The lipopolysaccharide model for the experimental induction of transient lameness and synovitis in Standardbred horses

An established lipopolysaccharide (LPS) model previously described in Warmbloods, was inconsistent in Standardbred horses, where lameness was not detected despite the presence of synovitis. The present study aimed to determine the dose of LPS from E. coli O55:B5 required to induce mild to moderate lameness following middle carpal joint injection in Standardbred horses and to quantitate the induced lameness over time, with and without anti-inflammatory pre-treatment. In a baseline trial, eight healthy, clinically sound Standardbred horses were used in a rule-based dose-escalation design trial, starting at a dose of 10 endotoxin units (EU). Lameness at trot was evaluated visually and quantitatively (using an inertial-sensor system and pressure plate analysis). Synovial fluid aspirates were analysed for total nucleated cell counts, total protein and prostaglandin E2 (PGE2). Following 2 months wash-out, the effective LPS-dose determined in the baseline trial was used to evaluate the effect of anti-inflammatory treatment. A mixed model for repeated measures with horse as random effect was used for analysis. After injection of 10 EU LPS, the desired degree of lameness was observed in the baseline trial, with maximal lameness at post-injection hour (PIH) 4, followed by a rapid decline and return to baseline by PIH 48. No lameness was observed following pre-treatment with meloxicam. In synovial fluid, PGE2 was significantly higher at PIH 8 and PIH 24 in the baseline trial compared with following meloxicam pre-treatment. In conclusion, injection of the middle carpal joint with 10 EU LPS consistently induces a transient lameness and synovitis in Standardbred horses

Human C-reactive protein aggravates osteoarthritis development in mice on a high-fat diet

Objective: C-reactive protein (CRP) levels can be elevated in osteoarthritis (OA) patients. In addition to indicating systemic inflammation, it is suggested that CRP itself can play a role in OA development. Obesity and metabolic syndrome are important risk factors for OA and also induce elevated CRP levels. Here we evaluated in a human CRP (hCRP)-transgenic mouse model whether CRP itself contributes to the development of ‘metabolic’ OA.Design: Metabolic OA was induced by feeding 12-week-old hCRP-transgenic males (hCRP-tg, n = 30) and wild-type littermates (n = 15) a 45 kcal% high-fat diet (HFD) for 38 weeks. Cartilage degradation, osteophytes and synovitis were graded on Safranin O-stained histological knee joint sections. Inflammatory status was assessed by plasma lipid profiling, flow cytometric analyses of blood immune cell populations and immunohistochemical staining of synovial macrophage subsets.Results: Male hCRP-tg mice showed aggravated OA severity and increased osteophytosis compared with their wild-type littermates. Both classical and non-classical monocytes showed increased expression of CCR2 and CD86 in hCRP-tg males. HFD-induced effects were evident for nearly all lipids measured and indicated a similar low-grade systemic inflammation for both genotypes. Synovitis scores and synovial macrophage subsets were similar in the two groups.Conclusions: Human CRP expression in a background of HFD-induced metabolic dysfunction resulted in the aggravation of OA through increased cartilage degeneration and osteophytosis. Increased recruitment of classical and non-classical monocytes might be a mechanism of action through which CRP is involved in aggravating this process. These findings suggest interventions selectively directed against CRP activity could ameliorate metabolic OA development

Apoptotic and immunological markers in idiopathic pulmonary fibrosis: evolving concepts of pathogenesis

Idiopathic pulmonary fibrosis (IPF) is a rare and devastating lung disease of unknown aetiology. Itis thought to be caused by repetitive damage to the epithelium and abnormal repair. The resulting fibrosis is defined by an overgrowth of fibroblasts and extracellular matrix deposition which results in fatal respiratory dysfunction. IPF mainly occurs in elderly white males and has an average survival time of less than 4 years. Currently, there is no simple test available to diagnose IPF. Clinical findings such as lung function tests and high resolution computed tomography (HRCT) are often considered sufficient, especially in typical cases of IPF. In other cases, lung biopsy will be needed. In many cases a bronchoalveolar lavage (BAL) will be performed to help exclude other diagnoses. The aim of this thesis was to find disease specific markers. In the search for molecular markers for diagnosis and prognosis, we focused especially on the immunological response to injury and on apoptosis, because of their emerging role in the pathogenesis of IPF. We hypothesized that the results might also contribute to new insights into the biological mechanisms involved in IPF. Genetic variations in TP53 and CDKN1A, the genes encoding p53 and p21, were associated with susceptibility to IPF and progression of the disease, in our cohort of 77 IPF patients. P53 and p21 are vital cell cycle regulators that influence apoptosis, cellular senescence and proliferation. Variations in these proteins could affect the damage and repair processes in the alveolar epithelium. Damaged epithelium is thought to send out signals that activated the immune system. Many inflammatory cytokines are elevated in IPF patients but their exact role in the disease is unknown. We measured levels of MRP14 and YKL-40 in serum and BAL fluid of IPF patients and in closely related interstitial lung diseases. These proteins are thought to be produced by macrophages, neutrophils and epithelial cells and may represent chronic inflammation and fibrosis. Both MRP14 and YKL-40 were elevated in IPF patients and were highest in diseases that are hallmarked by fibrosis. YKL-40 was significantly associated with the prognosis in IPF patients We also performed a meta-analysis of genetic variations in IL1RN that were determined in 5 IPF cohorts with a total of 302 IPF patients and 879 healthy controls. The VNTR*2 haploblock was significantly associated with IPF susceptibility and resulted in lower levels of the cytokine IL-1Ra. Our results suggest IL-1Ra plays a role in IPF pathogenesis. IL-1Ra is an inhibitor of the pro-inflammatory and pro-fibrotic cytokine IL-1. The findings in this thesis confirm that apoptosis and immune responses play a role in IPF susceptibility and progression. Our results suggest there may be a role for alternatively activated macrophages in the aetiology of IPF. These macrophages, also referred to as M2 macrophages, play a role in wound repair and fibrosis. They are thought to be elevated in IPF patients and may be associated with a poor prognosis. Further research is needed to identify the exact nature of the M2 macrophages and their role in IP