Ameliorating Mitochondrial Dysfunction of Neurons by Biomimetic Targeting Nanoparticles Mediated Mitochondrial Biogenesis to Boost the Therapy of Parkinson's Disease.

Mitochondrial dysfunction of neurons is the core pathogenesis of incurable Parkinson's disease (PD). It is crucial to ameliorate the mitochondrial dysfunction of neurons for boosting the therapy of PD. Herein, the remarkable promotion of mitochondrial biogenesis to ameliorate mitochondrial dysfunction of neurons and improve the treatment of PD by using mitochondria-targeted biomimetic nanoparticles, which are Cu2- x Se-based nanoparticles functionalized with curcumin and wrapped with DSPE-PEG2000 -TPP-modified macrophage membrane (denoted as CSCCT NPs), is reported. These nanoparticles can efficiently target mitochondria of damaged neurons in an inflammatory environment, and mediate the signaling pathway of NAD+ /SIRT1/PGC-1α/PPARγ/NRF1/TFAM to alleviate 1-methyl-4-phenylpyridinium (MPP+ )-induced neuronal toxicity. They can reduce the mitochondrial reactive oxygen species, restore mitochondrial membrane potential (MMP), protect the integrity of mitochondrial respiratory chain, and ameliorate mitochondrial dysfunction via promoting mitochondrial biogenesis, which synergistically improve the motor disorders and anxiety behavior of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice. This study demonstrates that targeting mitochondrial biogenesis to ameliorate mitochondrial dysfunction has a great potential in the treatment of PD and mitochondria-related diseases.

Induction of ferroxidase enzymatic activity by copper reduces MPP+-evoked neurotoxicity in rats.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by decreased dopamine, intracellular inclusions (Lewy bodies) and brain iron deposits. PD has also been related with reduced ferroxidase activity, diminished antioxidant defenses and lipid peroxidation. Striatal injection of 1-methyl-4-phenylpyridinium (MPP(+)) into rodents reproduces the major biochemical characteristics of PD, including oxidative stress. Copper (Cu) plays an important role as prosthetic group of several proteins involved in iron metabolism and antioxidant responses, such as ceruloplasmin (Cp). In the present study, intraperitoneal CuSO4 injection (10µmol/kg) produced an insignificant increase of Cu content in striatum and midbrain (17.5% and 7%, respectively). After 10 and 11h, Cu induced 6- and 4-fold increase Cp mRNA in midbrain and striatum, respectively. Cu-supplement also produced a time-dependent increase ferroxidase activity in striatal tissue, reaching a maximum 16h after Cu treatment in midbrain; while, ferrous iron content diminished 18% in striatum and 8% in midbrain. In regard the PD model, we found that MPP(+) (10µg/8µL, intrastriatal), induced a significant (P<0.05) reduction of striatal ferroxidase activity; this effect was reverted by Cu pre-treatment 16h before MPP(+). Likewise, Cu-supplement prevented lipid fluorescent products formation in striatum, evaluated (P<0.01) 6h after MPP(+). In the long term, apomorphine-evoked circling behavior was evaluated 6 days after MPP(+) injury; Cu pre-treatment significantly reduced (P<0.05) the apomorphine-induced ipsilateral turns in MPP(+)-lesioned rats. These results suggest that Cu-induced expression of Cp could be an interesting scope against the deleterious effects of iron deposits in PD.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced neuroblastic apoptosis in the subventricular zone is caused by 1-methy-4-phenylpiridinium (MPP(+)) converted from MPTP through MAO-B.

Intraperitoneal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration induces apoptosis of subventricular zone (SVZ) doublecortin (Dcx)-positive neural progenitor cells (migrating neuroblasts, A cells). Actually, a metabolite of MPTP, 1-methy-4-phenylpiridinium (MPP(+)), is responsible for neural progenitor cell toxicity. In the present study, to examine whether the MPTP-induced SVZ cell apoptosis is caused directly by MPP(+) metabolized through monoamine oxidase B (MAO-B), MPTP or MPP(+) was intracerebroventricularly (icv) injected into C57BL/6 mice. At Day 1 postinjection, many terminal deoxynucleotidyl transferase-mediated dUTP endlabeling (TUNEL)-positive cells were observed in the SVZ of both low (36 µg) and high (162 µg) dose MPTP- and MPP(+)-injected mice. The number of Dcx-positive A cells showed a significant decrease following high dose of MPTP- or MPP(+)-injection on Days 1 and 3, respectively, whereas that of EGFR-positive C cells showed no change in mice with any treatment. In addition, prior icv injection of a MAO-B inhibitor, R(-)-deprenyl (deprenyl), inhibited MPTP-induced apoptosis, but not MPP(+)-induced apoptosis. MAO-B- and GFAP-double positive cells were detected in the ependyma and SVZ in all mice. It is revealed from these results that icv injection of MPTP induces apoptosis of neural progenitor cells (A cells) in the SVZ via MPP(+) toxicity. In addition, it is suggested that the conversion from MPTP to MPP(+) is caused mainly by MAO-B located in ependymal cells and GFAP-positive cells in the SVZ.

