Levels Of Reactive Species Research Articles

Analysis of the antimelanogenic activity of zinc and selenium in vitro.

Melasma is an acquired chronic condition characterized by hyperchromic patches in photo-exposed areas. The search for new compounds for the treatment of melasma without side effects is constant. In this context, the aim of this study was to investigate the in vitro cytotoxic and antimelanogenic effects of the trace elements Zinc (Zn) and Selenium (Se). In this study, we evaluated the effects of 30µM hydroquinone, this concentration did not alter mitochondrial function (MTT assay), but increased the percentage of necrotic cells and levels of reactive species. Furthermore, it showed no influence on tyrosinase activity and melanin content. Unlike hydroquinone, exposure for 48h to 100µM Zn and 1 and 5µM Se had no significant influence on the analysis of reactive species, as well as on the percentage of necrotic cells. Still, specifically in relation to 100µM Zn, it decreased the melanin content. Given the above, the trace elements Zn and Se did not show toxicity at the concentrations tested and Zn showed a promising effect, however, the mechanism needs to be better explored in order to contribute to new and updated research in the fight against melasma with a perspective of therapeutic use.

Amelioration of oxidative kidney damage in offspring by maternal trans-fatty acid exposure in mice by secoisolariciresinol diglucoside.

Trans-fatty acids (TFAs), primarily derived from the food industry's production processes, have become a globally recognized public health issue due to the detrimental impact they have on human well-being. Secoisolariciresinol diglucoside (SDG) is a polyphenolic compound derived from flax lignans, possessing antioxidative properties. This study aims to investigate the protective effect of SDG on kidney oxidative damage in offspring of mice caused by maternal exposure to TFA during pregnancy and lactation. A total of 30 c57BL/6 female rats were randomly divided into 5 groups: a control group, a TFA-exposed group, a low-(TFA+LSDG) group, a medium-(TFA+MSDG) group, and a high-(TFA+HSDG) group (n=6 in each group). With the exception of the control group, the maternal mice in the remaining 4 groups received a daily oral gavage of TFA at a dosage of 60 mg/(kg·BW) throughout the experimental period. The mothers in the control group were administered physiological saline via oral gavage once daily. Meanwhile, the 3 SDG intervention groups were provided with ad libitum access to SDG feed containing 10 mg/kg (low), 20 mg/kg (medium), and 30 mg/kg (high) of SDG. The female mice were conceived overnight. If the vaginal plug appeared in the next morning, the female mice were conceived and included in the experimental stage until the end of the 21th day lactation period. The body weight and kidney mass of offspring were recorded, and the kidney coefficient was calculated. The kidney was detected by HE staining to observe the histopathological changes, and the level of reactive oxidative species (ROS) was detected by fluorescence probe-dihydroethidium (DHE) staining; the expression levels of total superoxide dismutase (T-SOD) and malondialdehyde (MDA) in renal homogenate were detected, and the expression of nuclear factor E2-related fator2 (Nrf2) and hemeoxygenase-1 (HO-1) protein was analyzed by immunohistochemistry (IHC) staining. The mRNA expressions of Nrf2 and HO-1 were detected by real-time PCR, and the protein expression of Cu/Zn-superoxide dismutase (Cu/Zn-SOD), Mn-superoxide dismutase (Mn-SOD), glutathione peroxidase-1 (GPx-1), Nrf2 and HO-1 were detected by Western blotting. Compared with the control group, the kidney coefficient in the TFA-exposed group was increased, the morphology and structure of kidney tissue was abnormal; the activity of T-SOD enzyme was decreased, and the content of MDA was increased, the level of ROS was increased; the expressions of Cu/Zn-SOD, Mn-SOD, GPx1 protein were decreased, and the mRNA and protein expressions of Nrf2 and HO-1 were decreased, there were all significant difference (all P<0.05). Compared with the TFA-exposed group, the ROS levels were reduced, and the T-SOD enzyme activity as well as the protein expression of Cu/Zn-SOD, GPx-1, Mn-SOD, Nrf2 and HO-1 were up-regulated in the low, middle and high dose SDG intervention groups; the kidney coefficient and MDA content were decreased in the middle and high dose SDG groups; the Nrf2 mRNA expression in the high dose SDG group was up-regulated, there were all significant difference (all P<0.05). Maternal exposure to TFA during pregnancy and lactation can lead to oxidative damage in the kidney of offspring, and the SDG intervention may alleviate TFA-induced oxidative damage by up-regulating the expression of Nrf2 and HO-1 signal pathway.

