Evaluating the TaqMan Jra-Genotyping Method for Rapidly Predicting the Presence of Anti-Jra Antibodies.

Background: The Jra antigen is a high-prevalence red blood cell (RBC) antigen. Reports on cases of fatal hemolytic disease of the fetus and newborn and acute hemolytic transfusion reactions suggest that antibodies against Jra (anti-Jra) have potential clinical significance. Identifying anti-Jra is challenging owing to a lack of commercially available antisera. We developed an alternative approach to rapidly predict the presence of anti-Jra using the TaqMan single-nucleotide polymorphism (SNP)-genotyping method. Methods: Residual peripheral blood samples from 10 patients suspected of having the anti-Jra were collected. Two samples with confirmed Jr(a-) RBCs and anti-Jra were used to validate the TaqMan genotyping assay by comparing the genotyping results with direct sequencing. The accuracy of the assay in predicting the presence of anti-Jra was verified through crossmatching with in-house Jr(a-) O+ RBCs. Results: The TaqMan-genotyping method was validated with two Jr(a-) RBC- and anti-Jra-confirmed samples that showed concordant Jra genotyping and direct sequencing results. Jra genotyping for the remaining samples and crossmatching the serum samples with in-house Jr(a-) O+ RBCs showed consistent results. Conclusions: We validated a rapid, simple, accurate, and cost-effective method for predicting the presence of anti-Jra using a TaqMan-based SNP-genotyping assay. Implementing this method in routine practice in clinical laboratories will assist in solving difficult problems regarding alloantibodies to high-prevalence RBC antigens and ultimately aid in providing safe and timely transfusions and proper patient care.

Effects of Cyclooxygenase-2 in Canine Melanoma-derived Extracellular Vesicles on Tumor Microenvironment In Vitro.

BACKGROUND/AIM: Tumor cell-derived extracellular vesicles (TEVs) promote tumor growth and metastasis; thus, they have drawn the attention of researchers. TEVs regulate the tumor microenvironment by facilitating crosstalk between immune and stromal cells. Macrophages are one of the key components involved in malignant behavior in melanomas. Generally, when activated, macrophages polarize into M1 (pro-inflammatory) or M2 (anti-inflammatory, pro-tumor) phenotypes. However, the role of canine melanoma-derived EVs in macrophage polarization is elusive. In this study, we aimed to analyze the pro- and anti-inflammatory cytokines that are common markers for M1 or M2 macrophages in vitro. MATERIALS AND METHODS: The analysis was performed under coculture conditions of canine melanoma-derived (LMeC) EVs with canine macrophages (DH82). Quantitative reverse transcription polymerase chain reaction, western blotting, and immunofluorescence were used. RESULTS: Canine melanoma-derived EVs polarized M1 macrophages (inducible nitric oxide synthase, tumor necrosis factor α) into M2 macrophages [cluster of differentiation (CD)206, interleukin-10] and cyclooxygenase-2 is a major factor in macrophage polarization in canine melanoma-derived EVs. Furthermore, we also found that melanoma-derived EVs induced the expression of angiogenic cytokines (vascular endothelial growth factor, transforming growth factor ß) in endothelial cells. CONCLUSION: Melanoma-derived EVs perform an immunomodulatory function and can be used as targets in anti-inflammatory treatment.

Isolated hypoaldosteronism managed by DOCP in a dog with chronic kidney disease and hypercortisolism.

A 13-year-old spayed female Schnauzer dog with chronic kidney disease (CKD; International Renal Interest Society stage 2, non-proteinuric, normotensive), diabetes mellitus, hypercortisolism and myxomatous mitral valve degeneration (American College of Veterinary Internal Medicine stage B2) presented with electrolyte imbalance that had progressed to hyperkalaemia and hyponatremia, with a sodium to potassium (Na:K) ratio of 19.6. Cortisol levels after the adrenocorticotropic hormone stimulation test were within the therapeutic range, but aldosterone levels were below the reference range; hence, isolated hypoaldosteronism was diagnosed. After administration of deoxycorticosterone pivalate (DOCP), the electrolyte imbalance improved with a Na:K ratio of 27.7. This is the first report of the management of isolated hypoaldosteronism and hypercortisolism using trilostane and DOCP in a dog. This case highlights the importance of recognizing isolated hypoaldosteronism after long-term treatment with trilostane in a canine patient with CKD.