The antioxidative effect of electro-acupuncture in a mouse model of Parkinson's disease.

Accumulating evidence indicates that oxidative stress plays a critical role in Parkinson's disease (PD). Our previous work has shown that 100 Hz electro-acupuncture (EA) stimulation at ZUSANLI (ST36) and SANYINJIAO (SP6) protects neurons in the substantia nigra pars compacta from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) toxicity in male C57BL/6 mice, a model of PD. In the present study we administered 100 Hz EA stimulation at the two acupoints to MPTP-lesioned mice for 12 sessions starting from the day prior to the first MPTP injection. We found that in the striatum of MPTP treated mice 100 Hz EA stimulation effectively inhibited the production of hydrogen peroxide and malonaldehyde, and increased glutathione concentration and total superoxide dismutase activity through biochemical methods. However, it decreased glutathione peroxidase activity via biochemical analysis and did not affect the level of 1-methyl-4-phenylpyridinium in the striatum revealed by high performance liquid chromatography with ultraviolet detection. These data suggest that 100 Hz EA stimulation at ST36 and SP6 has antioxidative effects in the MPTP model of PD. This data, along with our previous work, indicates that 100 Hz EA stimulation at ST36 and SP6 protects the nigrostriatal system by multiple mechanisms including antioxidation and antiapoptosis, and suggests that EA stimulation is a promising therapy for treating PD.

Effect of polyphenols on the intestinal and placental transport of some bioactive compounds.

Polyphenols are a group of widely distributed phytochemicals present in most foods of vegetable origin. A growing number of biological effects have been attributed to these molecules in the past few years and only recently has their interference with the transport capacity of epithelial barriers received attention. This review will present data obtained concerning the effect of polyphenols upon the transport of some compounds (organic cations, glucose and the vitamins thiamin and folic acid) at the intestinal and placental barriers. Important conclusions can be drawn: (i) different classes of polyphenols affect transport of these bioactive compounds at the intestinal epithelia and the placenta; (ii) different compounds belonging to the same phenolic family often possess opposite effects upon transport of a given molecule; (iii) the acute and chronic/short-term and long-term exposures to polyphenols do not produce parallel results and, therefore, care should be taken when extrapolating results; (iv) the effect of polyphenolics in combination may be very different from the expected ones taking into account the effect of each of these compounds alone, and so care should be taken when speculating on the effect of a drink based on the effect of one component only; (v) care should be taken in drawing conclusions for alcoholic beverages from results obtained with ethanol alone. Although most of the data reviewed in the present paper refer to in vitro experiments with cell-culture systems, these studies raise a concern about possible changes in the bioavailability of substrates upon concomitant ingestion of polyphenols.

Mitochondrial UCP4 attenuates MPP+ - and dopamine-induced oxidative stress, mitochondrial depolarization, and ATP deficiency in neurons and is interlinked with UCP2 expression.

Mitochondrial uncoupling proteins (UCPs) uncouple oxidative phosphorylation from ATP synthesis. We explored the neuroprotective role of UCP4 with its stable overexpression in SH-SY5Y cells, after exposure to either MPP(+) or dopamine to induce ATP deficiency and oxidative stress. Cells overexpressing UCP4 proliferated faster in normal cultures and after exposure to MPP(+) and dopamine. Differentiated UCP4-overexpressing cells survived better when exposed to MPP(+) with decreased LDH release. Contrary to the mild uncoupling hypothesis, UCP4 overexpression resulted in increased absolute ATP levels (with ADP/ATP ratios similar to those of controls under normal conditions and ADP supplementation) associated with increased respiration rate. Under MPP(+) toxicity, UCP4 overexpression preserved ATP levels and mitochondrial membrane potential (MMP) and reduced oxidative stress; the preserved ATP level was not due to increased glycolysis. Under MPP(+) toxicity, the induction of UCP2 expression in vector controls was absent in UCP4-overexpressing cells, suggesting that UCP4 may compensate for UCP2 expression. UCP4 function does not seem to adhere to the mild uncoupling hypothesis in its neuroprotective mechanisms under oxidative stress and ATP deficiency. UCP4 overexpression increases cell survival by inducing oxidative phosphorylation, preserving ATP synthesis and MMP, and reducing oxidative stress.

Modulation of MPP+ uptake by tea and some of its components in Caco-2 cells.

The entry of most xeno/endobiotics into the organism is limited by their intestinal absorption. The interference of certain foods with the therapeutic efficacy of drugs or with chemical toxicity is becoming evident and growing attention is being given to these subjects. The aim of this work was to study the effect of green tea (GT) and black tea (BT), as well as some of their components, on the transport of organic cation molecules. For this purpose, 3H-MPP+ (radiolabeled 1-methyl-4-phenylpyridinium) was used as a model organic cation and Caco-2 cells were used as an intestinal epithelial model. Our results showed that both GT and BT significantly increased 3H-MPP+ absorption in these cells. Additionally, we studied the effect of epigallocatechin-3-gallate (EGCG), myricetin, caffeine, and theophylline. Whereas EGCG (2 mM) increased, myricetin (50 microM) and caffeine (1 mM) decreased, and theophylline (1 mM) had no effect on the uptake of 3H-MPP+ into Caco-2 cells. When GT was supplemented with caffeine or theophylline, we observed a partial loss of its effect. When BT was supplemented with EGCG, its ability to increase 3H-MPP+ uptake was much more pronounced than that observed with BT alone. In conclusion, this study showed that GT and BT might interfere with the absorption of the model organic cation MPP+ by the intestinal epithelium. Since important compounds are organic cations, the consequences of this interference may have an impact on human health. Although this constitutes only preliminary work and further studies are needed, tea should be included in the growing list of foodstuffs that have the potential to be involved in food-drug interactions.