FeTPPS, a Peroxynitrite Decomposition Catalyst, Ameliorates Nitrosative Stress in Human Spermatozoa.

Excessive levels of reactive nitrogen species (RNS), such as peroxynitrite, promote nitrosative stress, which is an important cause of impaired sperm function. The metalloporphyrin FeTPPS is highly effective in catalyzing the decomposition of peroxynitrite, reducing its toxic effects in vivo and in vitro. FeTPPS has significant therapeutic potential in peroxynitrite-related diseases; however, its effects on human spermatozoa under nitrosative stress have not been described. This work aimed to evaluate the in vitro effect of FeTPPS against peroxynitrite-mediated nitrosative stress in human spermatozoa. For this purpose, spermatozoa from normozoospermic donors were exposed to 3-morpholinosydnonimine, a molecule that generates peroxynitrite. First, the FeTPPS-mediated peroxynitrite decomposition catalysis was analyzed. Then, its individual effect on sperm quality parameters was evaluated. Finally, the effect of FeTPPS on ATP levels, motility, mitochondrial membrane potential, thiol oxidation, viability, and DNA fragmentation was analyzed in spermatozoa under nitrosative stress conditions. The results showed that FeTPPS effectively catalyzes the decomposition of peroxynitrite without affecting sperm viability at concentrations up to 50 μmol/L. Furthermore, FeTPPS mitigates the deleterious effects of nitrosative stress on all sperm parameters analyzed. These results highlight the therapeutic potential of FeTPPS in reducing the negative impact of nitrosative stress in semen samples with high RNS levels.

The mechanism of trans-δ-viniferin inhibiting the proliferation of lung cancer cells A549 by targeting the mitochondria.

Trans-δ-viniferin (TVN), as a natural extract, is a resveratrol dimer with attractive biological activities, particularly its anti-tumor character. However, the mechanism of TVN interfering with cancerous proliferation has not been fully understood. Herein in this study, we found that TVN could trigger cancerous mitochondrial membrane potential (ΔΨm) reduction, with intracellular reactive oxidative species (ROS) level increasing, leading to apoptosis, which makes TVN a promising candidate for lung cancer cells A549 treatment. Therefore, this study provides TVN as an option to meet the demand for higher antitumor availability with lower biotoxicity and other clinical applications.

Mitochondrial impairment and downregulation of Drp1 phosphorylation underlie the antiproliferative and proapoptotic effects of alantolactone on oral squamous cell carcinoma cells

BackgroundOral squamous cell carcinoma (OSCC) is one of the most prevalent and fatal oral cancers. Mitochondria-targeting therapies represent promising strategies against various cancers, but their applications in treating OSCC are limited. Alantolactone (ALT) possesses anticancer properties and also regulates mitochondrial events. In this study, we explored the effects of ALT on OSCC and the related mechanisms.MethodsThe OSCC cells were treated with varying concentrations and duration of ALT and N-Acetyl-l-cysteine (NAC). The cell viability and colony formation were assessed. The apoptotic rate was evaluated by flow cytometry with Annexin V-FITC/PI double staining. We used DCFH-DA and flow cytometry to detect reactive oxygen species (ROS) production and DAF-FM DA to investigate reactive nitrogen species (RNS) level. Mitochondrial function was reflected by mitochondrial reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and ATP levels. KEGG enrichment analyses determined the mitochondrial-related hub genes involved in OSCC progression. Dynamin-related protein 1 (Drp1) overexpression plasmids were further transfected into the cells to analyze the role of Drp1 in OSCC progression. Immunohistochemistry staining and western blot verified the expression of the protein.ResultsALT exerted anti-proliferative and pro-apoptosis effects on OSCC cells. Mechanistically, ALT elicited cell injury by promoting ROS production, mitochondrial membrane depolarization, and ATP depletion, which were reversed by NAC. Bioinformatics analysis showed that Drp1 played a crucial role in OSCC progression. OSCC patients with low Drp1 expression had a higher survival rate. The OSCC cancer tissues presented higher phosphorylated-Drp1 and Drp1 levels than the normal tissues. The results further showed that ALT suppressed Drp1 phosphorylation in OSCC cells. Moreover, Drp1 overexpression abolished the reduced Drp1 phosphorylation by ALT and promoted the cell viability of ALT-treated cells. Drp1 overexpression also reversed the mitochondrial dysfunction induced by ALT, with decreased ROS production, and increased mitochondrial membrane potential and ATP level.ConclusionsALT inhibited proliferation and promoted apoptosis of oral squamous cell carcinoma cells via impairment of mitochondrial homeostasis and regulation of Drp1. The results provide a solid basis for ALT as a therapeutic candidate for treating OSCC, with Drp1 being a novel therapeutic target in treating OSCC.

