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Duchenne muscular dystrophy (DMD) is a progressive muscle-wasting disease. Recently approved molecular/gene treatments do not solve the downstream inflammation-linked pathophysiological issues such that supportive therapies are required to improve therapeutic efficacy and patients' quality of life. Over the years, a plethora of bioactive natural compounds have been used for human healthcare. Among them, plumbagin, a plant-derived analog of vitamin K3, has shown interesting potential to counteract chronic inflammation with potential therapeutic significance. In this work we evaluated the effects of plumbagin on DMD by delivering it as an oral supplement within food to dystrophic mutant of the fruit fly Drosophila melanogaster and mdx mice. In both DMD models, plumbagin show no relevant adverse effect. In terms of efficacy plumbagin improved the climbing ability of the dystrophic flies and their muscle morphology also reducing oxidative stress in muscles. In mdx mice, plumbagin enhanced the running performance on the treadmill and the muscle strength along with muscle morphology. The molecular mechanism underpinning these actions was found to be the activation of nuclear factor erythroid 2-related factor 2 pathway, the re-establishment of redox homeostasis and the reduction of inflammation thus generating a more favorable environment for skeletal muscles regeneration after damage. Our data provide evidence that food supplementation with plumbagin modulates the main, evolutionary conserved, mechanistic pathophysiological hallmarks of dystrophy, thus improving muscle function in vivo; the use of plumbagin as a therapeutic in humans should thus be explored further., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

In an initial screening, a series of novel Knoevenagel adducts were submitted to the National Cancer Institute for evaluation of antitumor activity in human cell lines. In particular, compound 5f showed remarkable selectivity against IGROV1, an ovarian cancer cell line, without affecting healthy human fibroblast cells. Analyses of cytotoxicity, cell proliferation, cell migration, epigenetic changes, gene expression, and DNA damage were performed to obtain detailed information about its antitumor properties. Our results show that 5f causes proliferation arrest, decrease in motility, histone hyperacetylation, downregulation of cyclin D1 and α5 subunit of integrin β1 gene transcription. In addition, 5f treatment reduces transcript and protein levels of cyclin D1, which increases sensitivity to ionizing radiation and results in DNA damage comparable to cyclin D1 gene silencing., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Alzheimer's disease (AD) and cancer are among the most devastating diseases of the 21st century. Although the clinical manifestations are different and the cellular mechanisms underlying the pathologies are opposite, there are different classes of molecules that are effective in both diseases, such as quinone-based compounds and histone deacetylase inhibitors (HDACIs). Herein, we investigate the biological effects of a series of compounds built to exploit the beneficial effects of quinones and histone deacetylase inhibition (compounds 1-8). Among the different compounds, compound 6 turned out to be a potent cytotoxic agent in SH-SY5Y cancer cell line, with a half maximal inhibitory concentration (IC 50 ) value lower than vorinostat and a pro-apoptotic activity. On the other hand, compound 8 was nontoxic up to the concentration of 100 μM and was highly effective in stimulating the proliferation of neural precursor cells (NPCs), as well as inducing differentiation into neurons, at low micromolar concentrations. In particular, it was able to induce NPC differentiation solely towards a neuronal-specific phenotype, without affecting glial cells commitment., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

γ-terpinene, α-terpinene, p-cymene, and myrcene are monoterpenes found in many essential oils extracted from a variety of plants and spices. Myrcene also occurs naturally in plants such as hops, cannabis, lemongrass, and verbena and is used as a flavoring agent in food and beverage manufacturing. In this research, the biological efficacy of γ-terpinene, α-terpinene, p-cymene, and myrcene was studied in human cell lines (HeLa, SH-SY5Y, and HDFa). Cytotoxicity, cell proliferation, cell migration, and morphology assays were performed to obtain detailed information on the anticancer properties. Our results show that myrcene has potential biological activity, especially in HeLa cells. In this cell line, it leads to an arrest of proliferation, a decrease in motility and morphological changes with loss of sphericity and thickness, and DNA damage. In addition, the interaction of γ-terpinene, α-terpinene, p-terpinene, and myrcene with calf thymus DNA (ct-DNA) was studied by UV-visible spectrophotometry. DNA binding experiments show that only myrcene can interact with DNA with an apparent dissociation constant ( K d ) of 29 × 10 -6 M.

A vaginal microbiota dominated by certain Lactobacillus species may have a protective effect against Chlamydia trachomatis infection. One of the key antimicrobial compounds produced is lactic acid, which is believed to play a central role in host defense. Lactobacillus strains producing the D(-)-lactic acid isomer are known to exert stronger protection. However, the molecular mechanisms underlying this antimicrobial action are not well understood. The aim of this study was to investigate the role of D(-)-lactic acid isomer in the prevention of C. trachomatis infection in an in vitro HeLa cell model. We selected two strains of lactobacilli belonging to different species: a vaginal isolate of Lactobacillus crispatus that releases both D(-) and L(+) isomers and a strain of Lactobacillus reuteri that produces only the L(+) isomer. Initially, we demonstrated that L. crispatus was significantly more effective than L. reuteri in reducing C. trachomatis infectivity. A different pattern of histone acetylation and lactylation was observed when HeLa cells were pretreated for 24 h with supernatants of Lactobacillus crispatus or L. reuteri , resulting in different transcription of genes such as CCND1, CDKN1A, ITAG5 and HER-1. Similarly, distinct transcription patterns were found in HeLa cells treated with 10 mM D(-)- or L(+)-lactic acid isomers. Our findings suggest that D(-) lactic acid significantly affects two non-exclusive mechanisms involved in C. trachomatis infection: regulation of the cell cycle and expression of EGFR and α5β1-integrin.

