Thermography is a method of imaging using an infrared radiation detection sensor to measure radiation emitted from a surface. After the acquisition, such images are organized as a distribution diagram with temperature information [15] so it is a non-invasive method. High sensitivity [8; 16; 17] is reported for such method and it allows the registration of the trophic conditions of the tissues, in areas with increased tissue metabolism or with an inflammatory response [18; 19]. By this method, the temperature is represented graphically (thermogram), with different colors for each temperature interval [7]. Each pixel in the thermogram represents a measured temperature of the surface of an object. Variations in the color pattern indicate thermal differences due to changes in surface temperature, which can be quantified by heat transfer principles [20; 21].
In the present study, using the thermographic camera, it was possible to observe an increase in tissue temperature in 24 hours and a further slow decrease until 21 days. From T7 onwards, the temperature values, in all groups, returned to baseline values (T0). To verify if detected modifications in temperature occurred simultaneously with other inflammatory signals, the paw's thickness and volume were also evaluated. A similar increase at 24 hours observed in the temperature through the thermography method, also were noticed in the thickness and volume paws. Such finding is stimulating since, for this specific type of inflammation model, this biological behavior is expected (the 24 hours peak).
The infrared technique has also been shown to be useful to differentiate the effectiveness of treatments with anti-inflammatory drugs with different mechanisms of action. Considering the animal’s paw thickness, differences were demonstrated between the initial time and all subsequent experimental times, in all groups. Considering data from the paw volume analysis for animals treated with topic diclofenac there was no difference at 30 minutes or 21 and 28 days, which demonstrated that in this group and times volume changes reached values similar to the baseline. This result could suggest that, for this group, the diclofenac topic treatment was more effective in volume change than in thickness. The formation of granuloma, as observed in the present study, could explain why the volume of the animals’ paws did not return to the initial values with exception of the animals treated with topic diclofenac.
The graphics curves demonstrated that temperature behavior followed edema (thickness and volume) behavior with an increase at 24 hours followed by a decrease reaching values similar to those of the baseline in a shorter experimental time, compared to volume and thickness parameters. A hypothesis considered for these outcomes is that temperature decrease could be solved faster in the inflammatory response than edema, however other studies must be performed with different animal models of inflammation.
The induction of chronic inflammation in rodents was achieved with injection of a suspension of inactive strains of Mycobacterium tuberculosis in Freund's adjuvant and it is expected a larger sensibilization period by the presence of non-metabolizable oils, such as paraffin that promotes the continuous release of antigens. With this, chronic inflammation is trigged inducing a strong and persistent inflammatory response that could achieve 35 days of duration [1; 22-24]. Some musculoskeletal disorders, related to chronic inflammation lack objective diagnostic and gold standards, then it is a challenge to effectively validate the present technique.
In histological sections, it was possible to qualitatively determine the presence of cellular infiltrate, consistent with chronic inflammation. Leukocyte differential counting informed the relative amount of different leukocyte types in blood cells (neutrophils, lymphocytes, basophils, eosinophils, and monocytes) according to their morphological characteristics. There is no reduction in the percentage of lymphocytes in the blood of groups that received the CFA injection and were treated with triamcinolone (26,6%) and diclofenac (21,1%) when compared to animals in the control group (33%), which could be related to the anti-inflammatory effect of the drugs used.
Inflammation is the process of recruitment of leukocytes and plasma proteins from the blood, their accumulation in tissues, and their activation to destroy the microorganisms. Many of these reactions involve cytokines, which are produced by dendritic cells, macrophages, and other types of cells during innate immune reactions. The major leukocytes that are recruited in inflammation are phagocytes neutrophils (which have a short life span in tissues) and monocytes (which develop into tissue macrophages) [12]. Therefore, a possible cause for the reduction in the percentage of lymphocytes in the blood cell counting could be due to their accumulation in tissues. In the present study, we also noticed a reduction, although not significant, of blood neutrophils in the control group (without treatment). However, we cannot say that these neutrophils would be more concentrated in the inflamed tissue since the characterization of the tissue cells was not carried out.
Drugs used were selected since topical treatments for inflammation disorders are frequently well-tolerated and preferred by many patients [25]. For these reasons, a topical corticosteroid was used. Another anti-inflammatory drug was used due to the current evidence that indicates that topical non-steroidal anti-inflammatory drugs may be effective for pain relief in osteoarthritis [26]. Diclofenac sodium is a potent inhibitor of cyclooxygenase-2 with analgesic and anti-inflammatory properties; however, it has little antipyretic action. It is recommended for the treatment of chronic inflammatory conditions such as rheumatoid arthritis and osteoarthritis [27]. Triamcinolone acetonide is a synthetic corticosteroid that has anti-inflammatory, antipruritic, and antiallergic action [27]. Components of the formula act as an adhesive vehicle to the active medication [28].
Our results suggest that thermography may also be useful to differentiate the anti-inflammatory efficacy of different drugs. When compared to the diclofenac sodium animal group (96 hours), the animals treated with triamcinolone acetonide returned faster (48 hours) to the initial temperature values. The pharmacology of triamcinolone as a corticosteroid drug could explain anti-inflammatory effects and also its vehicle since adhesive vehicles could improve drug substantively by prolonging the supply of the drug in the site as a result of the ability to adhere to the substrate and persist at effective drug concentration [29].
The right paws temperatures (injected with saline solution) were not different, as expected, since they are regions that did not receive pro-inflammatory stimulation.
The temperature of the extremities and skin depends on the blood flow dynamics and temperature. Additionally, individual variations at different times of the day can occur [19]. For this reason, all images were recorded at the same time, early in the morning in a controlled environment to prevent such aspects.
Thermography does not provide specific details of disease however it may be useful in defining the area affected by inflammation and also can assist the progression of the lesion. The temperature patterns can be associated with healthy or pathological situations [30].