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Cytokine Jul 2021A single nucleotide polymorphism (SNP), 251 bases upstream from the IL-8 transcription start (-251A>T, rs4073), has been extensively investigated in cancers and...
A single nucleotide polymorphism (SNP), 251 bases upstream from the IL-8 transcription start (-251A>T, rs4073), has been extensively investigated in cancers and inflammatory and infectious diseases in predominantly European and Asian populations. We sequenced the IL-8 gene of 109 black and 32 white South African (SA) individuals and conducted detailed characterization of gene variation and haplotype structure. IL-8 production in phytohaemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMCs) of a subset (black: N = 22; white: N = 32) of these individuals was measured using ELISA. Select variants were genotyped for additional black individuals (N = 141), and data from the 1000 Genomes Project were used for haplotype analysis and comparative purposes. In white individuals, the -251A>T SNP formed part of a prevalent six-variant haplotype [haplotype frequency (HF): 61%], Hap-1C, involving the following variants: -251A>T; +394T>G (rs2227307); +780C>T (rs2227306); +1240->A (rs2227541); +1635C>T (rs2227543) and +2770A>T (rs2227543). Hap-1C (-251T+394T+780C+1240+A+1635C+2770A) was composed of two three-variant sub-haplotypes [Hap-1Ca: -251T+394T+1240+A; Hap-1Cb: +780C+1635C+2770A) sharing similarities with haplotypes identified in the black population. Hap-1C was found to be present in European, East and South Asian populations. Four haplotypes were identified in the black population with the two prevalent haplotypes each comprised of two variants: Hap-1B [-251A>T and +1240->A; -251T+1240+A; HF: 14%] and Hap-2B [-743T>C (rs2227532) and +2452A>C (rs2227545); -743C+2452C; HF: 13%]. Populations did not differ in unstimulated PBMC IL-8 production. Upon PHA stimulation, PBMCs from white individuals produced more IL-8 (P = 0.04), suggesting the -251T allele is responsible for higher production, however further analysis revealed that Hap-1C (and constituent sub-haplotypes), did not associate with IL-8 production. Populations did however differ in monocyte number with the white population having significantly more monocytes compared to the black population (P = 0.025), and furthermore monocyte number strongly correlated with IL-8 production in both population groups (black: p = 0.0002, r = 0.71; white: P = 0.0005, r = 0.59). Hap-1B, Hap-2B, and a SNP located one base pair upstream of the IL-8 ATG start codon, +100C>T SNP (rs2227538), all associated with higher IL-8 production in the black population - individuals harbouring at least one of these haplotypes/variant associated with higher IL-8 production (P = 0.003) compared to individuals without. The black population was enriched for individuals harbouring Hap-1B and/or Hap-2B compared to the 1000 Genomes project sub-Saharan African population (P = 0.006), suggesting that SA black individuals may be high IL-8 producers. Given the paucity of IL-8-related studies that have been conducted in populations from sub-Saharan Africa, this study has significantly increased our understanding of this important chemokine in the South African population.
Topics: Adult; Africa South of the Sahara; Alleles; Black People; Ethnicity; Female; Gene Frequency; Genetic Variation; Genetics, Population; Haplotypes; Humans; Interleukin-8; Leukocytes, Mononuclear; Linkage Disequilibrium; Male; Middle Aged; Monocytes; Phytohemagglutinins; South Africa; White People; Young Adult
PubMed: 33814271
DOI: 10.1016/j.cyto.2021.155489 -
Journal of Immunological Methods Jan 2021Investigational cell-based therapeutics are rapidly heading towards pivotal clinical trials. The premise is that the scientific rationale is well defined, and that... (Comparative Study)
Comparative Study
Investigational cell-based therapeutics are rapidly heading towards pivotal clinical trials. The premise is that the scientific rationale is well defined, and that product quality reflects exactly this. In vitro potency assays are necessary tools for evaluating cell products, and with potency assays comes high demands for standardization and reproducibility of the methods involved. For demonstrating principles of cell therapeutics for allogeneic use or with claimed immunosuppressive efficacies, assays involving peripheral blood mononuclear cells (PBMC) are critical. Establishment of a cryopreserved bank of PBMC favors standardization, as it allows repeated use of a single donor and simultaneous testing of several donors. The first step to fulfil such potential is to ensure optimum conditions for preservation of PBMC function, and secondly to design assays which heightens the reproducibility. Emphasis should be put on application of the assay. The objective of the present study was to establish a methodological foundation for cell therapeutics to be tested, and several aspects were factored in, including cell concentrations and partial changes of medium. PBMC were isolated and cryopreserved in six formulations of cryoprotective medium consisting of fetal bovine serum (90%, 60%, and 30%) in combination with dimethyl sulfoxide (10% or 5%). The proliferative capacity of the cryopreserved cells was assayed by labeling with carboxyfluorescein succinimidyl ester and stimulation by phytohemagglutinin or in mixed lymphocyte reactions, analyzed by flow cytometry. To counter an eventual lag phase post thaw, the assays were designed to include two durations and to explore the possibility of reducing cell numbers, two cell concentrations. Qualitative and quantitative aspects of the staining were affected by formulation as well as design, stressing the importance of basic optimization for assay development. We conclude that the established methods allow for optimized preservation of function and will serve as a platform for further development of robust functional assays.
