL-4, IL-6, IL-10, IL-13, IL-35, and IL-37 of each participant were also measured. RESULTS: Patients with asthma demonstrated significantly higher IgE and cytokine (IL-1ß, IL-4, IL-6, IL-10, IL-13, IL-35, and IL-37) levels compared to the control group (p value < .001). The levels of IL-1ß, IL-4, IL-10, and IL-13 were consistently positively correlated with the serum levels of IgE among patients with asthma. However, the IgE levels in patients with asthma were consistently negatively correlated with IL-35 and IL-37. CONCLUSIONS: We found significantly higher levels of eosinophils, IgE, IL-1ß, IL-4, IL-6, IL-10, IL-13, IL-35, and IL-37 in patients with asthma compared to the controls, but no relationship between vitamin D and asthma.

Significant differences in FcγRIIa, FcγRIIIa and FcγRIIIb genes polymorphism and anti-malarial IgG subclass pattern are associated with severe Plasmodium falciparum malaria in Saudi children.

BACKGROUND: The FcγRs genotypes have been reported to play a key role in the defence against malaria parasites through both cellular and humoral immunity. This study aimed to investigate the possible correlation between FcγR (IIa, IIIa, and IIIb) genes polymorphism and the clinical outcome for anti-malarial antibody response of Plasmodium falciparum infection among Saudi children. METHODS: A total of 600 volunteers were enrolled in this study, including 200 malaria-free control (MFC) subjects, 218 patients with uncomplicated malaria (UM) and 182 patients with severe malaria (SM). The FcγR genotypes were analysed using PCR amplification methods, and measurements of immunoglobulin were determined using enzyme-linked immunosorbent assay (ELISA) technique. RESULTS: The data revealed that the FcγRIIa-R/R131 showed a statistically significant association with SM patients when compared to UM patients. Furthermore, higher levels of IgG1, IgG2, and IgG4 were associated with the FcγRIIa-H/H131 genotype among UM patients. Although the FcγRIIa-F/V176 genotype was not associated with UM, it showed a significant association with severe malaria. Interestingly, the FcγRIIIa-V/V176 genotype offered protection against SM. Moreover, SM patients carrying the FcγRIIIa-F/F genotype showed higher levels of AMA-1-specific IgG2 and IgG4 antibodies. The FcγRIIIb-NA1/NA1 and FcγRIIIb-NA2/NA2 genotypes did not show significant differences between the UM and the MFC groups. However, the genotype FcγRIIIb-NA2/NA2 was statistically significantly associated with SM patients. CONCLUSIONS: The data presented in this study suggest that the influence of the FcγRIIa-R/R131, FcγRIIIa-F/F176 and FcγRIIIb-NA2/NA2 genotypes are statistically significantly associated with SM patients. However, the FcγRIIa-H/H13 and FcγRIIIa-V/V176 genotypes have demonstrated a protective effect against SM when compared to UM patients. The impact of the FcyR (IIa, IIIa and IIIb) gene variants and anti-malaria IgG subclasses play an important role in susceptibility to malaria infection and disease outcome in Saudi children.

Association between cytokine genes polymorphisms and type 1 diabetes: a case-control study on Saudi population.

BACKGROUND: Association studies of genes encoding cytokines that play an important role in inflammatory response represent one approach to finding type 1 diabetes (T1D) disease genes. The aim of this study was to investigate the association of single nucleotide polymorphisms (SNPs) within cytokine genes with T1D in a cohort of Saudi subjects. METHODS: A total of 300 well-characterized type 1 diabetic patients and 300 T1D-free control subjects were enrolled in this investigation. Cytokine SNPs were genotyped by using Polymerase chain reaction (PCR) with sequence-specific primers. RESULTS: Our data revealed that IFN-γ +874T allele carriers [odds ratio (OR) = 1.87, p < 0.001] and TT homozygotes (OR = 1.28, p < 0.001) were significantly more susceptible to developing T1D than the A allele carriers. In addition, TNF-α -308A allele carriers (OR = 1.73, p < 0.001) and AA homozygotes (OR = 1.74, p < 0.001) were also overrepresented among the diabetics than G allele carriers. IL-4 -590C/T TT homozygotes (OR = 2.23, p < 0.001) were significantly more susceptible to develop T1D than CC genotypes, whereas CT heterozygotes were not significantly associated (OR = 1.43, p = 0.78) with T1D. Furthermore, IL-4 T allele was statistically associated with T1D patients compared to control group (OR = 2.24, p < 0.001). Similarly, IL-1ß -511C/T TT homozygotes (OR = 1.85, p = 0.012) and the T allele (OR = 1.85, p < 0.001) were significantly more susceptible to T1D than CC genotypes, whereas TC heterozygotes (OR = 1.04, p = 0.86) were not significantly associated with T1D. CONCLUSION: Our data concluded that IFN-γ +874T allele, TNF-α -308A allele, IL-1ß -511T allele, and IL-4 -590T allele could be considered risk factors for T1D development in Saudi subjects.

