Combined effects of Limonene and Ivermectin on P-glycoprotein-9 gene expression of lambs Infected with Haemonchus contortus.

Although ivermectin (IVM) has a wide spectrum and long half-life, its frequent use as an anthelmintic for the last 42 years led to its worldwide tolerance by Haemonchus contortus. We evaluated the combination of limonene (LIM), a P-glycoprotein (Pgp) modulator, with IVM in lambs infected with a multidrug-resistant H. contortus. Twenty-four male Dorper lambs were artificially infected with two doses (seven days apart) of 8000 infective larvae of a multidrug-resistant isolate of H. contortus. The infection was patent 25 days later. Fifteen days before treatment with IVM (DAY -15), animals were divided into 4 groups: Infected-untreated control (CTL), IVM, LIM, and LIM+IVM. From DAY -15 to DAY + 14, groups LIM and LIM+IVM received 200 mg/kg of body weight/day of LIM via oral. On DAY 0, a single dose of IVM at 200 µg/kg of body weight was administered orally to groups IVM and LIM+IVM. On DAY + 7 and DAY + 14, fecal egg counts (FEC) were performed and on DAY + 14 animals were euthanized for total worm count (TWC), worm length, fecundity of females, and Pgp-9 gene expression. On DAY + 7, group LIM+IVM had 96.29% efficacy based on Fecal Egg Count Reduction TEST (FECRT) and a highly significant reduction in FEC (P = 0.0005) when compared to CTL. On DAY + 14, the efficacy of LIM+IVM was 82.87% on FECRT, although no differences were found among groups for FEC, TWC, worm length, or Pgp-9 gene expression. Female worms from the CTL group had higher egg counts in their uterus when compared to LIM. No differences were found for hematological or biochemical parameters, body weight, or weight gain among groups. Thus, LIM given daily at 200 mg/kg was safe for animals and, when combined with IVM, decreased egg shedding and could reduce pasture contamination, although it was unable to kill multidrug-resistant H. contortus.

Effects of combined physical exercise on plasma lipid variables, paraoxonase 1 activity, and inflammation parameters in adults with obesity: a randomized clinical trial.

OBJECTIVE: We aimed to investigate whether combined physical exercise may affect plasma lipid variables, paraoxonase 1 (PON1) activity, and inflammation parameters in adults with obesity. METHODS: Thirty-six participants were recruited to complete the study protocol. The mean age was 37 ± 1 years, and the baseline body mass index was 33.0 ± 0.4 kg/m2. Participants were allocated to the control group (CG) and the exercise group (EG). The EG performed three weekly sessions of combined physical exercise for 16 weeks. Plasma lipid variables, PON1 activity, and inflammatory profile were determined before and after intervention. RESULTS: Total cholesterol levels decreased in both groups, without intergroup difference (time p = 0.001). Non-high-density lipoprotein cholesterol (HDL-C) levels decreased in both groups (time p = 0.001); however, they were lower in the EG than in the CG (p = 0.038). The EG had increased HDL-C levels, but the CG had decreased HDL-C levels (time*group p = 0.011). PON1 activity was reduced in both groups (time, p = 0.001). The Castelli risk Index I and II reduced in the EG and increased in the CG (time*group, p = 0.008 and p = 0.011, respectively). The inflammatory markers were not modified. CONCLUSION: Adults with obesity may benefit from regular practice of combined physical exercise training in many metabolic aspects that are related to protection against the development of cardiovascular disease.

Factors affecting executive function performance of Brazilian elderly in the Stroop test.

