Biology subjects

Baptista, M. S.

Publications and source records attributed to Baptista, M. S..

6 recordsLinked to original sources

bioRxiv2025

Transcriptomic and functional profiling reveal autophagy inhibition and persistent bioenergetic collapse following parallel photodamage to lysosomes and mitochondria

Photodynamic therapy (PDT) using 1,9-dimethyl methylene blue (DMMB) induces coordinated mitochondrial and lysosomal damage and results in strong cellular death induction. However, the underlying transcriptional regulation in response to DMMB remains elusive. We compared the transcriptome response of photoactivated DMMB (paDMMB) to the gene signature triggered by autophagy-modulating agents: rapamycin (an autophagy activator) and bafilomycin A1 (a lysosomal acidification inhibitor). Transcriptome analysis revealed a pronounced transcriptomic response to paDMMB, with 884 differentially expressed genes (DEGs), compared to 291 for bafilomycin and 154 for rapamycin. paDMMB treatment upregulated genes associated with autophagy, mitochondrial stress responses, and proteostasis, while downregulating genes involved in miRNA processing and lipid catabolism. Rapamycin treatment downregulated amino acid biosynthesis pathways, while upregulating processes associated with nutrient starvation. Conversely, bafilomycin treatment upregulated genes related to lipid metabolism, while suppressing cytoskeletal programs. Transcriptomic comparisons revealed a striking overlap (95%) between paDMMB and bafilomycin signatures. Among the several biological processes affected by paDMMB, mitochondrial-related processes were strongly enriched. To determine whether the acute transcriptome changes caused by paDMMB led to persistent functional effects, we stimulated cells with DMMB and assessed mitochondrial respiration after a recovery period. paDMMB reduced basal respiration, ATP production, proton leak, and maximal respiration. These effects were not further altered by bafilomycin co-treatment but were markedly exacerbated by rapamycin. Collectively, we show that paDMMB leads to a transcriptome rewiring, closely resembling autophagy inhibition with a sustained mitochondrial dysfunction. These findings provide a valuable resource to understand the interplay between DMMB-induced lysosomal stress, transcriptional regulation, and PDT.

cell biology↗

bioRxiv2024

Stage-specific phenotypic and transcriptional alterations in keratinocytes exposed to acute and chronic blue light

Despite evidence that visible light (VL) has similar effects on human skin as those of UVA, VL is often viewed as harmless. High SPF sunscreen prevents erythema but can lead to overexposure to UVA and VL, with unknown consequences. To explore the impact of chronic blue light exposure, we irradiated (50 J/cm{superscript 2}, {lambda}= 408 nm, three times a week) human immortalized keratinocytes under acute (3 irradiations), intermediate (14 irradiations), and chronic (42 irradiations) blue-light exposure, monitoring phenotypic and gene expression changes. Chronically exposed keratinocytes exhibit increased nuclei area, chromatin alterations, higher proliferation, and apoptosis resistance, mirroring the consequences of chronic UVA exposure. While acute exposure upregulated keratinization and downregulated tissue repair and apoptosis genes, chronically exposed cells had upregulated genes involved with energy metabolism and oxidative phosphorylation and downregulated genes were enriched for immune and inflammatory responses. Specific transcriptional factors were identified in the acute and chronic stages, some of them had been associated with UVB exposure. We identified some changes in chronically irradiated keratinocytes similar to the malignant transformation, emphasizing the need for further research on the long-term impacts of blue light exposure on human skin.

cell biology↗

bioRxiv2024

Mitochondrial Fatty Acid Oxidation is Stimulated by Red Light Irradiation

Keratinocytes are the primary constituents of sunlight-exposed epidermis. In these cells, UVA completely inhibited oxidative phosphorylation, while equivalent doses of blue and green light preserved metabolic fluxes, but reduced viability. In contrast, red light enhanced proliferation and elevated basal and maximal oxygen consumption rates for 48 h, without altering protein levels of the electron transport chain. Targeted flux analysis revealed that red light specifically activates AMPK-dependent mitochondrial fatty acid oxidation. This was accompanied by reduced the levels of free fatty acids and increased acetyl CoA carboxylase phosphorylation. Together, our results characterize wavelength- selective regulation of keratinocyte metabolism: UV/visible wavelengths induce damage, while red light triggers AMPK-dependent fatty acid oxidation, providing a mechanistic explanation for photobiomodulation in epidermal cells. HighlightsUVA (365 nm) abolishes oxidative phosphorylation, while blue/green light (450/517 nm) induce photosensitized toxicity, without metabolic disruption. Red light (660 nm) boosts keratinocyte proliferation and sustains elevated mitochondrial respiration for 48 h. Permeabilized cell electron transport remains unchanged, implicating cytosolic (not mitochondrial) regulation of fuel utilization. Red light activates AMPK and phosphorylates/inhibits acetyl CoA carboxylase (ACC). Photobiomodulation with red light occurs through AMPK/ACC-mediated lipid oxidation, not direct respiratory chain modulation.

biochemistry↗

bioRxiv2024

Plasmalogen oxidation induces the generation of excited molecules and electrophilic lipid species

