BRAF mutations in thyroid tumors from an ethnically diverse group

BACKGROUND: The molecular etiology of thyroid carcinoma (TC) and other thyroid diseases which may present malignant precursor lesions is not fully explored yet. The purpose of this study was to estimate frequency, type and clinicopathological value of BRAF exon 15 mutations in different types of cancerous and non-cancerous thyroid lesions originating in an ethnically diverse population. METHODS: BRAF exon 15 was sequenced in 381 cases of thyroid lesions including Hashimoto´s thyroiditis, nodular goiters, hyperplastic nodules, follicular adenomas (FA), papillary TC (PTC), follicular variant PTC (FVPTC), microcarcinomas of PTC (micro PTC; tumor size ≤ 1 cm), follicular TC (FTC), and non-well differentiated TC (non-WDTC). RESULTS: We identified BRAF mutations in one of 69 FA, 72 of 115 (63%) PTC, seven of 42 (17%) FVPTC, 10 of 56 (18%) micro PTC, one of 17 (6%) FTC, and one of eight (13%) non-WDTC. Most of the cases showed the common V600E mutation. One case each of PTC, FVPTC, and FTC harbored a K601E mutation. A novel BRAF mutation was identified in a FA leading to deletion of threonine at codon 599 (p.T599del). A rare 3-base pair insertion was detected in a stage III PTC resulting in duplication of threonine at codon 599 (p.T599dup). Patients with PTC harboring no BRAF mutation (BRAF(wt)) were on average younger than those with a BRAF mutation (BRAF(mut)) in the PTC (36.6 years vs. 43.8 years). Older age (≥ 45 years) in patients with PTC was significantly associated with tumor size ≥ 4 cm (P = 0.018), vessel invasion (P = 0.004), and distant metastasis (P = 0.001). Lymph node (LN) involvement in PTC significantly correlated with tumor size (P = 0.044), and vessel invasion (P = 0.013). Of notice, taken the whole TC group, family history of thyroid disease positively correlated with capsular invasion (P = 0.025). CONCLUSIONS: Older age is manifold associated with unfavorable tumor markers in our series. The K601E identified in a PTC, FVPTC, and FTC seems to be more distributed among different histological types of TC than previously thought. The T599del is a yet undescribed mutation and the rare T599dup has not been reported as a mutation in PTC so far

Pro-atherogenic actions of signal transducer and activator of transcription 1 serine 727 phosphorylation in LDL receptor deficient mice via modulation of plaque inflammation

Atherosclerosis is a chronic inflammatory disorder of the vasculature regulated by cytokines. We have previously shown that extracellular signal-regulated kinase-1/2 (ERK1/2) plays an important role in serine 727 phosphorylation of signal transducer and activator of transcription-1 (STAT1) transactivation domain, which is required for maximal interferon-γ signaling, and the regulation of modified LDL uptake by macrophages in vitro. Unfortunately, the roles of ERK1/2 and STAT1 serine 727 phosphorylation in atherosclerosis are poorly understood and were investigated using ERK1 deficient mice (ERK2 knockout mice die in utero) and STAT1 knock-in mice (serine 727 replaced by alanine; STAT1 S727A). Mouse Atherosclerosis RT² Profiler PCR Array analysis showed that ERK1 deficiency and STAT1 S727A modification produced significant changes in the expression of 18 and 49 genes, respectively, in bone marrow-derived macrophages, with 17 common regulated genes that included those that play key roles in inflammation and cell migration. Indeed, ERK1 deficiency and STAT1 S727A modification attenuated chemokine-driven migration of macrophages with the former also impacting proliferation and the latter phagocytosis. In LDL receptor deficient mice fed a high fat diet, both ERK1 deficiency and STAT1 S727A modification produced significant reduction in plaque lipid content, albeit at different time points. The STAT1 S727A modification additionally caused a significant reduction in plaque content of macrophages and CD3 T cells and diet-induced cardiac hypertrophy index. In addition, there was a significant increase in plasma IL-2 levels and a trend toward increase in plasma IL-5 levels. These studies demonstrate important roles of STAT1 S727 phosphorylation in particular in the regulation of atherosclerosis-associated macrophage processes in vitro together with plaque lipid content and inflammation in vivo, and support further assessment of its therapeutical potential