The Potential Role of PTPN-22 C1858T Gene Polymorphism in the Pathogenesis of Type 1 Diabetes in Saudi Population.

BACKGROUND: Recent investigations have reported an association between protein tyrosine phosphatase non-receptor type-22 (PTPN-22) gene polymorphism and susceptibility to the development of type 1 diabetes (T1D) in some populations and not in others. In this study, we aimed to investigate the association of PTPN-22 C1858T polymorphism with T1D in Saudi children. METHODS: A cohort of 372 type 1 diabetic children and 372 diabetes-free subjects was enrolled in the current investigation. The PTPN-22 C1858T polymorphism was identified using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. RESULTS: Our data showed that the frequency of CT and TT genotypes of PTPN-22 C1858T was higher in T1D children (17.7% and 4.3%, respectively) compared to healthy controls (4.8% and 1.6%, respectively), and both genotypes were statistically associated with T1D patients (OR = 4.4, 95% CI: 2.55-7.58, p < 0.001; and OR = 3.2, 95% CI: 1.23-8.28, p = 0.017, respectively). Moreover, the 1858T allele was significantly associated with T1D patients compared to the C allele (OR = 3.2, 95% CI: 1.59-6.88, p < 0.001). In addition, the T allele was significantly associated with elevated levels of HbA1c, anti-GAD, and anti-insulin antibodies (p < 0.001) and a lower concentration of C-peptide (p < 0.001) in T1D children. CONCLUSION: The data presented here suggests that the T allele of PTPN-22 C1858T polymorphism might be a risk factor for T1D development in Saudi children.

Comprehensive Analysis of the Chemical Composition and In Vitro Cytotoxic Mechanisms of Pallines Spinosa Flower and Leaf Essential Oils Against Breast Cancer Cells.

BACKGROUND/AIMS: In our quest for new natural anticancer agents, we studied the cytotoxicity of the essential oils extracted from flowers and leaves of Pallines spinosa. METHODS: The essential oils were extracted by hydrodistillation and solid phase microextraction (SPME) from flowers and leaves of the plant and their composition was determined by GC/GC-MS. The cytotoxicity of the oils was evaluated against MCF-7 and MDA-MB-231 breast adenocarcinomas, and the non-cancerous MCF-10-2A cells, using a flow cytometry-based assay Apoptosis was evaluated by flow cytometry, nuclear staining, caspases activation, and Western blotting techniques, and cell cycle by measuring DNA contents. RESULTS: The hydrodistilled flower oil contained mainly sesquiterpenes (96.39%), while the leaf sample was dominated by oxygenated-sesquiterpenes (51.60%) and sesquiterpene-hydrocarbons (34.06%). In contrast, the SPME oil contained mainly monoterpene-hydrocarbons (44.09%) and sesquiterpene-hydrocarbons (34.15%) in the flower and leaf samples, respectively. The cytotoxicity of the flower oil against MCF-7 (IC50 0.25 ± 0.03 µg/mL) and MDA-MB-231 (IC50 0.21 ± 0.03 µg/mL) was much stronger than the leaf oil (IC50 2.4 ± 0.5 µg/mL and 1.5 ± 0.1 µg/mL, respectively). The toxicity of the flower oil was ∼5 to 8-times less in normal MCF-10-2A (IC50 1.3 ± 0.2 µg/mL) and blood mononuclear cells (2.80 ± 0.45 µg/mL) as compared to breast and hematological cancer cells, respectively. Both oils induced a caspase-dependent and -independent apoptosis in MCF-7 and MDA-MB-231 cells, and altered the levels of Bcl-2 and Bax proteins. In addition, the oils arrested cell cycle in both cancer cell lines at G0/G1 phase by modulating the expression of cyclin D1, CDK4 and p21 proteins. CONCLUSION: The cytotoxicity of P. spinosa oils were mediated by apoptosis and cell cycle arrest, suggesting the potential use of their bioactive compounds as natural anticancer compounds.

Association of IL-37 gene polymorphisms with susceptibility to tuberculosis in Saudi subjects.

