Calcinosis Cutis with Large Extension and Uncommon Location in a Dog

Background: Calcinosis cutis is an uncommon dermatopathy characterized by the deposition of minerals in the skin, usually involving collagen and elastic fibers in the dermis. Usually, it results from dystrophic calcification and can be generalized or focal. The dermatopathy may be primary or secondary to certain disorders, especially chronic proliferative otitis, foreign body reactions, hyperadrenocorticism (HAC) and less frequently percutaneous penetration of calcium-rich products. The aim of this report is to describe a presentation of calcinosis cutis affecting the skin of the back, internal face of hind limbs and anal region of a 9-years-old bitch. Case: A 9-year-old, non-defined breed, bitch, ovariohysterectomized, weighing 9.45 kg, was attended at the Dermatological Service of companion animals at the Veterinary Hospital of the Federal University of Jataí (UFJ). The animal came in with the complaint of extensive dorsal alopecia, covered by firm lesions, with a 3-month evolution, additionally to polyuria and polydipsia. After physical examination, alopecic areas of great extension were confirmed on the dorsum, on the internal surface of the hind limbs and in the anal region. Also, an exudative and painful lesion located on the back was detected, plus loss of elasticity of the ventral abdomen skin and visible abdominal vessels. The screening tests showed a marked increase in the alanine aminotransferase enzyme (ALT), alkaline phosphatase (ALP) and total cholesterol. The specific urinary density was decreased. On the ultrasound examination, hepatomegaly and an increase in the caudal pole of the left adrenal were detected. Based on these findings, calcinosis cutis secondary to spontaneous hyperadrenocorticism (HAC) was suspected. For confirmation, skin biopsy and low dose dexamethasone suppression test (LDDS) were performed. LDDS test showed no reduction of serum cortisol after 8 h of dexamethasone dose administration and histopathological evaluation revealed multiple foci of calcinosis characterized by the deposition of basophilic material on the pre-existing collagen fibers, plus areas with pyogranulomatous inflammatory reaction and peripheral fibrosis with transepidermal elimination of minerals. Thus, trilostane and intense hydration of skin plaques were applied as treatment. Discussion: The dermatological alterations were compatible with those described in the consulted literature, with remarkable yellow-brown, firm, sandy-looking plaques, located on the back, internal face of hind limbs and anal region, possibly related to HAC later confirmed by LDDS test and biopsy. The management of the underlying disease and possible secondary bacterial infections are the basis of the treatment. Therefore, the patient was treated with trilostane, antibiotic therapy and intensive hydration of the mineralized plaques resulting in a satisfactory involution of the clinical signs. Even though there are reports of calcinosis cutis on the dorsum, in the consulted literature there was no evidence of dorsum large extension lesion due to HAC as in this case report, but secondary to exogenous corticosteroid treatment, systemic blastomycosis and leptospirosis. In this case report, the affected thorax portion was the dorsum, differently from a study that pointed the ventral thorax as the affected portion. Similarly, anus and ventral part of the tail were hardly affected together with secondary inflammation and ulceration. Thus, the existent literature shows areas of calcinosis cutis in dogs in different parts of the body, but neither extensive as in the back of this reported female dog, nor widely affected as in the anal area, additionally to the internal face of hind limbs as already reported in the literature.


INTRODUCTION
Calcinosis cutis is an uncommon dermatological alteration in dogs, in which insoluble mineral salts, such as calcium and phosphate are deposited in the dermis, subcutaneously and, more rarely, epidermis [7]. Mineral precipitation commonly affects collagen and elastic fibers in the dermis [13]. The occurrence generally results from dystrophic calcification and can occur in a generalized or localized manner [2]. The alteration may be primary or secondary to certain disorders, especially chronic proliferative otitis, foreign body reactions and hyperadrenocorticism (HAC) [1]. Less frequently, calcinosis cutis has been associated with percutaneous penetration of calcium-rich products [11] or as secondary to severe systemic diseases [2,6,10].