Aging is related to a decrease in physiological abilities, especially cognitive functions. To unravel further evidence of age-related cognitive decline, we analyzed which physical and functional variables are predictors of cognitive performance in a sample of 498 Brazilian elderly (67.26% women). To do so, we used the Stroop test as a tool to evaluate executive functions and the General functional fitness index (GFFI) to evaluate the functional fitness of the participants. A linear regression analysis revealed that female sex (ß=-0.097; t=-2.286; P=0.023), younger age (ß=0.205; t=4.606; P<0.0001), more years of education (ß=-0.280; t=-6.358; P<0.0001), and higher GFFI (ß=-0.101; t=-2.347; P<0.02) were predictors of better cognitive performance. Body mass index (kg/m2) and nutritional status (underweight, eutrophic, overweight, or obese) were not predictors of cognitive performance. Interestingly, among the GFFI tasks, muscle strength influenced the test execution time, both in upper and lower limbs (elbow flexion: ß=-0.201; t=-4.672; P<0.0001; sit-to-stand: ß=-0.125; t=-2.580; P<0.01). Our findings showed that: 1) women performed the Stroop test faster than men; 2) the older the person, the lower was the cognitive performance; 3) the higher the education, the better the test execution time; and 4) higher scores in the GFFI were associated with a better performance in the Stroop test. Therefore, gender, age, education, and functional fitness and capacity were predictors of cognitive performance in the elderly.

Factors affecting executive function performance of Brazilian elderly in the Stroop test

Aging is related to a decrease in physiological abilities, especially cognitive functions. To unravel further evidence of age-related cognitive decline, we analyzed which physical and functional variables are predictors of cognitive performance in a sample of 498 Brazilian elderly (67.26% women). To do so, we used the Stroop test as a tool to evaluate executive functions and the General functional fitness index (GFFI) to evaluate the functional fitness of the participants. A linear regression analysis revealed that female sex (β=-0.097; t=-2.286; P=0.023), younger age (β=0.205; t=4.606; P<0.0001), more years of education (β=-0.280; t=-6.358; P<0.0001), and higher GFFI (β=-0.101; t=-2.347; P<0.02) were predictors of better cognitive performance. Body mass index (kg/m2) and nutritional status (underweight, eutrophic, overweight, or obese) were not predictors of cognitive performance. Interestingly, among the GFFI tasks, muscle strength influenced the test execution time, both in upper and lower limbs (elbow flexion: β=-0.201; t=-4.672; P<0.0001; sit-to-stand: β=-0.125; t=-2.580; P<0.01). Our findings showed that: 1) women performed the Stroop test faster than men; 2) the older the person, the lower was the cognitive performance; 3) the higher the education, the better the test execution time; and 4) higher scores in the GFFI were associated with a better performance in the Stroop test. Therefore, gender, age, education, and functional fitness and capacity were predictors of cognitive performance in the elderly.

Corpses against science death: new Italian law titled 'Rules relating to the directives of one's body and tissues post-mortem for study, training, and scientific research purposes'.

The new Italian Law no. 10 of 2020, titled 'Rules relating to the directives of one's body and tissues post-mortem for study, training, and scientific research purposes' (10 February, 2020), has opened the doors to a new and clearer regimentation on body donation to science, according to the procedure already introduced by Law no. 219/2017 on the 'Rules relating to informed consent and advance directives of treatment'. The authors exposed some concerning regarding the application of the law, such as donation mistrust, also linked to our cultural heritage; the incomplete protection of minor and incompetent people; the complexity of an adequate information, which could undermine the informed consent; and finally, the fees overload for research institutes, with already exiguous budgets. Next, the authors analysed some positive aspects of the law and in primis the central role of corpses in scientifical knowledge and medical training. Moreover, a simple mechanism based on a conscious and pre-stated consent, registered in a database that follows European data protection Regulation, is discussed. Further, the procedure of the restitution of the bodies to their families, which represents a way to guarantee the relatives' commemoration, is also elucidated. In conclusion, the authors proposed a format of informed consent, to try a direct application of the law, thereby fastening the process of standardisation and centralisation of all the living wills. The authors optimistically investigated the opportunities offered by the law and encouraged a conscientious participation by citizens.

PERCUTANEOUS TRANSAXILLARY TRANSCATHETER AORTIC VALVE IMPLANTATION.

