OBJECTIVE: We aimed to resolve the uncertainty as to whether betulin exerted neuroprotection on early brain injury (EBI) caused by subarachnoid hemorrhage (SAH), and to investigate the related molecular mechanisms.
METHODS: Bioinformatic analysis was performed to pre-study the differently expressed genes (DEGs) and the possible signaling pathways. Rat and cellular model of SAH were introduced in this study, and betulin, an activator of DJ-1 protein, was administered to reveal the effect. Gross assessment regarding mortality, neurofunctions, SAH grade, brain water content (BWC) along with multiple cellular and molecular studies in vivo or/and in vitro such as immunofluorescence (IF) staining, western blot (WB), reactive oxygen species (ROS) assay, and flow cytometry (FCM) were all conducted after SAH induction to verify the protective effect and the relevant mechanisms of DJ-1 in diverse levels. In addition, MK2206 (selective inhibitor of Akt) and iRNADj-1 (interfering RNA to Dj-1) were utilized to confirm the mechanisms of the effect.
RESULTS: The data from our study showed that DJ-1 protein was moderately expressed in neurons, microglia, and astrocytes; its level in brain tissue elevated and peaked at 24-72 h after SAH induction. Betulin could efficaciously induce the expression of DJ-1 which in turn activated Akt and Bcl-2, and anti-oxidative enzymes SOD2 and HO-1, functioning to reduce the activation of cleaved caspase-3 (c-Casp-3) and reactive oxygen species (ROS). The induced DJ-1 could upregulate the expression of Nrf2. However, Akt seemed no direct effect on elevating the expression of Nrf2. DJ-1 alone could as well activate Akt-independent antiapoptotic pathway via suppressing the activation of caspase-8 (Casp-8).
CONCLUSIONS: Betulin which was a potent agonist of DJ-1 had the ability to induce its expression in brain tissue. DJ-1 had neuroprotective effect on EBI through comprehensive mechanisms, including facilitating intrinsic and extrinsic antiapoptotic pathway, and reducing oxidative injury by upregulating the expression of redox proteins. Betulin as an inexpensive drug showed the potential for SAH treatment.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory lung disease associated with high morbidity and mortality worldwide. Oxidative injury and mitochondrial dysfunction in the airway epithelium are major events in COPD progression.
RESULTS: The therapeutic effects of Progesterone (P4) were investigated in vivo and in vitro in this study. In vivo, in a cigarette smoke (CS) exposure-induced COPD mouse model, P4 treatment significantly ameliorated CS exposure-induced physiological and pathological characteristics, including inflammatory cell infiltration and oxidative injury, in a dose-dependent manner. The c-MYC/SIRT1/PGC-1α pathway is involved in the protective function of P4 against CS-induced COPD. In vitro, P4 co-treatment significantly ameliorated H2O2-induced oxidative injury and mitochondrial dysfunctions by promoting cell proliferation, increasing mitochondrial membrane potential, decreasing ROS levels and apoptosis, and increasing ATP content. Moreover, P4 co-treatment partially attenuated H2O2-caused inhibition in Nrf1, Tfam, Mfn1, PGR-B, c-MYC, SIRT1, and PGC-1α levels. In BEAS-2B and ASM cells, the c-MYC/SIRT1 axis regulated P4\'s protective effects against H2O2-induced oxidative injury and mitochondrial dysfunctions.
CONCLUSIONS: P4 activates the c-MYC/SIRT1 axis, ameliorating CS-induced COPD and protecting both airway epithelial cells and smooth muscle cells against H2O2-induced oxidative damage. PGC-1α and downstream mitochondrial signaling pathways might be involved.

