M.A. Pires

CD117 immunoexpression in canine mast cell tumours: correlations with pathological variables and proliferation markers

BackgroundCutaneous mast cell tumours are one of the most common neoplasms in dogs and show a highly variable biologic behaviour. Several prognosis tools have been proposed for canine mast cell tumours, including histological grading and cell proliferation markers. CD117 is a receptor tyrosine kinase thought to play a key role in human and canine mast cell neoplasms. Normal (membrane-associated) and aberrant (cytoplasmic, focal or diffuse) CD117 immunoexpression patterns have been identified in canine mast cell tumours. Cytoplasmic CD117 expression has been found to correlate with higher histological grade and with a worsened post-surgical prognosis. This study addresses the role of CD117 in canine mast cell tumours by studying the correlations between CD117 immunoexpression patterns, two proliferation markers (Ki67 and AgNORs) histological grade, and several other pathological variables.ResultsHighly significant (p < 0,001) correlations were found between CD117 immunostaining patterns and histological grade, cell proliferation markers (Ki67, AgNORs) and tumoral necrosis. Highly significant (p < 0,001) correlations were also established between the two cellular proliferation markers and histological grade, tumour necrosis and epidermal ulceration. A significant correlation (p = 0.035) was observed between CD117 expression patterns and epidermal ulceration. No differences were observed between focal and diffuse cytoplasmic CD117 staining patterns concerning any of the variables studied.ConclusionThese findings highlight the key role of CD117 in the biopathology of canine MCTs and confirm the relationship between aberrant CD117 expression and increased cell proliferation and higher histological grade. Further studies are needed to unravel the cellular mechanisms underlying focal and diffuse cytoplasmic CD117 staining patterns, and their respective biopathologic relevance.

Grade is an independent prognostic factor for feline mammary carcinomas: A clinicopathological and survival analysis

Feline mammary carcinomas (FMC) are highly infiltrative tumours which show a strong tendency for local recurrence and metastasis. Histological type assessment of these tumours is not sufficiently discriminatory in predicting prognosis and in this study the prognostic significance of the Elston and Ellis method of histological grading was evaluated. Ninety-two feline mammary carcinomas from 84 cats were graded and 64 queens were included in a follow-up study. Grade was significantly related to tumour size (P=0.006), clinical stage (P=0.005), lymphovascular invasion (P<0.0001), mitotic index (P<0.0001), Ki67 index (P=0.001), overall survival (P=0.0001) and disease-free survival (P<0.0001). Cox regression analysis identified grade as an independent prognostic factor. Multivariable analysis also showed regional lymph node metastasis and lymphovascular emboli as independent prognostic factors related to overall survival and to disease-free-survival, respectively. The study demonstrated that histological grading can be used as a prognostic factor to evaluate the biological behaviour of FMC.

Multioocyte follicles in domestic dogs: A survey of frequency of occurrence

Multioocyte follicles (MOFs) or polyovular follicles have been reported infrequent in the ovaries of the bitch, decreasing with the age. In this study, the routine observation of the ovaries allowed to verify that the existence of MOFs was far more frequent than previously reported. Ovaries from 150 genitalia, excised during ovariohysterectomy of bitches of different breeds and ages were used. The mean prevalence of MOFs was 40.7%, and the prevalence was higher in young animals (68.4 and 62.2%, in prepubertal and in bitches under 1 year, respectively). In 7-8 years old bitches, the occurrence decreased to 30.4%, and it decreased again to 14.3% in 10 or more years old bitches. It was also more frequent in mongrels than in pure-breeds (52.3-25.5%, respectively). Most follicles contained 2-3 oocytes, but follicles containing up to 10 oocytes were also observed. When the number of oocytes was higher than 3, oocytes of various morphological appearances could be noticed within the follicle. These observations show that the presence of MOFs can affect the number of oocytes recovered in assisted reproductive protocols and may also influence the ovulation rate and prolificity of these animals.

Lipid digestion, absorption and uptake in Solea senegalensis.

Dietary lipids are the major energy source for metabolic purposes in most fish species, and improve dietary protein utilization for growth. In a previous study we have reported a low tolerance of Senegalese sole juveniles to dietary lipid levels and suggested a maximal dietary inclusion level of 8% lipids for both optimal growth and nutrient utilization. The mechanisms behind this apparent poor utilization of the dietary lipids are still to be elucidated. The primary aim of the present study was to investigate the overall process of digestion and lipid absorption in relation to dietary lipid levels. Triplicate groups of twenty fish (mean initial mass 29g) were fed two isonitrogenous diets (54% of protein dry matter basis) with different lipid levels (L4 and L17, 4 and 17% lipids dry matter basis), for 88days. Protein and lipid apparent digestibility coefficients as well as lipase activity were similar in both groups suggesting that Solea senegalensis has the ability to digest equally well a low fat or a high fat diet. Plasma triglyceride concentrations were significantly higher 5 and 16h after feeding in fish fed the L17 compared to those fed L4, following dietary lipid supply, demonstrating effective lipid absorption. Expression of proteins related to lipid transport (microsomal triglyceride transfer protein), trafficking (Fatty acid binding protein 11) and fatty acid uptake (VLDL-r) was significantly higher in liver of fish fed the high fat diet 16h after the meal, but remained unchanged in muscle. In conclusion, it seems that high fat diets do not impair lipid digestion and absorption.

