Novel quinoxaline compound against extended-spectrum beta-lactamases producing bacteria.

OBJECTIVE: Extended-spectrum beta-lactamases (ESBLs) targeting beta-lactam antibiotics pose a major healthcare challenge. Carbapenems are known to be less impacted. However, the emergence of carbapenem-resistant strains can add further complexity to this existing challenge. With slow drug discovery and rapid resistance, repurposing existing drugs is crucial. This research study aims to provide insight into the antimicrobial effectiveness of 3-hydrazinoquinoxaline-2-thiol against diverse clinical ESBL-producing isolates. MATERIALS AND METHODS: The broth microdilution assay was conducted on a total of sixty-nine clinical ESBL-producing isolates to assess the minimum inhibitory concentrations (MICs) of 3-hydrazinoquinoxaline-2-thiol. The assay was conducted in triplicate, and the average MIC values were calculated. RESULTS: The most repeatedly observed MIC was 64 µg/ml (37.7%), followed by 256 µg/ml (23.2%) and 128 µg/ml (17.4%). Other MICs: 32 µg/ml (11.6%), 16 µg/ml (7.2%), 4-8 µg/ml (1.4%). CONCLUSIONS: This study demonstrated an effect of 3-hydrazinoquinoxaline-2-thiol on various ESBL-producing strains in vitro, indicating its promising therapeutic potential. To comprehensively understand the drug, rigorous testing, including pharmacokinetics, resistance assays, safety assessments, and exploration of potential synergies with other antibiotics against ESBL-producing organisms, is crucial.

Tests and estimates (transmission disequilibrium test) for allelic association of CCR5delta32 with asthma in high-risk families.

OBJECTIVE: To examine the possible associations between CCR5delta32 and asthma and related phenotypes in high-risk families. METHODS: A total of 154 families (453 individuals), with at least two affected children with physician-diagnosed asthma (PDA) and atopy defined as one or more skin prick test to common inhaled allergen (SPT wheal > or = 3 mm), were studied. Samples were genotyped using PCR assay and tested for possible associations by TDT and PDT and case control analyses. RESULTS: Overall allelic frequency for CCR5delta32 was 26.1%, and both TDT and PDT demonstrated similar nonsignificant associations (p=0.123) and (p=0.088). Analysis by the clinical categories of non atopic and atopic asthma and presence or absence of atopy without asthma failed to identify any significant associations. However there were strong associations of the mutant allele with the phenotypes of negative SPT, PC 20 less than 8 mg/ml, baseline FEV1 greater than the population median (83.5% predicted) and serum IgE less than 100 IU/l for child probands but only for negative SPT in unrelated parents. CONCLUSION: Non-significant association was seen with family based association tests (FBATs). The strong associations with the asthma related phenotypes in child probands support previous observations that CCR5 is in linkage disequilibrium with CCR2 or CCR3.

Investigation of adult-sera for the presence of anti-Cryptosporidium antibodies.

Cryptosporidiosis is an acute diarrhoeal disease caused by Cryptosporidium spp for both human and animals. Typically, the duration of diarrhoeal illness and ultimate outcome of intestinal cryptosporidiosis depend on the immune status of the patient. Random serology-based studies in humans and animals have suggested that infection with this organism is common during a life time. 130 serum samples of adults, (18-30 years) from two main public hospitals at Jeddah City, Saudi Arabia were recruited to participate in the study. The aim was to identify the sero-prevalence of cryptosporidiosis infection and determine factors associated with increased risk of the infection. Western Blot analysis (WB) with two different Cryptosporidium antigen markers; the 15-17 KDa and the 27 KDa, were used. Among participants, 8.5 % had antibodies to the 15 KDa, 23.8 % had antibodies to the 27 kDa, 34.6 % were sero-positive to both antigens, and 33.1 % were sero-negative to both antigens. Source of drinking water with a strong association with drinking tap water was the only factor significantly associated with seropositivity to cryptosporidiosis infection (OR= 37.33, P< 0.001).

The -17 T/C chemokine receptor 3 genetic polymorphism is associated asthma but not atopy: transmission and association studies.

We genotyped and identified the asthma and atopic status and related phenotypes of 154 nuclear families (453 individuals) each containing at least two affected children with physician-diagnosed asthma (PDA) in order to confirm or refute the possible relevance of known single nucleotide polymorphisms (SNPs) in the gene coding for the CCR3 receptor. Allelic quantification for each SNP by DNA pooling identified -17/TC as the only allele with a clinically relevant frequency in this population with a frequencies of 0.142 in cases of PDA and 0.035 in asymptomatic controls. The whole population frequency of the -17/TC polymorphism was 13.9% and the functional binding site analyses by MatInd and MatInspector programs found that it belonged to the same family as activating transcription factor 6 (ATF-6). The pedigree disequilibrium test (PDT) was applied in 34 informative families and the mutant allele was preferentially transmitted with PDA (P = 0.0001) with methacholine bronchial hyperresponsiveness (BHR) (0.002) but not with markers of atopy as assessed by allergen skin prick tests (SPT) or elevated serum IgE. Case-control analyses in 303 unrelated parents (34-61y [median 43y]) revealed a significant association with both atopic and non atopic asthma (P = 0.001), and in 150 unrelated child probands for non-atopic asthma (P = 0.001). The mutant allele was associated with BHR, with baseline Forced Expiratory Volume in the first second (FEV1) below the population median value but not with atopy defined SPT or elevated serum IgE (>100 IU/ml). The T17C chemokine receptor 3 polymorphism appears to be associated with asthma BHR and disease severity but not with atopy.