Topics: Cell Proliferation; Cells, Cultured; Cryopreservation; Cryoprotective Agents; Dimethyl Sulfoxide; Flow Cytometry; Humans; Leukocytes, Mononuclear; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Mitogens; Phytohemagglutinins; Reproducibility of Results; Serum Albumin, Bovine; Time Factors
PubMed: 33049298
DOI: 10.1016/j.jim.2020.112897 -
Croatian Medical Journal Feb 2019To assess the immunomodulatory effect of tonsil-derived mesenchymal stem cells (MSCs) on T-lymphocyte proliferation and cytokine production.
AIM
To assess the immunomodulatory effect of tonsil-derived mesenchymal stem cells (MSCs) on T-lymphocyte proliferation and cytokine production.
METHODS
Tonsils were obtained from children aged 3 to 12 years (n=15) who underwent tonsillectomy for obstructive sleep apnea from April 2012-October 2014 at the Merkur University Hospital, Zagreb. Tonsil-derived MSCs were co-cultured with peripheral blood mononuclear cells (PBMCs) and phytohemagglutinin as a mitogen. PBMCs were induced to differentiate into T helper 1 or T helper 2 cells in the presence or absence of tonsil-derived MSCs, after which the production of interferon-gamma in T helper 1 and interleukin-4 in T helper 2 cells was assessed.
RESULTS
Tonsil-derived MSC suppressed phytohemagglutinin-induced proliferation of PBMCs. Compared with controls, tonsil-derived MSC co-culture significantly decreased interferon-gamma production (P<0.001) and increased interleukin-4 production (P<0.001).
CONCLUSION
Tonsil-derived MSCs exert immunomodulatory effects on T lymphocyte proliferation and T helper 1- and T helper 2-specific cytokine production.
Topics: Cell Proliferation; Cells, Cultured; Child; Child, Preschool; Coculture Techniques; Cytokines; Female; Humans; Leukocytes, Mononuclear; Lymphocyte Activation; Male; Mesenchymal Stem Cells; Palatine Tonsil; Phytohemagglutinins; T-Lymphocytes
PubMed: 30825273
DOI: 10.3325/cmj.2019.60.12 -
APMIS : Acta Pathologica,... Oct 2017This study investigated phenotypic and functional characteristics of lymphocytes in children with common variable immunodeficiency (CVID) and unclassified...
This study investigated phenotypic and functional characteristics of lymphocytes in children with common variable immunodeficiency (CVID) and unclassified hypogammaglobulinemia (UH), as well as B-cell subsets in non-consanguineous parents. Blood samples of 30 children, CVID (n = 9), UH (n = 9), healthy donors HD (n = 12), and 19 adults (parents and controls) were labeled by a combination of surface markers to identify CD4, CD8 T-cell and B-cell subpopulations. T-cell cytokine production in children was analyzed in vitro after stimulation with phytohemagglutinin (PHA) and tetanus toxoid. We observed low percentages of switched memory B cells in children with CVID, increase in total CD4 T-cell counts, and high percentages of transitional B cells only in UH group. Analysis of T-cell immunity showed that CVID children had decreased percentages of CD8 IFN-γ-producing cells after stimulation with PHA and tetanus toxoid. Parent of children with CVID had low percentages of naive B cell and increased percentages of memory B cells in comparison with controls. These results suggest that (i) early combined immune defect in children with CVID and (ii) a possible familial B-cell disturbance in pediatric CVID.
Topics: Adolescent; Adult; B-Lymphocytes; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Child; Child, Preschool; Common Variable Immunodeficiency; Cytokines; Female; Humans; Immunologic Factors; Male; Middle Aged; Phytohemagglutinins; Tetanus Toxoid
PubMed: 28929596
DOI: 10.1111/apm.12738 -
PloS One 2019Mulberry leaf polysaccharide (MLP) was extracted and purified by DEAE-52 cellulose and Sephadex G-100 column chromatography to afford two major purified polysaccharides...
