Spray dried lipid nanoparticle formulations enable intratracheal delivery of mRNA.

RNA therapies have recently taken a giant leap forward with the approval of Onpattro™, a siRNA therapy delivered using a lipid nanoparticle (LNP), and the LNP-enabled mRNA vaccines against COVID-19, which are the first mRNA drugs to reach the marketplace. The latter medicines have illustrated that stability is a significant challenge in the distribution of RNA drugs using non-viral delivery systems, particularly in areas without cold chain storage. Here, we describe a proof-of-concept study on the engineering of an LNP mRNA formulation suitable for spray drying. This process produced a dry powder formulation that maintained stability and preserved mRNA functionality with increased performance compared to liquid formulations stored two weeks at 4 °C. Intratracheal delivery of spray dried LNPs loaded with eGFP mRNA to rats resulted in the production of the eGFP protein in a range of cell types including bronchiolar epithelial cells, macrophages and type II pneumocytes; cell types involved in adaptive immunity and which would be valuable targets for inhaled vaccines against respiratory pathogens. Together, these data show that spray drying of LNPs enhances their stability and may enable RNA delivery to the lung for protein replacement therapy, gene editing, vaccination, and beyond.

Falciform fat:femur length ratio provides a novel method for objective postmortem estimation of total body fat in overweight and obese cats.

Determination of the nutritional condition, including estimation of amounts of total body fat (tBF), at routine postmortem examination of cats is typically based on subjective visual assessment. Subjective assessment may result in uncertainties regarding degree of overweight, and objective methods that provide a numerical value reflecting the tBF could be valuable to accurately judge excess body fat. We investigated if the falciform fat pad weight (FFPW) was correlated to tBF and could be used to detect overweight and obesity in cats. The FFPW and the femur length (FL) were recorded at postmortem examination in 54 cats and the FFPW:FL ratio (FFR) calculated. Each cat was additionally assigned to a fat category (FC) according to subjective assessment. Computed tomography was used to determine tBF as the body fat percentage (%BF), the body fat volume (BFV), and BFV normalized to animal size (nBFV) in 39 cats. There was strong correlation between the FFPW and the BFV (r = 0.888) and between the FFR and the nBFV (r = 0.897). The correlation between the nBFV and %BF was very strong (r = 0.974). Using a lower FFR cutoff value of 3.5 for obesity and 1.6 for overweight, there was a discrepancy in FC between using the FFR and subjective assessment in 6 of 54 cats (11%). We conclude that the FFPW increases proportionally with tBF and that the FFR provides a method for objective tBF estimation. We suggest introducing the FFR to feline postmortem examination protocols as an objective estimate of tBF.

Computed tomography is superior to radiography for detection of feline elbow osteoarthritis.

Elbow osteoarthritis (OA) is common in cats and radiography is typically used for diagnosis. However computed tomography (CT), with its multiplanar three-dimensional characteristics, could have significant advantages for assessment of OA compared to radiography, particularly early in the disease process. The study objectives were to compare radiography and CT to histologic OA changes, investigate the stage of OA that radiography and CT detect, and search for specific changes in CT images strongly predictive for feline elbow OA. Right elbows from 29 cats were evaluated by radiography and CT, and articular cartilage lesions graded histologically and macroscopically. Three further joints were sampled to specifically evaluate the morphology of the anconeal process. Macroscopic, radiographic and CT OA diagnosis were compared to the reference standard histologic OA that was divided into mild, moderate and severe. Osteophytic spurs on the lateral margin of the anconeal process could be reliably measured in CT images (intra-class correlation 0.79) and when ≥0.5 mm had high sensitivity for moderate/severe histologic OA, moderate sensitivity for mild histologic OA and high specificity for all stages of OA. In moderate/severe histologic OA both radiography and CT subjective OA diagnosis had moderate to very high sensitivity. However, in mild histologic OA CT grading had low sensitivity and radiography did not detect OA. In conclusion, CT of the feline elbow including measurement of osteophytes on the anconeal process lateral margin is superior to radiography for OA detection and should be considered for OA diagnosis, particularly when mild OA changes are of interest.

Acetabular margin changes in feline hip joints - Implications for radiologic diagnosis and development of osteoarthritis.

