Solitary and multiple simultaneous malignant epithelial mammary tumours in dogs: An explorative retrospective study.

Canine mammary tumours represent a hard-prognostic task for veterinary clinicians. TNM staging and grading systems refer to a single tumour. Significant limits come to light when these systems are applied to multiple mammary tumours due to the arbitrary criterion in determining which single tumour is representative of the patient's prognosis. This study explored some clinical features of 50 dogs affected by at least one malignant mammary tumour. Clinical features and staging, together with histological classification and grading, have been related to disease-free survival (DFS) with the purpose to evaluate their impact on prognosis. The prognosis was worse in 10-11-year-old dogs (P < 0.05), in dogs affected by complex carcinoma (P < 0.05), and in patients assigned to Peña grade I (P < 0.05). The bodyweight was not linearly related to DFS (P < 0.01), and patients with a low number of neoformations (n ≤ 2) showed a better prognosis than dogs with 3-5 tumours (P < 0.05). Both the average and the total size of malignant tumours were related to DFS (P < 0.05). Dogs assigned with stage I had the best DFS (P < 0.05). In conclusion, the Peña grade I alone would not seem to guarantee a favourable prognosis when applied to mammary tumours in dogs affected by multiple simultaneous presentations. Different characteristics, besides tumour grading, such as tumour immunophenotype and expression of hormonal receptors, could in the future, contribute to elucidate the clinical behaviour of multiple canine mammary tumours.

High-density neutrophils in MGUS and multiple myeloma are dysfunctional and immune-suppressive due to increased STAT3 downstream signaling.

To understand neutrophil impairment in the progression from MGUS through active MM, we investigated the function of mature, high-density neutrophils (HDNs), isolated from peripheral blood. In 7 MM, 3 MGUS and 3 healthy subjects by gene expression profile, we identified a total of 551 upregulated and 343 downregulated genes in MM-HDN, involved in chemokine signaling pathway and FC-gamma receptor mediated phagocytosis conveying in the activation of STAT proteins. In a series of 60 newly diagnosed MM and 30 MGUS patients, by flow-cytometry we found that HDN from MM, and to a lesser extend MGUS, had an up-regulation of the inducible FcγRI (also known as CD64) and a down-regulation of the constitutive FcγRIIIa (also known as CD16) together with a reduced phagocytic activity and oxidative burst, associated to increased immune-suppression that could be reverted by arginase inhibitors in co-culture with lymphocytes. In 43 consecutive newly-diagnosed MM patients, who received first-line treatment based on bortezomib, thalidomide and dexamethasone, high CD64 could identify at diagnosis patients with inferior median overall survival (39.5 versus 86.7 months, p = 0.04). Thus, HDNs are significantly different among healthy, MGUS and MM subjects. In both MGUS and MM neutrophils may play a role in supporting both the increased susceptibility to infection and the immunological dysfunction that leads to tumor progression.

Evaluation of leukocyte counts and neutrophil-to-lymphocyte ratio as predictors of local recurrence of feline injection site sarcoma after curative intent surgery.

Local recurrence (LR) is the major concern in the treatment of feline injection-site sarcoma (FISS). Pretreatment leukocyte counts and ratios have been reported as diagnostic and/or prognostic markers in human and canine oncology. The aim of this retrospective study was to explore the prognostic impact on LR and overall survival time (OST) of pretreatment neutrophil-to-lymphocyte ratio (NLR), white blood cell count (WBCC), neutrophil count (NC) and lymphocyte count (LC) in cats with surgically excised FISS. Eighty-two cats with histologically confirmed FISS at first presentation, without distant metastases, and with available pretreatment haematological analyses were retrospectively enrolled. The correlation of NLR, WBCC, NC, LC with tumour variables and patient variables was explored. NLR was correlated with tumour size (P = .004), histological pattern of tumour growth (P = .024) and histotype (P = .029), while WBCC and NC were associated with ulceration (P = .007, P = .011) and pattern of growth (P = .028, P = .004). No significant relationships emerged between LC and any of the considered variables. The impact of NLR, WBCC, NC, LC on LR and OST was then estimated in univariate and multivariate analysis. In univariate analysis, NLR, WBCC and NC were significant prognostic factors for both LR and OST. NLR, WBCC and NC remained prognostic in multivariate analysis for LR but not for OST. When NLR, WBCC and NC were jointly analysed, WBCC was the marker with the greater impact on LR. Preoperative NLR, WBCC and NC may aid in identifying cats at higher risk of LR.