Investigating respiratory simplification caused by prenatal RAGE upregulation

Receptors for advanced glycation end-products (RAGE) are multi-ligand cell surface receptors of the immunoglobulin superfamily predominantly expressed by lung epithelium. Previous experiments showed that RAGE upregulation throughout pregnancy caused significant fetal lung hypoplasia via elevated apoptosis and misregulation of the transcription factors thyroid transcription factor 1 (TTF1) and forkhead box protein A2 (Fox-A2). Our current research focused on characterizing the contributors to these abnormalities. We confirmed poor development of murine lungs through a time course of RAGE upregulation from conception to sacrifice on embryonic day (E)15.5, E16.5, E17.5, E18.5, and specific upregulation from E15.5 to E18.5 only. H&amp;E staining confirmed significant simplification. We hypothesized that this malformation was in part due to aberrant levels of inflammation and apoptosis. Blotting revealed significantly increased levels of phosphorylated AKT and ERK1/2 in RAGE TG pups which cause NF-κB mediated transcription of inflammatory genes. Due to potential roles for these signaling intermediates, we sought to assess RAGE-mediated impact on mitochondrial efficiency. Oxidative phosphorylation was assessed in lungs from RAGE TG pups and controls by measuring oxygen consumption per unit mass when lungs were exposed to the substrates malate, glutamate, ADP, succinate, and FCCP which activate sequential components of the electron transport chain. We discovered significantly increased levels of oxidative phosphorylation in transgenic pups compared to controls. Levels of ATP and reactive oxidative species (ROS) in relation to altered oxidative phosphorylation and inflammation provided additional mechanistic clarity. These results support the importance of proper RAGE regulation in the development of lung tissue and elucidate mechanisms possibly causing such damage. This work was supported by funding from the National Institutes of Health (NIH 1R15-HL152257, PRR and JAA). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Hypoglycemic effect of the dry leaf extract of Myrcia multiflora in streptozotocin-induced diabetic mice.

Myrcia multiflora (Lam) DC. is a medicinal plant used in folk medicine for diabetes control, mainly in the Brazilian Amazon. The leaves of this species has already demonstrated antidiabetic properties; however, in mice with type 2 diabetes (DM2), the cumulative effect of the consumption of the dry extract of M. multiflora leaves (Mm) has not yet been reported. To investigate the effect of the dry extract obtained from the infusion of the dried leaves of M. multiflora on the blood glucose levels of diabetic mice. DM2 was induced in Swiss male mice by a single intraperitoneal injection of streptozotocin [150mg/kg body weight (bw)]. The animals were divided into two control groups (healthy and diabetic without treatment) and three sample groups that received Mm (25 and 50mg/kg bw) and acarbose (200mg/kg bw) by gavage once daily for 28 days (D28). Additionally, biochemical parameters, thiobarbituric acid reactive species (TBARS) levels in the liver, and histopathological analyses of the kidneys and liver were performed. On the seventh day of treatment, a 74.7% reduction in glucose levels were observed in the group of diabetic animals treated with Mm (50mg/kg bw) when compared to the beginning of the treatment. At D28, the hypoglycemic effect was maintained. The results of the biochemical and histopathological parameters and the TBARS levels suggest that this dry extract exerts nephro- and hepatoprotective effects. The findings demonstrate the potential that this extract has to inhibit the α-glucosidase enzyme, and it acts similarly to the positive control acarbose. Furthermore, this extract is nephro- and hepatoprotective. Therefore, this dry extract has the potential to be an adjuvant for DM2, which corroborates its use in folk medicine.