The Lab4P consortium of probiotics attenuates atherosclerosis in LDL receptor deficient mice fed a high fat diet and causes plaque stabilization by inhibiting inflammation and several pro-atherogenic processes

Scope Previous studies showed that Lab4 probiotic consortium plus Lactobacillus plantarum CUL66 (Lab4P) reduced diet-induced weight gain and plasma cholesterol levels in C57BL/6J mice fed a high fat diet (HFD). The effect of Lab4P on atherosclerosis is not known and was therefore investigated. Methods and results Atherosclerosis-associated parameters were analyzed in LDL receptor deficient mice fed HFD for 12 weeks alone or supplemented with Lab4P. Lab4P increased plasma HDL and triglyceride levels and decreased LDL/VLDL levels. Lab4P also reduced plaque burden and content of lipids and macrophages, indicative of dampened inflammation, and increased smooth muscle cell content, a marker of plaque stabilization. Atherosclerosis arrays showed that Lab4P altered the liver expression of 19 key disease-associated genes. Lab4P also decreased the frequency of macrophages and T-cells in the bone marrow. In vitro assays using conditioned media from probiotic bacteria demonstrated attenuation of several atherosclerosis-associated processes in vitro such as chemokine-driven monocytic migration, proliferation of monocytes and macrophages, foam cell formation and associated changes in expression of key genes, and proliferation and migration of vascular smooth muscle cells. Conclusion This study provides new insights into the anti-atherogenic actions of Lab4P together with the underlying mechanisms and supports further assessments in human trials

Anti-atherogenic actions of hydroxytyrosol

Introduction: Atherosclerosis is a chronic inflammatory disorder characterised by the accumulation of lipids in the arterial wall, and is considered to be a major contributor to cardiovascular disease (CVD). Worldwide, CVD is responsible for a third of all deaths. Current pharmacological therapeutic agents for CVD, such as statin therapy, are not fully effective and have considerable residual risk for the disease. Alternative agents for the prevention and treatment of the disease are therefore required. Hydroxytyrosol (HT) is a polyphenol compound found mainly in olive oil. HT has been reported to prevent CVD predominantly due to its antioxidant effects. HT is the only polyphenol recognised by the European Food Safety Authority as a protector against low-density lipoprotein-mediated oxidative damage. Previous investigations carried out in the host laboratory showed that HT altered multiple atherosclerosis-associated risk factors in wild-type mice fed a high-fat diet (HFD) for 3 weeks, as well as various anti-inflammatory and anti-atherogenic actions on human monocytes/macrophages in vitro. Unfortunately, the actions of HT in atherosclerosis in vivo are poorly understood. The aim of this study therefore was to elucidate its effects on atherosclerosis progression and regression in a mouse model system. Methods: In order to study the development of atherosclerosis in vivo, 8-week-old male or female low-density lipoprotein receptor-deficient (ldlr-/-) mice were given HFD alone or in combination with 10 mg/kg/day of HT for 12 weeks. For regression studies, the mice were fed a HFD for 12 weeks to promote the formation of established lesions, and then switched to normal chow diet (NCD) alone or in combination with HT. The two procedures were then followed up by an in-depth investigations of atherosclerosisassociated risk factors and the plaques that formed in the aortic root. RNA-sequencing and bioinformatic analyses was used to assess changes in gene expression and associated pathways in the thoracic aorta. The liver was also analysed in relation to non-alcoholic fatty liver disease (NAFLD) that is often associated with atherosclerosis and extended via the use of an in vitro hepatoma HepG2 cell culture model system. Results: As part of the progression study, female ldlr-/- mice that had received HT supplemented HFD for 12 weeks had attenuated weight gain, plaque size in the aortic root and neutrophil content in the peripheral blood, while male ldlr-/- mice had attenuated occlusion and T cells in the peripheral blood. In both cases, there was reduced plaque inflammation and improved plasma lipid profile and plaque stability. As part of the regression studies on male ldlr-/- mice, intervention with HT combined with NCD reversed hepatic injury and enhanced plaque stability (to a greater extent than NCD intervention alone). Conclusions: These findings provide support for the anti-atherogenic actions of HT as well as its possible use as an alternative nutraceutical agent for preventing the development of atherosclerosis. This is potentially possible due to the lack of adverse effects as well as the relatively low cost in comparison to that of typical pharmacological treatments. Additional research is necessary in order to determine the mechanisms that are responsible for these favourable anti-atherogenic and other beneficial changes