Metallodrugs represent a combination of multifunctionalities that are present concomitantly and can act differently on diverse biotargets. Their efficacy is often related to the lipophilic features exhibited both by long carbo-chains and the phosphine ligands. Three Ru(II) complexes containing hydroxy stearic acids (HSAs) were successfully synthesized in order to evaluate possible synergistic effects between the known antitumor activity of HSA bio-ligands and the metal center. HSAs were reacted with [Ru(H) 2 CO(PPh 3 ) 3 ] selectively affording O,O-carboxy bidentate complexes. The organometallic species were fully characterized spectroscopically using ESI-MS, IR, UV-Vis, and NMR techniques. The structure of the compound Ru-12-HSA was also determined using single crystal X-ray diffraction. The biological potency of ruthenium complexes (Ru-7-HSA, Ru-9-HSA, and Ru-12-HSA) was studied on human primary cell lines (HT29, HeLa, and IGROV1). To obtain detailed information about anticancer properties, tests for cytotoxicity, cell proliferation, and DNA damage were performed. The results demonstrate that the new ruthenium complexes, Ru-7-HSA and Ru-9-HSA, possess biological activity. Furthermore, we observed that the Ru-9-HSA complex shows increased antitumor activity on colon cancer cells, HT29.

Castanea sativa is very common in Italy, and the large amount of waste material generated during chestnut processing has a high environmental impact. Several studies demonstrated that chestnut by-products are a good source of bioactive compounds, mainly endowed with antioxidant properties. This study further investigates the anti-neuroinflammatory effect of chestnut leaf and spiny bur extracts, together with the deepest phytochemical characterisation (by NMR and MS) of active biomolecules contained in leaf extracts, which resulted in being more effective than spiny bur ones. BV-2 microglial cells stimulated with lipopolysaccharide (LPS) were used as a model of neuroinflammation. In BV-2 cells pre-treated with chestnut extracts, LPS signalling is partially blocked via the reduced expression of TLR4 and CD14 as well as the expression of LPS-induced inflammatory markers. Leaf extract fractions revealed the presence of specific flavonoids, such as isorhamnetin glucoside, astragalin, myricitrin, kaempferol 3-rhamnosyl (1-6)(2″-trans-p-coumaroyl)hexoside, tiliroside and unsaturated fatty acids, all of which could be responsible for the observed anti-neuroinflammatory effects. Interestingly, the kaempferol derivative has been identified in chestnut for the first time. In conclusion, the exploitation of chestnut by-products is suitable for the achievement of two goals: satisfaction of consumers' demand for new, natural bio-active compounds and valorisation of by-products.

Astaxanthin is a red orange xanthophyll carotenoid produced mainly by microalgae but which can also be chemically synthesized. As demonstrated by several studies, this lipophilic molecule is endowed with potent antioxidant properties and is able to modulate biological functions. Unlike synthetic astaxanthin, natural astaxanthin (NAst) is considered safe for human nutrition, and its production is considered eco-friendly. The antioxidant activity of astaxanthin depends on its bioavailability, which, in turn, is related to its hydrophobicity. In this study, we analyzed the water-solubility of NAst and assessed its protective effect against oxidative stress by means of different approaches using a neuroblastoma cell model. Moreover, due to its highly lipophilic nature, astaxanthin is particularly protective against lipid peroxidation; therefore, the role of NAst in counteracting ferroptosis was investigated. This recently discovered process of programmed cell death is indeed characterized by iron-dependent lipid peroxidation and seems to be linked to the onset and development of oxidative-stress-related diseases. The promising results of this study, together with the "green sources" from which astaxanthin could derive, suggest a potential role for NAst in the prevention and co-treatment of chronic degenerative diseases by means of a sustainable approach.

A series of naphthoquinones, namely, 1,4-naphthoquinone, menadione, plumbagin, juglone, naphthazarin, and lawsone, were reacted with N -acetyl- L -cysteine, and except for lawsone, which did not react, the related adducts were obtained. After the tuning of the solvent and reaction conditions, the reaction products were isolated as almost pure from the complex reaction mixture via simple filtration and were fully characterized. Therefore, the aim of this work was to evaluate whether the antitumor activity of new compounds of 1,4-naphthoquinone derivatives leads to an increase in ROS in tumor cell lines of cervical carcinoma (HeLa), neuroblastoma (SH-SY5Y), and osteosarcoma (SaOS2, U2OS) and in normal dermal fibroblast (HDFa). The MTT assay was used to assay cell viability, the DCF-DA fluorescent probe to evaluate ROS induction, and cell-cycle analysis to measure the antiproliferative effect. Compounds 8 , 9 , and 12 showed a certain degree of cytotoxicity towards all the malignant cell lines tested, while compound 11 showed biological activity at higher IC 50 values. Compounds 8 and 11 induced increases in ROS generation after 1 h of exposure, while after 48 h of treatment, only 8 induced an increase in ROS formation in HeLa cells. Cell-cycle analysis showed that compound 8 caused an increase in the number of G0/G1-phase cells in the HeLa experiment, while for the U2OS and SH-SY5Y cell lines, it led to an accumulation of S-phase cells. Therefore, these novel 1,4-naphthoquinone derivatives may be useful as antitumoral agents in the treatment of different cancers.

