This study aimed to explore whether chronic l-lactate exposure could affect the peripheral tissues of mice and to determine the underlying pathogenesis. Herein, male C57BL/6 mice were divided into control and l-lactate groups. After l-lactate treatment for eight weeks (1 g/kg), metabolic changes in liver, kidney, muscle, and serum samples were determined by 1H nuclear magnetic resonance (1H NMR)-based metabolomics. Additionally, organ function was evaluated by serum biochemical and histopathological examinations. Reactive oxygen species (ROS) levels were measured using dihydroethidium staining; levels of signals involved in lactate metabolism and ROS-related pathways were detected using western blotting or polymerase chain reaction. Apoptosis was detected by TUNEL-fluorescence staining. Metabolomic analysis revealed that l-lactate mice showed decreased levels of glutathione (GSH), taurine, ATP, and increased glucose content, compared to control mice. Furthermore, l-lactate mice presented significantly higher serum levels of alanine aminotransferase and aspartate aminotransferase and increased glycogen content in hepatic tissues, compared to control mice. l-lactate mice also had a greater number of apoptotic nuclei in the livers than controls. Moreover, l-lactate exposure reduced mRNA and protein levels of superoxide dismutase-2 and c-glutamylcysteine ligase, elevated levels of cytochrome P450 2E1 and NADPH oxidase-2, and increased the protein expressions of LDHB, Bax/Bcl-2, cleaved caspase-3, and sirtuin-1 in hepatic tissues. Together, these results indicate that chronic l-lactate exposure increases oxidative stress and apoptosis in hepatocytes via upregulation of Bax/Bcl-2 expression and the consequent mitochondrial cytochrome-C release and caspase-3 activation, which contributes to the pathogenesis of hepatic dysfunction.

UNASSIGNED: The incidence of ischemic cerebrovascular disease is increasing in recent years and has been one of the leading causes of neurological dysfunction and death. Ginsenoside Rg1 has been found to protect against neuronal damage in many neurodegenerative diseases. However, the effect and mechanism by which Rg1 protects against cerebral ischemia-reperfusion injury (CIRI) are not fully understood. Here, we report the neuroprotective effects of Rg1 treatment on CIRI and its possible mechanisms in mice.
UNASSIGNED: A bilateral common carotid artery ligation was used to establish a chronic CIRI model in mice. HT22 cells were treated with Rg1 after OGD/R to study its effect on [Ca2+]i. The open-field test and pole-climbing experiment were used to detect behavioral injury. The laser speckle blood flowmeter was used to measure brain blood flow. The Nissl and H&E staining were used to examine the neuronal damage. The Western blotting was used to examine MAP2, PSD95, Tau, p-Tau, NOX2, PLC, p-PLC, CN, NFAT1, and NLRP1 expression. Calcium imaging was used to test the level of [Ca2+]i.
UNASSIGNED: Rg1 treatment significantly improved cerebral blood flow, locomotion, and limb coordination, reduced ROS production, increased MAP2 and PSD95 expression, and decreased p-Tau, NOX2, p-PLC, CN, NFAT1, and NLRP1 expression. Calcium imaging results showed that Rg1 could inhibit calcium overload and resist the imbalance of calcium homeostasis after OGD/R in HT22 cells.
UNASSIGNED: Rg1 plays a neuroprotective role in attenuating CIRI by inhibiting oxidative stress, calcium overload, and neuroinflammation.

Although several artificial nanotherapeutics have been approved for practical treatment of metastatic breast cancer, their inefficient therapeutic outcomes, serious adverse effects, and high cost of mass production remain crucial challenges. Herein, we developed an alternative strategy to specifically trigger apoptosis of breast tumors and inhibit their lung metastasis by using natural nanovehicles from tea flowers (TFENs). These nanovehicles had desirable particle sizes (131 nm), exosome-like morphology, and negative zeta potentials. Furthermore, TFENs were found to contain large amounts of polyphenols, flavonoids, functional proteins, and lipids. Cell experiments revealed that TFENs showed strong cytotoxicities against cancer cells due to the stimulation of reactive oxygen species (ROS) amplification. The increased intracellular ROS amounts could not only trigger mitochondrial damage, but also arrest cell cycle, resulting in the in vitro anti-proliferation, anti-migration, and anti-invasion activities against breast cancer cells. Further mice investigations demonstrated that TFENs after intravenous (i.v.) injection or oral administration could accumulate in breast tumors and lung metastatic sites, inhibit the growth and metastasis of breast cancer, and modulate gut microbiota. This study brings new insights to the green production of natural exosome-like nanoplatform for the inhibition of breast cancer and its lung metastasis via i.v. and oral routes.