Immunohistochemical characterization of 13 canine renal cell carcinomas.

Canine renal cell carcinomas (RCCs) are uncommon aggressive tumors that occur mainly in middle-aged male dogs. Their histologic classification bears no relationship with prognosis, and little information is available concerning their immunohistochemical properties. In this retrospective study, formalin-fixed, paraffin-embedded tissues from 13 canine RCCs were retrieved from the archives, classified histologically, and evaluated immunohistochemically. The dogs were 7 males and 6 females (1 spayed) of 10 different breeds, averaging 8 years in age. The tumors were classified as papillary, tubulopapillary, papillary–cystic, solid, or sarcomatoid. All 13 tumors were immunohistochemically positive for uromodulin, 12 for c-KIT, 11 for vimentin, 9 for wide-spectrum-screening cytokeratins, 7 for cytokeratins AE1/AE3 and carcinoembryonic antigen, 4 for cytokeratins CAM 5.2, and 3 for CD10. All 3 solid RCCs expressed vimentin, c-KIT, and carcinoembryonic antigen and were negative for cytokeratins. All 7 papillary and tubulopapillary tumors expressed vimentin; 6 (86%), cytokeratins; and 6 (86%), c-KIT. Both papillary–cystic RCCs were positive for cytokeratins and c-KIT and negative for vimentin. These results indicate that the different histologic types of RCC have characteristic immunohistochemical profiles and that c-KIT may be involved in the pathogenesis of canine RCC.

Mammary Invasive Micropapillary Carcinoma in Cats: Clinicopathologic Features and Nuclear DNA Content

Invasive micropapillary carcinoma (IMC) is a variant of infiltrating ductal carcinoma of the breast associated with poor outcome. In this study, we report 16 carcinomas of the feline mammary gland displaying histologic features that correspond to IMC of the breast in women. The clinicopathologic findings, overall survival time, disease-free survival time, and nuclear DNA content of these cats were compared with 65 more common invasive mammary carcinomas (other feline mammary carcinoma [FMC]) of nonspecified type. IMC was associated with larger tumor size, higher histologic grade (P < .0001), deeper muscle invasion (P = .004), and more frequent lymphovascular invasion and nodal metastases (P = .009 and P = .001, respectively) than other FMCs. The aneuploid pattern was more frequent in IMC lesions. IMCs were also associated with lower survival rates. In summary, all cases of feline IMC were associated with clinicopathologic features of high biologic aggressiveness and should be classified as independent histologic types of FMC.

Hypospadias in a Male (78,XY; SRY-Positive) Dog and Sex Reversal Female (78,XX; SRY-Negative) Dogs: Clinical, Histological and Genetic Studies

Hypospadias is rarely reported in dogs. In this study we pre-sent 2 novel cases of this disorder of sexual development and, in addition, a case of hereditary sex reversal in a female with an enlarged clitoris. The first case was a male Moscow watchdog with a normal karyotype (78,XY) and the presence of the SRY gene. In this dog, perineal hypospadias, bilateral inguinal cryptorchidism and testes were observed. The second case, representing the Cocker spaniel breed, had a small penis with a hypospadic orifice of the urethra, bilateral cryptorchidism, testis and a rudimentary gonad inside an ovarian bursa, a normal female karyotype (78,XX) and a lack of the SRY gene. This animal was classified as a compound sex reversal (78,XX, SRY-negative) with the hypospadias syndrome. The third case was a Cocker spaniel female with an enlarged clitoris and internally located ovotestes. Cytogenetic and molecular analyses revealed a normal female karyotype (78,XX) and a lack of the SRY gene, while histology of the gonads showed an ovotesticular structure. This case was classified as a typical hereditary sex reversal syndrome (78,XX, SRY-negative). Molecular studies were focused on coding sequences of the SRY gene (case 1) and 2 candidates for monogenic hypospadias, namely MAMLD1 (mastermind-like domain containing 1) and SRD5A2 (steroid-5-alpha-reductase, alpha polypeptide 2). Sequencing of the entire SRY gene, including 5′- and 3′-flanking regions, did not reveal any mutation. The entire coding sequence of MAMLD1 and SRD5A2 was analyzed in all the intersexes, as well as in 4 phenotypically normal control dogs (3 females and 1 male). In MAMLD1 2 SNPs, including 1 missense substitution in exon 1 (c.128A>G, Asp43Ser), were identified, whereas in SRD5A2 7 polymorphisms, including 1 missense SNP (c.358G>A, Ala120Thr), were found. None of the identified polymorphisms cosegregated with the intersexual phenotype, thus, we cannot confirm that hypospadias may be associated with polymorphism in the coding sequence of the studied genes.