A series of regioisomers of the hydroxystearic acid ( HSA ) was prepared, and the effect of the position of the hydroxyl group along the chain on a panel of human cancer cell lines was investigated. Among the various regioisomers, those carrying the hydroxyl at positions 5, 7, and 9 had growth inhibitor activity against various human tumor cell lines, including CaCo-2, HT29, HeLa, MCF7, PC3, and NLF cells. 10-HSA and 11-HSA showed a very weak effect. 8-HSA did not show inhibitory activity in all cell lines. The biological role of 7-HSA and 9-HSA is widely recognized, while little is known about the effects of 5-HSA . Therefore, the biological effects of 5-HSA in HeLa, HT29, MCF7, and NLF cell lines were investigated using the Livecyte's ptychography technology, which allows correlating changes in proliferation, motility, and morphology as a function of treatment at the same time. 5-HSA not only reduces cell proliferation but also induces changes in cell displacement, directionality, and speed. It is important to characterize the biological effects of 5-HSA , this molecule being an important component of fatty acyl esters of hydroxy fatty acids (FAHFA), a class of endogenous mammalian lipids with noticeable anti-diabetic and anti-inflammatory effects.

Chlamydia persistence is a viable but non-replicative stage, induced by several sub-lethal stressor agents, including beta-lactam antibiotics. So far, no data about the connection between doxycycline and chlamydial persistence has been described in literature. We investigated the ability of doxycycline to induce C. trachomatis (CT) persistence in an in vitro model of epithelial cell infection (HeLa cells), comparing the results with the well-established model of penicillin-induced persistence. The effect of doxycycline was explored on 10 different CT strains by analysing (i) the presence of aberrant inclusions, (ii) chlamydial recovery, (iii) the expression of different chlamydial genes (omcB, euo, Ct110, Ct604, Ct755, HtrA) and (iv) the effects on epithelial cell viability. For each strain, the presence of foreign genomic islands responsible of tetracycline resistance was excluded. We found that low doses of doxycycline can induce a condition of CT persistence. For concentrations of doxycycline equal to 0.03-0.015 mg/L, CT inclusions are smaller and aberrant and CT cycle is characterized by the presence of viable but non-dividing RBs with the complete abolishment of chlamydial cytotoxic effect. Infectious EBs can be recovered after removal of the drug. During doxycycline-induced persistence, the expression of the late gene omcB is decreased, indicating the blocking of RB-to-EB conversion. Conversely, as for penicillin G, a significant up-regulation of the stress response HtrA gene is found in doxycycline-treated cells. This study provides a novel in vitro cell model to examine the characteristics of doxycycline-induced persistent CT infection., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Chlamydia persistence is a viable, but non-cultivable, growth stage, resulting in a long-term relationship with the infected host cell. In vitro, this condition can be induced by different stressor agents, including beta-lactam antibiotics, as penicillin. The aim of this study was to get new insights into the interactions between Chlamydia trachomatis (serovars D and L2) and the epithelial host cells (HeLa) during persistence condition. In particular, we evaluated the following aspects, by comparing the normal chlamydial development cycle with penicillin-induced persistence: (i) cell survival/death, (ii) externalization of phosphatidylserine, (iii) caspase 1 and caspase 3/7 activation, and (iv) reactive oxygen species (ROS) production by the infected cells. At 72 h post-infection, the cytotoxic effect displayed by CT was completely abolished for both serovars and for all levels of multiplicity of infection only in the cells with aberrant CT inclusions. At the same time, CT was able to switch off the exposure of the lipid phosphatidylserine on the surface of epithelial cells and to strongly inhibit the activation of caspase 1 and caspase 3/7 only in penicillin-treated cells. Forty-eight hours post-infection, CT elicited a significant ROS expression both in case of a normal cycle and in case of persistence. However, serovar L and penicillin-free infection activated a higher ROS production compared to serovar D and to penicillin-induced persistence, respectively. In conclusion, we added knowledge to the cellular dynamics taking place during chlamydial persistence, demonstrating that CT creates a suitable niche to survive, switching off signals able to activate phagocytes/leukocytes recruitment. Nevertheless, persistent CT elicits ROS production by the infected cells, potentially contributing to the onset of chronic inflammation and tissue damages., Competing Interests: Declaration of competing interest This study was conducted in the absence of any commercial or financial relationships that could be construed as potential conflict of interest. All the authors declare the absence of any dual or conflicting interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)