Tuberculosis (TB) is one of the most common infectious diseases worldwide. IL-37, a novel member of the IL-1 family, has anti-inflammatory activity. Various cytokine genes polymorphisms are reportedly associated with susceptibility to TB infection. However, an association between genetic variations in the IL-37 gene and susceptibility to TB infection has not been investigated. The aim of this case-control study was therefore to identify such an association in Saudi subjects, in which five single-nucleotide polymorphisms (SNPs) in the IL-37 gene were assessed. Serum concentrations of IL-37 were evaluated using ELISA, and genetic variants genotyped by multiplex PCR and ligase detection reaction. It was found that the C/C genotype of rs2723176 (-6962 A/C) occurs significantly more frequently in patients with active TB and that the C allele of this SNP is associated with TB. In addition, the C allele of rs2723176 SNP was associated with high circulating concentrations of IL-37. However, the genotype and allele frequency of the other four SNPs (rs3811046, rs3811047, rs2723186 and rs2723187) were not significantly associated with TB infection. In conclusion, the present data suggest that rs2723176 SNP of IL-37 is involved in the development of TB infection. Furthermore, high circulating concentrations of IL-37 may have a negative effect on protective immunity against TB infection.

The pyridone-annelated isoindigo (5'-Cl) induces apoptosis, dysregulation of mitochondria and formation of ROS in leukemic HL-60 cells.

BACKGROUND/AIMS: In our quest to develop an isoindigo with improved efficacy and bioavailability, we recently synthesized a series of novel substituted pyridone-annelated isoindigo and evaluated their antiproliferative effects. We identified the compound [(E)-1-(5'-Chloro-2'-oxoindolin-3'-ylidene)-6-ethyl-2,3,6,9-tetrahydro-2,9-dioxo-1H-pyrrolo[3,2-f] quinoline-8-carboxylic acid], abbreviated as 5'-Cl, which shows selective antiproliferative activities against various cancer cell lines mediated through apoptosis. Here we have investigated the molecular mechanisms underlying the apoptotic activity of 5'-Cl in the human promyelocytic leukemia HL-60 cells. METHODS: We employed different methods to determine the apoptotic pathways triggered by 5'-Cl in HL-60 cells, using flow cytometry, nuclear staining, caspases activation, mitochondria functioning, generation of reactive oxygen species (ROS) and Western blotting techniques. RESULTS: Low concentrations (1-8 µM) of 5'-Cl inhibited the growth of HL-60 cells in a dose and time-dependent manner. Cytotoxicity of this compound is found to be mediated by a caspase-dependent apoptosis. Also, there were indications of caspase independent apoptosis as z-VAD-FMK failed to fully rescue the cells from 5'-Cl-induced apoptosis. In addition, the compound triggered generation of Reactive Oxygen Species (ROS), caused depolarization of the mitochondrial inner membrane, decreased the level of cellular ATP, modulated the expression and phosphorylation of Bcl-2 leading to loss of its association with Bax and increased the release of cytochrome c to the cytosol of treated cells. The effects of 5'-Cl on mitochondria and apoptosis were substantially blocked in the presence of a combination between z-VAD-FMK and either of the ROS scavenger N-acetyl-L-cysteine (NAC) or pyrrolidine dithiocarbamate (PDTC). CONCLUSION: We demonstrated that the growth inhibitory effects of 5'-Cl in HL-60 cells involve multiple pathways of apoptosis and dysregulation of mitochondrial functions.

Neonatal infections in Saudi Arabia: Association with cytokine gene polymorphisms.

In recent years, many studies have reported potential associations between cytokine gene polymorphisms and the development, course, and outcome of sepsis, often with apparently conflicting results. The objective of this study was to investigate single nucleotide polymorphism (SNP) in the interleukin (IL)-1ß -31 T/C, IL-6 -174 G/C, tumor necrosis factor α (TNF-α) -308 G/A, and interferon γ (IFN-γ) +874 A/T genes for their possible association with susceptibility to early onset sepsis (EOS) in Saudi newborn infants. A total of 205 newborn infants aged 1-2 days were consecutively enrolled onto the study having met the inclusion criteria (as per the research protocol). DNA was extracted from filter papers using the Chelex-100 method. The cytokines SNP were genotyping using Taqman 5' nuclease allelic discrimination. For cytokine measurements we used the commercially available Enzyme-Linked Immunosorbent Assay (ELISA) kit. Our results show that the circulating IL-1ß, IL-6, TNF-α, and IFN-γ were significantly (p < 0.001) elevated in EOS patients compared to suspected and sepsis-free control groups; and IL-1ß -31C, IL-6 -174G, TNF-α -308G, and IFN-γ +874A alleles were associated with EOS in Saudi infants. In conclusion, analysis of cytokines concentrations and SNP for the four tested genes can be used as a predictor of sepsis outcome in newborns.

IFN-gamma and TNF associated with severe falciparum malaria infection in Saudi pregnant women.

