Mast cell tumors (MCTs) have been considered one of the most common tumors of the canine skin, comprising up to 20% of all cutaneous tumors in dogs (16). Trunks, perineal regions, and limbs are the most common locations for cutaneous MCTs (17). Surgical resection of the tumor is supposed to be an effective treatment option and complete tumor resection is the goal of the surgery (5,7). Surgical resection of the tumor is supposed to be an effective treatment option if possible, and complete tumor resection is the goal of the surgery (5,7) because inadequate surgical margins are related to a higher recurrence rate (17).

Intraoperative short-wave infrared (SWIR) imaging as image guided-surgery is an effective method for the detection of tumors and the evaluation of surgical margins (5). SWIR light (900-1700 nm) has many advantages, such as high tissue penetration with low autofluorescence of biological tissues, high signal-to-noise ratio (SNR), high signal contrast, and so on (10). Indocyanine green (ICG) is the first Food and Drug Administration approved fluorescent agent that responds to SWIR (8,9). Due to its safety for human use, ICG has been utilized to identify tumors and tumor margins and to locate residual tumor cells (9). In veterinary medicine, however, there are only a limited number of clinical reports about intraoperative ICG-guided resection of canine MCTs.

The following clinical case report demonstrates the application of ICG SWIR imaging method as fluorescence-guided surgery when identifying both the tumor and the surgical margin of a MCT located on the right carpus region in a dog.

A 6-year-old spayed, female Golden retriever dog, weighing 27 kg was referred for a skin mass on the dorsal region of the right carpus. The skin mass has been growing rapidly since it was discovered one month before the presentation by the owner. Fine-needle aspiration of the mass revealed clusters of well-differentiated mast cells suggestive of MCTs. Blood analysis including a chemistry profile and complete blood count, thoracic radiograph, abdominal ultrasonography, and Fine needle aspiration of popliteal lymph nodes were performed to evaluate oncologic stages. Neither specific evidence nor metastases were detected. Consequently, chemotherapy and surgical resection were recommended for curative intent because of the suspected MCTs.

Neoadjuvant chemotherapy was performed for debulking the mass size before tumor resection surgery by considering the location of the mass on the dorsal region of the distal right carpus. Prednisolone at a dosage of 1.0 mg/kg (0.45 mg/Ib) once daily and Toceranib at a dosage of 2.5 mg/kg once every other day was administered orally for two weeks. The volume of the mass was decreased from 2.6 * 3.5 * 1.7 (width * length * depth) cm to 1.8 * 2.1 * 1.2 cm. With the diminished volume of the mass, a SWIR fluorescence imaging system developed by MetabpleBIO Inc. and ICG as a fluorescence agent were exploited to determine the surgical margin.

ICG (25 mg ICG, Cellbiongreen, Cellbion Co., Ltd., Seoul, South Korea) was prepared by dissolving in 5 mL of sterile water (5 mg/mL of ICG). The solution was intravenously injected as a bolus at 5 mg/kg, followed by flushing with another 3 mL of saline water. The dog was hospitalized 24 hours before surgery for ICG injection and vital signs including systolic blood pressure, respiratory rate, heart rate, and capillary refilling time were carefully monitored after the injection. No significant change was observed during 24 hours and the surgery proceeded as planned.

The dog, diagnosed with MCT, was intravenously administered 2 mg/dog of chlorphenamine (Chlorpheniramine Maleate Inj., Huons) and 0.5 mg/kg of dexamethasone (Jeil Dexamethasone Inj., Jeil Pharm.) preoperatively. Under general anesthesia, induced with propofol intravenously (4 mg/kg; Provive, Myungmoon Pharm. Co.) and maintained with isoflurane (Terrell, Piramal Critical Care). SWIR image-guided surgery was performed. The 802.5-nm laser was exploited to excite ICG molecules. The output power of the laser can be controlled in a range from 0 to 5 W. The 808-nm band rejection filter with a bandwidth of -25 nm was implemented to suppress background noises. The exposing time and the gain value of the image sensor were controlled to improve the signal-to-noise ratio regarding the fluorescent images. The working distance in the fluorescence imaging system is defined as the distance between the laser output and the surgical site. The field of view at a working distance of -40 cm during surgery was measured by -15.1 cm². The optical intensity of laser output regarding a FOV of -15.1 cm during surgery was approximately estimated to be -14 mW/cm². During the operation, images of the surgical region (Fig. 1A) were obtained under ambient lights before surgical resection and the laser was turned on for fluorescence to evaluate the tumor and its margin (Fig. 2A). The activated ICG fluorescence of the MCT was visualized on the SWIR imaging system and the surgical margin was drawn around the fluorescence of the mass with a surgical marker. The tumor was resected using a 5 mm lateral margin and deep fascia as deep margins by dissecting fascial planes from the muscle (Fig. 1B). After the excision, the surgical site and normal tissues surrounding the removed tumor region were assessed again to evaluate the remaining fluorescent tissue by using the SWIR fluorescence imaging system (Fig. 2B). Residual fluorescent tissues were surgically excised. The removed tumor was immediately reimaged with the SWIR fluorescence imaging system ex vivo (Fig. 2C). The region of the resected mass was routinely closed by suturing with PDS (PDS II 3-0, Ethicon, USA) and Nylon (Blue nylon 4-0, Ailee Co., Korea) and the patient recovered successfully from the anesthesia.