Histopathological Terminology Standards for the Reporting of Prostatic Epithelial Lesions in Dogs.

The terminology applied to canine prostatic epithelial lesions, especially carcinomas, is currently not standardized and this hampers the ability of pathologists to study the biological and clinical significance of these lesions. The aim of this review is to present the essential histomorphological diagnostic attributes of a wide spectrum of prostatic epithelial lesions in dogs. In addition to the traditionally recognized prostatic hyperplasia, hormonal atrophy, prostatitis, squamous metaplasia, adenocarcinoma and transitional cell (urothelial) carcinoma, new entities are described and discussed in order to provide veterinary pathologists with a basic atlas of common histological lesions of the canine prostate that is comprehensive and easy to use.

The impact of extirpation of non-palpable/normal-sized regional lymph nodes on staging of canine cutaneous mast cell tumours: A multicentric retrospective study.

Metastasis to regional lymph nodes (RLNs) in dogs with cutaneous mast cell tumour (cMCT) has been correlated with shortened survival time and higher risk of spread to distant sites. In the present study, extirpation of non-palpable or normal-sized RLNs was included in the surgical management of cMCT in dogs. Correlations between histological nodal status (HN0-3) and tumour variables were analysed. Ninety-three dogs with single cMCT without distant metastasis that underwent wide surgical excision of the primary tumour and extirpation of non-palpable or normal-sized RLN were included. The association between HN (HN0 vs HN > 0; HN0-1 vs HN2-3) and tumour variables (site, longest diameter, ulceration, 3-tier and 2-tier histological grades) was analysed by a generalized linear model with multinomial error. Then, 33 (35.5%) RLNs were HN0, 14 (15%) were HN1, 26 (28%) were HN2 and 20 (21.5%) were HN3. The presence of positive (HN > 0) RLN was significantly associated with cMCT larger than 3 cm. No other association was statistically significant. Non-palpable/normal-sized RLN in dogs with cMCT can harbour histologically detectable metastatic disease in nearly half of the cases. Extirpation of the RLN should always perfomed to obtain a correct staging of the disease, even in the absence of clinical suspicion of metastasis. Further studies should evaluate the possible therapeutical effect of the tumour burden reduction obtained by exrtipartion of a positive RLN.

Diagnostic Utility of Cytokeratin-5 for the Identification of Proliferative Inflammatory Atrophy in the Canine Prostate.

Proliferative inflammatory atrophy (PIA), which is comprised of highly proliferative but atrophic prostate epithelial cells in association with chronic inflammation, is considered a risk lesion for prostate cancer in men, while its role in canine prostate carcinogenesis is still unknown. We evaluated the value of immunohistochemical labelling for the basal cell marker cytokeratin-5 (CK5) in identifying PIA lesions in 87 samples of formalin-fixed and paraffin wax-embedded canine prostate. Canine PIA showed cytological features identical to the human counterpart and in most cases was associated with chronic lymphoplasmacytic inflammation. PIA lesions were identified in a higher number of CK5-labelled slides (43 out of 87) compared with slides stained by haematoxylin and eosin (HE) (24 out of 87). This lesion was frequently present in normal, hyperplastic and neoplastic canine prostates, although it was underestimated on evaluation of HE-stained slides. Therefore, CK5 can be considered a useful basal cell marker with high sensitivity and specificity for PIA.

Ovarian Embryonal Carcinoma in a Dog.

