There have been reports regarding
There have been reports regarding the use of enolases as novel vaccine candidates for A. suum, S. suis, T. pisiformis and T. multiceps on account of this enzyme class's critical role in plasminogen activation and migration across tissue barriers (Chen et al., 2012; Zhang et al., 2009; Zhang et al., (Li et al., 2015). Seeing how membranal, secretory, and excretory proteins are frequently assessed for their use as potential immunodiagnostic antigens on account of their being highly exposed to the host immune system, our findings add credence to the idea that T. solium enolase may fit such a mold, regardless of its true in vivo functionality. Indeed, previous studies have attempted to utilize enolases in immunological techniques to diagnose human echinococcosis due to E. granulosus (Y. Wang et al., 2012) as well as to diagnose both P. falciparum and P. vivax variants of malaria in Southeast Asia (Sato et al., 2000). Unfortunately, in the case of T. solium enolase, one major hurdle to the development of a diagnostic assay invariably exists: given that the majority of the amino Milrinone receptor alignment between the enolases of T. solium, A. suum, and T. spiralis corresponded to conserved domains (∼75%), many epitopes contained within the enolases of these three species may well be identical. This would likely result in the generation of numerous cross-reactive antibodies that would be unable to distinguish one infection from another, thereby severely hampering specificity and potentially limiting the applicability of such an antigen to immunodiagnostics. Consequently, any efforts to develop an enolase-based immunodiagnostic for T. solium infection should be sure to focus on the regions that vary between these three species as to improve the specificity of any such assay, be it through the use of monoclonal antibodies against these regions or through the generation of recombinant proteins that only include non-conserved motifs.
As it happens, cross-reactivity may very well have occurred during our own evaluations; the high sensitivity (97.67%) and low specificity (46.51%) that we observed when applying our first cut-off value (i.e. the mean ODs of 6 negative samples + 3SD) to our rEnoTsBac-based immunodiagnostic is exactly what would have been expected if enolases were potent immunogens and if some of the T. solium-negative pigs had previously been exposed to A. suum or T. spiralis, both of which are known to infect pigs. Although T. spiralis has yet to have been reported in Peru, the presence of A. suum has been previously documented and its transmission in an industrialized farm is not out of the realm of possibility (Katakam et al., 2016). Such cross-reactivity could potentially be responsible for the high rate of false positives that we observed and might be able to be identified by assaying the sera of pigs known to have been exposed to these helminths.
In conclusion, we have successfully expressed a functional recombinant enolase from T. solium in both prokaryote and eukaryote expression systems and demonstrated its glycolytic activity. Moreover, we found that recombinant enolase expressed in Sf9 insect cells is a promising candidate for the development of future diagnostics tests for porcine cysticercosis. Further studies are required to explore the use of enolase as a vaccine candidate against porcine cysticercosis.
Acknowledgments This study was supported by the Programa Nacional de Innovación para la Competitividad y Productividad-Innóvate Perú (contract number 181-FINCyT-IB-2013) and the World Academy of Science award (No.14-233RG-BIO-LA UNESCO FR324028597).
Introduction The enzyme Enolase (ENO1, EC.22.214.171.124) acts in glycolysis to convert 2-phosphoglycerate to phosphoenolpyruvate. Twenty years ago, Shand and West had proposed that next to its glycolytic function Enolase can also inhibit cholesteryl ester hydrolases (CEHs) , . The transition of macrophages to “foam cells” in the atherosclerotic plaque is accompanied by storage of cholesterol esters in lipid droplets , . This process, which can be mimicked experimentally in vitro by loading macrophages with acetyl-LDL or oxidized-LDL, requires enhanced cholesterol esterification activity. Accordingly, cholesterol loading of macrophages results in a marked increase in ENO1 protein , , which can potentially inhibit CEHs on the surface of lipid droplets , .