Regenerative medicine approach to reconstruction of the equine upper airway.

Airway obstruction is a common cause of poor performance in horses. Structural abnormalities (insufficient length, rigidity) can be a cause for the obstruction. Currently, there are a few effective clinical options for reconstruction of the equine larynx. A regenerative medicine approach to reconstruction may provide the capability to stabilize laryngeal structures and to encourage restoration of site-appropriate, functional, and host-derived tissue. The purpose of this study was the histopathological evaluation of (1) decellularization of equine (horse) laryngeal cartilages (epiglottis and arytenoids); (2) the host response to decellularized laryngeal cartilages implanted subcutaneously in a donkey model as a test of biocompatibility; and (3) the use of decellularized laryngeal cartilages in a clinically relevant pilot study in the horse larynx. Equine laryngeal cartilages were found to be sufficiently decellularized and were subsequently implanted subcutaneously in donkeys to test biocompatibility. After 4 weeks, the implanted cartilage was harvested. In the subcutaneous model, the samples did not elicit a rejection or foreign body type reaction and were judged suitable for implantation in a clinically relevant equine model. Implants were placed in the upper airway (arytenoids and epiglottis) of one horse. At 4 weeks, the implants were observed to remodel rapidly and were replaced by dense connective tissue with signs of new hyaline cartilage formation in the arytenoids and by connective tissue containing glandular structures and an epithelial covering in the epiglottis. The results of the present study demonstrate the feasibility of a scaffold-based regenerative medicine approach to reconstruction of the equine upper airway; however, further studies investigating long-term integration, formation of new cartilage, and mechanical properties are needed.

Kudoa thunni from blackfin tuna (Thunnus atlanticus) harvested off the island of St. Kitts, West Indies.

Numerous myxozoan cysts (∼ 1 mm) were found in the musculature of blackfin tuna (Thunnus atlanticus) harvested off the Caribbean island of St. Kitts. Myxospores were consistent with quadrate members of the Kudoidae, measuring 8.8 (8.2-9.4) µm wide, 7.3 (6.6-8.3) µm thick, and 6.2 (5.8-6.9) µm long with 4 uniform drop-like polar capsules measuring 2.7 (2.2-3.2) µm long and 2.0 (1.7-2.2) µm wide. The 18S small-subunit (SSU) and 28S large-subunit (LSU) ribosomal DNA sequences did not result in direct matches to any published sequences. However, the SSU sequences (1,786 base pairs [bp]) obtained from 6 individual cysts were identical and demonstrated high homology to Kudoa thunni (99.0%) from albacore (Thunnus alalunga). Alternatively, 33 unique sequences were obtained for the LSU (∼ 800 bp), demonstrating 0.1 to 5.0% variability between them, although a majority of these sequences (60%) demonstrated high homology (>99%) to K. thunni. Morphologically, the case isolate was smaller than published descriptions of K. thunni; however, rDNA sequence homology, and phylogenetic placement based on concatenated SSU and LSU rDNA sequences suggests this case isolate and K. thunni are conspecific. To our knowledge this is the first report of K. thunni infection in blackfin tuna from the Caribbean.

Digital image analysis of rhodanine-stained liver biopsy specimens for calculation of hepatic copper concentrations in dogs.

OBJECTIVE: To evaluate the accuracy of digitally scanned rhodanine-stained liver biopsy specimens for determination of hepatic copper concentration and compare results with qualitatively assigned histologic copper scores in dogs. SAMPLE: 353 liver biopsy specimens from dogs. PROCEDURES: Specimens (n = 139) with quantified copper concentration ranging from 93 to 6,900 µg/g were allocated to group 1 (< 400 µg/g [37]), group 2 (401 to 1,000 µg/g [27]), group 3 (1,001 to 2,000 µg/g [34]), and group 4 (> 2,001 µg/g [41]); stained with rhodanine; and digitally scanned and analyzed with a proprietary positive pixel algorithm. Measured versus calculated copper concentrations were compared, and limits of agreement determined. Influence of nodular remodeling, fibrosis, or parenchymal loss on copper concentration was determined by digitally analyzing selected regions in 17 specimens. After method validation, 214 additional liver specimens underwent digital scanning for copper concentration determination. All sections (n = 353) were then independently scored by 2 naive evaluators with a qualitative scoring schema. Agreement between assigned scores and between assigned scores and tissue copper concentrations was determined. RESULTS: Linear regression was used to develop a formula for calculating hepatic copper concentration ≥ 400 µg/g from scanned sections. Copper concentrations in unremodeled specimens were significantly higher than in remodeled specimens. Qualitative scores widely overlapped among quantitative copper concentration groups. CONCLUSIONS AND CLINICAL RELEVANCE: Calculated copper concentrations determined by means of digital scanning of rhodanine-stained liver sections were highly correlated with measured values and more accurate than qualitative copper scores, which should improve diagnostic usefulness of hepatic copper concentrations and assessments in sequential biopsy specimens.

Molecular and immunohistochemical diagnosis of Francisella noatunensis subsp. orientalis from formalin-fixed, paraffin-embedded tissues.

Members of the genus Francisella (viz., F. noatunensis subsp. orientalis [Fno] and F. noatunensis subsp. noatunensis) have been described as causative agents of chronic granulomatous and pyogranulomatous lesions in wild and cultured fish species. In the present study, 68 archived formalin-fixed, paraffin-embedded (FFPE) tissues from several fish species, collected at different geographical locations from 2000 to 2011, were analyzed using a real-time polymerase chain reaction assay for the detection of the Fno intracellular growth loci C (iglC) gene and by immunohistochemistry for the demonstration of Fno antigens. The results revealed a high correlation between these 2 diagnostic techniques validating their use for the diagnosis of Fno infection in archived FFPE tissues and confirming the presence of Fno in fish species from the Cari y years of the present century.