CJVR - January 2022, Vol 86, No. 1



Modulation of mTOR signaling by radiation and rapamycin treatment in canine mast cell cancer cells

Morla Phan, Changseok Kim, Anthony Mutsaers, Valerie Poirier, Brenda Coomber (page 3)

Rapamycin has been reported to reduce cancer cell survival in certain tumors following radiation therapy, but the mechanisms driving this phenomenon are unclear. Rapamycin inhibits mTOR signaling, a pathway responsible for several essential cell functions. The objective of this study was to investigate the effects of rapamycin and radiation on the activation and inhibition of mTOR signaling and the relationship between mTOR signaling and DNA damage response in vitro using canine mast cell tumor (MCT) cancer cell lines. Rapamycin rapidly inhibited S6K phosphorylation in a dose-dependent manner. Ionizing radiation (3, 6, or 10 Gy) was able to activate mTOR signalling, but the combination of radiation and rapamycin maintained mTOR inhibition. The comet assay revealed that co-treatment with rapamycin induced modest increases in the severity of DNA damage to MCT cells, but that these differences were not statistically significant. Although the relationship between mTOR and DNA damage response in MCT cancer cell lines remains unclear, our findings suggest the possibility of interaction, leading to enhancement of radiation response.

Identification of the interleukin-8 (CXCL-8) pathway in feline oral squamous cell carcinoma — A pilot study

Leah H. Ackerman, Carlos H. de Mello Souza, Galaxia Cortés-Hinojosa, Marc E. Salute, Alexa A. Stephen, Elizabeth Anthony, Keijiro Shiomitsu, Rowan J. Milner (page 13)

The purpose of this pilot study was to detect the presence of interleukin-8 (IL-8) and the potential downstream effects of IL-8 receptor activation in 2 previously characterized feline oral squamous cell carcinoma cell lines (SCCF1 and SCCF2). Interleukin-8 messenger RNA (mRNA) was initially detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). A previously validated and commercially available enzyme-linked immunosorbent assay (ELISA) test was used to measure IL-8 production in the supernatant of the 2 cell lines. Western blot was used to detect phosphorylation of proteins (AKT, ERK1/2, JAK2, STAT3, and Src), known to be downstream of interleukin-8 receptor activation. The IL-8 receptor-specific antagonists, Reparixin and SCH527123, were used to identify effects on phosphorylation of these proteins. Interleukin-8 mRNA and protein were detected in both SCCF1 and SCCF2 by RT-PCR and ELISA, respectively. Phosphorylation of ERK1/2, STAT3, and Src was detected in both cell lines. Inhibition of the IL-8 receptor led to a decrease in phosphorylation of Src, but not ERK1/2 or STAT3. In conclusion, feline squamous cell carcinoma cell lines can produce IL-8. Phosphorylation of Src seems, at least in part, a consequence of IL-8 receptor activation. The phosphorylation of ERK1/2 and STAT3, although present, seems independent of IL-8 receptor activation. Due to its potential effects on the tumor microenvironment, in addition to its autocrine effects on Src phosphorylation, the inhibition of the IL-8 receptor may become a beneficial therapeutic tool. Evaluation of the presence of both IL-8 and Src in many cases should elucidate their importance.

Comparison of 2 blind approaches to the paravertebral brachial plexus regional block in canine cadavers

Maria Podsiedlik, Erik H. Hofmeister, Tanya Duke-Novakovski (page 20)

The paravertebral brachial plexus block (PVB) provides thoracic limb analgesia. The objective was to describe a blind craniocaudal (CC) approach to the PVB and compare its accuracy, time, and difficulty of performance with a blind dorsoventral (DV) approach. The operator was initially trained by experienced clinicians to perform both approaches on 5 cadavers. Next, a CC or DV approach to the PVB was performed on both thoracic limbs of 20 cadavers (20 for each approach). Methylene blue dye was equally divided into 4 aliquots to stain the ventral branches of the sixth to eighth cervical and first thoracic spinal nerves. Successfully stained (stain ≥ 1 cm) spinal nerves were counted. The time to perform each approach was recorded and ease of performance was scored using a numerical scale (1 “easy” to 4 “difficult”). The phrenic nerve was checked for stain. A Wilcoxon signed-rank test was used to compare approaches. The data are presented as median (interquartile range; minimum to maximum range). The number of stained nerves with the CC approach 3 (1; 2 to 4), was higher than the DV approach 2 (2; 0 to 4) (P = 0.002). The time (in seconds) to perform the CC approach 125 (79; 70 to 194), was not different from the DV approach 142 (54; 101 to 232) (P = 0.084). The CC approach 2 (2; 1 to 4) was easier to perform than the DV approach 3 (1; 2 to 4) (P = 0.024). No phrenic nerve staining was observed with either approach. The CC approach is an alternative to the DV approach for performing the PVB in dogs.

