Neural protection by naturopathic compounds—an example of tetramethylpyrazine from retina to brain

Given the advantages of being stable in the ambient environment, being permeable to the blood–brain and/or blood–eye barriers and being convenient for administration, naturopathic compounds have growingly become promising therapeutic candidates for neural protection. Extracted from one of the most common Chinese herbal medicines, tetramethylpyrazine (TMP), also designated as ligustrazine, has been suggested to be neuroprotective in the central nervous system as well as the peripheral nerve network. Although the detailed molecular mechanisms of its efficacy for neural protection are understood limitedly, accumulating evidence suggests that antioxidative stress, antagonism for calcium, and suppression of pro-inflammatory factors contribute significantly to its neuroprotection. In animal studies, systemic administration of TMP (subcutaneous injection, 50 mg/kg) significantly blocked neuronal degeneration in hippocampus as well as the other vulnerable regions in brains of Sprague–Dawley rats following kainate-induced prolonged seizures. Results from us and others also demonstrated potent neuroprotective efficacy of TMP for retinal cells and robust benefits for brain in Alzheimer’s disease or other brain injury. These results suggest a promising prospect for TMP to be used as a treatment of specific neurodegenerative diseases. Given the assessment of the distribution, metabolism, excretion, and toxicity information that is already available on most neuroprotective naturopathic compounds such as TMP, it would not take much preclinical data to justify bringing such therapeutic compounds to clinical trials in humans

Loss of Caveolin-1 Accelerates Neurodegeneration and Aging

The aged brain exhibits a loss in gray matter and a decrease in spines and synaptic densities that may represent a sequela for neurodegenerative diseases such as Alzheimer's. Membrane/lipid rafts (MLR), discrete regions of the plasmalemma enriched in cholesterol, glycosphingolipids, and sphingomyelin, are essential for the development and stabilization of synapses. Caveolin-1 (Cav-1), a cholesterol binding protein organizes synaptic signaling components within MLR. It is unknown whether loss of synapses is dependent on an age-related loss of Cav-1 expression and whether this has implications for neurodegenerative diseases such as Alzheimer's disease.We analyzed brains from young (Yg, 3-6 months), middle age (Md, 12 months), aged (Ag, >18 months), and young Cav-1 KO mice and show that localization of PSD-95, NR2A, NR2B, TrkBR, AMPAR, and Cav-1 to MLR is decreased in aged hippocampi. Young Cav-1 KO mice showed signs of premature neuronal aging and degeneration. Hippocampi synaptosomes from Cav-1 KO mice showed reduced PSD-95, NR2A, NR2B, and Cav-1, an inability to be protected against cerebral ischemia-reperfusion injury compared to young WT mice, increased Aβ, P-Tau, and astrogliosis, decreased cerebrovascular volume compared to young WT mice. As with aged hippocampi, Cav-1 KO brains showed significantly reduced synapses. Neuron-targeted re-expression of Cav-1 in Cav-1 KO neurons in vitro decreased Aβ expression.Therefore, Cav-1 represents a novel control point for healthy neuronal aging and loss of Cav-1 represents a non-mutational model for Alzheimer's disease

Human erythrocytes exposure to juglone leads to an increase of superoxide anion production associated with cytochrome b5 reductase uncoupling

Cytochrome b5 reductase is an enzyme with the ability to generate superoxide anion at the expenses of NADH consumption. Although this activity can be stimulated by cytochrome c and could participate in the bioenergetic failure accounting in apoptosis, very little is known about other molecules that may uncouple the function of the cytochrome b5 reductase. Naphthoquinones are redox active molecules with the ability to interact with electron transfer chains. In this work, we made an inhibitor screening against recombinant human cytochrome b5 reductase based on naphthoquinone properties. We found that 5-hydroxy-1,4-naphthoquinone (known as juglone), a natural naphthoquinone extracted from walnut trees and used historically in traditional medicine with ambiguous health and toxic outcomes, had the ability to uncouple the electron transfer from the reductase to cytochrome b5 and ferricyanide. Upon complex formation with cytochrome b5 reductase, juglone is able to act as an electron acceptor leading to a NADH consumption stimulation and an increase of superoxide anion production by the reductase. Our results suggest that cytochrome b5 reductase could contribute to the measured energetic failure in the erythrocyte apoptosis induced by juglone, that is concomitant with the reactive oxygen species produced by cytochrome b5 reductase.This work was supported by the Associate Laboratory for Green Chemistry-LAQV which is financed by national funds from FCT/MCTES (UID/QUI/50006/2019). Experimental work was also supported by funding from Ayuda a Grupos de la Junta de Extremadura (Group GR18118) co-financed by the European Funds for Structural Development (FEDER). The authors also thank FCT for funds to GHTM (UID/Multi/04413/2013) and AKSA acknowledges FCT/MCTES for their “Investigador Doutorado” contracts' funding and signed with FCT/UNL in accordance with DL 57/2016 e Lei 57/2017.Peer reviewe