Abstract 273: Synergistically suppressive effects by combination of glycolytic inhibitor with TRAP1-depletion in colorectal cancer

Abstract Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a mitochondrial homolog of HSP90 chaperones. It plays an important role in protection against oxidative stress and apoptosis by regulating reactive oxidative species (ROS). Recent studies have suggested TRAP1 as a regulator of metabolism and mitochondrial homeostasis in diverse pathological states, including cancer. To further elucidate the mechanistic role of TRAP1 in regulating tumor cell survival, we used gamitrinib-triphenylphosphonium (G-TPP) to inhibit TRAP1 signaling pathways in colon cancer. Inhibition of TRAP1 disrupted redox homeostasis and increased ER stress in colon cancer. To confirm the G-TPP effects of TRAP1 inhibition in colon cancer, we performed TRAP1 CRISPR/Cas9 to deplete TRAP1 expression in colon cancer cell line CT26. TRAP1-depleted cells were found to acquire several distinct phenotypes: induction of reactive oxidative species (ROS) level, reduction of cell growth, and lower pH value of culture medium. In addition, proteomic data showed that depletion of TRAP1 induced significant metabolism alteration in CT26 cells. Using seahorse assay to access mitochondrial respiration and glycolysis, we found that TRAP1-depleted cells were insensitive to Oxidative Phosphorylation (OXPHOS) inhibitor treatment and significantly increased extracellular acidification rate (ECAR) compared to parental cells. Furthermore, higher glucose consumption rate and higher lactic acid level of culture medium were observed in TRAP1-depleted cells. On the other hand, Glycolytic inhibitor (2DG) led to excessive ROS production, reduction of glucose uptake and exacerbated cell death in TRAP1-depleted cell compared to parental cells. Taken together, depletion of TRAP1 induces mitochondrial dysfunction and modulates metabolic switch toward aerobic glycolysis in TRAP1-depleted colon cancer cells. Dual inhibition of TRAP1 and glycolytic signaling may increase therapeutic efficiency through induction of excessive ROS production in colon cancer. Citation Format: Hong-Yuan Tsai, Miao-Hsueh Chen, Herath Lakmini Senavirathna, Jihye Yun, Sheng Pan, Ru Chen. Synergistically suppressive effects by combination of glycolytic inhibitor with TRAP1-depletion in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 273.

A Rat Model of Rheumatoid Arthritis: TDZD-8 is Associated with the Protection Against the Induction of the Synovium Knee Joint IL-17A/GSK3&amp;#946;/ROS/&amp;#945;-SMA Axis of Fibrosis

Rheumatoid arthritis (RA) that affects the synovial knee joint causes swelling of the synovial membrane and tissue damage. Interleukin-17A (IL-17A) and the enzyme glycogen synthase kinase-3β (GSK3β) are involved in the pathogenesis of RA. The link between IL-17A, GSK3β, the oxidative stress, and the profibrogenic marker alpha-smooth muscle actin (α-SMA) with and without TDZD-8, GSK3β inhibitor has not been studied before. Consequently, active immunization of rats was performed to induce RA after three weeks using collagen type II (COII) injections. The treated group received daily injection of 1 mg/kg TDZD-8 for 21 days following the immunization protocol (COII+TDZD-8). Blood and synovium tissue samples were harvested at the end of the experiment. RA development was confirmed as corroborated by a substantial increase in blood levels of the highly specific autoantibody for RA, anti-citrullinated protein antibody as well as augmentation of reactive oxidative species (ROS) levels measured as lipid peroxidation. RA induction also increased synovium tissue levels of IL-17A and the profibrogenic marker, α-SMA. All these parameters seemed to be significantly (p<0.0001) ameliorated by TDZD-8. Additionally, a significant correlation between IL-17A, ROS, and α-SMA and biomarkers of RA was observed. Thus, knee joint synovium RA induction augmented IL-17A/GSK3β/ROS/α-SMA axis mediated arthritis in a rat model of RA, which was inhibited by TDZD-8.

β-carotene-loaded nanoparticles protect against neuromotor damage, oxidative stress, and dopamine deficits in a model of Parkinson's disease in Drosophila melanogaster

β-carotene-loaded nanoparticles improves absorption by increasing bioavailability. The Drosophila melanogaster model of Parkinson's disease must be helpful in investigating potential neuroprotective effects. Four groups of four-day-old flies were exposed to: (1) control; (2) diet containing rotenone (500 μM); (3) β-carotene-loaded nanoparticles (20 μM); (4) β-carotene-loaded nanoparticles and rotenone for 7 days. Then, the percentage of survival, geotaxis tests, open field, aversive phototaxis and food consumption were evaluated. At the end of the behaviors, the analyses of the levels of reactive species (ROS), thiobarbituric acid reactive substances (TBARS), catalase (CAT) and superoxide dismutase (SOD) activity was carried out, as well as an evaluation of the levels of dopamine and acetylcholinesterase (AChE) activity, in the head of flies. Nanoparticles loaded with β-carotene were able to improve motor function, memory, survival and also restored the oxidative stress indicators (CAT, SOD, ROS and TBARS), dopamine levels, AChE activity after exposure to rotenone. Overall, nanoparticles loaded with β-carotene showed significant neuroprotective effect against damage induced by the Parkinson-like disease model, emerging as a possible treatment. Overall, β-carotene-loaded nanoparticles presented significant neuroprotective effect against damage induced by model of Parkinson-like disease, emerging as a possible treatment.