The medicinal uses of Calotropis procera are diverse, yet some of them are based on effects that still lack scientific support. Control of diabetes is one of them. Recently, latex proteins from C. procera latex (LP) have been shown to promote in vivo glycemic control by the inhibition of hepatic glucose production via AMP-activated protein kinase (AMPK). Glycemic control has been attributed to an isolated fraction of LP (CpPII), which is composed of cysteine peptidases (95%) and osmotin (5%) isoforms. Those proteins are extensively characterized in terms of chemistry, biochemistry and structural aspects. Furthermore, we evaluated some aspects of the mitochondrial function and cellular mechanisms involved in CpPII activity. The effect of CpPII on glycemic control was evaluated in fasting mice by glycemic curve and glucose and pyruvate tolerance tests. HepG2 cells was treated with CpPII, and cell viability, oxygen consumption, PPAR activity, production of lactate and reactive oxygen species, mitochondrial density and protein and gene expression were analyzed. CpPII reduced fasting glycemia, improved glucose tolerance and inhibited hepatic glucose production in control animals. Additionally, CpPII increased the consumption of ATP-linked oxygen and mitochondrial uncoupling, reduced lactate concentration, increased protein expression of mitochondrial complexes I, III and V, and activity of peroxisome-proliferator-responsive elements (PPRE), reduced the presence of reactive oxygen species (ROS) and increased mitochondrial density in HepG2 cells by activation of AMPK/PPAR. Our findings strongly support the medicinal use of the plant and suggest that CpPII is a potential therapy for prevention and/or treatment of type-2 diabetes. A common epitope sequence shared among the proteases and osmotin is possibly the responsible for the beneficial effects of CpPII.

Exposure to air pollution from traffic-generated sources is known to contribute to the etiology of inflammatory diseases, including cardiovascular disease (CVD) and obesity; however, the signaling pathways involved are still under investigation. Dysregulation of the renin-angiotensin system (RAS) can contribute to CVD and alter lipid storage and inflammation in adipose tissue. Our previous exposure studies revealed that traffic-generated emissions increase RAS signaling, further exacerbated by a high-fat diet. Thus, we investigated the hypothesis that exposure to engine emissions increases systemic and local adipocyte RAS signaling, promoting the expression of factors involved in CVD and obesity. Male C57BL/6 mice (6-8 wk old) were fed either a high-fat (HF, n = 16) or low-fat (LF, n = 16) diet, beginning 30d prior to exposures, and then exposed via inhalation to either filtered air (FA, controls) or a mixture of diesel engine + gasoline engine vehicle emissions (MVE: 100 μg PM/m3) via whole-body inhalation for 6 h/d, 7 d/wk, 30d. Endpoints were assessed via immunofluorescence and RT-qPCR. MVE-exposure promoted vascular adhesion factors (VCAM-1, ICAM-1) expression, monocyte/macrophage sequestration, and oxidative stress in the vasculature, associated with increased angiotensin II receptor type 1 (AT1) expression. In the kidney, MVE-exposure promoted the expression of renin, AT1, and AT2 receptors. In adipose tissue, both HF-diet and MVE-exposure mediated increased epididymal fat pad weight and adipocyte hypertrophy, associated with increased angiotensinogen and AT1 receptor expression; however, these outcomes were further exacerbated in the MVE + HF group. MVE-exposure also induced inflammation, monocyte chemoattractant protein (MCP)-1, and leptin, while reducing insulin receptor and glucose transporter, GLUT4, expression in adipose tissue. Our results indicate that MVE-exposure promotes systemic and local adipose RAS signaling, associated with increased expression of factors contributing to CVD and obesity, further exacerbated by HF diet consumption.

Infective endocarditis (IE) is an illness where the heart is invaded by bacteria, like Streptococcal and Staphylococcal species that contain lipoteichoic acid (LTA) related to an essential role in this disease. This study is the first in evaluating antioxidant enzyme levels in embryonic cardiomyocyte cell line (H9c2) induced by LTA from Streptococcus sanguinis. LTA increased reactive oxygen species (ROS) and reduced the levels of the antioxidant enzymes glutathione peroxidase, superoxide dismutase (SOD)-1 and catalase (CAT) but did not affect glutathione content. At the highest LTA concentration (15 μg/ml), SOD-1 and CAT levels did not change, and this effect was related to the induction of mRNA levels of Nrf2 induced by LTA. These results suggest that low antioxidant enzyme levels and ROS production could be related to IE.