Mulberry leaf polysaccharide (MLP) was extracted and purified by DEAE-52 cellulose and Sephadex G-100 column chromatography to afford two major purified polysaccharides (MLP-1 and MLP-2). The purified polysaccharides were characterized, and their immune-enhancing properties were investigated. MLP-1 had a molecular weight of 9.31×104 Da and was composed of mannose, rhamnose, glucose, galactose, xylose, and arabinose in a molar ratio of 0.71:1.00:2.76:1.13:3.70:2.81. The molecular weight of MLP-2 was 2.22×106 Da, and its monosaccharide constituents were mannose, rhamnose, glucose, galactose, and arabinose in a molar ratio of 1.31:8.45:6.94:1.00:11.96. Infrared spectroscopy showed that each MLP had a typical absorption peak characteristic of sugars, and ultraviolet (UV) spectroscopy showed that neither MLP contained nucleic acid or protein components. Then, the abilities of these polysaccharides to stimulate spleen lymphocyte proliferation in mice in vitro were compared by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay. MLP-2 was more effective than MLP-1; therefore, MLP-2 was chosen for the study of its immune-enhancing effects in vivo. For the in vivo experiments, 14-day-old chickens immunized with Newcastle disease (ND) vaccine were orally administered MLP-2, and Astragalus polysaccharide (APS) was used as the control. Each chicken was orally administered 4 mg or 8 mg of MLP-2 for seven consecutive days starting three days before ND vaccine immunization. MLP-2 significantly improved the ND serum antibody titer and interleukin-2 (IL-2), interferon-γ (IFN-γ) and immunoglobulin A (sIgA) concentrations in tracheal and jejunal wash fluids, and increasing numbers of immune globulin A-positive (IgA+) cells in cecal tonsils and increased body weight. These results indicated that MLP-2 could significantly enhance immune activity and could therefore be utilized as an immunopotentiator drug candidate.
Topics: Animals; Antibodies; B-Lymphocytes; Body Weight; Cell Count; Cell Proliferation; Chickens; Female; Immunity, Mucosal; Immunoglobulin A, Secretory; Lipopolysaccharides; Male; Mice, Inbred ICR; Monosaccharides; Morus; Phytohemagglutinins; Plant Leaves; Polysaccharides; Reference Standards; Spectrophotometry, Infrared; Spectrophotometry, Ultraviolet; Spleen; T-Lymphocytes; Vaccination
PubMed: 30601811
DOI: 10.1371/journal.pone.0208611 -
International Journal of Clinical... Apr 2021Given that there is no rapid and effective method for distinguishing active tuberculosis (ATB) from latent tuberculosis infection (LTBI), the discrimination between...
BACKGROUND
Given that there is no rapid and effective method for distinguishing active tuberculosis (ATB) from latent tuberculosis infection (LTBI), the discrimination between these two statuses remains challenging. This study sought to investigate the value of nutritional indexes and tuberculosis-specific antigen/phytohemagglutinin ratio (TBAg/PHA ratio) for distinguishing ATB from LTBI.
METHODS
Participants were consecutively recruited based on positive T-SPOT.TB results between January 2018 and January 2020. ATB was diagnosed by positive mycobacterial culture and/or positive GeneXpert MTB/RIF, with clinical symptoms and radiological characteristics suggestive of ATB. Individuals with positive T-SPOT.TB but without the evidence of ATB were defined as LTBI. Patients younger than 17 years and undergoing anti-TB treatment were excluded.
RESULTS
A total of 709 (312 ATB and 397 LTBI) and another 309 (120 ATB and 189 LTBI) subjects were respectively recruited from Tongji Hospital (Qiaokou cohort) and Sino-French New City Hospital (Caidian cohort). The level of prealbumin was significantly lower in ATB than in LTBI. With a cut-off value of 139 mg/L, the sensitivity and specificity of prealbumin in distinguishing ATB from LTBI were 50.96% (45.41%-56.51%) and 91.69% (88.97%-94.40%). Meanwhile, TBAg/PHA ratio was found statistically higher in ATB compared with LTBI. If using the threshold of 0.29, the sensitivity and specificity of TBAg/PHA ratio were 65.71% (60.44%-70.97%) and 90.93% (88.11%-93.76%), respectively. Moreover, the combination of prealbumin and TBAg/PHA ratio (obtaining by diagnostic model) yielded better specificity (90.18%, [87.25%-93.10%]) and sensitivity (87.18%, [83.47%-90.89%]), while the clinical utility index (CUI) positive and CUI negative were respectively 0.76 and 0.81. After anti-TB treatment, TBAg/PHA ratio was declined while the level of prealbumin was restored (Wilcoxon test, P < 0.001). Furthermore, the performance of diagnostic model obtained in Qiaokou cohort was confirmed in Caidian cohort.
