Afatinib triggers a Ni2+‐resistant Ca2+ influx pathway in A549 non‐small cell lung cancer cells

alexandrossfakianakis shared this article with you from Inoreader Afatinib triggers a Ni2+‐resistant Ca2+ influx pathway in A549 non‐small cell lung cancer cells via Fundamental & Clinical Pharmacology Abstract Afatinib is used to treat non-small cell lung cancer cells (NSCLC), and its mechanism involves irreversible inhibition of epidermal growth factor receptor (EGFR) tyrosine kinase. In this study, we examined if afatinib had cytotoxic action against NSCLC other than inhibition of tyrosine kinase. Afatinib (1-30 μM) caused apoptotic death in A549 NSCLC in a concentration-dependent manner. Afatinib triggered Ca2+ influx without causing Ca2+ release, and the Ca2+ influx was not unaffected by sodium orthovanadate (SOV, an inhibitor of tyrosine phosphatase), suggesting that afatinib-triggered Ca2+ response was unrelated to its inhibition of tyrosine kinase. Addition of afatinib also promoted Mn2+ influx. Ca2+ influx triggered by afatinib was resistant to SKF96365 and ruthenium red (two general blockers of TRP channels) and, unexpectedly, Ni2+ (a non-specific Ca2+ channel blocker). Afatinib caused an i ncrease in mitochondrial Ca2+ level, an initial mitochondrial hyperpolarization (4 h) and followed by mitochondrial potential collapse (24-48 h). Afatinib-induced cell death was slightly but significantly alleviated in low extracellular Ca2+ condition or under pharmacological block of mitochondrial permeability transition pore (MPTP) opening by cyclosporin A. Therefore, in addition to tyrosine kinase inhibition as a major anti-cancer mechanism of afatinib, stimulation of an atypical Ca2+ influx pathway, mitochondrial Ca2+ overload and potential collapse in part contribute to afatinib-induced cell death. View on Web

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Bupivacaine inhibits the TLR4 ‐ and TLR2 ‐ Myd88/NF‐κB pathways in human leukocytes

alexandrossfakianakis shared this article with you from Inoreader Bupivacaine inhibits the TLR4 ‐ and TLR2 ‐ Myd88/NF‐κB pathways in human leukocytes via Fundamental & Clinical Pharmacology Abstract Background Local anesthetics have anti-inflammatory effects. Because most previous experiments were performed with supra-therapeutic concentrations, we measured the effects of clinically relevant concentrations of bupivacaine on the TLR4- and TLR2-MyD88-NF-κB pathways. Methods We measured TNF-α and PGE2 release, p38 MAP-kinase phosphorylation and translocation of NF-κB in human peripheral blood mononuclear cells (hPBMC) and human monocytes challenged with LPS (lipopolysaccharide) or Pam3CSK4 (tripalmitoylated lipopeptide Pam3CysSerLys4) in the presence or absence of bupivacaine. Similarly, we measured the effect of bupivacaine on HEK293 cells expressing the hTLR4 and the hTLR2 genes and challenged with LPS or Pam3CSK4. Finally, molecular docking simulations of R(+)- and S(-)-bupivacaine binding to the TLR4-MD-2 complex and to the TLR2/TLR1 heterodimer were performed. Results In PBMCs, bupivacaine from 0.1 to 100 μM inhibited LPS-induced TNF-α and PGE2 secretion, phosphorylation of p38 and nuclear translocation of NF-κB in monocytes. Bupivacaine similarly inhibited the effects of Pam3CSK4 on TNF-α secretion. Bupivacaine inhibited the effect of LPS on HEK293 cells expressing the human TLR4 receptor and the effect of Pam3CSK4 on HEK293 cells expressing the human TLR2 receptor. Molecular docking showed that bupivacaine binds to the MD-2 co-receptor of TLR4 and to the TLR2 receptor. Conclusions Contrary to numerous experiments performed with supratherapeutic doses, our results were obtained with concentrations of bupivacaine as low as 0.1 μM. We conclude that bupivacaine modulates the inflammatory reactions such as those observed after surgery or trauma, at least partly by inhibiting the TLR4- and TLR2-NF-κB pathways. View on Web

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Immunomodulatory effect of IL‐1RA in LPS activated macrophage/dental pulp stem cells co‐culture