Computed tomography features of presumed normal mandibular and medial retropharyngeal lymph nodes in dogs

Alexandra F. Belotta, Sally Sukut, Candace Lowe, Cheryl Waldner, Elissa K. Randall, Valerie S. MacDonald, Jerome Gagnon, Monique N. Mayer (page 27)

Computed tomographic (CT) features of lymph nodes are used for diagnosis, staging, and response evaluation in veterinary patients. The objectives of this study were to describe maximum short- and long-axis transverse diameters (SATD and LATD, respectively), volume, and attenuation of presumed normal canine mandibular lymph nodes (MLNs) and medial retropharyngeal lymph nodes (MRLNs), to investigate the effect of variables on these features, and to describe inter-observer reliability of transverse diameter and attenuation measurement. Computed tomography studies of 161 dogs were retrospectively reviewed. Median values for SATD were 5.2 and 5.4 mm, median values for volume were 0.26 and 0.54 cm3, and median values for attenuation were 32.3 and 31.8 Hounsfield units (HU) for MLN and MRLN, respectively. Median LATD was 11 mm for MLNs. Height of the first cervical vertebra (HCV1) and weight were positively associated with transverse diameters of both MLNs and MRLNs (P < 0.001). Grade of dental disease was negatively associated with transverse diameters of both MLNs (P = 0.001) and MRLNs (P < 0.03). The intraclass correlation coefficient (ICC) was 0.94 [95% confidence interval (CI): 0.94 to 0.95] for transverse diameter measurement. This study provides descriptive data on the CT characteristics of presumed normal MLNs and MRLNs in dogs and describes a highly reliable method of measuring transverse diameter.

Effects of the number and the position of transfixation pins in acrylic connecting bars

Claire Leriquier, Florence Audet-Robin, Guy Beauchamp, Xavier Montasell (page 35)

Acrylic columns are commonly used in external skeletal fixators, especially for fracture management or trans-articular fixations. To the authors’ knowledge, there are no studies demonstrating if the number or position of the transfixation pins influence the ultimate strength and stiffness of the acrylic column. The objective of this study was to evaluate the effects of the number and position of transfixation pins (concentric versus eccentric) on the strength and stiffness of acrylic columns placed in axial compression. We hypothesized that strength and stiffness of acrylic columns under axial compression would not be affected by the number or position of the transfixation pins through the column. Three different groups of 12 acrylic columns were constructed with 4, 6, and 8 pins. In each group, 6 columns were constructed with the pins placed concentrically and the remaining 6 columns with the pins placed eccentrically. Each column was then placed under axial compression using a biomechanical testing machine. No significant differences were observed in ultimate strength regarding the number or position of transfixation pins (P = 0.83 and P = 0.27, respectively). However, stiffness was significantly decreased for columns with 4 eccentric pins compared with columns with 6 and 8 eccentric pins (P < 0.01) and with columns with 4 concentric pins (P < 0.001). Although the effects of transfixation pins on the rigidity of acrylic columns do not appear to be clinically significant, these tests were performed only in compression and results might differ if complete external fixator systems are used with different models of testing. Future studies are recommended.

Serum C-reactive protein and iron levels following gonadectomy are not modified by perioperative administration of robenacoxib to dogs

Verónica Vieitez, Victor López-Rámis, Rafael Barrera, Ignacio A. Gómez de Segura (page 40)

The aim of this study was to evaluate the perioperative effects of robenacoxib on serum C-reactive protein (CRP) and iron concentrations in dogs undergoing gonadectomy. In a prospective, blinded, controlled clinical trial, 60 healthy dogs were randomly assigned to receive preoperative subcutaneous injection of either robenacoxib [2 mg/kg body weight (BW)], meloxicam (0.2 mg/kg BW), or saline (0.04 mL/kg BW), followed by oral administration over 72 h (robenacoxib: 2 to 4 mg/kg BW; meloxicam: 0.1 mg/kg BW; saline: gelatin capsules). Blood samples were taken before surgery and 12, 24, 48, 72 h, and 7 d after surgery. Pain scores were assessed via the short-form Glasgow Composite Pain Scale over 72 h postoperatively. C-reactive protein (CRP) and iron serum levels increased and decreased (P < 0.01, both), respectively, after surgery and returned to baseline within 1 wk. No differences were observed among treatments (P > 0.05) or based on surgery/gender (P > 0.05). Pain assessment revealed a higher incidence of treatment failure in saline (6 females versus 2 and 1 female in robenacoxib and meloxicam, respectively). In conclusion, robenacoxib and meloxicam had no influence on postoperative CRP or iron in dogs, which suggests that these nonsteroidal anti-inflammatory drugs (NSAIDs) do not have a relevant effect on these biomarkers.

