MicroRNA-126 in dogs with immune complex-mediated glomerulonephritis.

BACKGROUND: Most proteinuric dogs with naturally occurring chronic kidney disease have amyloidosis (AMYL), glomerulosclerosis (GS), or immune complex-mediated glomerulonephritis (ICGN), each with different treatment and prognosis. A noninvasive and disease-specific biomarker is lacking. HYPOTHESIS: We hypothesized that the expression pattern of biofluid microRNA (miRNAs and miRs) would correlate with disease progression and categorization. ANIMALS: Archived serum and urine samples from 18 dogs with glomerular disease and 6 clinically healthy dogs; archived urine samples from 49 dogs with glomerular disease and 13 clinically healthy dogs. METHODS: Retrospective study. Archived biofluid samples from adult dogs with biopsy-confirmed glomerular disease submitted to the International Veterinary Renal Pathology Service between 2008 and 2016 were selected. Serum and urinary miRNAs were isolated and profiled using RNA sequencing. Urinary miR-126, miR-21, miR-182, and miR-486 were quantified using quantitative reverse transcription PCR. RESULTS: When comparing more advanced disease with earlier disease, no serum miRNAs were differentially expressed, but urinary miR-21 and miR-182 were 1.63 (95% CI: .86-3.1) and 1.45 (95% CI: .82-2.6) times higher in azotemic dogs, respectively (adjusted P < .05) and weakly correlated with tubulointerstitial fibrosis (miR-21: r = .32, P = .03; miR-182: r = .28, P = .05). Expression of urinary miR-126 was 10.5 (95% CI: 4.1-26.7), 28.9 (95% CI: 10.5-79.8), and 126.2 (95% CI: 44.7-356.3) times higher in dogs with ICGN compared with dogs with GS, AMYL, and healthy controls, respectively (P < .001). CONCLUSIONS AND CLINICAL IMPORTANCE: The miR-126 could help identify dogs that might benefit from immunosuppressive therapy in the absence of a biopsy. MiR-21 and miR-182 are potential markers of disease severity and fibrosis.

Longitudinal examination of urine pH, specific gravity, protein, culture, and antimicrobial resistance profiles in healthy dogs.

BACKGROUND: Urine is routinely evaluated in dogs to assess health. Reference ranges for many urine properties are well established, but the scope of variation in these properties over time within healthy dogs is not well characterized. OBJECTIVES: Longitudinally characterize urine properties in healthy dogs over 3 months. ANIMALS: Fourteen healthy client-owned dogs. METHODS: In this prospective study, dogs were evaluated for health; then, mid-stream free-catch urine was collected from each dog at 12 timepoints over 3 months. Urine pH, urine specific gravity (USG), protein, cultures, and antimicrobial resistance profiles were assessed at each timepoint. RESULTS: Urine pH varied within and between dogs over time (Friedman's test: within P = .03; between P < .005). However, USG, protein, and bacterial diversity of urine were consistent within dogs over time, and only varied between dogs (Kruskal-Wallis: between all P < .005). Antimicrobial resistant isolates were identified in 12 out of 14 dogs with 34 of 48 of the isolates demonstrating resistance to amoxicillin. CONCLUSIONS AND CLINICAL IMPORTANCE: Urine pH should be assessed at multiple timepoints via pH meter before making clinical decisions. Mid-stream free-catch urine with high concentrations of bacteria (>105 CFU/mL) should not be considered the only indicator of urinary tract infection. Bacterial isolates from dogs in this study had widespread resistance to amoxicillin/oxacillin underscoring the need for antimicrobial stewardship.

Clinical Utility of Cytology from Preoperative Percutaneous Fine Needle Aspirates of Solitary Liver Masses in 220 Dogs: A Retrospective Study (2009-2019).

