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PostPosted: Fri Dec 25, 2020 10:59 am 
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ABSTRACT ... tfgU0k5UZ0

Ozone (O3) therapy has been used for medical procedures for centuries; however, there are no extensive studies on its utilization in horses. This study aimed to evaluate the application of transrectal O3 on horses by physical and laboratorial evaluation, and production of reactive oxygen species (ROS). Sixteen healthy horses were separated in two groups: a control group (CG) and a group treated with O3 (TG). The TG animals received 1L of an oxygen and O3 mixture transrectally. The initial dose was 10µg/ml for the first two applications, 15μg/ml for the following two applications, and 20μg/ml for the next six applications. The CG animals received 1L of oxygen transrectally. In TG animals no variations in the physical examination were detected; furthermore, TG animals did not exhibit changes in biochemical evaluation results, fibrinogen concentrations, or ROS production. TG animals had increased red blood cell counts, hemoglobin concentrations, and packet cell volume values in comparison to the baseline and CG values. We could infer that O3 affected the red blood cell counts and improved rhetological properties of the blood. The transrectal application of O3 in horses is safe and can indirectly improve the oxygenation and metabolism of tissues.


Payr and Aubourg (1936) were the first to suggest a rectal insufflation of a gas mixture with O3 in the colon, because it is a simple route of administration with a low cost. Currently, this technique is commonly used in humans around the world because it can be easily performed, has a low cost, and is mostly safe. The O3 effect on red cells, including an increase in rheology proprieties and flexibility, were observed after an O3 application during autohemotherapy in humans (Bocci, 1994; Gérard, 2001; Menéndez, 2008; Sousa, 2009). Other O3 effects involve pH intracellular alterations and increased 2,3-diphosphoglycerate that create a right shift in the oxygen-hemoglobin dissociation curve, consequently increasing oxygen delivery to the tissues (Giunta et al., 2001; Sagai and Bocci, 2011).

The present study utilized the typical number of applications used in clinical treatments to test the safety of the technique in equines. The horses did not show any signs of discomfort or present behavioral changes during the rectal O3 applications. The only prominent observation was increased defecation, probably caused by the mild rectal ampoule gas distension that facilitated a motility stimulus. The physiological parameters were within normal ranges during the treatment, similar to what was observed by Haddad (2009) in a study that used 500 and 1000ml of physiological saline with O3 for an intravenous application on healthy horses.

In a study by López (2007) that used O3 autohemotherapy in doses of 500 and 1000ml in horses, a significant increase in red blood cells was detected; this result is similar to observations in the present study. Therefore, it can be suggested that the results obtained in equines resemble human studies; that is, humans and horses respond to O3 treatment in a similar way in both autohemotherapy and rectal insufflation (Menéndez, 2008; Sousa, 2009).

Besides the increased number of red blood cells, the TG presented an increased hematocrit and hemoglobin concentration concomitant with the maintenance of total protein and albumin values. On other hand, CG animals had decreased hematocrit levels, MCVs, and consequent increase in MCHC; however, measured values were within the reference parameters according to Carvalho et al. (2016). By generating red blood cells with a greater amount of hemoglobin and the consequent capacity of oxygen transport throughout the treatment, we can infer that O3 contributes to improved oxygenation (Sagai and Bocci, 2011). Similar results were described by Bocci (1994) and Re (2008), who also observed improvements in the rheological properties of the blood accompanied by an increase in the red blood cell flexibility.

López (2007) and Penido (2010) affirmed that O3 increased immunological system activity, stimulating the white cells and increasing leukocyte activity to detect pathogens. In our study, the number of leukocytes did not show any alterations, but this could be because the animals were healthy and without challenges during the experimental period. Thus, there was no stimulation before or during the experiment to activate defense mechanisms or rebalance them. It has also been shown that the effects of O3 metabolites may lead to increased white cell activity, helping to strengthen the body's defenses (Silva et al., 2009), which was not observed in the present study by flow cytometry analysis.

The utilized rectal insufflation doses did not provoke detectable inflammatory processes by the laboratorial exams since the plasma values of fibrinogen and protein remained unchanged throughout the experimental period. It also did not affect the renal function of the horses as the urea and creatinine values remained within the reference range for the species (Neves et al., 2014). We can also infer that transrectal O3 insufflation did not interfere with the hepatic profile. The values of total and direct bilirubin had a non-significant increase in relation to the baseline values and remained within the reference range (Zobel et al., 2012). This effect occurs due the action of the aldehyde-albumin complex that increases heme-oxygenase-1, thus generating the release of carbon monoxide and bilirubin responsible for vasodilation and antioxidant activity (Bocci et al., 2011). It may also be caused by lysis of the old and rigid red blood cells that may occur during O3 therapy, resulting in improved rheological properties of the blood.

AST remained stable in a study by Ajamieh et al. (2002) who used rectal O3 insufflation to treat ischemia/reperfusion and in a study by Haddad et al. (2009) who used O3 autohemotherapy. These results corroborate with the results of the present study. Flow cytometry was used to analyze leukocyte behavior and activation index changes. According to Bocci (2011), O3 therapy affects the neutrophils by increasing their function, making them more efficient and easier to activate. In the present study, this phenomenon was not observed as there was no change in neutrophil activity; in contrast, we observed that the control group was more susceptible to challenges with PMA. Therefore, we can suppose that O3 acted as a modulator of these reactions.


Transrectal O3 application in equines is safe; it does not lead to harmful clinical and laboratory alterations. In addition, we can infer that transrectal O3 therapy can indirectly improve tissue oxygenation and metabolism as well as modulate ROS production.

Keywords: antioxidant; red blood cell counts; horse; ozone therapy

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