Clinical significance of plasma PAF acetylhydrolase activity measurements as a biomarker of anaphylaxis: Cross-sectional study

Introduction Platelet-activating factor (PAF) has a direct role as a mediator in the pathogenesis of various disorders with an inflammatory component, including those with allergic aetiology. The peripheral blood concentration of PAF is dynamically regulated by plasma PAF acetylhydrolase (PAF-AH). Previous research suggest that low activity of plasma PAF-AH could be a predictive marker for increased severity of some types of allergic hypersensitivity reactions–especially anaphylaxis. The purpose of the study was to evaluate the association between plasma PAF-AH activity and severity in patients with anaphylactic reactions following a wasp or bee sting. Materials and methods The study group of 89 patients was divided into two subgroups depending on the increasing severity of the most severe anaphylactic reaction in the past, which was assessed according to the Müller’s scale. The first subgroup included participants with a history of hypersensitivity reactions up to grade II. The second subgroup consisted of patients who have experienced at least one grade III or IV reactions in the past. A control group of 20 people was established. Plasma PAF-AH activity was measured using a colorimetric method. Results It has been observed that plasma activity of platelet-activating factor acetylhydrolase was significantly lower in patients with anaphylaxis history compared to the control group with negative atopic history (on average 21.38 nmol/min/ml for the control group, 9.47 nmol/min/ml for the first subgroup and 10.16 nmol/min/ml for the second subgroup, in both cases p < 0.0001). Conclusion The plasma activity of PAF-AH is a promising parameter that can help to distinguish a group of patients not threatened with development of anaphylaxis and not requiring laborious or expensive prophylactic procedures.

Προσδιορισμός του παράγοντα ενεργοποίησης των αιμοπεταλίων (PAF) στο αίμα ασθενών με διαβητική αμφιβληστροειδοπάθεια

Εισαγωγή: Η διαβητική αμφιβληστροειδοπάθεια αποτελεί τη σοβαρότερη οφθαλμολογική επιπλοκή του σακχαρώδους διαβήτη, πλήττοντας 93 εκατομμύρια διαβητικούς ασθενείς παγκοσμίως. Ο Παράγοντας Ενεργοποίησης των Αιμοπεταλίων (Platelet activating factor, PAF) αποτελεί ένα φωσφολιπίδιο χαμηλού μοριακού βάρους και ενοχοποιείται για την αύξηση της αγγειακής διαπερατότητας και τη νεοαγγείωση, που παρατηρούνται στη διαβητική αμφιβληστροειδοπάθεια και σε άλλες οφθαλμικές παθήσεις.Στόχος: Η εκτίμηση του ρόλου της εξωκυττάριας (πλάσματος) ακετυλ-υδρολάσης του PAF (PAF acetylhydrolase, PAF-AH) στην παθογένεια και την εξέλιξη της διαβητικής αμφιβληστροειδοπάθειας.Μεθοδολογία: Εξήντα οκτώ διαβητικοί και 23 υγιή άτομα ανάλογης ηλικίας υπεβληθήκαν σε αιμοληψία και μετρήθηκε η δραστηριότητα της PAF-AH στο πλάσμα των συμμετεχόντων. Οι διαβητικοί ασθενείς ταξινομήθηκαν περεταίρω σε δύο ομάδες με βάσει το σύστημα ETDRS (Early Treatment Diabetic Retinopathy Study), με τη βοήθεια της έμμεσης οφθαλμοσκόπησης και της φλουοροαγγειογραφίας. Τριάντα επτά ασθενείς με μη παραγωγική διαβητική αμφιβληστροειδοπάθεια (non-proliferative diabetic retinopathy, NPDR) και 31 ασθενείς με παραγωγική διαβητική αμφιβληστροειδοπάθεια (proliferative diabetic retinopathy, PDR) συμπεριελήφθησαν τελικά στη μελέτη. Αποτελέσματα: Η δραστικότητα της PAF-AH του πλάσματος βρέθηκε αυξημένη στους διαβητικούς ασθενείς σε σχέση με την ομάδα των υγιών (υγιείς: 0.114µmol/min/ml, NPDR: 0.147µmol/min/ml and PDR: 0.206µmol/min/ml) και οι διαφορές των μέσω τιμών ήταν στατιστικά σημαντικές ανάμεσα στις τρεις ομάδες [One-Way ANOVA, p<0.0001, F= 10 (2,88)].Συμπεράσματα: Η δραστικότητα της PAF-AH του πλάσματος αυξάνεται παράλληλα με τη βαρύτητα της διαβητικής αμφιβληστροειδοπάθειας.