Plasmalogens are glycerophospholipids with a vinyl-ether linkage at the sn-1 position of the glycerol backbone. Despite being suggested as antioxidants due to the high reactivity of their vinyl ether groups with reactive oxygen species (ROS), our study reveals the generation reactive oxygen and electrophilic lipid species from oxidized plasmalogen intermediates. By conducting a comprehensive analysis of the oxidation products by liquid chromatography coupled to high-resolution mass spectrometry (LC-MS) we demonstrate that singlet molecular oxygen [O2 (1{Delta}g)] reacts with the vinyl ether bond, producing hydroperoxyl acetal as major primary product (97%) together with minor quantities of dioxetane (3%). Furthermore, we show that these primary oxidized intermediates lead to the formation of excited triplet carbonyls, O2 (1{Delta}g), and electrophilic phospholipid and fatty aldehyde species, as secondary reactive products. The generation of excited triplet carbonyls from dioxetane thermal decomposition was confirmed by light emission measurements in the visible region using dibromoantracene as a triplet enhancer. Moreover, O2 (1{Delta}g) generation from dioxetane and hydroperoxyacetal was evidenced by detection of near-infrared light emission at 1270 nm and chemical trapping experiments. Additionally, we have thoroughly characterized alpha-beta unsaturated phopspholipid and fatty aldehydes by LC-MS analysis using two probes that specifically reacts with aldehydes and alpha-beta unsaturated carbonyls. Overall, our findings demonstrate the generation of excited molecules and electrophilic lipid species from oxidized plasmalogen species unveiling the potential prooxidant nature of plasmalogen oxidized products. Significance StatementPlasmalogens, the most abundant subclass of ether lipids in mammalian cells, have traditionally been regarded as antioxidants. However, our study reveals a new perspective, shedding light on the generation of chemiexcited and reactive lipid species during plasmalogen photooxidation. We provide direct evidence revealing the production of excited triplet carbonyls and singlet molecular oxygen as secondary reactive products originating from dioxetane and hydroperoxyacetal intermediates. Importantly, we also demonstrate the generation of electrophilic alpha-beta unsaturated phospholipids and fatty aldehydes through plasmalogen oxidation. These findings highlight the production of excited states and reactive lipid species resulting from plasmalogen oxidation, which can potentially induce oxidative modifications in biological systems.

biochemistry↗

bioRxiv2020

Copper nanoparticles prompt the activity of estuarine denitrifying bacterial communities at relevant environmental concentrations

Effects of metallic nanoparticles (NPs) to the estuarine biota have mostly been shown for concentrations higher than those actually measured or predicted in these environments. To address this gap, a range of concentrations expected to occur in estuarine environments (from 0.01 to 1 μg g-1) was employed in microcosms studies to assess the impact of Cu NPs in the denitrification pathway. That was achieved by quantifying gene expression and the potential denitrification rate in estuarine sediments exposed to Cu NPs for up to six days. Expression of nitrite (nirS) and nitrous oxide (nosZ) reductase genes was enhanced in a timewise manner. For the highest Cu NPs (1 μg g-1) an increase in gene expression could be seen immediately after 1 h of exposure, and continuing to be enhanced up until 7 h of exposure. For the lowest Cu NPs (0.01 μg g-1) an increase in gene expression could only be seen after 4 h or 7 h of exposure; however it continued to rise up until 24 h of exposure. In any case, after 48 h the expression levels were no longer different from the non-exposed control. Concomitantly to increased gene expression the potential denitrification rate was increased by 30 %. Our results suggest that deposition and adsorption of Cu NPs to estuarine sediments promotes the immediate and transient expression of key genes of the denitrification pathway. The long term impact of continuous inputs of Cu NPs into estuaries deserves renewed analysis to account for their effects, not just on the biota, but especially on ecosystems services.Environmental significance Interactions of metallic nanoparticles with microbial communities of estuarine sediments are poorly characterized and its impact towards ecosystem services even less. By assessing the effect of copper nanoparticles on the expression of key genes of the denitrification pathway, an essential step for nitrogen (N) removal, we were able to show that denitrifying communities are immediately activated after exposure, increasing the denitrification rates in estuaries. The importance of denitrification lies in its release of dinitrogen (N2) to the atmosphere but also in the emissions of N2O (a potent greenhouse gas). The results obtained in this study gather data that contribute information on the denitrification dynamics in estuaries, invaluable for a timely response to the expected upcoming changes in coastal areas.Table of contentsIn estuaries the deposition upon the sediments of copper nanoparticles can contribute to change metal availability and promote the activity of denitrifying bacteriaCompeting Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗

bioRxiv2020

Distinct photooxidation-induced cell death pathways lead to selective killing of human breast cancer cells

Lack of effective treatments for aggressive breast cancer is still a major global health problem. We previously reported that Photodynamic Therapy using Methylene Blue as photosensitizer (MB-PDT) massively kills metastatic human breast cancer, marginally affecting healthy cells. In this study we aimed to unveil the molecular mechanisms behind MB-PDT effectiveness. Through lipidomic and biochemical approaches we demonstrated that MB-PDT efficiency and specificity relies on polyunsaturated fatty acids-enriched membranes and on the better capacity to deal with photooxidative damage displayed by non-tumorigenic cells. We found out that, in tumorigenic cells, lysosome membrane permeabilization is accompanied by ferroptosis and/or necroptosis. Our results broadened the understanding of MB-PDT-induced photooxidation mechanisms and specificity in breast cancer cells. Therefore, we demonstrated that efficient approaches could be designed on the basis of lipid composition and metabolic features for hard-to-treat cancers. The results further reinforce MB-PDT as a therapeutic strategy for highly aggressive human breast cancer cells.Competing Interest StatementThe authors have declared no competing interest.View Full Text

cancer biology↗