The transaxillary (TAX) approach for transcatheter aortic valve implantation (TAVI) results in comparable short and long-term clinical results compared to the transfemoral (TF) approach. However, adequate closure of the axillary artery is the most critical issue when performing the percutaneous approach. Compared to surgical transaxillary approach, the percutaneous approach was used only in selected cases due to this closure limitation. In the present paper, we aim to demonstrate the feasibility of implanting the CoreValve Evolut Pro transcatheter heart valve via percutaneous transaxillary approach and make a literature review of procedure particularities and outcome. We describe the case of a patient with severe aortic stenosis in the presence of small calibre and severely calcified femoral arteries. A CoreValve Evolut Pro 26 was successfully implanted percutaneously through the left axillary artery. Percutaneous transaxillary transcatheter aortic valve implantation is a feasible and safe alternative in patients who have suboptimal iliofemoral vessels.

Cardiac surgery in infective endocarditis and predictors of in-hospital mortality. / Cirurgia cardíaca na endocardite infeciosa e preditores da mortalidade intra­hospitalar.

INTRODUCTION: Infective endocarditis (IE) is a serious disease with significant in-hospital mortality (15-30%) despite advances in medical and surgical therapy. AIMS: To perform a clinical characterization of patients undergoing cardiac surgery for IE and to identify factors that predict in-hospital mortality. METHODS: We retrospectively analyzed 145 patients with IE admitted between January 2006 and October 2017. RESULTS: The median age was 72 years. IE was acquired mainly in the community (69%), and involved the native aortic valve in 54% of patients, biological prosthetic valves in 22.1% and mechanical valves in 10.3%. Staphylococcus spp. (31.0%) were the most frequent etiological agents. Cardiac surgery was emergent in 29 patients, urgent in 108, and elective in eight. The main indications were heart failure (57.9%), large vegetations (20%), systemic embolism (17.2%) and valve dysfunction (15.2%). Overall, biological valves were implanted in 62.1% of patients and mechanical valves in 37.2%. A total of 19 patients (13.1%) died. Predictors of mortality were preoperative atrial fibrillation and lower left ventricular ejection fraction, postoperative severe valve regurgitation associated with cardiogenic shock, sepsis, septic shock associated with cardiogenic shock, cardiac tamponade, need for renal replacement therapy and, although without statistical significance, emergent surgery. CONCLUSIONS: There is a need for better indicators to enable early identification of surgical candidates for IE, implementation of a heart team, and better surgical strategies, including more rapid intervention, more specific postoperative care, and optimal antibiotic therapy.

[Clinico-electroencephalographic variants in pharmacoresistant mesial temporal lobe epilepsy]. / Variantes clinicoelectroencefalograficas en la epilepsia del lobulo temporal mesial farmacorresistente.

INTRODUCTION: Focal epilepsy secondary to mesial temporal sclerosis (MTS) is one of the main causes of refractory epilepsy. It is typically associated with frontotemporal discharges in the electroencephalogram (EEG), a characteristic image in the magnetic resonance scan and a probability of post-operative remission above 70%. AIMS: To identify different patterns of ictal propagation in surface EEG recordings in patients with refractory epilepsy and MTS, and to analyse their relation with the post-operative outcome. PATIENTS AND METHODS: We conducted a retrospective review of the medical records of patients with refractory epilepsy secondary to MTS evaluated in the Epilepsy Surgery Programme of the Hospital de Clinicas, Montevideo (n = 30). The propagation of ictal rhythms was analysed in time windows of three seconds, and propagation maps were produced for each seizure. RESULTS: Six patterns were identified: ipsilateral temporal (type 1; 37%), bilateral frontotemporal with (type 2A; 22%) or without (type 2B; 17%) extension to suprasylvian regions, alternating temporal (type 3; 13%), unilateral suprasylvian (type 4; 7%) and bilateral at onset (type 5; 3%). The type 1 pattern was associated with classic clinical features and a favourable post-operative outcome. The clinical variants were associated with extratemporal EEG propagation. Patients with reflex seizures continued with post-operative seizures. Overall, no unambiguous relation was found between the ictal EEG pattern and the post-operative outcome. CONCLUSIONS: The ictal EEG pattern does not allow for a surgical prognosis in patients with epilepsy secondary to MTS. The history of reflex seizures in these patients may be a red flag suggesting a less favourable surgical outcome.

