%0 Journal Article %J Curr HIV Res %D 2018 %T Lopinavir/Ritonavir Treatment Induces Oxidative Stress and Caspaseindependent Apoptosis in Human Glioblastoma U-87 MG Cell Line. %A Gratton, Rossella %A Tricarico, Paola Maura %A Guimarães, Rafael Lima %A Celsi, Fulvio %A Crovella, Sergio %X

BACKGROUND: Lopinavir and Ritonavir (LPV/r) treatment is widely used to prevent HIV mother-to-child transmission. Nevertheless, studies related to the impact of these compounds on patients, in particular in the foetus and newborns, are strictly required due to the controversial findings reported in the literature concerning possible neurologic side effects following the administration of these drugs.

OBJECTIVES: In our study, we evaluated the impact of LPV/r treatment on the human glioblastoma U- 87 MG cell line.

METHODS: In order to evaluate the influence of Lopinavir and Ritonavir in terms of oxidative stress (ROS production), mitochondrial morphology and apoptotic cell death, the latter either in the presence or in the absence of caspase-3 and -9 inhibitors, we treated U-87 MG with increasing doses (0.1-1-10-25-50 µM) of Lopinavir and Ritonavir for 24h, either in single formulation or in combination. ROS production was measured by flow cytometry using H2DCFDA dye, mitochondrial morphology was evaluated using MitoRed dye and apoptotic cell death was monitored by flow cytometry using Annexin V-FITC and Propidium Iodide.

RESULTS: We observed that co-treatment with Lopinavir and Ritonavir (25 and 50 µM) promoted a significant increase in ROS production, caused mitochondrial network damage and induced apoptosis in a caspase-independent manner.

CONCLUSION: Based on our findings, concordant with others reported in the literature, we hypothesize that LPV/r treatment could not be entirely free from side effects, being aware of the need of validation in in vivo models, necessary to confirm our results.

%B Curr HIV Res %V 16 %P 106-112 %8 2018 %G eng %N 2 %1 http://www.ncbi.nlm.nih.gov/pubmed/29804534?dopt=Abstract %R 10.2174/1570162X16666180528100922 %0 Journal Article %J Oxid Med Cell Longev %D 2018 %T Photobiomodulation at Multiple Wavelengths Differentially Modulates Oxidative Stress and . %A Rupel, Katia %A Zupin, Luisa %A Colliva, Andrea %A Kamada, Anselmo %A Poropat, Augusto %A Ottaviani, Giulia %A Gobbo, Margherita %A Fanfoni, Lidia %A Gratton, Rossella %A Santoro, Massimo %A Di Lenarda, Roberto %A Biasotto, Matteo %A Zacchigna, Serena %K Adult %K Aged %K Aged, 80 and over %K Female %K Humans %K Keratinocytes %K Lasers, Semiconductor %K Low-Level Light Therapy %K Male %K Middle Aged %K Neutrophils %K Oxidation-Reduction %K Oxidative Stress %K Stomatitis %X

Photobiomodulation (PBM) is emerging as an effective strategy for the management of multiple inflammatory conditions, including oral mucositis (OM) in cancer patients who receive chemotherapy or radiotherapy. Still, the poor understanding of the mechanisms by which the light interacts with biological tissues and the heterogeneity of light sources and protocols employed worldwide significantly limits its applicability. Reactive oxygen species (ROS) are massively generated during the early phases of OM and play a major role in the pathogenesis of inflammation in general. Here, we report the results of a clinical and experimental study, aimed at evaluating the effect of laser light at different wavelengths on oxidative stress in oncologic patients suffering from OM and in two cell types abundantly present within the inflamed oral mucosa, neutrophil polymorphonuclear (PMN) granulocytes, and keratinocytes. In addition to standard ROS detection methods, we exploited a roGFP2-Orp1 genetically encoded sensor, allowing specific, quantitative, and dynamic imaging of redox events in living cells in response to oxidative stress and PBM. We found that the various wavelengths differentially modulate ROS production. In particular, the 660 nm laser light increases ROS production when applied either before or after an oxidative stimulus. In contrast, the 970 nm laser light exerted a moderate antioxidant activity both in the saliva of OM patients and in both cell types. The most marked reduction in the levels of ROS was detected in cells exposed either to the 800 nm laser light or to the combination of the three wavelengths. Overall, our study demonstrates that PBM exerts different effects on the redox state of both PMNs and keratinocytes depending on the used wavelength and prompts the validation of a multiwavelength protocol in the clinical settings.

%B Oxid Med Cell Longev %V 2018 %P 6510159 %8 2018 %G eng %1 http://www.ncbi.nlm.nih.gov/pubmed/30534349?dopt=Abstract %R 10.1155/2018/6510159 %0 Journal Article %J Cell Physiol Biochem %D 2017 %T Lack of Prenylated Proteins, Autophagy Impairment and Apoptosis in SH-SY5Y Neuronal Cell Model of Mevalonate Kinase Deficiency. %A Tricarico, Paola Maura %A Romeo, Alessandra %A Gratton, Rossella %A Crovella, Sergio %A Celsi, Fulvio %K Animals %K Apoptosis %K Autophagy %K Cell Line, Tumor %K Macrolides %K Mevalonate Kinase Deficiency %K Microtubule-Associated Proteins %K Models, Biological %K Mutation %K Phosphotransferases (Alcohol Group Acceptor) %K Protein Prenylation %K Rats %X

BACKGROUND/AIMS: Mevalonate Kinase Deficiency (MKD), is a hereditary disease due to mutations in mevalonate kinase gene (MVK). MKD has heterogeneous clinical phenotypes: the correlation between MVK mutations and MKD clinical phenotype is still to be fully elucidated. Deficiency of prenylated proteins has been hypothesized as possible MKD pathogenic mechanism. Based on this hypothesis and considering that neurologic impairment characterizes Mevalonic Aciduria (MA), the most severe form of MKD, we studied the effects of I268T and N301T MVK mutations on protein prenylation, autophagy and programmed cell death in SH-SY5Y neuroblastoma cell lines.

METHODS: SH-SY5Y cells were transiently transfected, with the pCMV-6 plasmid containing MVK wild type and the two mutated sequences. Protein prenylation levels were evaluated using GFP-RhoA-F to assess farnesylation, and GFP-RhoA to evaluate geranylgeranylation; autophagy was measured by evaluating LC3 and p62 protein levels, while Annexin V-FITC and Propidium Iodide staining allowed apoptosis detection.

RESULTS: MVK mutants' over-expression causes decreased levels of farnesylation and geranylgeranylation, and also increased LC3 lipidation in SH-SY5Y, with concomitant p62 accumulation. Treatment with bafilomycin A1 (an inhibitor of vacuolar H+-ATPase, a late autophagy inhibitor) further increase LC3-II and p62 levels, suggesting that degradation of autophagolysosome could be impaired. SH-SY5Y, with both MVK mutants, showed apoptosis increase; the presence of N301T associated with augmented cell death.

CONCLUSIONS: We hypothesize that mevalonate pathway impairment causes alteration of farnesylation and geranylgeranylation proteins and alteration of the autophagic flux; these changes can induce apoptosis, possibly more relevant in the presence of N301T mutation.

%B Cell Physiol Biochem %V 41 %P 1649-1660 %8 2017 %G eng %N 4 %1 http://www.ncbi.nlm.nih.gov/pubmed/28359055?dopt=Abstract %R 10.1159/000471235