Screen-printed electrodes testing for detection of potential stress biomarkers in sweat

Detection of stress biomarkers molecules, non-invasively, through (non-induced) sweat sampling is an important research field since sweat is a potential diagnostic fluid for early and continuous human health monitoring, not only for stress-related conditions directly but also for other pathologies (e.g., associated with chronic diseases). The simultaneous detection of multiple potential biomarkers in sweat samples, using simple and low-cost electrochemical methods (detecting patterns or “electrochemical fingerprints”) requiring no sample preparation and its correlation with physiological conditions, is an attractive methodology but not readily achievable. Several of these potential biomarkers’ electrochemical response presents very close oxidation potentials, among other factors that hamper the detection, such as pH dependence of the electrochemical response or partial adsorption on electrode surfaces. Disposable screen-printed electrode materials, with relatively low-cost, could be useful to overcome the difficulties. A set of selected potential stress-related (non-protein) biomarkers (tyrosine, phenylalanine, dopamine, serotonin, and hydrocortisone) was used in the current study for qualitative electrochemical detection on different screen-printed carbon-based electrodes. The detection was attained in simulated sweat solutions and real sweat samples. The goal was to evaluate the electrochemical response on the different surfaces and determine the most suitable carbon-based screen-printed electrodes that may be used in future sensing devices.This work was supported by the Associate Laboratory for Green Chemistry–LAQV which is fnanced by national funds from FCT/MCTES (UIDB/50006/2020 and UIDP/50006/2020). Fundação para a Ciência e Tecnologia provided fnancial support through Project PTDC/SAU-SOC/28390/2017 (STRESSSENSE)

Heterolytic reduction of fatty acid hydroperoxides by cytochrome c/cardiolipin complexes: antioxidant function in mitochondria

Cytochrome c (cyt c), a mitochondrial intermembrane electron shuttle between complexes III and IV, can, upon binding with an anionic phospholipid, cardiolipin (CL), act as a peroxidase that catalyzes cardiolipin oxidation. H(2)O(2) was considered as a source of oxidative equivalents for this reaction, which is essential for programmed cell death. Here we report that peroxidase cyt c/CL complexes can utilize free fatty acid hydroperoxides (FFA-OOH) at exceptionally high rates that are approximately 3 orders of magnitude higher than for H(2)O(2). Similarly, peroxidase activity of murine liver mitochondria was high with FFA-OOH. Using EPR spin trapping and LC-MS techniques, we have demonstrated that cyt c/CL complexes split FFA-OOH predominantly via a heterolytic mechanism, yielding hydroxy-fatty acids, whereas H(2)O(2) (and tert-butyl hydroperoxide, t-BuOOH) undergo homolytic cleavage. Computer simulations have revealed that Arg(38) and His(33) are important for the heterolytic mechanism at potential FFA-OOH binding sites of cyt c (but not for H(2)O(2) or t-BuOOH). Regulation of FFA-OOH metabolism may be an important function of cyt c that is associated with elimination of toxic FFA-OOH and synthesis of physiologically active hydroxy-fatty acids in mitochondria

Mitochondrial targeting of electron scavenging antioxidants: Regulation of selective oxidation vs random chain reactions

Effective regulation of highly compartmentalized production of reactive oxygen species and peroxidation reactions in mitochondria requires targeting of small molecule antioxidants and antioxidant enzymes into the organelles. This review describes recently developed approaches to mitochondrial targeting of small biologically active molecules based on: (i) preferential accumulation in mitochondria because of their hydrophobicity and positive charge (hydrophobic cations), (ii) binding with high affinity to an intra-mitochondrial constituent, and (iii) metabolic conversions by specific mitochondrial enzymes to reveal an active entity. In addition, targeted delivery of antioxidant enzymes via expression of leader sequences directing the proteins into mitochondria is considered. Examples of successful antioxidant and anti-apoptotic protection based on the ability of targeted cargoes to inhibit cytochrome c-catalyzed peroxidation of a mitochondria-specific phospholipid cardiolipin, in vitro and in vivo are presented. Particular emphasis is placed on the employment of triphenylphosphonium- and hemi-gramicidin S-moieties as two effective vehicles for mitochondrial delivery of antioxidants