When a solitary liver mass is identified in a dog, a fine-needle aspirate (FNA) is commonly employed to attempt to obtain a diagnosis. Little information is provided in the literature evaluating the sensitivity/specificity of FNA cytology for solitary liver masses. We hypothesized that liver lesion size nor the presence of cavitation would impact the success of cytological diagnosis. Medical records were obtained for 220 client-owned dogs. Inclusion criteria included preoperative abdominal imaging, percutaneous FNA of a solitary hepatic mass with cytologic interpretation by a board-certified pathologist, and a surgical biopsy or mass excision yielding a histopathological diagnosis. Six dogs (2.7%) experienced a complication after FNA, none considered severe. The agreement rate for correct cytologic diagnosis was 22.9% (49/220). Of the neoplastic masses 18.9% (35/185) were correctly diagnosed via cytology. The overall sensitivity was 60%, and the specificity was 68.6%. Neither institution (P = 0.16), lesion size (P = 0.88), cavitation (P = 0.34), or needle gauge (P = 0.20) had an association with correct diagnosis. This study demonstrates that, although there is a low risk of complications following FNA of a hepatic mass, overall success rate for correct cytologic diagnosis based on FNA was low compared to histopathologic diagnosis.

Urinary Biomarkers of Kidney Disease in Dogs and Cats.

A variety of urinary markers of the renal disease show promise for the identification of glomerular and tubular damage and monitoring treatment. Most of the markers are currently not widely available, and all could benefit from further study. This review summarizes recent studies on urinary biomarkers of renal disease in dogs and cats.

Use of doxorubicin chemotherapy following debulking surgery in the treatment of a cat with mesenteric hemangiosarcoma.

Case summary: A 7-year-old male castrated domestic shorthair cat presented with a 5-day history of inappetence. A mid-abdominal mass was palpated and, on exploratory laparotomy, a cystic mass arising from the root of the mesentery was observed. The mass was drained, debulked and omentalized. Histopathologic examination and immunohistochemistry supported a diagnosis of hemangiosarcoma. Adjuvant doxorubicin was started and, prior to the third of five doses of doxorubicin, repeat abdominal ultrasound showed complete response of the primary tumor. Continued monitoring 240 days following histopathologic diagnosis revealed suspected metastasis to local lymph nodes, though the primary tumor remained absent on abdominal ultrasound. A second course of five doses of doxorubicin chemotherapy was completed. Serial abdominal ultrasounds demonstrated stable disease in the locoregional lymph nodes with no visible recurrence of the primary tumor. The cat presented 430 days following diagnosis with lethargy and inappetence. Abdominal ultrasound revealed suspected metastatic mesenteric and ileocolic lymphadenopathy, hepatic metastasis and peritoneal effusion, and the owner elected for humane euthanasia. Necropsy findings and negative immunohistochemical staining for lymphatic vessel endothelial receptor-1 were consistent with a metastatic mesenteric hemangiosarcoma. Relevance and novel information: Hemangiosarcoma is an uncommon malignancy in cats, and few cases describing treatment have been reported. To our knowledge, this is the first report to describe the use of debulking surgery and adjuvant doxorubicin chemotherapy in the treatment of mesenteric hemangiosarcoma resulting in extended survival in a cat. Multimodal therapy can be considered for the management of cats with mesenteric hemangiosarcoma.

Utility of Spleen and Liver Cytology in Staging of Canine Mast Cell Tumors.

Abdominal ultrasound with spleen and liver cytology is part of routine staging for canine mast cell tumors (MCTs). However, such tests are associated with increased morbidity and cost. Therefore, the objectives of this study were to determine if spleen cytology was predictive of liver cytology in canine MCTs and if any patient or tumor variables were associated with spleen and/or liver metastasis. Records of dogs with MCTs and cytology of spleen and liver were reviewed. Two hundred five patients were included. Overall, 22 (10.7%) patients had metastasis, with 21 (10.2%) and 13 (6.3%) having spleen and liver metastasis, respectively, and 12 (5.9%) having both. For patients with a positive (or negative) spleen cytology, the odds ratio of having a positive (or negative) liver cytology was 233.49. However, a negative spleen cytology had a higher predictive value (0.99) than a positive cytology (0.54). Finally, the presence of local and systemic signs and tumor size were associated with spleen, liver, and/or spleen or liver metastasis. These results suggest that spleen cytology is predictive of liver cytology in staging of canine MCTs, and increasing tumor size and presence of local or systemic signs are associated with an increased risk of visceral metastasis.

Nasal colonization by Entamoeba gingivalis in a 13-year-old Italian Greyhound.