Transcranial direct current stimulation effects in the pain threshold and in oxidative stress parameters of neuropathic pain rats

Transcranial direct current stimulation (tDCS) can modulate cortical excitability and relieve neuropathic pain (NP), but the role of several biomarkers in this process is not well understood. This study aimed to analyze the effects of tDCS on biochemical parameters in rats with neuropathic pain (NP) induced by chronic constriction injury (CCI) of the right sciatic nerve. Eighty-eight male 60-day-old Wistar rats were divided into nine groups: control (C), control-electrode off (CEoff), control-tDCS (C-tDCS), sham-lesion (SL), sham-lesion electrode off (SLEoff), sham-lesion (SL-tDCS), lesion (L), lesion electrode off (LEoff), and lesion-tDCS (L-tDCS). After NP establishment, 20-minute bimodal tDCS for 8 consecutive days was applied to the rats. Fourteen days after the induction of NP, rats developed mechanical hyperalgesia with a decreased threshold, and at the end of treatment, an increase in the pain threshold was observed in NP rats. In addition, NP rats had increased levels of reactive species (RS) in the prefrontal cortex, while superoxide dismutase (SOD) activity was decreased in NP rats. In the spinal cord, nitrite levels and glutathione-S-transferase (GST) activity decreased in the L-tDCS group, and it was observed that increased levels in total sulfhydryl content for neuropathic pain rats were reversed by tDCS. In serum analyses, the neuropathic pain model increased the levels of RS and thiobarbituric acid-reactive substances (TBARS) and decreased the activity of butyrylcholinesterase (BuChE). In conclusion, bimodal tDCS increased total sulfhydryl content in the spinal cord of rats with neuropathic pain, positively modulating this parameter.

Irinotecan-gut microbiota interactions and the capability of probiotics to mitigate Irinotecan-associated toxicity

BackgroundIrinotecan is a chemotherapeutic agent used to treat a variety of tumors, including colorectal cancer (CRC). In the intestine, it is transformed into SN-38 by gut microbial enzymes, which is responsible for its toxicity during excretion.ObjectiveOur study highlights the impact of Irinotecan on gut microbiota composition and the role of probiotics in limiting Irinotecan-associated diarrhea and suppressing gut bacterial β-glucuronidase enzymes.Material and methodsTo investigate the effect of Irinotecan on the gut microbiota composition, we applied 16S rRNA gene sequencing in three groups of stool samples from healthy individuals, colon cancer, and Irinotecan treated patients (n = 5/group). Furthermore, three Lactobacillus spp.; Lactiplantibacillus plantarum (L. plantarum), Lactobacillus acidophilus (L. acidophilus), Lacticaseibacillus rhamnosus (L. rhamnosus) were used in a single and mixed form to in-vitro explore the effect of probiotics on the expression of β-glucuronidase gene from E. coli. Also, probiotics were introduced in single and mixed forms in groups of mice before the administration of Irinotecan, and their protective effects were explored by assessing the level of reactive oxidative species (ROS) as well as studying the concomitant intestinal inflammation and apoptosis.ResultsThe gut microbiota was disturbed in individuals with colon cancer and after Irinotecan treatment. In the healthy group, Firmicutes were more abundant than Bacteriodetes, which was the opposite in the case of colon-cancer or Irinotecan treated groups. Actinobacteria and Verrucomicrobia were markedly present within the healthy group, while Cyanobacteria were noted in colon-cancer and the Irinotecan-treated groups. Enterobacteriaceae and genus Dialister were more abundant in the colon-cancer group than in other groups. The abundance of Veillonella, Clostridium, Butryicicoccus, and Prevotella were increased in Irinotecan-treated groups compared to other groups. Using Lactobacillus spp. mixture in mice models significantly relieved Irinotecan-induced diarrhea through the reduction of both β-glucuronidase expression and ROS, in addition to guarding gut epithelium against microbial dysbiosis and proliferative crypt injury.ConclusionsIrinotecan-based chemotherapy altered intestinal microbiota. The gut microbiota participates greatly in determining both the efficacy and toxicity of chemotherapies, of which the toxicity of Irinotecan is caused by the bacterial ß-glucuronidase enzymes. The gut microbiota can now be aimed and modulated to promote efficacy and decrease the toxicity of chemotherapeutics. The used probiotic regimen in this study lowered mucositis, oxidative stress, cellular inflammation, and apoptotic cascade induction of Irinotecan.