CONCLUSIONS
The diagnostic model based on combination of prealbumin and TBAg/PHA ratio is a rapid and accurate tool for discriminating ATB from LTBI.
Topics: Antigens, Bacterial; Humans; Latent Tuberculosis; Mycobacterium tuberculosis; Phytohemagglutinins; Prealbumin; Tuberculosis
PubMed: 33175465
DOI: 10.1111/ijcp.13831 -
Biomarkers : Biochemical Indicators of... Mar 2018Oral lichen planus (OLP) is one of the most common oral mucosal lesions affecting 0.5-2% of the adult population. It is difficult to distinguish between OLP and other...
BACKGROUND
Oral lichen planus (OLP) is one of the most common oral mucosal lesions affecting 0.5-2% of the adult population. It is difficult to distinguish between OLP and other oral mucosal diseases. Structural changes in the glycans of saliva proteins might be reliable indicators of OLP. However, little is known about the alteration of salivary glycopatterns during OLP.
OBJECTIVE
We aimed to investigate the alterations of salivary protein glycosylation related to OLP.
MATERIAL AND METHODS
Twenty-eight patients with OLP and 30 age- and sex-matched healthy volunteers (HVs) were enrolled in the test group to probe the difference of salivary glycopatterns using lectin microarrays. The lectin blotting were further utilized to validate the expression of certain glycans.
RESULTS
The glycoproteins recognized by three lectins [Aleuria aurantia lectin (AAL); Phytolacca americana (PWM); Phaseolus vulgaris agglutinin (E + L), (PHA-E + L)] were mainly increasing in the saliva of OLP. Meanwhile, these glycoproteins also exhibited significant age-associated alterations.
CONCLUSIONS
This study provided a new basic insight into salivary glycopatterns in OLP and helped to develop new potential biomarkers for diagnosis of OLP.
Topics: Adult; Biomarkers; Female; Glycoproteins; Glycosylation; Humans; Lectins; Lichen Planus, Oral; Male; Middle Aged; Phytohemagglutinins; Pokeweed Mitogens; Saliva; Salivary Proteins and Peptides; Young Adult
PubMed: 29130773
DOI: 10.1080/1354750X.2017.1405284 -
The Journal of Comparative Neurology Feb 2023The orbital cortex (ORB) of the rat consists of five divisions: the medial (MO), ventral (VO), ventrolateral (VLO), lateral (LO), and dorsolateral (DLO) orbital...
The orbital cortex (ORB) of the rat consists of five divisions: the medial (MO), ventral (VO), ventrolateral (VLO), lateral (LO), and dorsolateral (DLO) orbital cortices. No previous report has comprehensively examined and compared projections from each division of the ORB to the thalamus. Using the anterograde anatomical tracer, Phaseolus vulgaris leucoagglutinin, we describe the efferent projections from the five divisions of the ORB to the thalamus in the rat. We demonstrated that, with some overlap, each division of the ORB distributed in a distinct (and unique) manner to nuclei of the thalamus. Overall, ORB projected to a relatively restricted number of sites in the thalamus, and strikingly distributed entirely to structures of the medial/midline thalamus, while completely avoiding lateral regions or principal nuclei of the thalamus. The main termination sites in the thalamus were the paratenial nucleus (PT) and nucleus reuniens (RE) of the midline thalamus, the medial (MDm) and central (MDc) divisions of the mediodorsal nucleus, the intermediodorsal nucleus, the central lateral, paracentral, and central medial nuclei of the rostral intralaminar complex and the submedial nucleus (SM). With some exceptions, medial divisions of the ORB (MO, VO) mainly targeted "limbic-associated" nuclei such as PT, RE, and MDm, whereas lateral division (VLO, LO, DLO) primarily distributed to "sensorimotor-associated" nuclei including MDc, SM, and the rostral intralaminar complex. As discussed herein, the medial/midline thalamus may represent an important link (or bridge) between the orbital cortex and the hippocampus and between the ORB and medial prefrontal cortex. In summary, the present results demonstrate that each division of the orbital cortex projects in a distinct manner to nuclei of the thalamus which suggests unique functions for each division of the orbital cortex.