TITLE: Variantes clinicoelectroencefalograficas en la epilepsia del lobulo temporal mesial farmacorresistente.Introduccion. La epilepsia focal secundaria a esclerosis temporal mesial (ETM) constituye una de las principales causas de epilepsia refractaria al tratamiento farmacologico. Tipicamente asocia descargas frontotemporales en el electroencefalograma (EEG), imagen caracteristica en la resonancia magnetica y probabilidad de remision posquirurgica mayor del 70%. Objetivos. Identificar diferentes patrones de propagacion ictal en los registros electroencefalograficos de superficie en pacientes con epilepsia farmacorresistente y ETM, y analizar su relacion con la evolucion postoperatoria. Pacientes y metodos. Se revisaron retrospectivamente los registros de pacientes con epilepsia refractaria secundaria a ETM evaluados en el Programa de Cirugia de Epilepsia del Hospital de Clinicas, Montevideo (n = 30). La propagacion de ritmos ictales se analizo en ventanas temporales de tres segundos, y se elaboraron mapas de propagacion para cada crisis. Resultados. Se identificaron seis patrones: temporal ipsilateral (tipo 1; 37%), frontotemporal bilateral con (tipo 2A; 22%) o sin (tipo 2B; 17%) extension a regiones suprasilvianas, temporal alternante (tipo 3; 13%), suprasilviano unilateral (tipo 4; 7%) y bilateral de inicio (tipo 5; 3%). El patron de tipo 1 asocio un cuadro clinico clasico y buena evolucion posquirurgica. Las variantes clinicas se asociaron a propagacion EEG extratemporal. Los pacientes con crisis reflejas persistieron con crisis postoperatorias. En su conjunto, no se hallo una relacion univoca entre el patron EEG ictal y la evolucion postoperatoria. Conclusiones. El patron EEG ictal no permite predecir el pronostico quirurgico en pacientes con epilepsia secundaria a ETM. La historia de crisis reflejas en estos pacientes puede constituir una 'bandera roja' y sugerir un peor pronostico quirurgico.

Different outcomes of a cardiac rehabilitation programme in functional parameters among myocardial infarction survivors according to ejection fraction.

INTRODUCTION: Exercise-based cardiac rehabilitation (EBCR) is part of the management of patients who have suffered an acute myocardial infarction (AMI). Patients with a reduced ejection fraction (EF) comprise a higher-risk subgroup and are referred less often for these programmes. This study aimed at assessing the impact of the baseline EF on the functional benefits, as assessed by peak oxygen uptake (pVO2) and exercise duration, of an EBCR programme in AMI survivors. METHODS: Observational, retrospective cohort study including all patients admitted to a tertiary centre due to an AMI who completed a phase II EBCR programme after discharge, between November 2012 and April 2017. Functional parameters were assessed by a symptom-limited cardiopulmonary exercise test. RESULTS: A total of 379 patients were included [40.9% with reduced EF (<50%) at discharge]. After the programme, pVO2 and exercise duration increased significantly (p < 0.001). Patients with a reduced EF had a lower pVO2 and completed a shorter duration of exercise at the beginning and end of the programme. This group presented a higher increase in pVO2 (p = 0.001) and exercise duration (p = 0.007). This was maintained after adjusting for age, gender, history of coronary artery disease, number of sessions, Killip classification, arterial hypertension, dyslipidaemia, diabetes mellitus, smoking status and baseline pVO2. CONCLUSION: A phase II EBCR programme was associated with significant improvements in pVO2 and exercise duration among AMI survivors, irrespective of baseline EF classification. Those with a reduced baseline EF derived an even greater improvement, highlighting the importance of EBCR in this subgroup of patients.

The Impact of the Right Coronary Artery Geometric Parameters on Hemodynamic Performance.