alexandrossfakianakis shared this article with you from Inoreader Immunomodulatory effect of IL‐1RA in LPS activated macrophage/dental pulp stem cells co‐culture via International Endodontic Journal Abstract Aims: Lipopolysaccharides (LPS) activated human dental pulp stem cells (hDPSCs) and macrophage co-cultures showed downregulated TNF-α secretion that is modulated by hDPSCs through IDO axis, whereas the secretory levels of IL-1β remained unchanged. Therefore, sustained production of IL-1β could contribute to progressive dental pulp inflammation. However, the role of IL-1 receptor antagonists (IL-1RA) in downregulating the secretion of IL-1β and TNF-α in LPS activated M0/M1/M2 macrophage and hDPSCs co-culture has not been studied yet. Therefore, the aim of the present study is to determine the immunomodulatory role of blocking IL-1 receptors in DPSCs macrophage co-culture activated with LPS. Methodology: Human monocytic cell line THP-1 was polarised to M0, M1, and M2 macrophages and co-cultured with hDPSCs. The viability of the co-cultured cells was assessed by apoptosis assay. Co-cultures were activated with LPS followed by the assessment of gene expression and protein levels of IL-1β and TNF-α with and without IL-1RA blocking via qRT-PCR and cytokine flex assay by flow cytometry. Data from three separate experiments was analysed using One-way ANOVA followed by Tukey's Post-hoc test and a p value of <0.05 was considered statistically significant. Results: THP-1 derived M0, M1, and M2 macrophages co-cultured with hDPSCs showed spindle and round-shaped cells, with > 90% viability when assessed by apoptosis assay. Inflammatory TNF-α and IL-1β profile in stimulated co-cultures showed upregulated IL-1β whereas TNF-α was downregulated (p< 0.05). Anti-inflammatory gene expression levels of IL-10, TGF-β were downregulated (p< 0.05). Blocking with IL-1RA resulted in remarkable decrease of IL-1β at the gene expression and protein production levels while TNF-α levels remained low (p< 0.05). Levels of anti-inflammatory cytokine IL-10 showed no significant difference. Conclusion: Blocking the IL-1 receptor in hDPSCs and macrophage (M0, M1, M2) co-cultures activated with LPS resulted in down regulation of inflammatory cytokines IL-1β and TNF-α. These findings highlight the immunomodulatory effect of IL-1RA in inflammatory conditions of dental pulp infections. View on Web

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Estimation of the time-varying incremental effect of low dose aspirin on incidence of pregnancy

alexandrossfakianakis shared this article with you from Inoreader Estimation of the time-varying incremental effect of low dose aspirin on incidence of pregnancy via Epidemiology - Published Ahead-of-Print Background. In many research settings, the intervention implied by the average causal effect of a time-varying exposure is impractical or unrealistic, and we might instead prefer a more realistic target estimand. Instead of requiring all individuals to be always exposed vs. unexposed, incremental effects quantify the impact of merely shifting each individual's probability of being exposed. Methods. We demonstrate estimation of incremental effects in the time-varying setting, using data from the Effects of Aspirin in Gestation and Reproduction trial, which assessed the effect of preconception low-dose aspirin on pregnancy outcomes. Compliance to aspirin or placebo was summarized weekly and was affected by time-varying confounders such as bleeding or nausea. We sought to estimate what the incidence of pregnancy by 26 weeks post-randomization would have been if we shifted each participant's probability of taking aspirin or placebo in each week by odds ratios (OR) between 0.30 and 3.00. Results. Under no intervention (OR=1), the incidence of pregnancy was 77% (95% CI: 74%, 80%). Decreasing women's probability of complying with aspirin had little estimated effect on pregnancy incidence. When we increased women's probability of taking aspirin, estimated incidence of pregnancy increased, from 83% (95% CI: 79%, 87%) for OR=2 to 89% (95% CI: 84%, 93%) for OR=3. We observed similar results when we shifted women's probability of complying with placebo. Conclusions. These results estimated that realistic interventions to increase women's probability of taking aspirin would have yielded little to no impact on the incidence of pregnancy, relative to similar interventions on placebo. Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved. View on Web

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Polyethylenglycol – Freund oder Feind?

alexandrossfakianakis shared this article with you from Inoreader Polyethylenglycol – Freund oder Feind? via Laryngo-Rhino-Otologie Laryngorhinootologie 2022; 101: 784-786 DOI: 10.1055/a-1861-7047 Anaphylaktische Reaktionen nach Impfungen sind selten. Zwei Fälle von Reaktionen nach der Pfizer-BioNTech-Covid-19-Vakzine, BNT162b2, wurden aus dem UK berichtet, weitere 19 aus den USA. Als möglicher Auslöser wurde Polyethylenglycol (PEG) angeschuldigt. Pegylierung ist ein üblicher Vorgang in der Impfstoffherstellung, um die Substanzen vor schnellem immunologischem Abbau zu schützen. PEG ist ein Polyether (Synonyme z. B. Laureth-9, Polidocanol, Thesit, Macrogol) und findet sich in Kosmetika, Körperpflegeprodukten, Nahrungsmitteln, Liposomen und Nanopartikeln als Drug-Delivery-Systeme für Medikamente und als osmotisches Laxans vor Gastro-/Koloskopien. IgM-Antikörper auf PEG werden bei 70% der Bevölkerung nachgewiesen. Bei 2 Fällen von Soforttyp-Reaktionen auf PEG fanden sich IgE-Antiköper, bei anderen Medikamentengaben mit PEG geht man von pseudoallergischen Reaktionen über die Komplement-Aktivierungs-related Pseudoallergie (CARPA) aus. In Relation zum weit verbreiteten Einsatz von PEG sind die Impfreaktionen sehr selten und aktuell kein Grund, Allergiker oder Menschen mit Anaphylaxien bei entsprechenden Vorsichtsmaßnahmen auf bekannte Auslöser auszuschließen. Aufgrund der Meldepflicht (Gesundheitsamt und Arzneimittelkommission der deutschen Ärzteschaft) ist bald mit weiteren Erkenntnissen zu rechnen. [...] Georg Thieme Verlag KG Rüdigerstraße 14, 70469 Stuttgart, Germany Article in Thieme eJournals: Table of contents  |  Abstract  |  Full text View on Web

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