Oxidative stress and cardiomyocyte apoptosis are involved in the pathogenesis of doxorubicin (DOX)-induced cardiotoxicity. Matrine is well-known for its powerful anti-oxidant and anti-apoptotic capacities. Our present study aimed to investigate the effect of matrine on DOX-induced cardiotoxicity and try to unearth the underlying mechanisms. Mice were exposed with DOX to generate DOX-induced cardiotoxicity or normal saline as control. H9C2 cells were used to verify the effect of matrine in vitro. DOX injection triggered increased generation of reactive oxygen species (ROS) and excessive cardiomyocyte apoptosis, which were significantly mitigated by matrine. Mechanistically, we found that matrine ameliorated DOX-induced uncoupling protein 2 (UCP2) downregulation, and UCP2 inhibition by genipin could blunt the protective effect of matrine on DOX-induced oxidative stress and cardiomyocyte apoptosis. Besides, 5\'-AMP-activated protein kinase α2 (Ampkα2) deficiency inhibited matrine-mediated UCP2 preservation and abolished the beneficial effect of matrine in mice. Besides, we observed that matrine incubation alleviated DOX-induced H9C2 cells apoptosis and oxidative stress level via activating AMPKα/UCP2, which were blunted by either AMPKα or UCP2 inhibition with genetic or pharmacological methods. Matrine attenuated oxidative stress and cardiomyocyte apoptosis in DOX-induced cardiotoxicity via maintaining AMPKα/UCP2 pathway, and it might be a promising therapeutic agent for the treatment of DOX-induced cardiotoxicity.

In this work, we measured the effect of cytochrome c on the NADH-dependent superoxide anion production by synaptic plasma membrane vesicles from rat brain. In these membranes, the cytochrome c stimulated NADH-dependent superoxide anion production was inhibited by antibodies against cytochrome b5 reductase linking the production to this enzyme. Measurement of the superoxide anion radical generated by purified recombinant soluble and membrane cytochrome b5 reductase corroborates the production of the radical by different enzyme isoforms. In the presence of cytochrome c, a burst of superoxide anion as well as the reduction of cytochrome c by cytochrome b5 reductase was measured. Complex formation between both proteins suggests that cytochrome b5 reductase is one of the major partners of cytochrome c upon its release from mitochondria to the cytosol during apoptosis. Superoxide anion production and cytochrome c reduction are the consequences of the stimulated NADH consumption by cytochrome b5 reductase upon complex formation with cytochrome c and suggest a major role of this enzyme as an anti-apoptotic protein during cell death.

This study explores and characterizes the toxicity of two closely related carcinogenic dinitro-pyrenes (DNPs), 1,3-DNP and 1,8-DNP, in human bronchial epithelial BEAS-2B cells and mouse hepatoma Hepa1c1c7 cells. Neither 1,3-DNP nor 1,8-DNP (3-30 μM) induced cell death in BEAS-2B cells. In Hepa1c1c7 cells only 1,3-DNP (10-30 μM) induced a mixture of apoptotic and necrotic cell death after 24 h. Both compounds increased the level of reactive oxygen species (ROS) in BEAS-2B as measured by CM-H2DCFDA-fluorescence. A corresponding increase in oxidative damage to DNA was revealed by the formamidopyrimidine-DNA glycosylase (fpg)-modified comet assay. Without fpg, DNP-induced DNA damage detected by the comet assay was only found in Hepa1c1c7 cells. Only 1,8-DNP formed DNA adduct measured by 32P-postlabelling. In Hepa1c1c cells, 1,8-DNP induced phosphorylation of H2AX (γH2AX) and p53 at a lower concentration than 1,3-DNP and there was no direct correlation between DNA damage/DNA damage response (DR) and induced cytotoxicity. On the other hand, 1,3-DNP-induced apoptosis was inhibited by pifithrin-α, an inhibitor of p53 transcriptional activity. Furthermore, 1,3-DNP triggered an unfolded protein response (UPR), as measured by an increased expression of CHOP, ATF4 and XBP1. Thus, other types of damage possibly linked to endoplasmic reticulum (ER)-stress and/or UPR could be involved in the induced apoptosis. Our results suggest that the stronger carcinogenic potency of 1,8-DNP compared to 1,3-DNP is linked to its higher genotoxic effects. This in combination with its lower potency to induce cell death may increase the probability of causing mutations.

Abdominal aortic aneurysm (AAA) is an inflammatory vascular disorder with high mortality. Accumulating evidence shows that toll-like receptor 2 (TLR2) plays a critical role in the regulation of wound-repairing process after tissue injury. We wondered if TLR2 signaling contributed to the pathogenesis of AAA and that targeting TLR2 would attenuate AAA development and progression. In this study, enhanced expression of TLR2 and its ligands were observed in human AAA tissue. Neutralization of TLR2 protected against AAA development and caused established AAA to regress in mouse models of AAA. In addition, TLR2-deficient mice also failed to develop AAA. The prophylactic and therapeutic effects of blocking TLR2 were accompanied by a significant resolution of inflammation and vascular remodeling, as indicated by the decreased expression or activity of MMP-2/9, α-SMA, inflammatory cytokines, and transcription factors NF-κB, AP-1 and STAT1/3 in AAA tissue. Mechanistically, blocking TLR2 decreased the expression and interaction of TLR2 and several endogenous ligands, which diminished chronic inflammation and vascular remodeling in the vascular tissue of AAA. Our studies indicate that the interactions between TLR2 and its endogenous ligands contribute to the pathogenesis of AAA and that targeting TLR2 offers great potential toward the development of therapeutic agents against AAA.