Benzeneseleninic Acid Promoting the Selenofunctionalization of 2- Naphthol Derivatives: Synthesis and Antioxidant Activity of 1- Organoselanyl-naphthalen-2-ols

Background: We report the synthesis and antioxidant evaluation of 1-organoselanyl-naphthalen-2-ols. The title compounds were selectively prepared through the selenofunctionalization of 2-naphthol derivatives using benzeneseleninic acids as a selenium source. Objective: We aimed at synthesizing 1-organoselanyl-naphthalen-2-ol by functionalizing 2-naphthol with arylseleninic acids using glycerol as the solvent and further evaluating the pharmacological activity. Methods: The products were synthesized at 70 °C using glycerol as a solvent in an oil bath. Reactive species (RS) tests, DPPH radical scavenging activity assay, and ABTS+ radical scavenging activity assay were performed. A statistical analysis of the data was performed. Results and Discussion: A total of fifteen 1-organoselanyl-naphthalen-2-ols were selectively obtained in yields of 58-95% in 0.25-48 h of reaction. Additionally, all the synthesized seleno-derivatives exhibited antioxidant activity, as revealed by their ability to scavenge DPPH and ABTS+ radicals and reduce the reactive species (RS) levels. Conclusion: The synthesis of 1-arylselanyl-naphthalen-2-ols was developed under mild reaction conditions using benzeneseleninic acid derivatives in reactions with 2-naphthol. All synthesized 2-naphthol derivatives exhibited antioxidant activity, as revealed by their DPPH and ABTS+ radical scavenging activity and reduced RS levels.

Ferulic acid-loaded nanostructure prevents morphine reinstatement: the involvement of dopamine system, NRF2, and ΔFosB in the striatum brain area of rats.

Morphine is among the most powerful analgesics and pain-relieving agents. However, its addictive properties limit their medical use because patients may be susceptible to abuse and reinstatement. Morphine addiction occurs because of dopamine release in the mesolimbic brain area, implying in an increase in oxidative stress. Ferulic acid (FA), a phenolic phytochemical found in a variety of foods, has been reported to exert antioxidant and neuroprotective effects; however, its low bioavailability makes its nano-encapsulated form a promising alternative. This study aimed to evaluate the protective effects of a novel nanosystem with FA on morphine reinstatement and the consequent molecular neuroadaptations and oxidative status in the mesolimbic region. Rats previously exposed to morphine in conditioned place preference (CPP) paradigm were treated with ferulic acid-loaded nanocapsules (FA-Nc) or nonencapsulated FA during morphine-preference extinction. Following the treatments, animals were re-exposed to morphine to induce the reinstatement. While morphine-preference extinction was comparable among all experimental groups, FA-Nc treatment prevented morphine reinstatement. In the dorsal striatum, while morphine exposure increased lipid peroxidation (LP) and reactive species (RS), FA-Nc decreased LP and FA decreased RS levels. Morphine exposure increased the dopaminergic markers (D1R, D3R, DAT) and ΔFosB immunoreactivity in the ventral striatum; however, FA-Nc treatment decreased D1R, D3R, and ΔFosB and increased D2R, DAT, and NRF2. In conclusion, FA-Nc treatment prevented the morphine reinstatement, promoted antioxidant activity, and modified the dopaminergic neurotransmission, NRF2, and ΔFosB, what may indicate a neuroprotective and antioxidant role of this nanoformulation.

Rosmarinic Acid-Crosslinked Supramolecular Nanoassembly with Self-Regulated Photodynamic and Anti-Metastasis Properties for Synergistic Photoimmunotherapy.