Diabetic peripheral neuropathy (DPN) is caused by several factors, including reactive free oxygen radicals (ROS)-induced excessive Ca2+ influx. Transient receptor potential (TRP) vanilloid 4 (TRPV4) is a member of the Ca2+-permeable TRP superfamily. Resveratrol (RESV) has been extensively utilized in TRP channel regulation due to its pharmacological properties, which include antioxidant and TRP inhibitory effects. The protective function of RESV and the contribution of TRPV4 to streptozotocin (STZ)-induced neuropathic pain in mice are still unclear. Here, we evaluated the effects of RESV through the modulation of TRPV4 on Ca2+ influx, ROS-mediated pain, apoptosis, and oxidative damage in the mouse dorsal root ganglion (DRGs). From the 32 mice, four groups were induced: control, RESV, STZ, and STZ + RESV. We found that the injection of RESV reduced the changes caused by the STZ-induced stimulation of TRPV4, which in turn increased mechanical/thermal neuropathic pain, cytosolic Ca2+ influx, TRPV4 current density, oxidants (lipid peroxidation, mitochondrial ROS, and cytosolic ROS), and apoptotic markers (caspase-3, -8, and -9). The RESV injection also increased the STZ-mediated reduction of viability of DRG and the amounts of glutathione, glutathione peroxidase, vitamin A, β-carotene, and vitamin E in the brain, erythrocytes, plasma, liver, and kidney. All of these findings suggest that TRPV4 stimulation generates oxidative neurotoxicity, neuropathic pain, and apoptosis in the STZ-induced diabetic mice. On the other hand, neurotoxicity and apoptosis were reduced due to the downregulation of TRPV4 carried out through the RESV injection.

Ferulic acid (Fer) and geraniol (Ger) are natural compounds whose antioxidant and anti-inflammatory activity confer beneficial properties, such as antibacterial, anticancer, and neuroprotective effects. However, the short half-lives of these compounds impair their therapeutic activities after conventional administration. We propose, therefore, a new prodrug (Fer-Ger) obtained by a bio-catalyzed ester conjugation of Fer and Ger to enhance the loading of solid lipid microparticles (SLMs) designed as Fer-Ger delivery and targeting systems. SLMs were obtained by hot emulsion techniques without organic solvents. HPLC-UV analysis evidenced that Fer-Ger is hydrolyzed in human or rat whole blood and rat liver homogenates, with half-lives of 193.64 ± 20.93, 20.15 ± 0.75, and 3.94 ± 0.33 min, respectively, but not in rat brain homogenates. Studies on neuronal-differentiated mouse neuroblastoma N2a cells incubated with the reactive oxygen species (ROS) inductor H2O2 evidenced the Fer-Ger ability to prevent oxidative injury, despite the fact that it appears ROS-promoting. The amounts of Fer-Ger encapsulated in tristearin SLMs, obtained in the absence or presence of glucose, were 1.5 ± 0.1%, allowing the control of the prodrug release (glucose absence) or to sensibly enhance its water dissolution rate (glucose presence). These new \"green\" carriers can potentially prolong the beneficial effects of Fer and Ger or induce neuroprotection as nasal formulations.

The small intestine is important to the digestion and absorption of rumen undegradable nutrients, as well as the barrier functionality and immunological responses in ruminants. Oxidative stress induces a spectrum of pathophysiological symptoms and nutritional deficits, causing various gastrointestinal ailments. Previous studies have shown that nicotinamide (NAM) has antioxidant properties, but the potential mechanism has not been elucidated. The aim of this study was to explore the effects of NAM on hydrogen peroxide (H2O2)-induced oxidative injury in bovine intestinal epithelial cells (BIECs) and its potential mechanism. The results showed that NAM increased the cell viability and total antioxidant capacity (T-AOC) and decreased the release of lactate dehydrogenase (LDH) in BIECs challenged by H2O2. The NAM exhibited increased expression of catalase, superoxide dismutase 2, and tight junction proteins. The expression of autophagy-related proteins was increased in BIECs challenged by H2O2, and NAM significantly decreased the expression of autophagy-related proteins. When an autophagy-specific inhibitor was used, the oxidative injury in BIECs was not alleviated by NAM, and the T-AOC and the release of LDH were not affected. Collectively, these results indicated that NAM could alleviate oxidative injury in BIECs by enhancing antioxidant capacity and increasing the expression of tight junction proteins, and autophagy played a crucial role in the alleviation.

