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Evaluation of opioid and TLR-4 receptors in the mechanism of opioid effects on heart muscle cells
Biriczová, Lilla ; Novotný, Jiří (advisor) ; Alánová, Petra (referee)
It has been reported that opioid receptor activation mimics ischemic preconditioning, which may protect the heart from the development of infarction. Toll-like receptor 4 (TLR-4) during infarction stimulates cytokine production leading to inflammation and injury of the heart tissue. Our aim was to study the effect of morphine in vitro on the viability and oxidative state of H9c2 cells (rat cardiomyoblasts) and the role of TLR-4 during oxidative stress. Our experiments showed that pretreatment with morphine before tert-butylhydroperoxide (t-BHP)-, 2,2'-bipyridyl (BP)- and lipopolysaccharide (LPS)-induced oxidative stess had protective effect on the viability of H9c2 cells and markedly reduced the production of reactive oxygen species (ROS). The protective effect of morphine was diminished after naloxone treatment, which confirms the role of opioid receptors in preconditioning. TLR-4 inhibition by TAK-242 pretreatment and silencing TLR-4 by RNA interference resulted in a partial increase in cell viability but significant attenuation of ROS production after t-BHP and BP treatment. The action of LPS was reduced in response to TLR-4 silencing. Interestingly, naloxone pretreatment and suppression of TLR-4 markedly alleviated oxidative stress and resulted in a significant improvement of cell viability. We...
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Beta-adrenergic receptors and their desensitization
Biriczová, Lilla ; Novotný, Jiří (advisor) ; Kolář, David (referee)
β-Adrenergic receptors (β-ARs) are G-protein-coupled receptors (GPCR), widely present in the animal organism and mediate catecholamine pathways leading to diverse physiological responses. The family of β-ARs consists of β1-AR, β2-AR and β3-AR, which are distinguished by their affinity to adrenaline and noradrenaline. A typical model of β-AR signalling includes binding of the ligand, G-protein coupling, activation of adenylyl cyclase (AC) resulting in production of the second messenger cAMP and activation of protein kinase A (PKA) that phosphorylates downstream proteins leading to physiological responses. Beacause excessive catecholamine signalling can cause undesirable consequences, a mechanism has evolved, which attenuates the function of β-ARs in spite of further stimulation, so called desensitization. The classic course of desensitization consists of characteristic steps including phosphorylation of the receptor, β-arrestin attachement and uncoupling of the G-protein from β-AR. Restoration of the signalling ability is allowed through resensitization of β-AR when the receptor is sequestrated and dephosphorylated. Given that β-ARs are structurally and genetically different, it is reasonable to consider that each step of the desenstization process may happen differently among the different subtypes...
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