PURPOSE: Coronary artery geometry can have a significant impact in the hemodynamic behavior of coronary blood flow, influencing atherosclerotic plaque formation. The present work focuses on, through a statistical study, the connection between several geometric parameters of the right coronary artery-ostium cross-sectional area, angles between the common trunk and the side-branches, tortuosity, curvature and cross-sectional area in each side-branch-and their influence on hemodynamic descriptors. Parameters such as low wall shear stress and local disturbed flow, which are associated with atherosclerosis formation, were analysed. METHODS: Computed tomography images of ten healthy individuals were selected to reconstruct in vivo three-dimensional models of right coronary arteries. Blood flow was simulated through a compliant model with realistic boundary conditions. Calculated hemodynamic descriptors values were correlated with the geometric parameters using the Pearson correlation coefficient (r) and the p value. RESULTS: The strongest correlations were found in the middle and distal segments of the right coronary artery. A decrease in the ostium area promotes a decrease in the WSS magnitude from the proximal to the distal segment (r = 0.82). Very strong correlations (r > 0.90) were achieved between geometric parameters (cross-sectional area, angle, tortuosity) of the right-ventricular branch and the wall shear stress magnitude in the middle and distal segments. CONCLUSIONS: Low values of tortuosity, smaller cross-sectional area and higher angle of the right-ventricular branch leads to a hemodynamic behavior more propitious to atherosclerosis formation, within the study cases. The right-ventricular branch seems to have the highest influence in the hemodynamic behavior of the right coronary artery.

Adrenergic receptor ß3 is involved in the memory consolidation process in mice.

Attention and emotion have a positive impact on memory formation, which is related to the activation of the noradrenergic system in the brain. The hippocampus and amygdala are fundamental structures in memory acquisition, which is modulated by noradrenaline through the noradrenergic receptors. Pharmacological studies suggest that memory acquisition depends on the action of both the ß3 (ß3-AR) and ß2 (ß2-AR) receptor subtypes. However, the use of animal models with specific knockout for the ß3-AR receptor only (ß3-ARKO) allows researchers to more accurately assess its role in memory formation processes. In the present study, we evaluated short- and long-term memory acquisition capacity in ß3-ARKO mice and wild-type mice at approximately 60 days of age. The animals were submitted to the open field test, the elevated plus maze, object recognition, and social preference. The results showed that the absence of the ß3-AR receptor caused no impairment in locomotion and did not cause anxious behavior, but it caused significant impairment of short- and long-term memory compared to wild-type animals. We also evaluated the expression of genes involved in memory consolidation. The mRNA levels for GLUT3, a glucose transporter expressed in the central nervous system, were significantly reduced in the amygdala, but not in the hippocampus of the ß3-ARKO animals. Our results showed that ß3-AR was involved in the process of acquisition of declarative memory, and its action may be due to the facilitation of glucose absorption in the amygdala.

Identification of Staphylococcus epidermidis with transferrable mupirocin resistance from canine skin.

Resistance to mupirocin was analysed in Staphylococcus spp. isolated from healthy dogs (n=21) and dogs with pyoderma (n=47) or otitis externa (n=52). Isolates were identified to species level by MALDI-TOF and PCR-RFLP of the groEL gene. One isolate of Staphylococcus epidermidis from the skin of a healthy dog, which harboured a plasmid carrying the mupA gene, was resistant to mupirocin.

Adrenergic receptor ß3 is involved in the memory consolidation process in mice

Attention and emotion have a positive impact on memory formation, which is related to the activation of the noradrenergic system in the brain. The hippocampus and amygdala are fundamental structures in memory acquisition, which is modulated by noradrenaline through the noradrenergic receptors. Pharmacological studies suggest that memory acquisition depends on the action of both the β3 (β3-AR) and β2 (β2-AR) receptor subtypes. However, the use of animal models with specific knockout for the β3-AR receptor only (β3-ARKO) allows researchers to more accurately assess its role in memory formation processes. In the present study, we evaluated short- and long-term memory acquisition capacity in β3-ARKO mice and wild-type mice at approximately 60 days of age. The animals were submitted to the open field test, the elevated plus maze, object recognition, and social preference. The results showed that the absence of the β3-AR receptor caused no impairment in locomotion and did not cause anxious behavior, but it caused significant impairment of short- and long-term memory compared to wild-type animals. We also evaluated the expression of genes involved in memory consolidation. The mRNA levels for GLUT3, a glucose transporter expressed in the central nervous system, were significantly reduced in the amygdala, but not in the hippocampus of the β3-ARKO animals. Our results showed that β3-AR was involved in the process of acquisition of declarative memory, and its action may be due to the facilitation of glucose absorption in the amygdala.