Control of eosinophil toxicity in the lung.

The inappropriate accumulation of eosinophils and the subsequent release of their potent pro-inflammatory mediator arsenal are thought to be important contributors to the pathogenesis of asthma and other allergic diseases. It is also becoming apparent that eosinophils may play a role in the orchestration of immune responses in the asthmatic lung. There is therefore much interest in the development of strategies to limit or prevent eosinophil-induced toxicity. The mechanisms by which eosinophils accumulate in the peribronchial tissues of the lung are complex and include enhanced differentiation and release from the bone marrow, selective adhesion and transendothelial migration, directed movement in response to specific chemotactic mediators and finally prolonged survival as a consequence of delayed apoptosis. Thus it can be appreciated that there are many points at which the toxicity of eosinophils can be limited or even prevented. Important areas for potential advances in glucocorticoid (GC) development include efforts to dissociate their anti-inflammatory effects from unwanted side effects. Other areas include the development of humanized monoclonal antibodies against IL-4, IL-13 and IL-5 together with the inhibition of adhesion pathways and/or chemokines responsible for eosinophil accumulation in the asthmatic lung. Several avenues of research are currently underway in an attempt to define mechanisms by which pro-inflammatory cells such as eosinophils can be safely removed from the asthmatic lung through apoptosis induction and their subsequent ingestion by phagocytes. This review will discuss both the potential and shortcomings of these diverse approaches to limit eosinophil toxicity in the asthmatic lung.

Phagocytosis of apoptotic eosinophils but not neutrophils by bronchial epithelial cells.

BACKGROUND: We have previously demonstrated that human bronchial epithelial cells engulf apoptotic eosinophils. OBJECTIVES: To compare and contrast the phagocytic capabilities of monocyte-derived macrophage and primary airway epithelial cells for apoptotic granulocytes. RESULTS: Here we compared phagocytosis of human apoptotic eosinophils and neutrophils by small and large airway epithelial cells (SAEC and LAEC) and monocyte-derived macrophages. Confocal microscopy of F-actin staining and scanning and transmission electron microscopy revealed phagocytic cup formation around apoptotic eosinophils by airway epithelial cells (AEC) membranes with evidence of their digestion. Resting and cytokine-stimulated AEC did not recognize and ingest apoptotic neutrophils. The latter were phagocytosed by macrophages that exhibited greater ingestion of and higher capacity for, apoptotic eosinophils over apoptotic neutrophils. Cytochalasin D completely abolished uptake of apoptotic eosinophils by SAEC, LAEC or macrophage monolayers. Ligation of epithelial cell CD44 receptors for 24 h increased phagocytosis of apoptotic eosinophils by SAEC and LAEC with a potency comparable with that of IL-1. Phagocytosis was a specific receptor-mediated process involving integrin- (alphavbeta3, alphavbeta5, CD36), phosphatidylserine receptor- and lectin-dependent mechanisms. No significant differences were observed in avarice for apoptotic eosinophils by SAEC or LAEC either resting, CD44 monoclonal antibodies- or cytokine- stimulated, or in their usage and expression of recognition receptors. CONCLUSION: These findings further suggest and define an important role for the bronchial epithelium in the selective removal of apoptotic eosinophils from the airways in asthma.

Granule protein changes and membrane receptor phenotype in maturing human eosinophils cultured from CD34+ progenitors.

BACKGROUND: Eosinophils are now recognized as major effector cells in allergic and asthmatic disease with a potent armoury of mediators whose release makes a major contribution to the inflammation underlying these conditions. OBJECTIVES: The purpose of this study was to compare cultured eosinophils (CE) with normal-density peripheral blood eosinophils (PBE) in terms of their membrane receptor expression and to analyse the expression and storage of the eosinophil granule proteins major basic protein (MBP) and eosinophil cationic protein (ECP) during eosinophil maturation in vitro. METHODS: Purified human peripheral blood CD34+ cells were cultured in the presence of recombinant human IL-3, IL-5, rhGM-CSF, SCF, and FLT-3 ligand. PBE were isolated by density gradient centrifugation and negative immunomagnetic selection. Expression of CD11b, CD18, CD45, CD45RA, CD45RB, CD45RO, CD69, CD95, IL-5Ralpha, IL-9Ralpha, CCR1, CCR3, and CXCR4 by CE as they matured in culture were assessed by immunostaining and flow cytometry and expression of these receptors compared with freshly isolated PBE. Immunohistochemical staining and labophot-2TM light microscopy determined expression of MBP, ECP, and CD69 during eosinophil maturation. RESULTS: Positive immunostaining for MBP and ECP was detectable in a proportion (15-20%) of CE as early as 3 days of culture even though these cells were mononuclear in appearance. The numbers of CE positive for both granule proteins increased in rhIL-3 and rhIL-5 treated cells to a maximum of approximately 80% by day 28. Maturing eosinophils exhibited positive immunostaining for CD69 after 14, 21 and 28 days of culture. Compared with PBE, CE had lower expression of pan-CD45 and CD45 isoforms, CD95 and CD11b. In contrast, the specific mean fluorescence for CD69, CD18, IL-5Ralpha, and IL-9Ralpha was significantly elevated for CE compared with PBE. CCR3 expression by CE and PBE was similar with no expression of CXCR4 detected by either CE or PBE. No significant difference in expression of CCR1 was found between CE and PBE. CONCLUSION: These data suggest that CE and PBE share many phenotypic properties and both MBP and ECP appear early in eosinophil development in vitro. However, there are quantitative differences that may be a consequence of their immaturity and/or the influence of the cytokines used in their culture.