A 13-year-old 4.6 kg castrated male Italian Greyhound was presented for evaluation of sneezing, bilateral epistaxis and mucoid nasal discharge of approximately 3 months duration. Severe periodontal disease was noted on physical exam. The patient's CBC was unremarkable. His biochemical profile revealed mild azotemia as well as mild electrolyte abnormalities. Rhinoscopy revealed a tan plaque present in the left dorsal meatus of the nasal cavity. Vigorous flushing yielded granular white particulates suspended in the saline, which were cytologically examined. Marked neutrophilic inflammation, a diverse bacterial population, and organisms consistent with amoebic trophozoites were identified. The remaining fluid was analyzed by PCR and sequencing, and Entamoeba gingivalis was identified. The dog's clinical signs improved dramatically after initiating treatment with clindamycin. Six weeks after diagnosis, a dental cleaning with multiple extractions was performed. Four weeks after the dental procedure, the dog was clinically normal. This report describes the clinical, hematologic, cytologic, and gross findings in a case of Entamoeba gingivalis infection in a dog.

Pathology in Practice.

In collaboration with the American College of Veterinary Pathologists.

Pathology in Practice.

In collaboration with the American College of Veterinary Pathologists.

Interobserver reliability of canine urine specific gravity assessed by analog or digital refractometers.

Refractometry is utilized routinely to evaluate canine urine specific gravity (USG) in veterinary clinical settings. We aimed to determine if the magnitude of interobserver reliability when assessing canine USG via refractometry could impact clinical judgment. USG was determined in 38 dogs by 3 registered veterinary technicians (RVTs) using both an optical analog refractometer and a digital refractometer. Summary statistics were reported, interobserver reliability was assessed via intraclass correlation coefficient (ICC) analysis through a 2-way mixed-effects model, and agreement between RVT pairs was compared through Bland-Altman plots. The median analog refractometer USG measurement was 1.018 (range: 1.004-1.040) and for the digital refractometer was 1.0176 (1.0035-1.0357). The analog refractometer average measure ICC was 0.995 (95% CI: 0.992, 0.997; p < 0.001). The digital refractometer average measure ICC was 0.999 (95% CI: 0.999, 1.000; p < 0.001). Strong agreement between all pairs of RVTs was seen via Bland-Altman plots for both analog and digital refractometers, with 95% CIs spanning no more than 0.002 in either the positive or negative direction for all pairings. The interobserver variability in canine USG measurements by RVTs was trivial and did not impact clinical judgment and decision-making.

Comparison of pathologist review protocols for cytologic detection of prostatic and urothelial carcinomas in canines: A bi-institutional retrospective study of 298 cases.

Urothelial carcinoma, also known as transitional cell carcinoma, is the most common primary bladder tumour in dogs, and can also involve the prostate gland. Cytology is a common diagnostic tool utilized for dogs with bladder or prostate gland lesions. The objectives of this retrospective study were to compare the sensitivity and specificity of cytologic evaluation for urothelial or prostatic carcinoma between two institutions with different cytology review protocols as well as determine if certain collection methods resulted in higher cytologic accuracy. A total of 298 cases met inclusion criteria. The overall sensitivity and specificity for institution 1 were 91.8% and 50%, respectively, compared to 31.1% and 97.4%, respectively, for institution 2. When the urine sample review protocol at institution 2 was matched to that of institution 1, sensitivity and specificity were more similar to institution 1 (71.2% and 88.9%, respectively). Our results show that the sensitivity and specificity of cytology are affected by screening and review protocols implemented by different institutions. The data also demonstrate that sensitivity and specificity vary by collection method. Diagnostic catheterization had the highest performance: of the 11 cases between two institutions, it had 100% sensitivity and specificity. In contrast, examination of urine sediment not collected via diagnostic catheterization had low sensitivity and specificity that varied greatly by institution. In summary, cytologic interpretation should be undertaken with consideration given to both processing and collection protocols.

Fibrosarcoma with sarcomatosis and metastasis in a FeLV-negative cat.

