manganese(III) Tetrakis(1-methyl-4-pyridyl)porphyrin Research Articles

A consequence of immature breathing induces persistent changes in hippocampal synaptic plasticity and behavior: a role of prooxidant state and NMDA receptor imbalance.

Underdeveloped breathing results from premature birth and causes intermittent hypoxia during the early neonatal period. Neonatal intermittent hypoxia (nIH) is a condition linked to the increased risk of neurocognitive deficit later in life. However, the mechanistic basis of nIH-induced changes to neurophysiology remains poorly resolved. We investigated the impact of nIH on hippocampal synaptic plasticity and NMDA receptor (NMDAr) expression in neonatal mice. Our findings indicate that nIH induces a prooxidant state that leads to an imbalance in NMDAr subunit composition favoring GluN2B over GluN2A expression and impairs synaptic plasticity. These consequences persist in adulthood and coincide with deficits in spatial memory. Treatment with an antioxidant, manganese (III) tetrakis (1-methyl-4-pyridyl)porphyrin (MnTMPyP), during nIH effectively mitigated both immediate and long-term effects of nIH. However, MnTMPyP treatment post-nIH did not prevent long-lasting changes in either synaptic plasticity or behavior. In addition to demonstrating that the prooxidant state has a central role in nIH-mediated neurophysiological and behavioral deficits, our results also indicate that targeting the prooxidant state during a discrete therapeutic window may provide a potential avenue for mitigating long-term neurophysiological and behavioral outcomes that result from unstable breathing during early postnatal life.

A HIF1a-Dependent Pro-Oxidant State Disrupts Synaptic Plasticity and Impairs Spatial Memory in Response to Intermittent Hypoxia.

Sleep apnea causes cognitive deficits and is associated with several neurologic diseases. Intermittent hypoxia (IH) is recognized as a principal mediator of pathophysiology associated with sleep apnea, yet the basis by which IH contributes to impaired cognition remains poorly defined. Using a mouse model exposed to IH, this study examines how the transcription factor, hypoxia inducible factor 1a (HIF1a), contributes to disrupted synaptic physiology and spatial memory. In wild-type mice, impaired performance in the Barnes maze caused by IH coincided with a loss of NMDA receptor (NMDAr)-dependent long-term potentiation (LTP) in area CA1 and increased nuclear HIF1a within the hippocampus. IH-dependent HIF1a signaling caused a two-fold increase in expression of the reactive oxygen species (ROS) generating enzyme NADPH oxidase 4 (NOX4). These changes promoted a pro-oxidant state and the downregulation of GluN1 within the hippocampus. The IH-dependent effects were not present in either mice heterozygous for Hif1a (HIF1a+/−) or wild-type mice treated with the antioxidant manganese (III) tetrakis(1-methyl-4-pyridyl) porphyrin (MnTMPyP). Our findings indicate that HIF1a-dependent changes in redox state are central to the mechanism by which IH disrupts hippocampal synaptic plasticity and impairs spatial memory. This mechanism may enhance the vulnerability for cognitive deficit and lower the threshold for neurologic diseases associated untreated sleep apnea.

Pro-oxidant effect of ALA is implicated in mitochondrial dysfunction of HepG2 cells.

Heme biosynthesis begins in the mitochondrion with the formation of delta-aminolevulinic acid (ALA). In acute intermittent porphyria, hereditary tyrosinemia type I and lead poisoning patients, ALA is accumulated in plasma and in organs, especially the liver. These diseases are also associated with neuromuscular dysfunction and increased incidence of hepatocellular carcinoma. Many studies suggest that this damage may originate from ALA-induced oxidative stress following its accumulation. Using the MnSOD as an oxidative stress marker, we showed here that ALA treatment of cultured cells induced ROS production, increasing with ALA concentration. The mitochondrial energetic function of ALA-treated HepG2 cells was further explored. Mitochondrial respiration and ATP content were reduced compared to control cells. For the 300 μM treatment, ALA induced a mitochondrial mass decrease and a mitochondrial network imbalance although neither necrosis nor apoptosis were observed. The up regulation of PGC-1, Tfam and ND5 genes was also found; these genes encode mitochondrial proteins involved in mitochondrial biogenesis activation and OXPHOS function. We propose that ALA may constitute an internal bioenergetic signal, which initiates a coordinated upregulation of respiratory genes, which ultimately drives mitochondrial metabolic adaptation within cells. The addition of an antioxidant, Manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), resulted in improvement of maximal respiratory chain capacity with 300 μM ALA. Our results suggest that mitochondria, an ALA-production site, are more sensitive to pro-oxidant effect of ALA, and may be directly involved in pathophysiology of patients with inherited or acquired porphyria.