A primary concern about photodynamic therapy (PDT) is its inability to regulate thegeneration levels of reactive oxidative species (ROS) based on the complex microenvironment, resulting in theimpairment toward normal tissues and immunosuppression. Besides, tumor metastasis also compromises PDT's efficacy and drives mortality. However, it is very challenging to achieve such two goals within one nanosystem. Here, the nanoassembly (CPR) with self-regulated photodynamic and antimetastasis properties comprises three parts: chlorin e6-conjugated β-cyclodextrin (CD-Ce6) acts as the main PDT agent and ferrocene (Fc)-terminated phenylboronic acid-containing conjugates entering into the cavity of CD-Ce6, as well as rosmarinic acid (RA)-boronic acid crosslinked shell. Compared with non-crosslinked counterpart, CPR displays better stability and enhanced tumor accumulation. Under laser irradiation, CPR generates plenty of ROS to damage tumor cells and induce immunogenic cell death. Mildly acidic TME partly cleaves the crosslinkers to dissociate antioxidant RAs from micelles, which together with Fc in CPR scavenge PDT-induced ROS in the TME. By contrast, under acidic lysosomal conditions, Fc catalyzes abundant H2 O2 in tumor cells to produce highly cytotoxic •OH, while RA continuously reduces ferroptosis-generated Fc+ into Fc, both to augment the PDT efficacy in tumor cells. CPR also remarkably hinders the epithelial-mesenchymal transition to prevent the lung metastasis.

Elevated Fibroblast Growth Factor 23 Impairs Endothelial Function through the NF-κB Signaling Pathway.

Roles of fibroblast growth factor 23 (FGF23) in endothelial dysfunction remain controversial, and evidence from population-based studies is lacking. The present study aimed to explore the effects of FGF23 on endothelial dysfunction on the basis of both clinical data of patients with coronary artery disease (CAD) and the in vitro research in human umbilical vein endothelial cells (HUVECs). A total of 321 CAD patients were enrolled after coronary angiography, brachial artery flow-mediated dilation (FMD) was assessed using ultrasound equipment. Serum FGF23, nitric oxide (NO), and endothelin-1 (ET-1) were detected via enzyme-linked immunosorbent assay. Apoptosis was determined using the annexin V-fluorescein isothiocyanate/propidium lodide apoptosis detection kit. Cell migration was evaluated by wound healing and transwell migration assays. Reactive oxide species levels were determined using fluorescent probes, and NF-κB p65 nuclear translocation was assessed via immunofluorescence. Serum FGF23 was significantly increased in CAD patients combined with severe endothelial dysfunction (FMD ＜2%) compared to those with FMD ≥ 2% (P＜0.001). Furthermore, the levels of FGF23 were negatively correlated with NO, whereas positively correlated with ET-1 both in unadjusted analysis and multivariate-adjusted analysis. In HUVECs, FGF23 interfered with the bioavailability of NO via increased oxidative stress. Moreover, FGF23 directly impaired the endothelium by promoting HUVECs apoptosis and attenuating the migration of HUVECs. Additional experiments showed that FGF23 induced endothelial injury through activation of the NF-κB signaling pathway. Elevated FGF23 is clinically associated with endothelial dysfunction in CAD patients, and FGF23 impairs endothelial function through activation of the NF-κB signaling pathway.

Redox-Dependent Modulation of Human Liver Progenitor Cell Line Fate.

Redox homeostasis is determinant in the modulation of quiescence/self-renewal/differentiation of stem cell lines. The aim of this study consisted of defining the impact of redox modifications on cell fate in a human hepatic progenitor line. To achieve this, the HepaRG cell line, which shows oval ductular bipotent characteristics, was used. The impact of redox status on the balance between self-renewal and differentiation of HepaRG cells was investigated using different methodological approaches. A bioinformatic analysis initially proved that the trans-differentiation of HepaRG toward bipotent progenitors is associated with changes in redox metabolism. We then exposed confluent HepaRG (intermediate differentiation phase) to oxidized (H2O2) or reduced (N-acetylcysteine) extracellular environments, observing that oxidation promotes the acquisition of a mature HepaRG phenotype, while a reduced culture medium stimulates de-differentiation. These results were finally confirmed through pharmacological modulation of the nuclear factor (erythroid-derived 2)-like 2 (NRF2), a principal modulator of the antioxidant response, in confluent HepaRG. NRF2 inhibition led to intracellular pro-oxidative status and HepaRG differentiation, while its activation was associated with low levels of reactive species and de-differentiation. In conclusion, this study shows that both intra- and extracellular redox balance are crucial in the determination of HepaRG fate. The impact of redox status in the differentiation potential of HepaRG cells is significant on the utilization of this cell line in pre-clinical studies.