A 17-month-old female doberman pinscher was referred for an abdominal mass and ascites. Exploratory laparotomy revealed the presence of a large neoplastic mass replacing the right ovary and associated with multiple mesovarian, mesometrial and peritoneal nodules. An ovariohysterectomy was performed. Grossly, the tumour was soft and multilocular with large areas of haemorrhage and necrosis. Microscopically, it was infiltrative and composed of round and polygonal cells arranged respectively in solid sheets or forming distorted tubular structures separated by thick fibrovascular septae. Tubules contained necrotic debris, proteinaceous fluid or small endoluminal papillary structures. Marked cellular atypia, multiple neoplastic emboli and high mitotic count were observed. Immunohistochemically, the round cells uniformly expressed placental alkaline phosphatase, while the polygonal cells arranged in tubules and papillae expressed cytokeratin (CK) AE1/AE3 and CK7. A final diagnosis of metastasizing ovarian embryonal carcinoma (EC), a primitive germ cell tumour characterized by rudimentary epithelial differentiation was made. Canine ovarian EC should be considered as a differential diagnosis for undifferentiated aggressive ovarian tumours in young dogs.

Immunohistochemical Expression of Placental Alkaline Phosphatase in Five Cases of Seminoma in Rabbits.

Testicular seminoma is reported in the rabbit but data about the immunophenotype of these tumours are lacking. The classification of human testicular germ cell tumours includes spermatocytic tumour (ST) originating from the post-pubertal spermatogonia/spermatocytes, which metastasizes rarely, and seminoma (SE), originating from gonocytes, which is malignant and metastasizes frequently. Gonocytes express placental alkaline phosphatase (PLAP) and are stained with periodic acid-Schiff (PAS). We report five cases of seminoma in pet rabbits. Microscopically, all the cases were diffuse seminoma and in one case there was metastasis to a sublumbar lymph node. Immunohistochemical expression of PLAP was diffuse in this metastatic tumour, in two other cases it was multifocal, in another it was limited to rare cells and in the remaining case was negative. PAS-positive cells were detected only in the four cases that expressed PLAP. These four cases were therefore classified as SE and the tumour without PLAP labelling or PAS staining was defined as ST. Both forms of human germ cell tumour therefore occur in the rabbit. SE appears to be well represented and may show metastasis, paralleling the human counterpart. The results of this study provide a basis for further evaluations of the rabbit as a possible animal model for the study of human SE.

Clinical and computed tomography tumour dimension assessments for planning wide excision of injection site sarcomas in cats: how strong is the agreement?

In injection site sarcoma (ISS) in cats lateral as well as deep margins should be correctly planned for a successful surgical outcome. The discrepancy between clinical and computed tomography (CT) measurements of dimension in resectable tumour has led to possible bias that affects the subsequent surgical dose. The aim of this study was to prospectively investigate the agreement between clinical and CT measurements of dimension in newly diagnosed ISS in cats. Fifty-three client-owned cats that underwent both clinical and CT measurements of the length and width of ISS were included. CT measurements showed a tendency towards being larger than clinical dimensions, and this difference increased with increasing tumour size. Based on our results, in further studies focusing on ISS in cats, the kind of assessment used to define tumour dimensions (CT versus clinic) should be declared and specified to properly consider surgical results and prognostic impact of this variable.

Immunophenotyping of Rabbit Testicular Germ and Sertoli Cells Across Maturational Stages.

During testicular maturation, both Sertoli cells (SCs) and germ cells (GCs) switch from an immature to a mature immunophenotype. The reexpression of markers of immaturity in adults has been reported in cancer and in other testicular pathologies, in men as well as in animal species. Naturally affected with testicular cancer, rabbits have long been used in human reproductive research, but reports on the expression of testicular cell markers in this species are few and data about the immunophenotype of normal postnatal SCs and GCs are lacking. The aim of this study was to investigate the immunophenotype of SCs and GCs in the rabbit, from neonatal to adult age, using the antibodies anti-Müllerian hormone (AMH), vimentin (VIM), CKAE1/AE3 (cytokeratins [CKs]), desmin (DES), inhibin alpha (INH-α), placental alkaline phosphatase (PLAP), and periodic acid-Schiff (PAS) staining. In SCs, VIM was constantly expressed, and AMH and CKs expression was limited to neonatal and prepubertal age, whereas DES, INH-α, PLAP, and PAS were constantly negative. GCs were negatively stained for PLAP, PAS, and for the other markers. Results revealed analogies with human testicular immunophenotype, suggesting that rabbits could represent a potential experimental model for the study of human testicular pathology.

