Leukotrienes are among the most potent mediators of inflammation, and inhibition of their biosynthesis, is becoming increasingly important in the treatment of many pathologies. In this work, we demonstrated that preincubation of human neutrophils with the mitochondria targeted antioxidant SkQ1 (100 nM) strongly inhibits leukotriene synthesis induced by three different stimuli: the Ca²⁺ ionophore A23187, the chemotactic formyl-peptide fMLP in combination with cytocholasin B, and opsonized zymosan. The SkQ1 analogue lacking the antioxidant quinone moiety (C12TPP) was ineffective, suggesting that mitochondrial production of reactive oxygen species (ROS) is critical for activating of leukotriene synthesis in human neutrophils. The uncoupler of oxidative phosphorylation FCCP also inhibits leukotriene synthesis, indicating that a high membrane potential is a prerequisite for stimulating leukotriene synthesis in neutrophils. Our data show that activation of mitogen-activated protein kinases p38 and ERK1/2, which is important for leukotriene synthesis in neutrophils is a target for SkQ1: 1) the selective p38 inhibitor SB203580 inhibited fMLP-induced leukotriene synthesis, while the ERK1/2 activation inhibitor U0126 suppressed leukotriene synthesis induced by any of the three stimuli; 2) SkQ1 effectively prevents p38 and ERK1/2 activation (accumulation of phosphorylated forms) induced by all three stimuli. This is the first study pointing to the involvement of mitochondrial reactive oxygen species in the activation of leukotriene synthesis in human neutrophils. The use of mitochondria-targeted antioxidants can be considered as a promising strategy for inhibiting leukotriene synthesis and treating various inflammatory pathologies.

Genotoxic and cytotoxic drugs, widely used in anticancer therapy, target proliferating cells and induce cell death through a variety of cell cycle-dependent mechanisms. The mechanisms of the delayed toxicity induced by chemotherapy are not fully understood. The accumulation of senescent cells may underlie some of the mechanisms for the development of late adverse effects of chemotherapy on muscle tissue. Cellular models are necessary for the development of therapeutic approaches to these side effects. In our study we used human immortalized myoblast MB135 to optimize the protocol for obtaining the senescent phenotype of muscle cells under the influence of chemotherapeutic drugs such as doxorubicin, cisplatin and arsenic trioxide (As₂O₃). We evaluated the dynamics of changes in senescence proteins pRb, p21 and p53 and SASP-associated proteins such as TNF, IL-1b, IL-6, IL-8, CXCL2, GDF15 using Western blot, RT-PCR and ELISA. Cell senescence was confirmed by the measurement of cell senescence index by flow cytometry after 7 days of exposure to chemotherapeutic agents. The obtained results indicate that all three investigated chemotherapeutic compounds induce the appearance of senescence markers, but the dynamics of these changes are somewhat different for them, which may reflect differences in the mechanisms of senescence phenotype induction.

Fluoroquinolones are effective synthetic broad-spectrum antibiotics, but their use can be associated with persistent adverse effects, sometimes described as fluoroquinolone-induced disability. The molecular mechanisms underlying these effects remain incompletely understood, although accumulating evidence implicates mitochondrial dysfunction, oxidative stress, and DNA damage. We hypothesized that the convergence of these stress responses may promote cellular senescence and thereby contribute to persistent fluoroquinolone-associated adverse effects. We examined three clinically used fluoroquinolones—ciprofloxacin, moxifloxacin, and levofloxacin—in primary human fibroblasts in vitro. Exposure for 7 days revealed that 200 µM ciprofloxacin, but not 200 µM moxifloxacin or 500 µM levofloxacin, induced a senescence-like phenotype characterized by increased senescence-associated β-galactosidase (SA-β-Gal) activity and an elevated senescence index (SI), which integrates increased cell size and autofluorescence measured by imaging flow cytometry. At 7 days, ciprofloxacin did not induce a broad canonical senescence-associated secretory phenotype (SASP) at either the transcriptional or secretory level, consistent with a non-canonical senescence-like state; only a modest increase in IL-1β secretion emerged after 14 days. The ciprofloxacin-induced phenotype was not accompanied by a robust increase in the canonical p53, p16, or p21 protein markers, but was associated with reduced retinoblastoma protein (RB) phosphorylation at Ser795. However, ciprofloxacin removal from the culture medium after 1 or 2 weeks of incubation resulted in the recovery of the resazurin signal to approximately control values by the end of the 3-week experimental period. Ciprofloxacin also increased oxidative stress, as assessed using dihydroethidium and dichlorodihydrofluorescein diacetate. Both the antioxidant Trolox and the mitochondria-targeted antioxidant SkQ3 attenuated the ciprofloxacin-induced increase in SA-β-Gal activity and SI. These findings identify oxidative stress as a putative mediator of ciprofloxacin-induced senescence-like remodeling in human fibroblasts and support further investigation of drug-induced cellular senescence as a potential contributor to persistent fluoroquinolone-associated adverse effects.