Comparison of the Results of Living Donor Liver Transplantation Due to Acute Liver Failure and Biliary Atresia in a Quaternary Center.

OBJECTIVE: The objective of this study was to compare the complications, outcomes, and survival prevalence in patients undergoing living donor liver transplantation due to biliary atresia (BA) or acute liver failure (ALF). RESULTS: In the period of June 1998-July 2016, 199 children underwent living transplantation due to BA or ALF. Of these 199, 184 were included in the analysis. The average age, weight, and body mass index of BA patients were lower than those of ALF (P < .001). The chi-square test showed a higher prevalence of infection in transplant recipients due to BA (P = .0001) and a higher prevalence of hepatic artery stenosis in those who underwent transplantation due to ALF (P = .001). In the multivariate analysis, the infection remains statistically more prevalent in the BA group (95% confidence interval [CI], 0.20-0.60), while hepatic artery stenosis loses significance. The mortality rate was similar in both groups and the survival in 5 years also. The prevalence of hepatic artery thrombosis, portal vein thrombosis/stenosis, biliary stenosis, and acute and chronic cellular rejection showed no statistical difference between the two groups. CONCLUSION: Living donor liver transplantation should be a valid option in cases of fulminant hepatitis with an indication for liver transplantation, especially in places where the number of cadaverous donors is low and the length of time on the waiting list is high.

Epithelial cells from oral mucosa: How to cultivate them?

Epithelial cells from oral mucosa (EOM) are responsible for important functions, like the primary protection of oral mucosa against external aggressions building a mechanical barrier against microorganisms, mechanical damage, toxic material, thermal regulation and secretion of different classes of inflammatory mediators. EOM could be an interesting tool for cellular and molecular biology research. Usually, EOM are collected by a painful and invasive process. In this study, we propose an alternative method to cultivate EOM collected by non-invasive scraping method of oral mucosa. Papanicolaou staining showed mainly two kinds of epithelial cell population after EOM scraping. As result of the five culture methods tested here, our results revealed that the EOM were successfully cultured on a murine feeder layer. In addition, EOM could be frozen and thawed, without morphology changes and loss of viability. Our findings suggest that EOM can be considered as a good cell source for many purposes, such as genetic studies, diagnosis and cell therapy.

Fibroblast sources: Where can we get them?

Fibroblasts are cells widely used in cell culture, both for transient primary cell culture or permanent as transformed cell lines. Lately, fibroblasts become cell sources for use in disease modeling after cell reprogramming because it is easily accessible in the body. Fibroblasts in patients will maintain all genetic background during reprogramming into induced pluripotent stem cells. In spite of their large use, fibroblasts are obtained after an invasive procedure, a superficial punch skin biopsy, collected under patient's local anesthesia. Taking into consideration the minimum patient's discomfort during and after the biopsy procedure, as well as the aesthetics aspect, it is essential to reflect on the best site of the body for the biopsy procedure combined with the success of getting robust fibroblast cultures in the lab. For this purpose, we compared the efficiency of four biopsy sites of the body (skin from eyelid, back of the ear, abdominal cesarean scar and groin). Cell proliferation assays and viability after cryopreservation were measured. Our results revealed that scar tissue provided fibroblasts with higher proliferative rates. Also, fibroblasts from scar tissues presented a higher viability after the thawing process.

Neonatal hyperthyroidism on rat heart: interrelation with nitric oxide and sex.

PURPOSE: To clarify the mechanism mediating the effect of hyperthyroidism on cardiac function during the second month of life in rats. METHODS: Male and female Sprague-Dawley rats were assigned to a control or to a triiodothyronine (T3)-treated group. Treatment of each group was started on the third day after birth. Control rats (Eut) received 0.9 NaCl [0.1 ml/100 g body weight (BW)] every second day during 60 days and T3-treated rats (Hyper) received subcutaneous (SC) T3 injections every second day during 60 days. RESULTS: Hyperthyroidism decreased left ventricle volume only in male rats. Female euthyroid rats presented higher atrial nitric oxide synthase (NOS) activity than male rats and hormonal treatment decreased this enzyme's activity in both sexes. Euthyroid male and female rats had similar atrial NOS protein levels, but females had higher caveolin (cav) 3 protein levels. T3 treatment increased this protein only in males. Female rats had lower ventricular NOS activity than male rats; hyperthyroidism increased NOS activity in both sexes but this effect was associated with lower cav 3 protein levels. Hyperthyroidism did not change cav 1 protein levels in both male and female rats. CONCLUSIONS: The results of this study demonstrating clinically relevant sex-related differences in the pathophysiology of the hyperthyroid heart have raised new questions regarding the mechanisms responsible for the observed differences. This study suggests that sex-related intrinsic factors such as nitric oxide may modulate the response to hyperthyroidism that leads to cardiovascular dysfunction.