A dynamic distention protocol for whole-organ bladder decellularization: histological and biomechanical characterization of the acellular matrix.

A combined physical-chemical protocol for whole full-thickness bladder decellularization is proposed, based on organ cyclic distention through repeated infusion/withdrawal of the decellularization agents through the urethra. The dynamic decellularization was intended to enhance cell removal efficiency, facilitating the delivery of detergents within the inner layers of the tissue and the removal of cell debris. The use of mild chemical detergents (hypotonic solution and non-ionic detergent) was employed to limit adverse effects upon matrix 3D ultrastructure. Inspection of the presence of residual DNA and RNA was carried out on decellularized matrices to verify effective cell removal. Histological investigation was focused on assessing the retention of adequate structural and functional components that regulate the biomechanical behaviour of the acellular tissue. Biomechanical properties were evaluated through uniaxial tensile loading tests of tissue strips and through ex vivo filling cystometry to evaluate the whole-organ mechanical response to a physiological-like loading state. According to our results, a dynamic decellularization protocol of 17 h duration with a 5 ml/min detergent infusion flow rate revealed higher DNA removal efficiency than standard static decellularization, resulting in residual DNA content < 50 ng/mg dry tissue weight. Furthermore, the collagen network and elastic fibres distribution were preserved in the acellular ECM, which exhibited suitable biomechanical properties in the perspective of its future use as an implant for bladder augmentation.

Different Growth Patterns of Canine Prostatic Carcinoma Suggests Different Models of Tumor-Initiating Cells.

Controversies remain regarding the cell type from which human prostate cancer originates, and many attempts have been made to identify the cellular origin of canine prostate cancer but without definitive proof. This study aims to evaluate the expression of luminal (androgen receptor [AR], cytokeratin [CK]8/18) and basal (CK14, CK5) cell markers in different histologic subtypes of canine prostatic carcinoma (PC) and to suggest the most likely tumor-initiating cells. Normal prostates (n = 8) were characterized by AR+CK8/18+ luminal cells and few CK5+ basal cells, while CK14 was absent. Similar pattern was observed in all 35 prostates with benign prostatic hyperplasia, except few scattered CK14+ basal cells in 13 samples (37.14%). AR was localized in the nucleus of both normal and hyperplastic cells. In 34 samples of PC, the following growth patterns were identified: cribriform (44.12%), solid (32.35%), small acinar/ductal (20.59%), and micropapillary (2.94%). Most PCs expressed AR and CK8/18, while CK5 and CK14 expression was observed in 25% and 20% of cases, respectively. AR revealed a variable intracellular distribution, both nuclear and cytoplasmic. Solid PC was characterized by an undifferentiated or aberrant phenotype with a reduced expression of AR and CK8/18, increased number of CK14+ cells, and 7 antigen expression patterns. This study demonstrated a predominance of differentiated luminal cell types in canine prostatic tumors, although the role of basal cells in prostate carcinogenesis should also be considered. Moreover, few scattered CK5+ cells in AR+CK8/18+ tumors identified the existence of intermediate cells, from which neoplastic transformation may alternatively commence.

Congenital abnormality of the vagina complicated by haemato-pyocolpos in a 1-year labrador retriever.

A 1-year-old female Labrador retriever was referred with a few days history of haematic-like vulvar discharge. Physical examination, vaginal inspection and palpation did not reveal any remarkable finding. Transabdominal ultrasound showed echogenic fluid accumulation in the vagina suggesting haemato-pyocolpos. An exploratory laparotomy was performed: a well-delimited ectasic vagina was identified. Ovariohysterectomy and partial vaginectomy and vaginoplasty were performed to spay the bitch and to remove the ectasic vagina. Post-operative recovery and 12-month follow-up were uneventful. Clinical, morphological and histological findings were consistent with a congenital abnormality of the muscular layer of the vagina complicated by haemato-pyocolpos. The disorganization of the vaginal tunica muscularis may have acted as locus minoris resistentiae in the vaginal wall. The organ was dilated and atonic due to the gradual accumulation of physiological fluids complicated by an overgrowth of genital bacteria. This congenital disorder has to be taken into account as differential diagnosis of haemato-pyocolpos with vaginal discharge in young bitches.