A 6-year-old, spayed female, mixed shorthair cat presented to the emergency service at The Ohio State University Veterinary Medical Center for evaluation of hypercalcemia, a right eye mass, and multiple intrathoracic and intra-abdominal masses. Cytologic evaluation of one of the abdominal masses revealed a uniform population of large, anaplastic mesenchymal cells found individually, in loose aggregates, and occasionally associated with pink, extracellular matrix. The cytology was consistent with a malignant mesenchymal neoplasm, with primary consideration given to fibrosarcoma and hemangiosarcoma. The cat was euthanized and histopathology confirmed disseminated fibrosarcoma. Fibrosarcoma comprises 12%-41% of feline cutaneous tumors and affects cats at a mean age of 9.6 years. Three manifestations of fibrosarcoma predominate in cats: spontaneous solitary fibrosarcoma, vaccine-induced/injection site fibrosarcoma, and oncogene-induced (FSV) fibrosarcoma. The history, signalment, and results from diagnostics performed did not support solitary fibrosarcoma or injection-induced sarcoma. Although some criteria fit with virally induced fibrosarcoma, such as age and the presence of multiple fibrosarcomas, the neoplastic population was negative for FeLV IHC. The presence of fibrosarcomas throughout the pleural and peritoneal cavity was most compatible with sarcomatosis and the distant metastasis of an unidentified primary neoplasm. To the authors' knowledge, this is the first reported case of sarcomatosis in a FeLV-negative cat.

Persistent nonregenerative anemia in a 4-year-old cat.

A 4-year-old male neutered domestic shorthair cat was presented to The Ohio State University College of Veterinary Medicine for a 2-month history of severe weight loss, lethargy, anemia, and bilaterally hyperechoic kidneys with loss of corticomedullary distinction as reported by the referring veterinarian. Relevant initial laboratory results included severe non-regenerative normocytic hypochromic anemia, increased blood urea nitrogen, minimally concentrated urine, proteinuria, and an increased urine protein:creatinine ratio. Cytologic evaluation of a bone marrow aspirate revealed a markedly hypocellular marrow with abundant mucinous material. Gelatinous marrow transformation (GMT) was confirmed histologically by the presence of mucinous material in the bone marrow that stained positive for Alcian blue but negative for periodic acid-Schiff. The cat died despite repeated blood transfusions and supportive care. Gelatinous marrow transformation, immune complex-mediated membranoproliferative glomerulonephritis, and gastrointestinal hemorrhage were observed on autopsy and histology. It is likely that the development of GMT was secondary to chronic kidney disease (CKD) and that CKD, GMT, and gastrointestinal hemorrhage contributed to the cat's non-regenerative anemia.

Author Correction: RNA-seq of serial kidney biopsies obtained during progression of chronic kidney disease from dogs with X-linked hereditary nephropathy.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Clinicopathologic characteristics, pathology, and prognosis of 77 dogs with focal segmental glomerulosclerosis.

BACKGROUND: Focal segmental glomerulosclerosis (FSGS) is a common cause of nonimmune complex glomerulopathy and the prognosis and clinicopathologic findings associated with this condition have not been described in dogs. OBJECTIVE: To characterize the presentation and identify clinical factors associated with the survival of dogs with FSGS. ANIMALS: Seventy-seven dogs diagnosed with FSGS based on evaluation of renal biopsy samples submitted to the International Veterinary Renal Pathology Service. METHODS: Retrospective review of medical records of dogs biopsied for evaluation of proteinuria between January 2015 and May 2017. RESULTS: The incidence of FSGS among all dogs biopsied for proteinuria was 26%. Significantly more females (48; 62.3%) than males (29; 37.7%) were affected (P = .04). At the time of biopsy, median serum creatinine concentration (SCr) was 1.2 mg/dL (range, 0.3-8.7), median serum albumin concentration (Alb) was 2.8 g/dL (range, 1.1-4.6), median systolic blood pressure was 153.5 mm Hg (range, 95-260), and median urine protein : creatinine ratio was 5.9 (range, 1.4-22). Median survival time after biopsy was 258 days (range, 26-1003) for dogs that died from all causes (n = 32). Factors that were associated with a shorter survival time included SCr ≥ 2.1 mg/dL (P < .01) and Alb < 2 g/dL (P < .01). CONCLUSIONS AND CLINICAL IMPORTANCE: Most dogs with FSGS were female, and although commonly hypertensive, azotemia, severe hypoalbuminemia and ascites or edema were observed infrequently. Variables significantly associated with survival time were SCr and Alb.