BACKGROUND: Tumour necrosis factor (TNF) and interferon gamma (IFN-γ), encoded by TNF-836 C/A (rs 1800630) and IFN-γ -1616 C/T (rs2069705) genes, are key immunological mediators that are believed to both play protective and pathological roles in malaria. The aim of this study was to investigate the relationship between TNF-836 C/A and IFN-γ-1616 C/T polymorphism and susceptibility to severe malaria in pregnant women. METHODS: A prospective cohort (cross-sectional) study was conducted in pregnant women attending the out-patient clinic in King Fahad Specialist Hospital in Jazan (KFSHJ), with a clinical diagnosis of malaria. A total of one hundred and eighty six pregnant women were genotyped for single nucleotide polymorphism (SNP) for TNF and IFN-γ using Taqman® MGB Probes. Serum cytokine concentrations were measured by sandwich ELISA method. RESULTS: A hospital case-control study of severe malaria in a Saudi population identified strong associations with individual single-nucleotide polymorphisms in the TNF and IFN-γ genes, and defined TNF-836 C and IFN-γ-1616 T genotypes and alleles which were statistically significantly associated with severe malaria infection. Furthermore, TNF-836 CC and IFN-γ-1616 TT genotypes were associated with higher serum concentration of TNF and IFN-γ, respectively, and with susceptibility to severe malaria. CONCLUSIONS: This data provides a starting point for functional and genetic analysis of the TNF and IFN-γ genomic region in malaria infection affecting Saudi populations.

In vitro cytotoxicity of Artemisia vulgaris L. essential oil is mediated by a mitochondria-dependent apoptosis in HL-60 leukemic cell line.

BACKGROUND: The essential oil (EO) of Artemisia vulgaris L. has been traditionally used worldwide for treating a large number of diseases. Although major components in A. vulgaris EO have been shown to inhibit growth of different cancer cells, as pure compounds or part of other plants extracted oil, no information is known about its anti-proliferative activities. Therefore, the current investigation has evaluated the toxicity of the plant extracted oil from buds (AVO-b) and leaves (AVO-l) and characterized their growth inhibitory effects on cancer cells. METHODS: AVO-b and AVO-l from A. vulgaris L. were extracted by hydrodistillation, and their effect on the viability of human HL-60 promyelocytic leukemia and various other cancer cell lines was tested using MTT assay. Flow cytometric analysis of apoptosis, DNA fragmentation assay, caspases enzymatic activities and Western blotting were used to determine the apoptotic pathway triggered by their action on HL-60 cells. RESULTS: Low concentrations of AVO-b and AVO-l inhibited the growth of HL-60 cells in a dose- and time-dependent manner. Employing flow cytometric, DNA fragmentation and caspase activation analyses, demonstrated that the cytotoxic effect of the oils is mediated by a caspase-dependent apoptosis. Kinetic studies in the presence and absence specific caspase inhibitors showed that activation of caspase-8 was dependent and subsequent to the activation of caspases-9 and -3. In addition, the essential oil caused a disruption of the mitochondrial transmembrane potential (ΔΨm), increased the release of cytochrome c to the cytosol, and altered the expression of certain members of Bcl-2 family (Bcl-2, Bax and Bid), Apaf-1 and XIAP. Interestingly, low doses of AVO-b and AVO-1 also induced apoptosis in various cancer cell lines, but not in noncancerous cells. CONCLUSIONS: The results demonstrate that the EO-induced apoptosis in HL-60 cells is mediated by caspase-dependent pathways, involving caspases-3, -9, and -8, which are initiated by Bcl-2/Bax/Bid-dependent loss of ΔΨm leading to release of cytochrome c to the cytoplasm to activate the caspase cascade. The finding that AVO-b and AVO-l are more efficient to induce apoptosis in different cancer cell lines than noncancerous cells, suggests that A. vulgaris might be a promising source for new anticancer agents.

Durability of COVID-19 humoral immunity post infection and different SARS-COV-2 vaccines.