Immunohistochemical study of mixed germ cell sex cord stromal tumours in 13 canine testes.

Mixed germ cell sex cord stromal tumours (MGSCTs) are composed of seminiferous tubules, filled with admixed neoplastic Sertoli cells (SCs) and germ cells (GCs). The aim of the present study was to describe 13 canine testicular MGSCTs and to investigate the histochemical features and the immunophenotype of the neoplastic GCs and SCs. Neoplastic SCs were always diffusely labelled for vimentin (VIM), neuron specific enolase (NSE), inhibin (INH)-α and anti-Müllerian hormone (AMH). Cytokeratins AE1/AE3 (CK) and desmin (DES) were expressed in 6/13 and 8/13 cases, respectively. Neoplastic GCs were labelled for placental alkaline phosphatase (PLAP) in 7/13 cases and for CD117 (KIT) in 8/13 cases, while 10 cases were stained uniformly by periodic acid-Schiff (PAS). Immature canine SCs are known to express CK, DES, INH-α and AMH, while immature GCs are stained by PAS and express PLAP and KIT. This GC phenotype also distinguishes between classical and spermatocytic seminoma, with the latter being negative for these markers. The results of the present study show that both neoplastic SCs and GCs in MGSCTs have a de-differentiated phenotype.

A retrospective analysis of 111 canine prostatic samples: histopathological findings and classification.

The purpose of this retrospective study is to evaluate the frequency and further characterize the pathological features of common and uncommon histological lesions in 111 canine prostatic samples. Benign prostatic hyperplasia, suppurative and non-suppurative prostatitis, and prostate cancer were observed individually or in combination in 45, 11, 68 and 50 samples, respectively. Six growth patterns of prostatic carcinoma were differentiated: papillary, cribriform, solid, small acinar/ductal, signet ring, mucinous. In a few cases, perineurial invasion and collagenous micronodules were observed. Lesions considered preneoplastic in men, such as high-grade prostatic intraepithelial neoplasia (HGPIN) and prostatic inflammatory atrophy (PIA), were observed in 27 and 21 histological samples, respectively. This study represents a detailed characterization of the different histological subtypes of canine prostate cancer. The awareness of the unusual patterns might be critical in avoiding diagnostic misinterpretation. The high prevalence of PIA and HGPIN underlines the reasonable chance of their detection in routine biopsy specimens.

Immunohistochemical expression of markers of immaturity in sertoli and seminal cells in canine testicular atrophy.

During maturation from fetal to adult testis, both Sertoli cells (SCs) and germ cells (GCs) switch from an immature to a mature immunophenotype. Immature canine SCs express cytokeratins (CKs), desmin (DES), vimentin (VIM), anti-Müllerian hormone (AMH) and inhibin (INH)-α, while mature SCs retain only expression of VIM. Immature GCs express placental alkaline phosphatase (PLAP), which is lost in spermatocytes. Re-expression of markers of immaturity has been observed in human atrophic testes and in human and canine testicular tumours. In human medicine, testicular atrophy is considered a risk factor for testicular cancer. In the present study 13 canine atrophic testes were examined immunohistochemically. VIM was expressed in the SCs of all cases, while CK, DES, INH-α and AMH were expressed in a variable percentage of SCs in two, five, five and eight cases, respectively. PLAP was expressed by single GCs in one case. Markers of immaturity are therefore expressed by SCs and GCs in canine atrophic testes. Similar results were reported previously in canine testicular neoplasia, suggesting that testicular atrophy may represent a risk factor for tumour development in the dog.

Medium term endoscopic assessment of the surgical outcome following laryngeal saccule resection in brachycephalic dogs.