Monitoring cellularity and expression of key in atherosclerotic plaques

Atherosclerotic plaques are highly diverse and heterogeneous structures, even within the same individual, and can vary depending on its anatomical location within the vascular bed. Early in the disease and throughout its progression, immune cells infiltrate the lesion, contributing to the plaque phenotype via different mechanisms. Detailed characterization of constituent cell populations within plaques is hence required for more accurate assessment of disease severity and inflammatory burden. A wide range of fluorophore-conjugated antibodies targeted to key cell types implicated in all stages of the disease are commercially available, enabling visualization of the dynamic cellular landscape present within lesions. This chapter describes the use of immunofluorescence staining of atherosclerotic plaque sections to study plaque cellularity and expression of key markers

Evaluation of plaque burden and in atherosclerotic plaques

Atherosclerosis is a chronic inflammatory disease characterized by the formation of lipid-rich, fibrous plaques within the arterial wall of medium and large arteries. Plaques prone to rupture are typically rich in lipids and pro-inflammatory markers. Cells within the plaque can take up lipids via different mechanisms leading to the formation and accumulation of lipid-rich foam cells, a key hallmark of the disease. Evaluation of plaque burden and lipid content is hence important to determine disease progression and severity. This chapter describes the most commonly used staining methods that enable visualization and analysis of mouse atherosclerotic plaques. These methods include en face preparation of mouse aorta, and staining sections of arteries using hematoxylin and eosin, Oil Red O, and Masson’s Trichrome

Survey of approaches for investigation of atherosclerosis in vivo

Although in vitro model systems are useful for investigation of atherosclerosis-associated processes, they represent simplification of complex events that occur in vivo, which involve interactions between many different cell types together with their environment. The use of animal model systems is important for more in-depth insights of the molecular mechanisms underlying atherosclerosis and for identifying potential targets for agents that can prevent plaque formation and even reverse existing disease. This chapter will provide a survey of such animal models and associated techniques that are routinely used for research of atherosclerosis in vivo

Epidemiology of Healthcare-Associated Infections and Adherence to the HAI Prevention Strategies

Healthcare-associated infections are widely considered one of the most common unfavorable outcomes of healthcare delivery. Ventilator-associated pneumonia, central line-associated bloodstream infections, and catheter-associated urinary tract infections are examples of healthcare-associated infections. The current study was a retrospective study conducted at a public hospital in Unaizah, Saudi Arabia, to investigate the frequency of healthcare-associated illnesses and adherence to healthcare-associated infection prevention techniques in the year 2021. Surgical site infections occurred at a rate of 0.1%. The average number of catheter-associated urinary tract infections per 1000 catheter days was 0.76. The average number of central line-associated bloodstream infections per 1000 central line days was 2.6. The rate of ventilator-associated pneumonia was 1.1 per 1000 ventilator days on average. The average number of infections caused by multidrug-resistant organisms per 1000 patient days was 2.8. Compliance rates were 94%, 100%, 99%, and 76% for ventilator-associated pneumonia, central line-associated bloodstream infections, catheter-associated urinary tract infections, and hand hygiene bundles, respectively. It is critical to participate in more educational events and workshops, particularly those that emphasize hand cleanliness and personal safety equipment