In dogs, dystrophic calcification usually occurs with hyperglucocorticoidism as calcinosis cutis, or as a result of focal trauma as calcinosis circumscripta [3]. Calcinosis cutis due to primary HAC has a high correlation with the average age of diagnosed patients, which varies from 9 to 11 years [1]. On the other hand, no sex predilections have been reported [3]. Clinically, calcinosis characterizes an isolated to multicentric, firm, erythematous and painful papules, nodules or plaques which can become ulcerated or crusted [9]. Skin points susceptible to repetitive flexure are more likely to the occurrence of calcinosis cutis, such as flexural surfaces of the groin, back of the neck and axillae [3].
The aim of this report is to describe a case of calcinosis cutis in a 9-years-old bitch with coalescing erythematous papules affecting the skin of the back, the internal face of hind limbs and anal region.

CASE
A 9-year-old, non-defined breed bitch, ovariohysterectomized, weighing 9.45 kg, was attended at the Dermatological Service of companion animals at the Veterinary Hospital of the Federal University of Jataí (UFJ). The main complaint was the development of dorsal alopecia (Figure 1), covered by firm lesions, with a three-month evolution and the recent appearance of wet and painful lesions in the region. When asked, the pet owner reported that in the last few months the patient had lost weight, despite having her appetite preserved. The owner also mentioned that the animal had polyuria and polydipsia. According to him, no medication had been administered.
On the physical examination, there was evident muscular atrophy, especially in the temporal region and on the spine. In the dermatological inspection, alopecic areas of great extension were observed on the dorsum, on the internal surface of the hind limbs and in the anal area. In these areas, there were erythematous plaques with mineral deposition (Figure 2). The fur adjacent to these areas was opaque, dry and easily removable. In addition, an exudative and painful lesion located on the back was detected during examination. The skin of the ventral abdomen was thin, with loss of elasticity and visible abdominal vessels.
Among the laboratory changes detected in the screening tests, there was a marked increase in the alanine aminotransferase enzyme (450 IU/L) and alkaline phosphatase (2,800 IU/L), in addition to increased total cholesterol (350 mg/dL). Triglycerides and HDL cholesterol were within normal values. The urinary density resulted in 1,011. On the ultrasound examination, hepatomegaly and an increase in the caudal pole of the left adrenal (0.83 cm) were detected.
As for the complementary dermatological evaluation, both parasitological tests such as, skin scraping and mycological tests were negative. In the cytological evaluation of the exudative lesion, squamous epithelial cells and a moderate presence of leukocytes infiltrate (predominantly neutrophils) and coccus bacteria were observed, indicating secondary bacterial infection of the plaques.
In view of the clinical and laboratory findings, the suspected diagnosis was calcinosis cutis secondary to spontaneous hyperadrenocorticism (HAC). For confirmation, skin biopsy and low dose dexamethasone suppression test -LDDS (SNAP® Cortisol Test) 1 were performed, with cortisol measured by the radioimmunoassay technique (RIA), as recommended by Herrtage & Ramsey [4].
Two wedge specimens of skin were collected from better-preserved lesions through punch biopsy 2 (4 mm) and fixed in 10% buffered-formaldehyde solution. Four micrometers cuts were performed using a mechanical rotating microtome (RM 2245) 3 , arranged in histological slides, and stained with hematoxylin 4 and eosin 5 . The slides were evaluated in light microscopy (O400S opticam) 6 . Grossly, the samples showed irregular, firm, yellowish-tan, gritty material. Microscopically, there were dermal deposition of multifocal, basophilicintense material (mineral), sometimes associated with In order to perform the LDDS test, a blood sample was collected and centrifuged to obtain 1.0 mL of serum in which basal cortisol was measured, resulting in 6.79 µg/dL (reference interval: 1.0 to 4.6 µg/dL). Then, 0.01 mg/kg of dexamethasone (Decadron ® ) 7 was intravenously administered. After 8 h, a new dosage of cortisol levels was carried out, resulting in 3.64 µg/ dL (reference interval: < 0.90 µg/dL). The LDDS test confirmed HAC because it did not reveal suppression of cortisol 8 h after the application of the glucocorticoid.
