Sirilak Disatian Surachetpong

Plasma and platelet serotonin concentrations in healthy dogs and dogs with myxomatous mitral valve disease.

OBJECTIVES Serotonin has been implicated in canine myxomatous mitral valve disease (MMVD); however, the sources of serotonin have not been fully elucidated. This study compared the concentration of serotonin in plasma and platelets of normal healthy small breed dogs with predisposition to MMVD and dogs with naturally occurring MMVD. ANIMALS 43 small-breed client-owned dogs with an approximate weight of <10 kg and age of 6 years or above were divided into 2 groups: a healthy control group (n = 20) and a group with echocardiographic evidence of MMVD (n = 23). METHODS 5 ml samples of blood were collected. Plasma and platelets were separated by centrifugation and assayed for serotonin measured by enzyme linked immunosorbent assay (ELISA). RESULTS Median plasma serotonin concentration was not significantly different (p = 0.3630) between normal healthy dogs (3.7 ng/ml) and dogs with MMVD (4.3 ng/ml). Males had higher plasma serotonin concentration than females (4.7 and 2.9 ng/ml respectively, p = 0.0043). Platelet serotonin concentration was not different between healthy dogs and dogs with MMVD (128.6 ng/10⁹ platelets and 176.6 ng/10⁹ platelets respectively, p = 0.4575). Age, echocardiographic indices and platelet count showed no correlation with plasma or platelet serotonin concentration. CONCLUSIONS Circulating plasma serotonin is unlikely a major source of serotonin signaling in canine MMVD. Platelets could be a source of serotonin in canine MMVD through platelet adhesion to the mitral valve; however, the amount of serotonin stored in platelets of healthy dogs and dogs with MMVD is not different.

Apoptosis and abundance of Bcl-2 family and transforming growth factor β1 signaling proteins in canine myxomatous mitral valves.

OBJECTIVES To determine the percentage of cells undergoing apoptosis within canine myxomatous valves and to evaluate whether TGFβ1 can be implicated as an anti-apoptosic signal through the Bcl-2 family of signaling proteins. ANIMALS Post-mortem mitral valve leaflets harvested from 5 normal dogs, 5 dogs with early-stage myxomatous mitral valve disease (MMVD), and 5 dogs with late-stage MMVD. MATERIALS AND METHODS The number of cells expressing cleaved caspase-3, DNA fragmentation (TUNEL marker) and apoptotic bodies were evaluated as a measure of apoptosis. To evaluate the relationship between TGFβ1 signaling and apoptosis, the abundance of activated TGFβ1 signaling protein, phosphorylated Smad 2/3 (p-Smad 2/3), and Bcl-2 family proteins (pro-apoptotic Bax and anti-apoptotic Bcl-2) was determined by immunohistochemistry. RESULTS Cells in normal and both stages of MMVD expressed the TUNEL marker and cleaved caspase-3, but not apoptotic bodies. The percentage of TUNEL marker and cleaved caspase-3 positive nuclei was not significantly different between groups of dogs (p > 0.05). P-Smad 2/3 and Bax were more abundant in myxomatous mitral valves while Bcl-2 was less abundant. P-Smad 2/3 primarily increased in the atrialis layer and was abundantly increased only in late-stage MMVD. CONCLUSIONS These data suggest that interstitial cells in MMVD are in a pro-apoptotic condition; however, they do not execute apoptosis. Thus, apoptosis does not explain differences in cellular density in canine MMVD. TGFβ1 signaling through the canonical SMAD pathway is increased in myxomatous mitral valves, but does not apparently mediate interstitial cell apoptosis in canine MMVD.

Histopathological changes and apoptosis detection in canine myxomatous mitral valve disease using tissue microarray technique

Myxomatous mitral valve disease (MMVD) is the most common cardiac disease in dogs. The etiology of MMVD is currently unclear. The cell density has been found increasingly in myxomatous mitral valves. The proliferation is unlikely a major mechanism of increased cell density in diseased valves. Other mechanisms such as anti-apoptosis or cell migration may be involved. The objective of this study was to determine the apoptotic cells within MMVD valves by assessing the immunoreactivity of cleaved caspase-3 and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assay using tissue microarray technique. Fifteen normal and 40 MMVD valves were included in the study. TUNEL marker and cleaved caspase-3 expressions were significantly decreased in myxomatous valves compared to normal valves (p < 0.05). Apoptotic bodies were hardly seen in normal and diseased valves. These findings suggest a relationship between the anti-apoptotic mechanism and increased cellularity in canine MMVD.