BACKGROUND: The global challenge posed by Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) has been a major concern for the healthcare sector in recent years. Healthcare workers have a relatively high risk of encountering COVID-19 patients, making protective immunity against SARS-CoV-2 is a priority for them. This study aims to evaluate the longitudinal measurement of SARS-CoV-2 IgG spike protein antibodies in healthcare workers (HCWs) after COVID-19 infection and after receiving the first and second doses of SARS-CoV-2 vaccines, including Pfizer-BioNTech (BNT162b2) and Oxford-AstraZeneca (AZD1222). METHODS: This longitudinal cohort study involved 311 healthcare workers working in two tertiary hospitals in Saudi Arabia. All participants were followed between July 2020 and July 2022 after completing the study questionnaire. A total of 3 ml of the blood samples were collected at four intervals: before/after vaccination. RESULTS: HCWs post-infection had lower mean SARS-CoV-2 IgG levels three months post-infection than post-vaccination. 92.2% had positive IgG levels two weeks after the first dose and reached 100% after the second dose. Over 98% had positive antibodies nine months after the second dose, regardless of vaccine type. The number of neutralizing antibodies decreased and was around 50% at nine months after the second dose. CONCLUSION: The results show different antibody patterns between infected and vaccinated HCWs. A high proportion of participants had positive antibodies after vaccination, with high levels persisting nine months after the second dose. Neutralizing antibodies decreased over time, with only about 50% of participants having positive antibodies nine months after the second dose. These results contribute to our understanding of immunity in healthcare workers and highlight the need for the continuous monitoring and possible booster strategies.

Neonatal infections in Saudi Arabia: association with C-reactive protein, CRP -286 (C>T>A) gene polymorphism and IgG antibodies.

BACKGROUND: C-reactive protein (CRP) is a nonspecific, acute-phase protein that rises in response to infectious and non-infectious inflammatory processes. Infections are the single largest cause of neonatal deaths globally.The primary aim of this study is to examine the association between CRP gene polymorphism and serum levels of CRP in correlation with early onset sepsis (EOS) infection in newborns living in Taif city, Saudi Arabia. The second aim is to examine the relationship between specific IgG/IgG subclasses and early onset sepsis (EOS) infection among these newborns. METHODS: Staphylococcus aureus (S. aureus) is one of the most common organisms related to sepsis infection in the newborn at King Abdel Aziz Specialist Hospital (KAASH). This study was conducted in Taif city, at KAASH's neonatal intensive care unit between March and August 2012. Neonates were consecutively enrolled onto the study having met our inclusion criteria (as per our research protocol).The CRP concentration level was analysed using NycoCard CRP Single Test. CRP -286 (C>T>A) A polymorphisms were analyzed using Pyrosequencing technology for CRP genotyping. IgG subclasses were analysed in the study population using ELISA. RESULT: Logistic regression analyses showed that the AA and AC genotypes were negatively associated amongst EOS neonates compared to suspected neonates. The frequency of CC and CT were significantly associated with the EOS neonates compared to the suspected group. The levels of specific IgG1, IgG2 and IgG3 antibodies were significantly lower amongst EOS compared to the suspected group. CONCLUSIONS: Taken together, the CRP-286 (C>T>A) A genotype polymorphism and specific IgG antibodies isotype levels can contribute to a reduced risk of EOS. Furthermore, CRP has a potential use in detecting EOS in neonates, which may mean earlier detection and management of EOS and subsequently better clinical outcome.

Anti-malarial IgG subclasses pattern and FcγRIIa (CD32) polymorphism among pregnancy-associated malaria in semi-immune Saudi women.

BACKGROUND: Pregnant women remain are at an increased risk of malaria with primigravidae being at the highest risk. Genetic polymorphism of the Fc receptor IIa for immunologlobulin (Ig) G (FcγRIIa) determines IgG subclass binding. Protection against pregnancy-associated malaria (PAM) is associated with the production of IgG specific for apical membrane antigen-1 (AMA-1). The present study was undertaken to examine the relationship between specific IgG/IgG subclasses and malaria infection. The second aim of the study is to examine the association between FcγRIIa R/H131 polymorphism in correlation with specific anti-malarial IgG antibodies of AMA-1 distribution and asymptomatic malaria infection among Saudi women living in the southern part of Saudi Arabia. METHODS: One hundred and twenty pregnant women living in an area of meso-endemic Plasmodium falciparum malaria infection were consecutively enrolled onto the study. These pregnant women were asymptomatic and attending routine antenatal clinics. The levels of plasma antibodies (IgG and subclasses AMA-1) were measured using indirect enzyme-linked immunosorbent assays (ELISA). Genotyping of FcγRIIa-R/H131 dimorphism was performed using gene-specific polymerase chain reaction (PCR) amplification with allele-specific restriction enzyme digestion (BstU1) of the PCR product. RESULTS: A total of sixty-two (52%) pregnant women was diagnosed with asymptomatic malarial infection (ASM) compared with 58 (48%) malaria free controls (MFC). In the ASM group, there were high levels of anti-malarial IgG1 and IgG3, when compared to MFC (P value <0.001, respectively). The FcγRIIa-R/R131 genotype and R131 were found to be statistically significantly more prevalent in the ASM group when compared to the MFC group [55% for ASM versus 12% for MFC, odds ratio (OR) 5.62, 95% confidence interval (CI)= (2.03- 15.58), P value= 0.001]. However, the H/H131 genotype showed statistically significant association with MFC [14% for ASM versus 50% for MFC, OR(0.36), 95% CI= (0.14- 0.95), P value= 0.03]. CONCLUSIONS: The study revealed that the ASM patients had higher anti-malarial IgG and IgG subclasses antibody levels when compared to the MFC. The FcγRIIa-R/R131 genotype and R131 allele were found to be statistically prevalent in the ASM when compared to the MFC group. The individuals carrying H/H131 were consistently associated with higher levels of anti-malarial IgG subclasses.

