Proteogenomics of selective susceptibility to endotoxin using circulating acute phase biomarkers and bioassay development in sheep: a review

Abstract

Scientists have injected endotoxin into animals to investigate and understand various pathologies and novel therapies for several decades. Recent observations have shown that there is selective susceptibility to Escherichia coli lipopolysaccharide (LPS) endotoxin in sheep, despite having similar breed characteristics. The reason behind this difference is unknown, and has prompted studies aiming to explain the variation by proteogenomic characterisation of circulating acute phase biomarkers. It is hypothesised that genetic trait, biochemical, immunological and inflammation marker patterns contribute in defining and predicting mammalian response to LPS. This review discusses the effects of endotoxin and host responses, genetic basis of innate defences, activation of the acute phase response (APR) following experimental LPS challenge, and the current approaches employed in detecting novel biomarkers including acute phase proteins (APP) and micro-ribonucleic acids (miRNAs) in serum or plasma. miRNAs are novel targets for elucidating molecular mechanisms of disease because of their differential expression during pathological, and in healthy states. Changes in miRNA profiles during a disease challenge may be reflected in plasma. Studies show that gel-based two-dimensional electrophoresis (2-DE) coupled with either matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) or liquid chromatography–mass spectrometry (LC-MS/MS) are currently the most used methods for proteome characterisation. Further evidence suggests that proteomic investigations are preferentially shifting from 2-DE to non-gel based LC-MS/MS coupled with data extraction by sequential window acquisition of all theoretical fragment-ion spectra (SWATH) approaches that are able to identify a wider range of proteins. Enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qRT-PCR), and most recently proteomic methods have been used to quantify low abundance proteins such as cytokines. qRT-PCR and next generation sequencing (NGS) are used for the characterisation of miRNA. Proteogenomic approaches for detecting APP and novel miRNA profiling are essential in understanding the selective resistance to endotoxin in sheep. The results of these methods could help in understanding similar pathology in humans. It might also be helpful in the development of physiological and diagnostic screening assays for determining experimental inclusion and endpoints, and in clinical trials in future.

Determination of the Efficiencies of the Prokinetics in Ruminants with Postoperative Ileus Using Pro-Inflammatory Markers

“…HPT, with SAA in particular, is a major APP in lambs [4], and has antioxidant, anti-inflammatory, immune regulatory and antibacterial properties. Therefore, it can be used to detect various diseases [4,13].…”

“…The area between the umbilicus and pelvic region was shaved for surgery. General anesthesia was performed by injecting 6 mg kg -1 ketamine hydrochloride (Ketasol ® ) 4 intramuscularly following the injection of 0.2 mg kg -1 xylazine hydrochloride (Rompun ® ) 5 as intramuscularly.…”

The recognition of LpxC inhibitors as potential antibiotics could revolutionise the management of sepsis in veterinary patients if their unknown biological properties are widely evaluated in suitable animal models

“…Established large animal studies such as these on LPS could set a foundation for testing MDRGNB challenge through incremental and validation experiments. Work in this area could embrace objectives such as the determination of physiological, haematological, biochemical, immunological, histopathological, and inflammation effects, and of proteogenomic studies [26] of LpxC-2 in vivo. The studies should able to determine the kinetics of LpxC-2 and related molecules in blood, tissues and urine.…”

The implications of brain death in donor lung injury: investigation and blockade of the endothelin axis

“…An ongoing challenge in developing new models is a relative paucity of validated, species specific analytical methods. Our group continues to develop and validate ovine specific tests, [331,355] and the presented model will provide a platform to further investigate cytokine expression after BD in future studies.…”

“…Ventricular dysfunction that is unresponsive to intervention and leads to death is reported to occur in 10-40% of cases of brain death. [121] Although the application of ovine models is gaining increased acceptance, [294,315,331] industry development of sheep specific investigational modalities is slow. Antibodies and ELISA kits aimed at identifying a wide range of targets in rodent and human tissue/blood samples have been developed, however not all of these are appropriate for use in ovine models.…”