After confirmation of the diagnosis of HAC, treatment with trilostane (Vetoryl ® ) 8 was applied at a dosage of 0.5 mg/kg in each 12 h, with dose adjustments every 21 days in order to minimize the chances of side effects. Amoxicillin with clavulanic acid (Agemoxi CL ® ) 9 at a dose of 25 mg/kg in each 12 h for 21 days was used to control secondary bacterial infection. Also, topic treatment with moisturizing baths and antiseptic spray twice a week were considered. The antiseptic spray was manipulated with chlorhexidine pyogranulomatous inflammation and perilesional fibrosis. Furthermore, epidermis reveals transepidermal elimination of mineral and mild pyogranulomatous inflammatory reaction. Together, the findings led to the final diagnosis of calcinosis cutis (Figure 3). 4%, hydroviton 4% and aqueous solution 10 . Plus, the application of oily lotion (Dersani ® ) 11 in every 12 h composed of essential fatty acids, medium chain triglycerides and vitamins A and E. Two weeks after the onset of the treatment, there was a clinical resolution of the secondary infection, in addition to an initial improvement in calcinosis cutis and signs of HAC.

DISCUSSION
Calcinosis cutis is an uncommon dermatological alteration, usually secondary to other disorders [1,7]. Considering the correlation with HAC, especially in animals between 9 and 11 years old, the patient in this report fits in, reinforcing the age target when it comes to calcinosis cutis [1,8,14]. In this case report, calcinosis cutis reached large proportions and was accompanied by clinical signs suggestive of endocrinopathy, such as alopecia, muscular atrophy, thin skin, polyuria and polydipsia. The dermatological alterations were compatible with those described in the consulted literature, with remarkable yellow-brown, firm, sandy--looking plaques, located on the back. The patient also developed well marked inflammatory plaques in the perineal region and internal face of hind limbs, as a possible syndrome related to HAC [5].
The screening tests reinforced the suspicion of HAC, which could be confirmed by the the technique recommended by Herrtage & Ramsey [4]. To perform skin biopsy, the areas of preference were those calcified without concurrent occurrence of trauma or ulceration, following recommendations [3]. The macroscopic and microscopic characteristics were compatible with those described by the mentioned authors, thus confirming the diagnosis of calcinosis cutis.
Microscopically, the differential diagnosis is simple if mineralization is obvious in tissue as it was in the present case [3]. In early lesions, when mineralization is not evident, Von Kossa staining may aid to the diagnosis as the presence of fractured collagens fibers. Calcinosis circumscripta is characterized by large focal or multifocal masses, which is highly different from calcinosis cutis pattern [1]. Another type of mineralization, as metastatic, could be easily differentiated from calcinosis cutis, since the first was secondary to elevated calcium levels in the blood, associated with chronic kidney disease and is manifested as calcinosis circumscripta [3,10].
The management of the underlying disease and possible secondary bacterial infections are the basis of treatment [12]. Thus, the patient was treated with trilostane, antibiotic therapy and intensive hydration of the mineralized plaques resulting in a satisfactory involution of the clinical signs.
Even though, there are reports of calcinosis cutis on the dorsum, in the consulted literature there was no evidence of dorsum large extension lesion due to HAC, but secondary to exogenous corticosteroid treatment, systemic blastomycosis and leptospirosis [2,7,10]. Lesions are usually described as multiple lesions distributed in areas such as flank, axilla, ventral thorax, extremities, inguinal region, head and perigenital region [1]. In this case report, the thorax portion affected was the dorsum, differently from that related in the aforementioned study. Similarly, anus and ventral part of the tail were affected together with secondary inflammation and ulceration as previously shown.