Structural characterization and anti-osteoporosis effect of an arabinomannan from Anemarrhena asphodeloides Bge

To discover the polysaccharide with anti-diabetic osteoporosis (DOP) activity and clarify its structure, an arabinomannan (PAAP-1B) with a molecular weight of 14.0 kDa was isolated from Anemarrhena asphodeloides Bge. using column chromatography. It consists of arabinose, mannose, and galactose in a molar ratio of 6:3:1. PAAP-1B has a backbone composed of 1,5-α-Araf, 1,4-β-Manp, and 1,6-β-Galp residues that are branched at C3 of α-Araf and β-Galp residues. The side chains are T-α-Araf, T-α-Manp, T-β-Galp, and 1,6-β-Galp. PAAP-1B attenuated DOP and reduced ferroptosis in the femurs and tibias of alloxan-induced mice. It also suppressed ferroptosis in advanced glycation end product-induced osteoblasts by decreasing 4-hydroxynonenal, malondialdehyde, mitochondrial reactive oxidative species levels, and lipid peroxidation, while reversing the downregulation of solute carrier family 7 membrane 11 and glutathione expression.

Connexin 43 hemichannels regulate mitochondrial ATP generation, mobilization, and mitochondrial homeostasis against oxidative stress.

Oxidative stress is a major risk factor that causes osteocyte cell death and bone loss. Prior studies primarily focus on the function of cell surface expressed Cx43 channels. Here, we reported a new role of mitochondrial Cx43 (mtCx43) and hemichannels (HCs) in modulating mitochondria homeostasis and function in bone osteocytes under oxidative stress. In murine long bone osteocyte-Y4 cells, the translocation of Cx43 to mitochondria was increased under H₂O₂-induced oxidative stress. H₂O₂ increased the mtCx43 level accompanied by elevated mtCx43 HC activity, determined by dye uptake assay. Cx43 knockdown (KD) by the CRISPR-Cas9 lentivirus system resulted in impairment of mitochondrial function, primarily manifested as decreased ATP production. Cx43 KD had reduced intracellular reactive oxidative species levels and mitochondrial membrane potential. Additionally, live-cell imaging results demonstrated that the proton flux was dependent on mtCx43 HCs because its activity was specifically inhibited by an antibody targeting Cx43 C-terminus. The co-localization and interaction of mtCx43 and ATP synthase subunit F (ATP5J2) were confirmed by Förster resonance energy transfer and a protein pull-down assay. Together, our study suggests that mtCx43 HCs regulate mitochondrial ATP generation by mediating K⁺, H⁺, and ATP transfer across the mitochondrial inner membrane and the interaction with mitochondrial ATP synthase, contributing to the maintenance of mitochondrial redox levels in response to oxidative stress.

Bioreactor expansion reconfigures metabolism and extracellular vesicle biogenesis of human adipose-derived stem cells in vitro

Human mesenchymal stem cells (hMSCs), including human adipose tissue-derived stem cells (hASCs), as well as the secreted extracellular vesicles (EVs), are promising therapeutics in treating inflammatory and neural degenerative diseases. However, prolonged expansion can lead to cellular senescence characterized by a gradual loss of self-renewal ability while altering secretome composition and EV generation. Additionally, hMSCs are highly sensitive to biophysical microenvironment in bioreactor systems utilized in scaling production. In this study, hASCs grown on Plastic Plus or Synthemax II microcarriers in a spinner flask bioreactor (SFB) system were compared to traditional 2D culture. The SFB microenvironment was found to increase the expression of genes associated with hASC stemness, nicotinamide adenine dinucleotide (NAD+) metabolism, glycolysis, and the pentose phosphate pathway as well as alter cytokine secretion (e.g., PGE2 and CXCL10). Elevated reactive oxidative species levels in hASCs of SFB culture were observed without increasing rates of cellular senescence. Expression levels of Sirtuins responsible for preventing cellular senescence through anti-oxidant and DNA repair mechanisms were also elevated in SFB cultures. In particular, the EV biogenesis genes were significantly upregulated (3–10 fold) and the EV production increased 40 % per cell in SFB cultures of hASCs. This study provides advanced understanding of hASC sensitivity to the bioreactor microenvironment for EV production and bio-manufacturing towards the applications in treating inflammatory and neural degenerative diseases.