Antioxidant activity of hyaluronic acid investigated by means of chemiluminescence of equine neutrophil bursts and electron paramagnetic resonance spectroscopy.

Activated neutrophils (PMNs), the ROS/RNS released by PMNs and the derived inflammatory processes are involved in the pathogenesis and progression of human inflammatory airway diseases. Similar diseases are also present in horses which suffer from recurrent airway obstruction (RAO), exercise-induced pulmonary haemorrhage (EIPH) and inflammatory airway diseases (IAD). Hyaluronic acid (HA) plays numerous roles in modulating inflammatory processes. The aim of this study was to examine whether a preparation of HA (MW 900 000 Da) interferes with ROS/RNS during the course of equine PMN respiratory bursts, and to establish the lowest concentration at which it still has antioxidant activity by means of luminol-amplified chemiluminescence (LACL). Electron paramagnetic resonance (EPR) spectroscopy was also used to investigate the direct antiradical activity of HA. The hydroxyl radical was significantly scavenged in a concentration-dependent manner at HA concentrations ranging from 2.5 to 0.16 mg/mL. Superoxide anion, Tempol radical and the ABTS(•+) were significantly inhibited at concentrations ranging from 2.5 to 0.62 mg/mL. The LACL of stimulated equine neutrophils showed that HA induced a statistically significant concentration-effect reduction from 5 mg/mL to 1.25 mg/mL. These findings were confirmed also when l-Arg was added to investigate the inhibition of the resulting peroxynitrite anion. Our findings indicate that, in addition to the human use, HA can also be used to antagonize the oxidative stress generated by free radicals in horses peripheral blood mononuclear cells (PBMCs). In order to achieve therapeutic concentrations, a direct aerosol administration to horses with horse respiratory diseases can be considered, as this route of application is also recommended in human medicine.

Diclofenac-Choline Antioxidant Activity Investigated by means of Luminol Amplified Chemiluminescence of Human Neutrophil Bursts and Electron Paramagnetic Resonance Spectroscopy.

A new diclofenac salt called diclofenac-choline (DC) has recently been proposed for the symptomatic treatment of oropharyngeal inflammatory processes and pain because its greater water solubility allows the use of high concentrations, which are useful when the contact time between the drug and the oropharyngeal mucosa is brief, as in the case of mouthwashes or spray formulations. The antioxidant activity of DC has not yet been investigated, and so the aim was to use luminol-amplified-chemiluminescence (LACL) to verify whether various concentrations of DC (1.48, 0.74 and 0.37 mg/mL for incubation times of 2, 4 and 8 min) interfere with oxygen and nitrogen radicals during the course of human neutrophils respiratory bursts; electron paramagnetic resonance (EPR) spectroscopy was used to investigate its direct antiradical (scavenger) activity. The EPR findings showed that DC has concentration-dependent scavenging activity against the ABTS, the DPPH, and the hydroxyl radicals, but no activity on superoxide anion, as has been previously reported in the case of other NSAIDs. LACL revealed an inhibitory effect that was statistically significant after only 2 min of incubation, and similar after 4 and 8 min. The effects on the peroxynitrite radical paralleled those observed in the previous test. High concentrations and short incubation times showed that there is no interference on PMN viability, and so the inhibitory findings must be attributed to the effect of the drug. The anti-inflammatory effects of DC cannot be attributed solely to the inhibition of prostaglandin synthesis, but its effects on free radicals and neutrophil bursts suggest that they may contribute to its final therapeutic effect.