Cloning and functional expression of a mouse liver organic cation transporter.

Hepatic uptake of organic cations is essential for the metabolism and secretion of numerous endobiotics and drugs. Several hepatic organic cation transporters have been kinetically defined, yet have not been isolated or cloned. We have isolated a complementary DNA (cDNA) from both murine liver and kidney cDNA libraries (mOct1/Slc22a1), and have functionally expressed it in Xenopus laevis oocytes. Although mOct1/Slc22a1 is homologous to previously cloned rat and human organic cation transporters, organic cation transport kinetics differed markedly. mOct1/Slc22a1-RNA injection of oocytes resulted in the saturable, time- and temperature-dependent uptake of the quaternary organic cation [14C]-tetraethylammonium ([14C]-TEA), with a Km of 38 micromol/L. TEA uptake was inhibited by several other organic cation drugs, but was not inhibited by the organic cation n-methyl-nicotinamide (NMN), being instead stimulated by it (fourfold). [14C]-TEA uptake was also stimulated by an inside-outside proton gradient. mOct1/Slc22a1-injected oocytes transported the organic cations [3H]-1-methyl-4-phenylpyridium and [3H]-choline chloride, but did not transport other classes of organic compounds. mOct1/Slc22a1 encodes for a hepatic and renal organic cation transporter which may be important for the uptake and secretion of cationic drugs and endobiotics.

Effect of MPTP on brain mitochondrial H2O2 and ATP production and on dopamine and DOPAC in the striatum.

An experimental rat model of Parkinson's disease was established by injecting rats directly in the striatum with the neurotoxic agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In order to study the action mechanism of this neurotoxic agent, MPTP and its main metabolite 1-methyl-4-phenylpyridinium (MPP+) were also added to suspensions of pyruvate/malate-supplemented nonsynaptic brain mitochondria, and the rates of hydrogen peroxide and ATP production were measured. Intrastriatal administration of MPTP produced a pronounced decrease in striatal dopamine levels (p < 0.005) and a strong increase in 3,4-hydroxiphenylacetic acid/dopamine ratio (an indicator of dopamine catabolism; p < 0.005) in relation to controls, as evaluated by in situ microdialysis. MPTP addition to rat brain mitochondria increased hydrogen peroxide production by 90%, from 1.37+/-0.35 to 2.59+/-0.48 nanomoles of H2O2/minute . mg of protein (p < 0.01). The metabolite MPP+ produced a marked decrease on the rate of ATP production of brain mitochondria (p < 0.005). These findings support the mitochondria-oxidative stress-energy failure hypothesis of MPTP-induced brain neurotoxicity.

The neurotoxin 1-methyl-4-phenylpyridinium is a substrate for the canalicular organic cation/H+ exchanger.

Hepatic organic cation transport consists, in part, of carrier-mediated sinusoidal uptake stimulated by an inside-negative membrane potential and canalicular excretion driven by electroneutral organic cation/H+ exchange. Intracellular organic cation transport involves sequestration into acidified organelles, also mediated by organic cation/H+ exchange. A sinusoidal organic cation transporter has been cloned; however, canalicular organic cation transport has not been characterized at the molecular level. On the assumption that hepatic organic cation/H+ exchange resembles monoamine transport in synaptic vesicles, we examined, using canalicular rat liver plasma membrane vesicles, the transport of 1-methyl-4-phenylpyridinium (MPP+), a neurotoxin taken up by a synaptic vesicular monoamine transporter that has been cloned. Under voltage-clamped conditions, an outwardly directed H+ gradient stimulated [3H]MPP+ uptake, compared with uptake under pH-equilibrated conditions, consistent with electroneutral MPP+/H+ exchange. Substrates for canalicular organic cation/H+ exchange cis-inhibited pH-dependent MPP+ uptake. Equilibrium exchange of [14C]tetraethylammonium was inhibited by MPP+ in a concentration-dependent manner, consistent with a direct interaction of MPP+ with the organic cation carrier. Carrier-mediated MPP+ uptake exhibited saturability, with kinetic parameters similarto those described for canalicular tetraethylammonium+/H+ exchange. Canalicular [3H]MPP+ uptake was ATP-independent and, thus, distinct from P-glycoprotein-mediated efflux. The finding that MPP+ is a substrate for canalicular organic cation/H+ exchange is applicable to studies, using degenerate oligonucleotides complementary to sequences conserved in neurotransmitter transporters, aimed at cloning this transporter.