The effects of superoxide dismutase mimetic MnTMPyP on the altered blood–brain barrier integrity in experimental preeclampsia with or without seizures in rats

We investigated the effects of a cell-permeable superoxide dismutase mimetic, manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP) on blood–brain barrier (BBB) integrity following pentylenetetrazole (PTZ)-induced seizures in experimental preeclampsia symptoms induced by N(omega)-nitro-l-arginine methyl ester (l-NAME) in pregnant rats. To show the functional and morphological alterations in BBB integrity, quantitative analysis of sodium fluorescein (NaFlu) extravasation, immunohistochemistry and electron microscopic assessment of horseradish peroxidase (HRP) permeability were performed. Varying degrees of proteinuria were seen and arterial blood pressure increased in l-NAME-treated pregnant rats (p<0.01). MnTMPyP pretreatment and convulsive PTZ challenge significantly decreased the immunoreactivity of occludin in hippocampal capillaries in l-NAME-treated pregnant rats (p<0.01). BBB permeability to NaFlu significantly increased in pregnant rats treated with l-NAME plus PTZ (p<0.01), but MnTMPyP pretreatment did not significantly decrease NaFlu penetration into the brain parenchyma in these animals. Ultrastructurally, frequent vesicles containing HRP reaction products were observed in the capillary endothelial cells in the cerebral cortex and hippocampus of pregnant rats treated with l-NAME and l-NAME plus PTZ with the abundance being more in the latter group. MnTMPyP pretreatment caused a marked reduction in the frequency of HRP reaction product containing vesicles in both experimental settings. In conclusion, the results of the present study provide evidence that MnTMPyP plays an important role in limiting the enhanced vesicle-mediated transcellular transport in BBB endothelium in a rat model of preeclampsia and the differences in the way of transports of NaFlu and HRP might be responsible for the different effects of MnTMPyP on the BBB permeability to these two tracers.

Minocycline, levodopa and MnTMPyP induced changes in the mitochondrial proteome profile of MPTP and maneb and paraquat mice models of Parkinson's disease

Mitochondrial dysfunction is the foremost perpetrator of the nigrostriatal dopaminergic neurodegeneration leading to Parkinson's disease (PD). However, the roles played by majority of the mitochondrial proteins in PD pathogenesis have not yet been deciphered. The present study investigated the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and combined maneb and paraquat on the mitochondrial proteome of the nigrostriatal tissues in the presence or absence of minocycline, levodopa and manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin (MnTMPyP). The differentially expressed proteins were identified and proteome profiles were correlated with the pathological and biochemical anomalies induced by MPTP and maneb and paraquat. MPTP altered the expression of twelve while combined maneb and paraquat altered the expression of fourteen proteins. Minocycline, levodopa and MnTMPyP, respectively, restored the expression of three, seven and eight proteins in MPTP and seven, eight and eight proteins in maneb- and paraquat-treated groups. Although levodopa and MnTMPyP rescued from MPTP- and maneb- and paraquat-mediated increase in the microglial activation and decrease in manganese-superoxide dismutase expression and complex I activity, dopamine content and number of dopaminergic neurons, minocycline defended mainly against maneb- and paraquat-mediated alterations. The results demonstrate that MPTP and combined maneb and paraquat induce mitochondrial dysfunction and microglial activation and alter the expression of a bunch of mitochondrial proteins leading to the nigrostriatal dopaminergic neurodegeneration and minocycline, levodopa or MnTMPyP variably offset scores of such changes.

Electrochemical Sensor for Simultaneous Measurement of Nitrite and Superoxide Anion Radical Using Superoxide Dismutase-Mimetic Manganese(III) Tetrakis(1-methyl-4-pyridyl)Porphyrin on Polypyrrole Matrix

An electrochemical highly sensitive nitrite (NO2 and superoxide anion radical (O •− 2 sensor was fabricated and developed based on an electrochemical deposition of manganese(III) tetrakis(1methyl-4-pyridyl)porphyrin (MnTMPyP) in polypyrrole (PPy) matrix on Pt electrode. The surface morphological image of MnTMPyP-PPy and PPy matrix on Pt electrode was obtained by scanning electron microscopy exhibiting microporous structure. The electrochemical behavior of the MnTMPyP-PPy-Pt electrode as sensor investigated by cyclic voltammetry revealed that the characteristic of MnTMPyP reversible redox peaks obtained at −0.12 V and −0.38 V versus Ag/AgCl respectively. The sensor electrode showed an excellent electrocatalytic nitrite oxidase and superoxide dismutase (SOD) activities. This electrochemical sensor exhibited a linear current response over the concentration range from 0.8 to 1000 M, with a detection limit of 0 8±0 03 M for NO2 and a corresponding linear range from 0.6 to 1000 M, with a detection limit of 0 6± 0 02 M for O•− 2 . In addition, the sensor displayed high sensitivity, good reproducibility and retained stability over a period of ∼4 weeks. This SOD-mimetic electrode was proved to be effective not only in detecting NO2 and O •− 2 independently but also in determining the concentrations of NO − 2 and O •− 2 simultaneously in in vitro systems.