Deoxynivalenol (DON) is a prevalent contaminant in feed and food, posing a serious threat to the health of both humans and animals. The pig stands as an ideal subject for the study of DON due to its recognition as the most susceptible animal to DON. In this study, the IPEC-J2 cells were utilized as an in vitro model to explore the potential of SeMet in alleviating the intestinal toxicity and oxidative injury in intestinal epithelial cells when exposed to DON. Cells were treated either with or without 4.0 μM SeMet, in combination with or without a simultaneous treatment with 0.5 μg/mL DON, for a duration of 24 h. Then, cells or related samples were analyzed for cell proliferation, lactate dehydrogenase (LDH) release, reactive oxygen species (ROS) level, gene expressions, and protein expressions. The results showed that SeMet mitigated the cellular toxicity caused by DON, evidenced by elevated cell proliferation and the reduced LDH release of IPEC-J2 cells in the SeMet + DON group vs. the DON group. Moreover, the SeMet treatment markedly promoted antioxidant functions and decreased the oxidative injury in IPEC-J2 cell, which is indicated by the decreased ROS level and up-regulated mRNA levels of GPX1, TXNRD1, Nrf2, and GCLC in IPEC-J2 cells in the SeMet + DON group vs. the DON group. However, in both the absence and presence of exposure to DON, the SeMet treatment did not affect the protein expression of MAPK (JNK, Erk1/2, and P38) and phosphorylated MAPK (p-JNK, p-Erk1/2, and p-P38) in IPEC-J2 cells. Collectively, SeMet alleviated the DON-induced oxidative injury in porcine intestinal epithelial cells independent of the MAPK pathway regulation.

This research evaluated the impacts of selenomethionine (Se-Met) on hepatic functions, oxidative stress, mitochondrial function, and apoptosis of piglets fed deoxynivalenol (DON)-contaminated diets. Twenty-four piglets were allocated four dietary treatments (n = 6) in a 28-day feeding trial. The four treatments included the control group, which received 0.3 mg/kg of Se (as Se-Met) without DON treatment, and the DON treatment groups received 0, 0.3, or 0.5 mg/kg Se as Se-Met. A dietary addition of 0.5 mg/kg Se improved liver pathology and reduced serum aspartate aminotransferase and lactate dehydrogenase levels in piglets fed DON-contaminated diets. Furthermore, 0.5 mg/kg Se mitigated the oxidative stress and apoptosis of piglets fed DON-contaminated diets, as indicated by the decreased reactive oxygen species level, and the down-regulated mRNA levels of NRF-1, Bax, and CASP9 in the liver. Importantly, 0.5 mg/kg Se enhanced the hepatic antioxidant capacity, as evidenced by increased hepatic total antioxidant capacity, catalase, glutathione peroxidase, and total superoxide dismutase activities, as well as the up-regulated mRNA levels of Nrf2, Gclm, NQO1, SOD1, and GPX1 in the liver. Moreover, 0.5 mg/kg Se down-regulated the p-JNK protein level in the liver of piglets fed DON-contaminated diets. Collectively, Se-Met supplementation mitigated liver dysfunction, oxidative injury, and apoptosis through enhancing antioxidant capacity and inhibiting the JNK MAPK pathway in piglets fed DON-contaminated diets.

Atherosclerosis constitutes a proverbial pathogenic mechanism for cardio-cerebrovascular disease that accounts for the most common cause of disability and morbidity for human health worldwide. Endothelial dysfunction and inflammation are the key contributors to the progression of atherosclerosis. Glutaredoxin 2 (GLRX2) is abundantly existed in various tissues and possesses a range of pleiotropic efficacy including anti-oxidative and anti-inflammatory responses. However, its role in atherosclerosis is still undefined. Here, down-regulation of GLRX2 was validated in lipopolysaccha (LPS)-induced vascular endothelial cells (HUVECs). Moreover, elevation of GLRX2 reversed the inhibition of cell viability in LPS-treated HUVECs and decreased LPS-induced increases in cell apoptosis and caspase-3 activity. Additionally, enhancement of GLRX2 expression antagonized oxidative stress in HUVECs under LPS exposure by inhibiting ROS, lactate dehydrogenase and malondialdehyde production and increased activity of anti-oxidative stress superoxide dismutase. Notably, GLRX2 abrogated LPS-evoked transcripts and releases of pro-inflammatory cytokine (TNF-α, IL-6, and IL-1β), chemokine MCP-1 and adhesion molecule ICAM-1 expression. Furthermore, the activation of Nrf2/HO-1 signaling was demonstrated in LPS-stimulated HUVECs. Importantly, blockage of the Nrf2 pathway counteracted the protective roles of GLRX2 in LPS-triggered endothelial cell injury, oxidative stress and inflammatory response. Thus, these data reveal that GLRX2 may alleviate the progression of atherosclerosis by regulating vascular endothelial dysfunction and inflammation via the activation of the Nrf2 signaling, supporting a promising therapeutic approach for atherosclerosis and its complications.
UNASSIGNED: The online version contains supplementary material available at 10.1007/s10616-023-00606-x.