Correlation of electrophoretic urine protein banding patterns with severity of renal damage in dogs with proteinuric chronic kidney disease.

BACKGROUND: Urine protein loss is common in dogs with chronic kidney disease (CKD). Currently available noninvasive means of evaluating CKD in dogs cannot accurately predict the severity of glomerular and tubulointerstitial damage. Electrophoretic analysis of urine proteins can indicate the compromised renal compartment (glomerular vs tubular), but extensive evaluation of protein banding pattern associations with histologic damage severity has not been performed in dogs. OBJECTIVES: We aimed to evaluate electrophoretic banding patterns as indicators of the presence and severity of glomerular and tubulointerstitial damage in dogs with naturally occurring, predominantly proteinuric CKD. METHODS: We performed a retrospective study using urine and renal tissue from 207 dogs with CKD. Urine protein banding patterns were correlated with histologic severity of renal damage. Sensitivity and specificity of banding patterns for the detection of glomerular and tubulointerstitial damage were determined. RESULTS: Banding patterns were 97% sensitive and 100% specific for the detection of glomerular damage and 90% sensitive and 100% specific for the detection of tubulointerstitial damage. Correlations between composite banding patterns and the severity of renal damage were strong, while glomerular banding patterns correlated moderately with glomerular damage severity, and tubular gel scores correlated weakly to moderately with the severity of tubulointerstitial damage. CONCLUSIONS AND CLINICAL IMPORTANCE: Urine protein banding patterns are useful for the detection of glomerular and tubulointerstitial damage in dogs with proteinuric CKD.

RNA-seq of serial kidney biopsies obtained during progression of chronic kidney disease from dogs with X-linked hereditary nephropathy.

Dogs with X-linked hereditary nephropathy (XLHN) have a glomerular basement membrane defect that leads to progressive juvenile-onset renal failure. Their disease is analogous to Alport syndrome in humans, and they also serve as a good model of progressive chronic kidney disease (CKD). However, the gene expression profile that affects progression in this disease has only been partially characterized. To help fill this gap, we used RNA sequencing to identify differentially expressed genes (DEGs), over-represented pathways, and upstream regulators that contribute to kidney disease progression. Total RNA from kidney biopsies was isolated at 3 clinical time points from 3 males with rapidly-progressing CKD, 3 males with slowly-progressing CKD, and 2 age-matched controls. We identified 70 DEGs by comparing rapid and slow groups at specific time points. Based on time course analysis, 1,947 DEGs were identified over the 3 time points revealing upregulation of inflammatory pathways: integrin signaling, T cell activation, and chemokine and cytokine signaling pathways. T cell infiltration was verified by immunohistochemistry. TGF-ß1 was identified as the primary upstream regulator. These results provide new insights into the underlying molecular mechanisms of disease progression in XLHN, and the identified DEGs can be potential biomarkers and therapeutic targets translatable to all CKDs.

Advances in the evaluation of canine renal disease.

Many recent advances in the evaluation of dogs with kidney disease have improved our diagnostic algorithms and have impacted our therapeutic strategies. Non-invasive techniques, such as urinary and serologic biomarker evaluation, can help a clinician diagnose and treat a patient that cannot undergo a renal biopsy for clinical or financial reasons. Some biomarkers might help localize the affected structure (glomerulus vs. tubule) and indicate the type or severity of injury present. Although more research is needed, studies indicate that some biomarkers (e.g. urine protein to creatinine ratio and urinary immunoglobulins) can be useful in predicting adverse outcomes. Importantly, the sensitivity and specificity of biomarkers for renal injury should be established and clinicians need to understand the limitations of these assays. If a renal biopsy is performed, then it should be evaluated by a specialty diagnostic service with expertise in nephropathology. A panel of special stains, immunofluorescence for the detection of immunoglobulins and complement factors, and transmission electron microscopy can be routinely employed in cases of glomerular disease. These advanced diagnostics can be used to detect immune deposits in order to definitively diagnose immune complex mediated glomerular disease. Integrating the results of biomarker assays and comprehensive renal biopsy evaluation, the clinician can make informed therapeutic decisions, such as whether or not to immunosuppress a patient.