Correlation analysis between cytokines' profile, autoimmune antibodies and the duration of type 1 diabetes: A case control study in a specialized children's centre in Riyadh.

OBJECTIVE: The aim of this study was to investigate the role of cytokines in children with T1D living in Saudi Arabia and their correlation with disease duration and autoimmune antibody markers. METHODS: A case-control study was conducted in the endocrine clinic of King Abdullah Specialized Children's Hospital in Riyadh. A total of 274 T1D and healthy control children were enrolled in the study. 5 mL of venous blood samples were collected in the morning after 9 to 12 h of fasting in BD Vacutainer® EDTA tubes and centrifuged at 250g for 15 min at. Plasma was then stored at -20°C for detection of anti-islet, anti-GAD antibodies (Abs), and C-peptide using commercial ELISA kits from Thermo Fisher Scientific. The levels of cytokines were measured using commercial sandwich ELISA kits from Abcam. RESULTS: Median differences in cytokine levels (IFN-γ, TNF-α, IL-1ß, IL-2, IL-4, IL-6, IL-10, IL-13, IL-18, IL-21, IL-35, and IL-37) were significantly higher in T1D patients compared with healthy controls (p-value < .001). Spearman's Rho correlation indicated that TNFα, IL-1ß, IL-4, IL-10, IL-13, and IL-21 correlated significantly with T1D Abs (p-value = .01). HbA1C correlated negatively with IL-35 and IL-37, and positively with IL-18 (p-value = .01). Linear regression analysis showed a significant increase in anti-glutamic acid antibodies (GAD) in patients with >3 years of T1D duration. CONCLUSION: Autoantibodies remained positive at high levels in our patients over a 3-year duration of the disease and correlated with specific cytokines. The clear correlations with disease duration and profile of specific cytokines could be targets for future therapeutic interventions.

Analysis of the Origin of Emiratis as Inferred from a Family Study Based on HLA-A, -C, -B, -DRB1, and -DQB1 Genes.

In this study, we investigated HLA class I and class II allele and haplotype frequencies in Emiratis and compared them to those of Asian, Mediterranean, and Sub-Saharan African populations. METHODS: Two-hundred unrelated Emirati parents of patients selected for bone marrow transplantation were genotyped for HLA class I (A, B, C) and class II (DRB1, DQB1) genes using reverse sequence specific oligonucleotide bead-based multiplexing. HLA haplotypes were assigned with certainty by segregation (pedigree) analysis, and haplotype frequencies were obtained by direct counting. HLA class I and class II frequencies in Emiratis were compared to data from other populations using standard genetic distances (SGD), Neighbor-Joining (NJ) phylogenetic dendrograms, and correspondence analysis. RESULTS: The studied HLA loci were in Hardy-Weinberg Equilibrium. We identified 17 HLA-A, 28 HLA-B, 14 HLA-C, 13 HLA-DRB1, and 5 HLA-DQB1 alleles, of which HLA-A*02 (22.2%), -B*51 (19.5%), -C*07 (20.0%), -DRB1*03 (22.2%), and -DQB1*02 (32.8%) were the most frequent allele lineages. DRB1*03~DQB1*02 (21.2%), DRB1*16~DQB1*05 (17.3%), B*35~C*04 (11.7%), B*08~DRB1*03 (9.7%), A*02~B*51 (7.5%), and A*26~C*07~B*08~DRB1*03~DQB1*02 (4.2%) were the most frequent two- and five-locus HLA haplotypes. Correspondence analysis and dendrograms showed that Emiratis were clustered with the Arabian Peninsula populations (Saudis, Omanis and Kuwaitis), West Mediterranean populations (North Africans, Iberians) and Pakistanis, but were distant from East Mediterranean (Turks, Albanians, Greek), Levantine (Syrians, Palestinians, Lebanese), Iranian, Iraqi Kurdish, and Sub-Saharan populations. CONCLUSIONS: Emiratis were closely related to Arabian Peninsula populations, West Mediterranean populations and Pakistanis. However, the contribution of East Mediterranean, Levantine Arab, Iranian, and Sub-Saharan populations to the Emiratis' gene pool appears to be minor.