Hematology of dairy cows with purulent-necrotic processes in the digital area treated with Subtilin ointment

Valiyan Gimranov, Evgeny Skovorodin, Ilshat Giniyatullin, Azamat Mirzin, Oleg Dyudbin (page 48)

The objective of this study was to investigate the effect of Subtilin ointment on the hematology of dairy cows when used to treat inflammation and necrotic tissue in the digital area. Holstein-Friesian cows were allocated to 2 groups of 10 animals each. The first group consisted of clinically healthy cows and the second group consisted of cows with a purulent-necrotic lesion in the digital area. To make a comparative analysis, blood samples of healthy and diseased cows were taken and tested before treatment with Subtilin ointment and 10 days after treatment. The blood of the diseased cows had more leukocytes (53.38%), lymphocytes (46.81%), erythrocytes (15.78%), granulocytes (64.56%), and platelets (34.76%), and a higher mean cell volume (of 8.37%) and increased level of monocytes/eosinophils (65.12%) than did the blood of clinically healthy cows. At the same time, there were no evident changes in the level of erythrocytes, hemoglobin, mean cell hemoglobin, mean cell hemoglobin concentration, and percentage of lymphocytes in either clinically healthy or diseased cows. The hematology of diseased cows treated with Subtilin ointment demonstrated positive dynamics in the healing of ulcerative processes.

Evaluation of bexagliflozin in cats with poorly regulated diabetes mellitus

Suzanne L. Benedict, Orla M. Mahony, Talon S. McKee, Philip J. Bergman (page 52)

The aim of this study was to investigate the effect of bexagliflozin on glycemic control in poorly regulated diabetic cats and to evaluate for adverse events associated with this medication.

Sodium-glucose cotransporter 2 inhibitors are a newer class of drugs used in the management of humans with type 2 diabetes mellitus. The objective of this study was to evaluate the effect of the orally administered drug, bexagliflozin in a group of poorly regulated diabetic cats over a 4-week study period. Five client-owned cats with poorly controlled diabetes mellitus receiving insulin therapy were enrolled. Bexagliflozin was administered once daily. Serum fructosamine, serum biochemistry profile, and 10-hour blood glucose curves were assessed at baseline (Day 0), Day 14, and Day 28. All cats had a significant reduction in insulin dose requirement (P = 0.015) and insulin was discontinued in 2 cats. There was a significant decrease in blood glucose concentration obtained from blood glucose concentration curves during the study period (P = 0.022). Serum fructosamine decreased in 4 of the 5 cats with a median decrease of 152 µmol/L (range: 103 to 241 µmol/L), which was not statistically significant (P = 0.117). No cats had any documented episodes of hypoglycemia. Adverse effects were mild. The addition of bexagliflozin significantly improved diabetic management in this group of cats.

In vitro antimicrobial activity of equine platelet lysate and mesenchymal stromal cells against common clinical pathogens

Haileigh K. Avellar, John D. Lutter, Charan K. Ganta, Warren Beard, Joseph R. Smith, Naveen Jonnalagadda, Sarah Peloquin, Qing Kang, Kanwal Ayub (page 59)

Septic arthritis is considered a medical emergency. Disease following bacterial colonization can lead to significant morbidity and mortality and requires costly treatment. Antimicrobial properties of regenerative therapies, including mesenchymal stromal cells and platelet products, have been researched extensively in human medicine. Although fewer studies have been conducted in veterinary species, they have shown promising results. The purpose of this study was to evaluate bacterial suppression by equine platelet lysate (EPL) and adipose-derived mesenchymal stromal cells (ASCs) in vitro. We hypothesized that both products would significantly inhibit the growth of Staphylococcus aureus and Escherichia coli. Pooled blood from 10 horses was used for production of EPL. Mesenchymal stromal cells were isolated from adipose tissue harvested from the gluteal region of 3 horses. The study evaluated 3 treatment groups: 10 × EPL, 1.6 million ASCs, and a control, using an incomplete unbalanced block design with repeated measurements. Optical density readings and colony-forming units/mL were calculated at 0, 3, 6, 9, 12, 18, and 24 hours. Decreased bacterial growth was seen at multiple time points for the S. aureus-ASC and S. aureus-EPL treatments, supporting our hypothesis. Increased bacterial growth was noticed in the E. coli-EPL group, with no difference in the E. coli-ASC treatment, which opposed our hypothesis. A clear conclusion of antimicrobial effects of EPL and ASCs cannot be made from this in vitro study. Although it appears that ASCs have a significant effect on decreasing the growth of S. aureus, further studies are needed to explore these effects, particularly in Gram-positive bacteria.

