Binding of lipopeptide to CD14 induces physical proximity of CD14, TLR2 and TLR1

Lipoproteins or lipopeptides (LP) are bacterial cell wall components detected by the innate immune system. For LP, it has been shown that TLR2 is the essential receptor in cellular activation. However, molecular mechanisms of LP recognition are not yet clear. We used a FLAG-labeled derivative of the synthetic lipopeptide N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2R,S)-propyl]-(R)-cysteinyl-seryl-(lysyl)(3)-lysine (Pam(3)CSK(4)) to study the roles of CD14, TLR2 and TLR1 in binding and signaling of LP and their molecular interactions in human cells. The activity of Pam(3)CSK(4)-FLAG was TLR2 dependent, whereas the binding was enabled by CD14, as evaluated by flow cytometry and confocal microscopy. Using FRET and FRAP imaging techniques to study molecular associations, we could show that after Pam(3)CSK(4)-FLAG binding, CD14 and Pam(3)CSK(4)-FLAG associate with TLR2 and TLR1, and TLR2 is targeted to a low-mobility complex. Thus, LP binding to CD14 is the first step in the LP recognition, inducing physical proximity of CD14 and LP with TLR2/TLR1 and formation of the TLR2 signaling complex.     

Effectiveness of diaphragmatic stimulation with single-channel electrodes in rabbits

Every year, a large number of individuals become dependent on mechanical ventilation because of a loss of diaphragm function. The most common causes are cervical spinal trauma and neuromuscular diseases. We have developed an experimental model to evaluate the performance of electrical stimulation of the diaphragm in rabbits using single-channel electrodes implanted directly into the muscle. Various current intensities (10, 16, 20, and 26 mA) produced tidal volumes above the baseline value, showing that this model is effective for the study of diaphragm performance at different levels of electrical stimulation     

Współwystępowanie dwóch rzadkich chorób genetycznych: fenyloketonurii oraz zespołu Pradera i Williego. Opis przypadku

Phenylketonuria (PKU) is an autosomal recessive metabolic disease caused by an error of phenylalanine metabolism. Delayed treatment or treatment performed unsystematically might lead to neurological disorders and progressive intellectual disability. In the Prader-Willi syndrome (PWS) clinical manifestations change with age. Feeding difficulties resulting from the poor suck and hypotonia are typical in the neonatal period and early infancy. As patients grow their activities increase, muscle tone improves, the extreme hyperphagia appears – the main cause of obesity. Mental development of patients is usually mildly retarded. We report the case of a patient affected by two genetic diseases: PKU and PWS. Variety of clinical symptoms and abnormal results of laboratory analyses make the correct diagnosis difficult. Elevated phenylalanine level in screening newborn test, disorders of muscle tone, poor suck, and low urine biopterin concentration did not allow to clearly rule out the hyperphenylalaninemia caused by a tetrahydrobiopterin deficiency (BH₄). Data from obstetric anamnesis and the early postnatal clinical findings suggested the PWS. Hyperphagia and the increased risk of carbohydrates tolerance disorders in patients with PWS make it very difficult to balance properly the low-phenylanine diet necessary in PKU treatment. The patient presented by us needs well-coordinated multidisciplinary medical care which aims to provide proper physical development and to support the boy's mental potential as well as appropriate functioning in the society.     

Contemporary use of P2Y12 inhibitors in patients with ST-segment elevation myocardial infarction referred to primary percutaneous coronary interventions in Poland: Data from ORPKI national registry

Journal of Thrombosis and Thrombolysis - According to guidelines, it is recommended to give P2Y12 inhibitors (preferably ticagrelor or prasugrel) at the time of first medical contact in patients...     

Correction to: Henoch-Schönlein purpura nephritis: initial risk factors and outcomes in a Latin American tertiary center

Clinical Rheumatology - One of the author’s name on this article was incorrectly spelled as “Sylvia C. L. Fahrat” . The correct spelling is “Sylvia C. L. Farhat” and...     

Role of EP3 and EP4 prostaglandin receptors in reorganization of the cytoskeleton in mature human osteoclasts

OBJECTIVE: Osteoclasts are central to the pathophysiology of several bone diseases. Prostaglandin E2 (PGE2) is well known to influence osteoclasts indirectly, but its direct action on osteoclasts is still controversial and the relevant receptors are unknown. We investigated the distribution and function of EP receptors in human mature osteoclasts. METHODS: Osteoclasts were extracted from femurs and tibias of human fetuses obtained from legal abortions. In situ hybridization and immunohistochemistry were used to detect the presence of EP1, EP2, EP3, and EP4 receptors on these cells. Actin staining and fluorescent microscopy were used to detect the effects of receptor activation on the cytoskeleton. RESULTS: Only EP3 and EP4 receptors were detected at the RNA and protein level in osteoclasts. These receptors were functional: PGE2 decreased the number of osteoclasts presenting an actin ring; 11-deoxy-PGE1, an EP2 and EP4 agonist, also decreased the number of tartrate-resistant acid phosphatase-positive cells with an actin ring; sulprostone, an EP3-specific agonist, had no effect on this variable but increased the number of cells with lamellipodia. CONCLUSION: Mature human osteoclasts present 2 subtypes of EP receptors, namely EP3 and EP4, that mediate different actions of PGE2 on these cells: activation of the EP4 receptors inhibits actin ring formation and activation of the EP3 receptors increases the number of lamellipodia. Activation or inhibition of these receptors by specific agents could be used to study and influence osteoclast function.     

Gentamicin affects melanogenesis in normal human melanocytes

Background: Aminoglycoside antibiotics, including gentamicin, despite their ability to induce adverse effects on pigmented tissues, remain valuable and sometimes indispensable for the treatment of various infections. It is known that gentamicin binds to melanin biopolymers, but the relation between this drug affinity to melanin and its toxicity is not well documented. The aim of this work was to examine the impact of gentamicin on viability and melanogenesis in HEMa-LP (light pigmented) and HEMn-DP (dark pigmented) normal human melanocytes.

Methodology/principal findings: The effect of gentamicin on cell viability was determined by 4-[3-(4-iodophenyl)-2-(4-nitrophenyl)-2H-5-tetrazolio]-1,3-benzene disulfonate (WST-1) assay; melanin content and tyrosinase activity were measured spectrophotometrically. It has been demonstrated that gentamicin induces concentration-dependent loss in melanocytes viability. The application of antibiotic in concentration of 10?mM causes higher reduction in viability of the light pigmented melanocytes (by about 74%) when compared with the dark pigmented ones (by about 62%). The value of the concentration of a drug that produces loss in cell viability by 50% (EC₅₀) for both cell lines was found to be ～7.5?mM. It has been shown that gentamicin causes inhibition of tyrosinase activity and reduces melanin content in light pigmented melanocytes significantly more than in the dark pigmented cells.

Conclusion/significance: We have found that gentamicin modulates melanization process in melanocytes in vitro, what may explain the potential role of melanin biopolymer in the mechanisms of undesirable toxic effects of this drug in vivo, as a result of its accumulation in pigmented tissues. We have also stated that the melanogenesis process in light pigmented melanocytes is more sensitive to the inhibitory effect of gentamicin than in the dark pigmented cells.