Growth, immune responses and intestinal morphology of rainbow trout (Oncorhynchus mykiss) supplemented with commercial probiotics

The influence of two commercial probiotics on the growth, innate immune parameters and intestinal morphology of rainbow trout (Oncorhynchus mykiss) juveniles (initial weight: 16.4 ± 0.4 g) was evaluated. Two probiotic types: A, multi-species (Bacillus sp., Pedicoccus sp., Enterococcus sp., Lactobacillus sp.) and B, mono-species (Pediococcus acidilactici) were tested at two levels each (A1: 1.5 g kg(-1), 8.6 × 10(5) CFU g(-1); A2: 3 g kg(-1), 1.6 × 10(6) CFU g(-1); B1: 0.1 g kg(-1), 2.6 × 10(4) CFU g(-1); B2: 0.2 g kg(-1), 7.2 × 10(4) CFU g(-1)) versus an unsupplemented diet (C). Diets were distributed to sextuplicate tanks, three times a day to visual satiation for 8 weeks. Growth performance and immune responses (plasma lysozyme, ACH50, peroxidase and head kidney respiratory burst) were determined at 4 and 8 weeks of feeding. Body composition and intestine morphology were determined at the end of the feeding trial. At 8 weeks, the lower dose of multi-species probiotic (A1) improved growth rate, while both probiotic types improved feed conversion rate compared to the control animals, at the lower dose of multi-species (A1) and at the higher dose of mono-species (B2) probiotics. Body composition did not vary between treatments. At 4 weeks, ACH50 activity was significantly higher in fish fed higher dose of B probiotic (B2, 123.7 ± 50.6 vs 44.1 ± 7.7 U.ml(-1) in control). At 8 weeks, lysozyme activity was higher in fish fed A1 (13.1 ± 5.2 μg ml(-1)) diet compared to fish fed control diet (7.8 ± 1 μg ml(-1)). Plasma peroxidase and head-kidney respiratory burst did not differ among the dietary treatments. Villi length and integrity and goblet cell counting of a cross section of the anterior intestine were not significantly different between groups. Results suggest benefits in zootechnical performance and immune humoral responses using both probiotic types, in a dose dependent manner, without apparent alterations in intestinal morphology.

Tumour necrosis factor in the canine endometrium: an immunohistochemical study.

Tumour necrosis factor (TNF), a pleiotropic cytokine that regulates cell growth and differentiation as well as the synthesis of other cytokines, has been identified in the uterus of several species describing a cyclic pattern, eventually under ovarian steroid regulation. Information is yet limited on the presence of TNF protein in the canine endometrium during the oestrous cycle and early pregnancy. This study depicts the temporal immunolocalization of TNF in the bitch endometrium along the oestrous cycle and changes associated with the early steps of embryo invasion. TNF immunolabelling was found in both the stromal fibroblasts and epithelial components of the canine endometrium in all stages studied. Stromal immunostaining was more intense than that of the epithelia, in all the stages of the oestrous cycle. In addition, a tendency for a decrease in the surface epithelium intensity score was found in early dioestrus. A positive glandular content was only observed in anoestrus and proestrus stages. In early pregnancy (days 13-16), TNF immunolabelling was detected at the embryo-maternal surface, in the syncytium cords and the trophoblast, as well in the endometrial stroma and the basal endometrial glands, but not in the lacunar epithelium. The overall TNF immunoreactivity was higher in early pregnancy samples in comparison with those of the early dioestrus and dioestrus stages, suggesting it plays a role during implantation.

An Update on Canine, Feline and Bovine Papillomaviruses

Over recent years, a growing number of papillomaviruses have been identified, which cause a wide range of lesions in domestic and wild animals. Papillomavirus-induced lesions may have a great impact on animal health, and some diseases observed in farm animals are associated with significant economic losses. This concise review brings together recent advancements on animal papillomavirus research, providing the scientific community and veterinary practitioners with an update on this rapidly evolving field. Among others, bovine, canine and feline papillomaviruses (BPV, CPV and FcaPV) are most extensively discussed, in view of the recent discovery of new viral types and their worldwide importance for animal health. Feline papillomaviruses 2 is an emerging, highly prevalent pathogen in domestic cats, associated with a subset of malignant skin lesions. Aspects related to cross-species infection by BPV and its environmental co-factors are also addressed. Animal papillomaviruses are also fascinating models for studying molecular and cell biology and have recently inspired some major breakthroughs. Overall, it is clear that additional, international and systematic efforts are needed to clarify which lesions are caused by which viral types and to develop experimental models for studying animal papillomavirus.