Laryngeal saccule eversion has been widely reported as an important component of brachycephalic airway obstructive syndrome (BAOS). The authors hypothesised that saccules affected by acute histological changes in patients showing marked improvement following palate and nares surgery might spontaneously return to normal; moreover, spontaneous resolution of the eversion in patients with fibrotic saccules and/or without clinical improvement following BAOS surgery might be impossible and, on the contrary, the persistence of turbulent airflow and associated ongoing inflammation might lead to aberrant tissue proliferation after resection. In order to demonstrate our hypotheses, the authors decided to perform a unilateral sacculectomy and to postpone and assess the need for the execution of the contralateral saccule resection according to the findings of a second-look laryngoscopy. Ten dogs were enrolled. None of the saccules left in situ underwent spontaneous resolution of the eversion. In one dog, after sacculectomy, proliferation of a soft tissue lesion endoscopically similar to a newly formed saccule occurred. The results of the present study suggest that spontaneous resolution of saccule eversion is uncommon, even after the correction of the primary abnormalities (palate, nares). Resection of the saccules can relieve ventral rima glottidis obstruction; however, secondary intention healing might occasionally result in the recurrence of the obstruction.

Immunohistochemical evaluation of the expression of anti-Müllerian hormone in mature, immature and neoplastic canine Sertoli cells.

In mammals, the earliest specific protein expressed by Sertoli cells (SCs) is the anti-Müllerian hormone (AMH), which induces the regression of Müllerian ducts and is produced by SCs until the functional maturation of the testes. The aim of this study was to evaluate the expression of AMH by canine SCs during testicular maturation and neoplastic transformation. Testes from two fetuses, 18 newborn puppies, five puppies aged 43-180 days and six adult dogs, and 24 canine Sertoli cell tumours (SCTs) were studied immunohistochemically for expression of AMH. Fifteen of the 24 SCTs were classified as typical, eight as lipid-rich and one was considered malignant based on evidence of lymph node metastasis. SCs from fetuses and neonatal puppies and puppies up to 45 days old expressed AMH, while SCs from older puppies and adults were negative. All SCTs expressed AMH, suggesting that AMH expression is a useful marker of immature and neoplastic canine SCs.

Inhibin-α immunohistochemical expression in mature and immature canine Sertoli and Leydig cells.

Formalin-fixed, paraffin-embedded sections from 32 canine pairs of testes were immunohistochemically examined for Inhibin-α (INHα). Samples were subdivided into two groups (group 1, neonates; group 2, puppies and adults) and results statistically compared. Inhibin-α was significantly expressed only in Sertoli cells of neonatal testes, while in Leydig cells it was expressed without significant difference between groups. These results suggest that, in dogs, INHα expression switches from Sertoli to Leydig cells during testicular maturation and that, in adult, Leydig cells represent the main source of INHα.

Performances of different diagnostic tests for feline infectious peritonitis in challenging clinical cases.

OBJECTIVES: Feline infectious peritonitis (FIP) can be difficult to diagnose. Histopathology is considered the gold standard test but immunohistochemistry (IHC) is mandatory to confirm/exclude the disease. This study aimed to assess the performances of tests carried out in vivo or at postmortem examination in challenging cases in which FIP was confirmed or excluded based on IHC or on adequate follow-up. METHODS: Twelve cases (four without FIP, eight with FIP) were retrospectively studied. Clinical findings, serum protein electrophoresis (SPE), analysis of the effusions (AE), antifeline coronavirus serology, serum concentration of α1-acid glycoprotein (AGP) and histopathology were classified as consistent, doubtful or non-consistent with FIP. Sensitivity, specificity and concordance (κ) with the final diagnosis were calculated. RESULTS: Concordance was absent for serology (κ=-0·08) and AE (κ=-0·52), poor for histopathology (κ=0·09), fair for SPE (κ=0·25) and perfect for AGP (κ=1·00). Sensitivity was high for AGP (100%) and low for AE (50%), SPE (37·5%) and histopathology (37·5%). Specificity was high for AGP or histopathology (100%) and low for SPE (50%) and AE (0%). CLINICAL SIGNIFICANCE: IHC must always be performed to confirm FIP. If this is not possible, when histopathology is controversial, elevated AGP concentrations may support the diagnosis of FIP.