Histological and histochemical characteristics of lacrimal glands in beluga whales (Delphinapterus leucas)

Bactelius Turicea, Chantale L. Pinard, Emily R. Brouwer, Stéphane Lair, Geoffrey A. Wood, Marie-Odile Benoit-Biancamano (page 65)

The objective of this study was to describe the histological and histochemical characteristics of the lacrimal glands of beluga whales. The study was carried out on the formalin-fixed ocular globes from 96 carcasses of beluga whales found stranded in the St. Lawrence estuary in Quebec, Canada. Hematoxylin and eosin (H&E) stained slides from the eyes of each whale were examined for lacrimal glands. Histological description was done with H&E and Masson Trichrome (MT) stains. Period Acid-Schiff (PAS), Alcian blue (AB) pH 1.0 and 2.5, and High Iron Diamine (HID) stains were used for histochemical characterization of glycoproteins. Thirteen ocular samples from animals ranging from neonate to 48 y included sections of lacrimal glands. The H&E stain revealed a tubuloalveolar gland architecture, separated into lobules by dense connective tissue. Each lobule contained a mixture of acini and tubules with ductules. Small and large acini were composed of low and tall columnar cells, respectively. Acinar cells contained basophilic cytoplasmic granules. The ductules were lined with a bi-layered cuboidal-to-squamous epithelium. The MT stain highlighted the connective tissue separating ductules and acini. Large acini were positive for PAS and some small acini had patchy uptake. Positive staining for AB pH 1.0 and 2.5 was mainly seen in tall columnar cells as compared to small acini that had faint to no stain uptake. High Iron Diamine stain revealed 90% staining of all acinar cells, with 10% exhibiting a mixed blue-black tinge. It was concluded that the lacrimal glands of beluga whales have similar histological and histochemical findings to those of artiodactyla and carnivora orders.

Intrauterine iodine administration is superior to antibiotics for reconception after late-embryonic losses in dairy cows

Mehmet Cengiz, Sarper Ozbek, Muhammad Furqan Asghar Chacher (page 74)

This field study was conducted to compare conception and insemination efficiency responses to intrauterine polyvinylpyrrolidone-iodine (PVP-I) and antibiotic (AB) treatments in dairy cows that experienced pregnancy loss. Data were collected from lactation cows with a history of pregnancy loss 27 to 70 days post-insemination (n = 97) during the 1st to 3rd lactation (days in milk = 196 ± 28). Cows were subjected to 1 of 3 treatments: i) 50 mL saline solution intrauterine infusion (S; n = 23); ii) 2% PVP-I (n = 42); or iii) 150 mg of amoxicillin trihydrate and 40 mg/mL gentamicin sulphate (n = 32). All cows followed the progesterone-based fixed-time insemination protocol. Data were analyzed by Chi-square test and 1-way analysis of variance. The PVP-I treatment (n = 25; 59.5%) was as effective as the AB treatment (n = 19; 59.4%), compared to the S treatment (n = 5; 21.7%) to achieve a new conception. The cows in group PVP-I conceived in a shorter time than those in group AB (46.0 ± 8.7 days versus 105.0 ± 10.0 days; P < 0.05) with a more efficient insemination to conception ratio (2.32 ± 0.43 versus 4.10 ± 0.32; P < 0.05). Data suggest that intrauterine PVP-I administration is superior to intrauterine AB administration in rescheduling reproductive protocol upon late embryonic and fetal losses.

Brief Communication

Distribution and characterization of Streptococcus suis serotypes isolated from January 2015 to June 2020 from diseased pigs in Québec, Canada

Sonia Lacouture, Yaindrys Rodriguez Olivera, Segura Mariela, Marcelo Gottschalk (page 78)

Streptococcus suis is one of the most important swine bacterial pathogens causing economic losses. This report presents the serotype distribution of S. suis recovered from diseased pigs in Québec from January 2015 to June 2020. Serotypes 1/2 and 2 predominated, followed by serotypes 7, 3, 5, 4, 9, 1, and 14. Compared to previously reported data, very few changes could be observed concerning the serotype distribution, indicating a relative stability. Half of the untypable isolates did not belong to the species S. suis sensu stricto, as determined by recN polymerase chain reaction. Less than 10% of “real S. suis” isolates were untypable. The genetic diversity of S. suis serotypes 1, 2, and 14, as analyzed by multilocus sequence typing, was mainly represented by sequence type (ST)1, ST28, ST25, and ST94. All ST1 isolates (considered highly virulent) belonged to either serotype 1 or 14.