HLA-DRB1 and -DQB1 Alleles, Haplotypes and Genotypes in Emirati Patients with Type 1 Diabetes Underscores the Benefits of Evaluating Understudied Populations.

Background: HLA class II (DR and DQ) alleles and antigens have historically shown strong genetic predisposition to type 1 diabetes (T1D). This study evaluated the association of DRB1 and DQB1 alleles, genotypes, and haplotypes with T1D in United Arab Emirates. Materials and Methods: Study subjects comprised 149 patients with T1D, and 147 normoglycemic control subjects. Cases and controls were Emiratis and were HLA-DRB1 and -DQB1 genotyped using sequence-based typing. Statistical analysis was performed using Bridging Immunogenomic Data-Analysis Workflow Gaps R package. Results: In total, 15 DRB1 and 9 DQB1 alleles were identified in the study subjects, of which the association of DRB1*03:01, DRB1*04:02, DRB1*11:01, DRB1*16:02, and DQB1*02:01, DQB1*03:02, DQB1*03:01, and DQB1*06:01 with altered risk of T1D persisted after correcting for multiple comparisons. Two-locus haplotype analysis identified DRB1*03:01∼DQB1*02:01 [0.44 vs. 0.18, OR (95% CI) = 3.44 (2.33-5.1), Pc = 3.48 × 10-10]; DRB1*04:02∼DQB1*03:02 [0.077 vs. 0.014, OR = 6.06 (2.03-24.37), Pc = 2.3 × 10-3] and DRB1*04:05∼DQB1*03:02 [0.060 vs. 0.010, OR = 6.24 (1.79-33.34), Pc = 0.011] as positively associated, and DRB1*16:02∼DQB1*05:02 [0.024 vs. 0.075, OR = 0.3 (0.11-0.74), Pc = 0.041] as negatively associated with T1D, after applying Bonferroni correction. Furthermore, the highest T1D risk was observed for DR3/DR4 [0.104 vs. 0.006, OR = 25.03 (8.23-97.2), Pc = 2.6 × 10-10], followed by DR3/DR3 [0.094 vs. 0.010, OR = 8.72 (3.17-25.32), Pc = 3.18 × 10-8] diplotypes. Conclusion: While DRB1 and DQB1 alleles and haplotypes associated with T1D in Emiratis showed similarities to Caucasian and non-Caucasian populations, several alleles and haplotypes associated with T1D in European, African, and Asian populations, were not observed. This underscores the contribution of ethnic diversity and possible diverse associations between DRB1 and DQB1 and T1D across different populations.

Seroprevalence of SARS-CoV-2 among high-risk healthcare workers in a MERS-CoV endemic area.

INTRODUCTION: Healthcare workers (HCWs) in Saudi Arabia are a unique population who have had exposures to the Middle East Respiratory Syndrome coronavirus (MERS-CoV) and Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). It follows that HCWs from this country could have pre-existingMERS-CoV antibodies that may either protect from coronavirus disease 2019 (COVID-19) infection or cause false SARS-CoV-2 seropositive results. In this article, we report the seroprevalence of MERS-CoV and SARS-CoV-2 among high-risk healthcare workers in Riyadh city, Saudi Arabia. METHODS: This is a cross-sectional study enrolling 420 high-risk HCWs who are physically in contact with COVID-19 patients in three tertiary hospitals in Riyadh city. The participants were recruited between the 1st of July to the end of December 2020. A 3 ml of the venous blood samples were collected and tested for the presence of IgG antibodies against the spike proteins of SARS-CoV-2 and MERS-CoV using enzyme-linked immunosorbent assay (ELISA). RESULTS: The overall prevalence of SARS-CoV-2 in high-risk HCWs was 14.8% based on SARS-CoV-2 IgG testing while only 7.4% were positive by Polymerase Chain Reaction (PCR) for viral RNA. Most of the SARS-CoV-2 seropositive HCWs had symptoms and the most frequent symptoms were body aches, fever, cough, loss of smell and taste, and headache. The seroprevalence of MERS-CoV IgG was 1% (4 participants) and only one participant had dual seropositivity against MERS-CoV and SARS-CoV-2. Three MERS-CoV positive samples (75%) turned to be negative after using in-house ELISA and none of the MERS-CoV seropositive samples had detectable neutralization activity. CONCLUSION: Our SARS-CoV-2 seroprevalence results were higher than reported regional seroprevalence studies. This finding was expected and similar to other international findings that targeted high-risk HCWs. Our results provide evidence that the SARS-CoV-2- seropositivity in Saudi Arabia similar to other countries was due to exposure to SARS-CoV-2 rather than MERS-CoV antibody.