BACKGROUND: Liver fibrosis is a formidable global medical challenge, with no effective clinical treatment currently available. Yinhuang granule (YHG) is a proprietary Chinese medicine comprising Scutellariae Radix and Lonicerae Japonicae Flos. It is frequently used for upper respiratory tract infections, pharyngitis, as well as acute and chronic tonsillitis.
OBJECTIVE: To investigate the potential of YHG in alleviating carbon tetrachloride (CCl4)-induced liver fibrosis in mice.
METHODS: To induce a hepatic fibrosis model in mice, this study involved intraperitoneal injections of 2 mL/kg of CCl4 twice a week for 4 wk. Meanwhile, liver fibrosis mice in the low dose of YHG (0.4 g/kg) and high dose of YHG (0.8 g/kg) groups were orally administered YHG once a day for 4 wk. Serum alanine/aspartate aminotransferase (ALT/AST) activity and liver hydroxyproline content were detected. Sirius red and Masson\'s trichrome staining assay were conducted. Real-time polymerase chain reaction, western-blot and enzyme-linked immunosorbent assay were conducted. Liver glutathione content, superoxide dismutase activity level, reactive oxygen species and protein carbonylation amount were detected.
RESULTS: The administration of YHG ameliorated hepatocellular injury in CCl4-treated mice, as reflected by decreased serum ALT/AST activity and improved liver histological evaluation. YHG also attenuated liver fibrosis, evident through reduced liver hydroxyproline content, improvements in Sirius red and Masson\'s trichrome staining, and lowered serum hyaluronic acid levels. Furthermore, YHG hindered the activation of hepatic stellate cells (HSCs) and ameliorated oxidative stress injury and inflammation in liver from CCl4-treated mice. YHG prompted the nuclear accumulation of nuclear factor erythroid 2-related factor 2 (Nrf2) and upregulated the expression of Nrf2-dependent downstream antioxidant genes. In addition, YHG promoted mitochondrial biogenesis in liver from CCl4-treated mice, as demonstrated by increased liver adenosine triphosphate content, mitochondrial DNA levels, and the expression of peroxisome proliferator-activated receptor gamma coactivator 1 alpha and nuclear respiratory factor 1.
CONCLUSIONS: YHG effectively attenuates CCl4-induced liver fibrosis in mice by inhibiting the activation of HSCs, reducing inflammation, alleviating liver oxidative stress damage through Nrf2 activation, and promoting liver mitochondrial biogenesis.

Antioxidant peptides were purified from Hydrilla verticillata (Linn. f.) Royle (HVR) protein hydrolysate by ultrafiltration, gel filtration chromatography, and semipreparative reversed-phase HPLC and identified by UPLC-ESI-MS/MS. Therein, TCLGPK and TCLGER were selected to be synthesized, and they displayed desirable radical-scavenging activity to ABTS (99.20 ± 0.56-99.20 ± 0.43%), DPPH (97.32 ± 0.59-97.56 ± 0.97%), hydroxyl radical (54.32 ± 1.27-70.42 ± 2.01%), and superoxide anion (42.93 ± 1.46-52.62 ± 1.11%) at a concentration of 0.96 μmol/mL. They possessed a cytoprotective effect against H2O2-induced oxidative stress in HepG2 cells in a dose-dependent manner. 1.6 μmol/mL of the two peptides could perfectly protect HepG2 cells from H2O2-induced injury. The TCLGPK exhibited higher antioxidant activity and cytoprotective effect than TCLGER. Western blot and molecular docking results indicated that the two peptides achieved antioxidant ability and cytoprotective effect by combining with Kelch-like ECH-associated protein 1 (Keap1) to activate the Keap1-nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response elements signaling pathway, leading to the activity and expression of the related antioxidases in the pathway significantly up-regulating and the intracellular reactive oxygen species level, lipid peroxidation, and cell apoptosis rate significantly down-regulating.