A Neutralizing Monoclonal IgG1 Antibody of Platelet-Activating Factor Acetylhydrolase SsE Protects Mice against Lethal Subcutaneous Group A Streptococcus Infection

Group AStreptococcus(GAS) can cause life-threatening invasive infections, including necrotizing fasciitis. There are no effective treatments for severe invasive GAS infections. The platelet-activating factor (PAF) acetylhydrolase SsE produced by GAS is required for invasive GAS to evade innate immune responses and to invade soft tissues. This study determined whether the enzymatic activity of SsE is critical for its function in GAS skin invasion and inhibition of neutrophil recruitment and whether SsE is a viable target for immunotherapy for severe invasive GAS infections. An isogenic derivative of M1T1 strain MGAS5005 producing SsE with an S178A substitution (SsES178A), an enzymatically inactive SsE mutant protein, was generated. This strain induced higher levels of neutrophil infiltration and caused smaller lesions than MGAS5005 in subcutaneous infections of mice. This phenotype is similar to that of MGAS5005ssedeletion mutants, indicating that the enzymatic activity of SsE is critical for its function. An anti-SsE IgG1 monoclonal antibody (MAb), 2B11, neutralized the PAF acetylhydrolase activity of SsE. Passive immunization with 2B11 increased neutrophil infiltration, reduced skin invasion, and protected mice against MGAS5005 infection. However, 2B11 did not protect mice when it was administered after MGAS5005 inoculation. MGAS5005 induced vascular effusion at infection sites at early hours after GAS inoculation, suggesting that 2B11 did not always have access to infection sites. Thus, the enzymatic activity of SsE mediates its function, and SsE has the potential to be included in a vaccine but is not a therapeutic target. An effective MAb-based immunotherapy for severe invasive GAS infections may need to target virulence factors that are critical for systemic survival of GAS.

Enhanced breast cancer cell adherence to the lung endothelium via PAF acetylhydrolase inhibition: a potential mechanism for enhanced metastasis in smokers

Cancer deaths are primarily caused by distant metastases, rather than by primary tumor growth; however, the role of smoking in metastasis remains unclear. We demonstrated previously that endothelial cell platelet-activating factor (PAF) production results in enhanced inflammatory cell recruitment to the lung. We propose that endothelial cell PAF accumulation plays a role in cancer cell migration to distal locations. We used cigarette smoke extract (CSE) to inhibit the activity of endothelial cell PAF acetylhydrolase (PAF-AH), which hydrolyzes and inactivates PAF, and determined whether this results in increased endothelial cell PAF accumulation and breast cancer adherence. Incubation of human lung microvascular endothelial cells (HMVEC-L) with CSE resulted in a significant inhibition of PAF-AH activity that was accompanied by increased PAF production and adherence of highly invasive MDA-MB-231 breast cancer cells. Pretreatment of HMVEC-L with ( S)-bromoenol lactone to inhibit calcium-independent phospholipase A2β (iPLA2β, which initiates endothelial cell PAF production) prior to CSE exposure resulted in complete inhibition of MDA-MB-231 cell adherence. Similarly, pretreatment of MDA-MB-231 cells with the PAF receptor antagonist Ginkgo biloba resulted in inhibition of adherence to the endothelium. Immunoblot analysis indicated an increase in MDA-MB-231 cell PAF receptor expression with CSE exposure. Taken together, our data indicate that CSE exposure increases endothelial cell PAF production, resulting in enhanced adherence of tumor cells to the endothelium. Our in vitro data indicate that increased tumor cell adherence would lead to enhanced metastasis formation in smokers. Potential therapeutic targets include endothelial cell iPLA2β or the tumor cell PAF receptor.

Characterization of Streptococcal Platelet-Activating Factor Acetylhydrolase Variants That Are Involved in Innate Immune Evasion

ABSTRACTHuman pathogen group A streptococcus (GAS) has developed mechanisms to subvert innate immunity. We recently reported that the secreted esterase produced by serotype M1 GAS (SsEM1) reduces neutrophil recruitment by targeting platelet-activating factor (PAF). SsEM1and SsE produced by serotype M28 GAS (SsEM28) have a 37% sequence difference. This study aims at determining whether SsEM28is also a PAF acetylhydrolase and participates in innate immune evasion. We also examined whether SsE evolved to target PAF by characterizing the PAF acetylhydrolase (PAF-AH) activity and substrate specificity of SsEM1, SsEM28, SeE, the SsE homologue inStreptococcus equi, and human plasma PAF-AH (hpPAF-AH). PAF incubated with SsEM28or SeE was converted into lyso-PAF. SsEM1and SsEM28hadkcatvalues of 373 s−1and 467 s−1, respectively, that were ≥30-fold greater than that of hpPAF-AH (12 s−1). The comparison of SsEM1, SsEM28, and hpPAF-AH inkcatandKmin hydrolyzing triglycerides, acetyl esters, and PAF indicates that the SsE proteins are more potent hydrolases against PAF and have high affinity for PAF. SsEM28possesses much lower esterase activities against triglycerides and other esters than SsEM1but have similar potency with SsEM1in PAF hydrolysis. Deletion ofsseM28in acovSdeletion mutant of GAS increased neutrophil recruitment and reduced skin infection, whereas intransexpression of SsEM28in GAS reduced neutrophil infiltration and increased skin invasion in subcutaneous infection of mice. These results suggest that the SsE proteins evolved to target PAF for enhancing innate immune evasion and skin invasion.