Ruggerone / Paltrinieri

Beatrice Ruggerone, DVM, PhD Student

She graduated in May 2013 in Veterinary Medicine at the University of Milan (110/100 cum laude), with a thesis entitled “Cystine calculi in English Bulldog: clinical examination and DNA test” (Supervisor: Prof. Michele Polli).
After graduating and once acquired the professional license, she started her career in a private clinic where she was in charge of internal medicine and recovery management. At the same time, she continued to work as a volunteer assistant at the Institute of Clinical Medicine and at the Laboratory of the Veterinary Science and Public Health Department (former Institute of General Patology), where she had the opportunity to acquire more skills about internal medicine, the execution and the interpretation of hematological and biochemical analysis.
Her main research interests are pathogenesis and diagnosis of endocrinopathies, nephropathies, infectious diseases of domestic animals and the analytical validation of innovative diagnostic markers of inflammation.


Saverio Paltrinieri, DVM, PhD, Dipl. ECVCP

After his degree in Veterinary Medicine at the University of Milan, Saverio Paltrinieri obtained a PhD in Comparative pathology of domestic animals at the same University. Currently he is associate professor of Veterinary general pathology and pathophysiology at the Department of Veterinary sciences and Public Health of the University of Milan and vicedirector of the Clinial Pathology Unit of the Central Lab of the veterinary teaching hospital of the University of Milan. Saverio Paltrinieri is a charter member of the European College of Veterinary Clinical Pathology (ECVCP) and he is the current ECVCP Past-President. Saverio Paltrinieri is also the current president of the Italian association of feline medicine (SIMEF) and a member of the working group on canine leishmaniasis. Research interest are focused on the pathogenesis and diagnosis of the main clinico-pathological abnormalities of domestic animals, with special emphasis on biomarkers of inflammation and on infectious diseases of dogs and cats.


RESEARCH PROJECT
Oxidative-antioxidative compounds and lipid parameters as potential diagnostic and prognostic markers in animals with SIRS

SIRS is a systemic inflammatory response that could be triggered by a variety of infectious and noninfectious conditions. Signs of systemic inflammation can and do occur, for example, in the absence of infection among patients with burns, pancreatitis, and other disease states.
Sepsis is a systemic inflammatory response caused by an infection. For example, the most common causes of sepsis in dogs and cats are pyometra, endocarditis, peritonitis, pyelonephritis and pneumonia; in horses, SIRS in often associated to colics, laminitis and pneumonia. To make diagnosis, it is necessary to find directly the involved pathogens with PCR or culture and/or to evaluate their indirect effects on the organism (leukogram, acute phase proteins, serum protein electrophoresis, temperature, heart and respiratory rate); but it’s uneasy to diagnose sepsis because these parameters are not enough sensible and specific and useful markers in Human medicine (e.g. procalcitonin) aren’t reliable in Veterinary medicine.

The inflammatory process associated with SIRS or sepsis is characterized by oxidative phaenomena, induced by pathogens or, more frequently, by phagocytes through the release of oxidant compounds from cytoplasmatic granules or of enzymes that damage cell membranes or tissue molecules generating oxidant compounds. The efficiency of phagocytes depends on their ability to produce reactive oxygen species (ROS).
Oxidative stress (OS) is the consequence of an imbalance between oxidants and antioxidants in which oxidant activity exceeds the neutralizing capacity of antioxidants.
The d-ROMs (Reactive Oxygen Metabolites - derived compounds) test provides a measure of the whole oxidant capacity of plasma. Alternatively, the presence of oxidation is indirectly measured by assessing the amount of oxidated molecules (independently on the type of molecule responsible for this oxidation). This may be done using different molecules (MDA, TBARS).


Another approach to evaluate the presence of oxidation is the measurement of anti-oxidant compounds (such as the BAP, or the total antioxidant capacity – TAC – in blood), that decrease when oxidation is present. Moreover, since some antioxidative compound is strictly linked to lipid parameters such as lipoproteins, the measurement of lipid parameters may provide similar information.

[Schematic representation of the interactions between inflammatory cells, acute phase proteins, antixodant molecules and lipids (designed by Dr. Gabriele Rossi).]

PON1 is an enzyme associated with high-density lipoproteins (HDL) that protects low-density lipoproteins (LDL) and HDL from peroxidation. Moreover, PON1 possesses anti-inflammatory properties, as it reduces the production of proinflammatory. PON1 is mainly expressed in the liver and is transported in plasma bound to HDL. During an acute phase response, HDL molecules lose apolipoprotein A1, esterified cholesterol, and most of the HDL-associated enzymes, including PON1, which is replaced mainly by serum amyloid A and ceruloplasmin. Altogether, this results in reduced antioxidative properties of HDL.

High density lipoproteins (HDLs), produced in the liver, distribute cholesterol and triglycerides to peripheral tissues and have antioxidant properties. In people, HDL levels negatively correlate with the severity of clinical signs and with the concentration of cytokines in several diseases.

In this project, enrolled patients will be divided into 3 groups on the basis of clinical findings and tests indicating SIRS (controls, sick without SIRS and sick with SIRS). A variety of conditions that may induce SIRS will be considered. These include sepsis or other conditions on which infectious agents (e.g. intracellular parasites) may induce a systemic inflammatory response).

Segmented neutrophils with toxic changes, immature neutrophils (one band neutrophil and one metamyelocyte) and a lymphocyte in a blood smear from a dogs with acute inflammation.

In every group, we will measure traditional markers and the new ones (dROMs, TBARS, TAC, PON and HDL) at enrollment and during therapy.
Based on our knowledge about the role of oxidative-antioxidative compounds during inflammation and on the relationship between the redox status and HDL lipoproteins, is very likely that oxidants and LDL increase and anti-oxidants or HDLs decrease in both groups of animals with inflammation compared to controls. However, being the magnitude of inflammation (and therefore oxidation) higher in subjects with SIRS that in those sick but asymptomatic, it is expected that the magnitude of biochemical changes will be higher in subjects with SIRS than in the other groups. Therefore, this, or future studies on larger cohorts of animals, will allow to select a potential cut-off to differentiate patients with SIRS from those without SIRS. Moreover, results of sequential samples collected after treatment will allow to identify which parameters may work as best indicators of response to treatments.
In other words, the use in routine practice of dROMS, TBARS, TAC , PON, and HDL will may be a complementary approach to conventional markers of inflammation: results at admission higher or lower than a given cut-off will be useful to predict the presence of SIRS and to draw prognostic informations. Similarly, repeated measurements of these indicators during the follow up may be used in practice to early identify subjects that successfully respond (or not) to treatments.