Topics: Animals; Rats; Prefrontal Cortex; Thalamus; Midline Thalamic Nuclei; Hippocampus; Intralaminar Thalamic Nuclei; Phytohemagglutinins; Neural Pathways
PubMed: 36226328
DOI: 10.1002/cne.25419 -
European Cytokine Network 2014Psoriasis is one of the most common, immune-mediated, chronic inflammatory skin diseases. Proinflammatory cytokines play an important pathogenetic role at a local level.
BACKGROUND
Psoriasis is one of the most common, immune-mediated, chronic inflammatory skin diseases. Proinflammatory cytokines play an important pathogenetic role at a local level.
OBJECTIVE
To assess whether the proinflammatory cytokines IL-1β, IL-6, IL-17, IL-22 and TNF-α are released systemically during psoriasis.
METHODS
Peripheral blood mononuclear cells (PBMCs) were isolated from 30 patients with psoriasis and 30 healthy volunteers. Cytokine production was assessed in supernatants using an enzyme immunoassay after stimulation of PBMCs with microbial stimuli. In addition, flow cytometry was used to determine the subsets of monocytes involved and the intracellular TNF-α production in monocytes.
RESULTS
IL-17 levels were significantly higher in the supernatants of PBMCs from psoriatic patients after stimulation with phytohemagglutinin. TNF-α production was also significantly higher in cells from psoriatic patients after stimulation with all stimuli, as compared with health volunteers. Similar changes were not found for the other cytokines. A statistically significant difference was observed between patients and controls for inflammatory CD14(+)/CD16(+) monocytes (p<0.0001) and patrolling CD14-/CD16(+) monocytes.
CONCLUSION
Hyper-production of TNF-α is documented in psoriasis. These results support the concept that there is a systemic, proinflammatory component in psoriasis.
Topics: Adult; Aged; Antigens, Bacterial; Case-Control Studies; Female; GPI-Linked Proteins; Gene Expression; Humans; Interleukin-17; Interleukin-1beta; Interleukin-6; Interleukins; Leukocytes, Mononuclear; Lipopolysaccharide Receptors; Lipopolysaccharides; Lipoproteins; Macrophages; Male; Middle Aged; Phytohemagglutinins; Primary Cell Culture; Psoriasis; Receptors, IgG; Tumor Necrosis Factor-alpha; Interleukin-22
PubMed: 25679113
DOI: 10.1684/ecn.2014.0358 -
Genes and Immunity Mar 2015Th1/Th17-type T-cell responses are upregulated in Behcet's disease (BD). However, signaling pathways associated with this aberrant immune response are not clarified....
Th1/Th17-type T-cell responses are upregulated in Behcet's disease (BD). However, signaling pathways associated with this aberrant immune response are not clarified. Whole-genome microarray profiling was performed with human U133 (Plus 2.0) chips using messenger RNA of isolated CD14(+) monocytes and CD4(+) T cells from peripheral blood mononucleated cell (PBMC) in patients with BD (n = 9) and healthy controls (HCs) (n = 9). Flow cytometric analysis of unstimulated (US) and stimulated (phytohaemagglutinin) signal transducer and activator of transcription (STAT3) and pSTAT3 expressions of PBMCs were also analyzed (BD and HC, both n = 26). Janus family of kinase (JAK1) was observed to be upregulated in both CD14(+) monocytes (1.95-fold) and CD4(+) T lymphocytes (1.40-fold) of BD patients. Using canonical pathway enrichment analysis, JAK/STAT signaling was identified as activated in both CD14(+) monocytes (P = 9.55E-03) and in CD4(+) lymphocytes (P =8.13E-04) in BD. Interferon signaling was also prominent among upregulated genes in CD14(+) monocytes (P = 5.62E-05). Glucocorticoid receptor signaling and interleukin (IL-6) signaling were among the most enriched pathways in differentially expressed genes in CD14+ monocytes (P = 2.45E-09 and 1.00E-06, respectively). Basal US total STAT3 expression was significantly higher in BD (1.2 vs 3.45, P < 0.05). The JAK1/STAT3 signaling pathway is activated in BD, possibly through the activation of Th1/Th17-type cytokines such as IL-2, interferon (IFN-γ), IL-6, IL-17 and IL-23.
Topics: Adult; Behcet Syndrome; CD4-Positive T-Lymphocytes; Female; Gene Expression Profiling; Genome-Wide Association Study; Humans; Interleukins; Janus Kinase 1; Lipopolysaccharide Receptors; Male; STAT3 Transcription Factor; Signal Transduction
PubMed: 25410656
DOI: 10.1038/gene.2014.64