The development and early morphological features of feline hip osteoarthritis (OA) are largely unknown. Tears in the acetabular labrum and at the chondrolabral transition zone are suggested to be important in the pathogenesis of human hip OA, but in cats such lesions have not been described. We investigated associations between computed tomography (CT)-detected joint changes and microscopic articular cartilage lesions, the distribution of detected changes, and histologically evaluated the acetabular margin (AM) in hip joints from 20 cats. Histologic evaluation was undertaken on at least one joint from each cat. CT-detected joint changes and articular cartilage lesions were graded and the histological appearance of CT-detected osteophytes evaluated. The majority of CT-detected lesions and cartilage lesions were mild. Whole-joint CT scores and AM osteophyte CT scores showed moderate to strong correlation with cartilage scores. The odds were higher for presence of CT-detected osteophytes in craniodorsal, cranial, cranioventral, ventral and dorsal AM regions. Peripheral acetabular regions showed higher cartilage lesion grades than central regions. Tears, seen as fissures/clefts, in labral and perilabral tissues were common. CT-detected AM osteophytes morphologically presented as pointed sclerotic bone, spur-shaped bone or rounded chondro-osteophytes. The results suggest that CT is a valuable tool for diagnosing early feline hip OA. CT-detected osteophytes showed variable histologic morphologies, which may implicate different disease mechanisms and/or disease stages. Tears in the AM could represent an early event in feline hip OA and this warrants further investigation.

Cardiac Anomalies in Two White-Tailed Eagles (Haliaeetus albicilla) in Sweden.

Severe cardiomegaly with an atrial septal defect was discovered during necropsy of a subadult White-tailed Eagle (Haliaeetus albicilla) found dead in the wild. A thin membrane composed of fibromuscular tissue separated the left atrium into two chambers, most consistent with that described for cor triatriatum sinister (CTS) in other species. Seventeen months later, necropsy of an adult White-tailed Eagle again revealed CTS. This lesion has not been reported previously in raptors.

Thoracolumbar meningeal fibrosis in pugs.

BACKGROUND: Thoracolumbar myelopathies associated with spinal cord and vertebral column lesions, with a similar clinical phenotype, but different underlying etiologies, occur in pugs. OBJECTIVES: To further characterize the clinical and neuropathological characteristics of pugs with longstanding thoracolumbar myelopathy. ANIMALS: Thirty client-owned pure-bred pugs with a history of more than a month of ataxia and paresis of the pelvic limbs, suggesting a myelopathy localized to the thoracolumbar spinal cord, were included in the study. METHODS: Prospective clinicopathological study. Included pugs underwent a complete neurological examination and gross and histopathologic postmortem studies with focus on the spinal cord. Computed tomography (n = 18), magnetic resonance imaging (n = 17), and cerebrospinal fluid analysis (n = 27) were performed before or immediately after death. RESULTS: Twenty male and 10 female pugs had a median age at clinical onset of 84 months (interquartile range, 66-96). Affected pugs presented with a progressive clinical course and 80% were incontinent. There was circumferential meningeal fibrosis with concomitant focal, malacic, destruction of the neuroparenchyma in the thoracolumbar spinal cord in 24/30 pugs. Vertebral lesions accompanied the focal spinal cord lesion, and there was lympho-histiocytic inflammation associated or not to the parenchymal lesion in 43% of the pugs. CONCLUSIONS AND CLINICAL IMPORTANCE: Meningeal fibrosis with associated focal spinal cord destruction and neighboring vertebral column lesions were common findings in pugs with long-standing thoracolumbar myelopathy.

Cartilage lesions in feline stifle joints - Associations with articular mineralizations and implications for osteoarthritis.

Feline stifle osteoarthritis (OA) is common, however little is known about the early stages of the disease. Furthermore, the importance of small articular mineralizations (AMs) in feline stifle OA is controversial. This study aimed to describe microscopic articular cartilage lesions and to investigate associations between cartilage lesions and AMs, synovitis, osteochondral junction findings and subchondral bone sclerosis. Stifles of 29 cats, aged 1-23years and euthanized for reasons other than stifle disease, were examined. Osteochondral tissue and synovial membrane changes were histologically evaluated. Computed tomography and radiography were used for evaluation of AMs. Global cartilage scores (GCS, n=28) were summarized and joints assigned a histologic OA grade. Minimal to mild histologic OA was seen in 24/28 joints. In 27/29 joints tibial cartilage lesions were seen, whereas femoral lesions were only seen in two joints. Articular mineralizations were detected in 13/29 joints, 11 were small and 12 were located entirely within the medial meniscus. There was no association between GCS and presence or volumes of AMs. However, higher GCS was associated with synovitis (P=0.001) and age (P<0.0005). Presence of subchondral bone sclerosis (P<0.0005) and disruption of the calcified cartilage or tidemark (P<0.0005) were associated with cartilage lesions. We conclude that the tibial articular cartilage is a common location for histologic OA lesions in cats. Synovitis and changes in the subchondral bone and calcified cartilage may be important in the pathogenesis of feline stifle OA, whereas small AMs likely represent incidental findings.