Metformin alters peripheral blood mononuclear cells (PBMC) senescence biomarkers gene expression in type 2 diabetic patients.

BACKGROUND: Although there is increasing evidence showing that cell senescence is increased in circulating PBMC in type 2 diabetes mellitus (T2DM), the data are contradictory. This study examined several senescence biomarkers, including LMNA/C transcript variants, p16INK4a, p53, and p21Cip1/WAF, in PBMC of T2DM patients and the effect of Metformin on these senescence markers. METHODS: Blood samples were obtained from 30 lean, 30 obese, 20 newly diagnosed type 2 diabetes mellitus (T2DM), and 30 T2DM on Metformin. PBMC were isolated and mRNA expression of the senescence biomarkers were quantified by RT-qPCR. The effect of ectopic expression of LMNA and LMNC in human monocytic cells lines (THP-1 and U937) on several inflammatory mediators were also examined. RESULTS: LMNA expression was significantly higher in PBMC of obese and T2DM patients. LMNC expression was significantly inhibited in T2DM patients. LMNAΔ10 and Progerin mRNA expression was not detected in PBMC of all groups. Expression of p16INK4a, p21Cip1/WAF and p53 were inhibited significantly in T2DM. Metformin treatment reverted LMNA, LMNC, and p53 expression levels to normal levels. Upregulation of LMNA in monocytic THP-1 and U937 cell lines induced CD68, TNFα, CCL2, IL-6 and NOS2. CONCLUSIONS: These data support the notion that LMNA may mediate senescence in PBMCs of T2DM by upregulating inflammatory pathways. Metformin may exert its anti-inflammatory property by modulation of senescence mediator LMNA.

Association of a single nucleotide polymorphism in the C-reactive protein gene (-286) with susceptibility to Plasmodium falciparum malaria.

The role of inflammation in malaria pathogenesis is not fully understood, although C-reactive protein (CRP) may have a negative influence on host immunity to infections. An upstream polymorphism, -286 (C > T > A), in the CRP gene is known to influence CRP levels. In this study, a cohort of 192 Sudanese donors, followed for malaria infection for 9 years, had their CRP -286 gene locus genotyped by pyrosequencing. The number of malaria episodes experienced by each individual over the study period was used as an index for malaria susceptibility. The prevalence of the CRP alleles A, C and T were 21%, 52% and 27%, respectively. Importantly, the A-allele, unlike the C- and T-alleles or CRP genotypes, was significantly associated with an increased number of malaria episodes, P = 0.007. The proportion of A-allele carriers among donors not known to have had malaria during the study period was 18%, whereas it was 43% and 63% among donors who had experienced 1-4 and > or =5 malaria episodes, respectively, over the same period (P = 0.002). Furthermore, the A-allele was associated with higher parasite counts. In conclusion, the CRP -286 A-allele was associated with an increased susceptibility to uncomplicated Plasmodium falciparum malaria.

Marked differences in CRP genotype frequencies between the Fulani and sympatric ethnic groups in Africa.

BACKGROUND: C-reactive protein (CRP) is an acute phase protein that can activate various immune cells and bind to certain Fcgamma receptors. The latter may compete with the binding of IgG antibodies to these receptors and could thereby interfere with the antigen-specific immune response. Polymorphisms in the promoter region of the CRP gene have been strongly associated with the plasma concentration of CRP. The known lower susceptibility to malaria in the Fulani ethnic group, as compared to their sympatric neighbours in Africa, has been linked to different genetic backgrounds. The present study was performed to investigate if polymorphisms in the CRP gene could contribute to the lower susceptibility to malaria seen in the Fulani ethnic group. METHODS: The CRP -717 T>C, -286 C>T>A, and +1444 C>T polymorphisms were analysed in asymptomatic Fulani and non-Fulani individuals from Mali and Sudan using Pyrosequencing T and TaqMan r MGB probes. RESULTS: The rare -286 A allele, previously shown to be associated with increased CRP expression and plasma levels, was shown to be more frequent in the non-Fulani ethnic groups as compared to the sympatric Fulani ethnic group both in Mali and Sudan. The common -717 T allele was more prevalent in the non-Fulani ethnic group compared to the sympatric Fulani ethnic group, but only in Mali. The parasite prevalence was increased for the -286 A allele, but not for the -717 T allele. No differences regarding genotype frequency or parasite prevalence were seen for +1444 C>T. CONCLUSION: This study indicate that CRP may play an important role in the immune responses to malaria, and that the -286 C/T/A CRP polymorphism may be a contributing factor to the lower susceptibility to malaria seen in the Fulani.