Figure 1. Images of (A) a mast cell tumor on the right region of the carpus preoperatively, and (B) the surgical site after the resection of the tumor.

Figure 2. Representative brightfield of the tumor and surroundings (A) before resection and (B) after resection. The green dotted line indicates the tumor. The residual fluorescence tissues (the yellow dotted line) surrounding the tumor were additionally resected. (C) A brightfield image of the resected tumor (red arrow) and the wound bed after additional removal of residual fluorescence tissues.

The surgical ink was applied to indicate the margins of the resected tumors. Both the removed tumor and the additional resected tissues were submitted for histopathologic examination (IDEXX Laboratories, Inc., USA). Pathological evaluation of the lesion at the right carpus regions revealed the characteristics of low-grade cutaneous MCTs (Fig. 3), in which the mitotic count was less than 1 per 10 high power fields. The tumor excision was narrowly completed within a peripheral margin of less than 1 mm. The vascular and lymphatic invasions were not observed in this case. The additional resected tissues consisted of well-differentiated fibrovascular tissues in which residual neoplastic cells were not noticed.

Figure 3. Histopathology of a mass on the right carpus region shows the characteristics of a cutaneous mast cell tumor. (A) Image showing an unencapsulated and poorly circumscribed round cell neoplasm expanding the dermis and extending into the subcutis (H&E stain, original magnification ×10. Scale bar 100 μm). (B) Magnified image of neoplastic cells that have discrete individual margins and a moderate amount of cytoplasm which contains basophilic granules (H&E stain, original magnification ×40. Scale bar 50 μm).

The patient was discharged seven days postoperatively and no significant clinical symptoms were shown. Skin wound healing was well processed and skin sutures were removed 20 days after surgery. Since the MCTs after complete excision have low-grade characteristics without lymphatic or vascular invasions, adjuvant chemotherapy was not required. Up until the time of writing, 2 months after surgery, the dog had not developed any further complications or recurrence of the disease. Regular clinical follow-up will be done for evaluating the state of metastasis.

The approach to MCT resections in dogs generally involves a lateral margin of 2 cm and a depth of one fascial plane for the excision of all grade I and II cutaneous tumors (3). However, tumors located on limbs or in other anatomically challenging regions are not amenable to complete surgical resection due to limited skin availabilities or options for reconstruction (1,15). In certain anatomic sites, limb amputation might be the only practical way to achieve a radical margin (11). Wide and aggressive surgical resections of tumors in patients, which are located on the right carpus, seem to be challenging. The owner rejected the amputation and wished to preserve the limb function, even if the result of excisional margins is incomplete. Then, the mass after diagnosis with MCT was treated with neoadjuvant prednisolone and toceranib phosphate to reduce the tumor volume and complete resection margins were obtained (4).

In this case, SWIR ICG fluorescence-guided surgery was performed to identify both the tumor and its surgical margin, which can make a minimally invasive resection of the mass. In this patient’s case, the mass on the right carpus was initially imaged using the SWIR fluorescence imaging system and the edge of the tumor was incised based on the fluorescence margin assessment. The resection margin was successfully reduced to 5 mm lateral to the tumor, which was much smaller than that based on the standard MCT (- laterally 2-3 cm) (3) and the histopathological result showed the complete margin. The use of the SWIR fluorescence imaging method during surgery can successfully eliminate unnecessary resection of normal tissues. This would be crucial to the surgical removal of tumors in anatomically challenging locations, such as on limbs or heads.

The results in this case report should be considered with some limitations. It is reported that one of the contributions of the SWIR fluorescence imaging method is investing residual cancer cells after the resection of wound beds (5,12). The surgical site and the removed tumor were imaged again with the SWIR fluorescence system in this case and the removed tumor region showed some residual fluorescent tissues. On the other hand, additional resected tissue surrounding the wound bed didn’t include neoplastic cells according to its histopathological result. Some studies reported that the SWIR imaging method shows false positive fluorescence from inflammatory tissues (8). This is due to clinical limitations of ICG drug delivery to tissues with leaky capillaries (2). Once ICG is injected into the blood, it quickly binds to proteins like albumin and these complexes particularly accumulate in tumor tissue (13). However, ICG is not a receptor-specific agent, it can be collected in others like inflammatory cells than tumor tissues (6,14). The resection of inflammatory tissues is often likely to be required in many cases because the remaining tissues have lost their functions and might continue to harm the patient (8). Therefore, residual fluorescent tissues, in this case, were additionally removed after the resection of the mast cell tumor.

The SWIR fluorescent imaging method incorporating ICG can be feasible for the identification of tumors and the evaluation of the surgical margins for complete resection of canine cutaneous mast cell tumors. The use of SWIR fluorescence imaging system during surgery can remove unnecessary excision of normal tissues, which would be essential for surgically removing tumors on limbs or other anatomically challenging locations. Further clinical studies are required to assess more applications of SWIR ICG imaging methods in veterinary medicine.

This work was supported by the National Research Foundation (NRF) grant funded by the Korea government (MSIT; No. 2021R1A2C1012058) and the Global Research and Development Center (GRDC) Program through NRF of Korea funded by the Ministry.

The authors have no conflicting interests.