Attenuated vasodilatation in lambs with endogenous and exogenous activation of cGMP signaling: Role of protein kinase G nitration

Pulmonary vasodilation is mediated through the activation of protein kinase G (PKG) via a signaling pathway involving nitric oxide (NO), natriuretic peptides (NP), and cyclic guanosine monophosphate (cGMP). In pulmonary hypertension secondary to congenital heart disease, this pathway is endogenously activated by an early vascular upregulation of NO and increased myocardial B-type NP expression and release. In the treatment of pulmonary hypertension, this pathway is exogenously activated using inhaled NO or other pharmacological agents. Despite this activation of cGMP, vascular dysfunction is present, suggesting that NO-cGMP independent mechanisms are involved and were the focus of this study. Exposure of pulmonary artery endothelial or smooth muscle cells to the NO donor, Spermine NONOate (SpNONOate), increased peroxynitrite (ONOO(-) ) generation and PKG-1α nitration, while PKG-1α activity was decreased. These changes were prevented by superoxide dismutase (SOD) or manganese(III)tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP) and mimicked by the ONOO(-) donor, 3-morpholinosydnonimine N-ethylcarbamide (SIN-1). Peripheral lung extracts from 4-week old lambs with increased pulmonary blood flow and pulmonary hypertension (Shunt lambs with endogenous activation of cGMP) or juvenile lambs treated with inhaled NO for 24 h (with exogenous activation of cGMP) revealed increased ONOO(-) levels, elevated PKG-1α nitration, and decreased kinase activity without changes in PKG-1α protein levels. However, in Shunt lambs treated with L-arginine or lambs administered polyethylene glycol conjugated-SOD (PEG-SOD) during inhaled NO exposure, ONOO(-) and PKG-1α nitration were diminished and kinase activity was preserved. Together our data reveal that vascular dysfunction can occur, despite elevated levels of cGMP, due to PKG-1α nitration and subsequent attenuation of activity.

Reactive oxygen species differently regulate renal tubular epithelial and interstitial cell proliferation after ischemia and reperfusion injury

Reactive oxygen species (ROS) function as an inducer of cell death and survival or proliferative factor, in a cell-type-specific and concentration-dependent manner. All of these roles are critical to ischemia-induced renal functional impairment and progressive fibrotic changes in the kidney. In an effort to define the role of ROS in the proliferation of tubular epithelial cells and of interstitial cells in kidneys recovering after ischemia and reperfusion (I/R) injury, experimental mice were subjected to 30 min of bilateral kidney ischemia and administered with manganese(III) tetrakis(1-methyl-4-pyridyl) porphyrin (MnTMPyP), a superoxide dismutase mimetic, from 2 to 15 days after I/R for 14 days daily (earlier and longer) and from 8 to 15 days after I/R for 8 days daily (later and shorter). Cell proliferation was assessed via 5'-bromo-2'-deoxyuridine (BrdU) incorporation assays for 20 h before the harvest of kidneys. After I/R, the numbers of BrdU-incorporating cells increased both in the tubules and interstitium. MnTMPyP administration was shown to accelerate the proliferation of tubular epithelial cells, presenting tubule-specific marker proteins along tubular segments, whereas it attenuated the proliferation of interstitial cells, evidencing α-smooth muscle actin, fibroblast-specific protein-1, F4/80, and NADPH oxidase-2 proteins; these results indicated that ROS attenuates tubular cell regeneration, but accelerates interstitial cell proliferation. Earlier and longer MnTMPyP treatment more effectively inhibited tissue superoxide formation, the increment of interstitial cells, and the decrement of epithelial cells compared with later and shorter treatment. After I/R, apoptotic cells appeared principally in the tubular epithelial cells, but not in the interstitial cells, thereby indicating that ROS is harmful in tubule cells, but is not in interstitial cells. In conclusion, ROS generated after I/R injury in cell proliferation and death performs a cell-type-specific and concentration-dependent role, even within the same tissues, and timely intervention of ROS is crucial for effective therapies.

Biochemical Activity of Reactive Oxygen Species Scavengers Do Not Predict Retinal Ganglion Cell Survival

Retinal ganglion cells (RGCs) die as a result of axonal injury in a variety of optic neuropathies, including glaucoma. Reactive oxygen species (ROS) act as intracellular signaling molecules and initiate apoptosis in nerve growth factor-deprived sympathetic neurons and axotomized RGCs. Determination of the role of specific ROS relies on the use of small molecule or protein scavengers with various degrees of specificity. The pro- or anti-cell-death effect of several ROS generating and scavenging systems in cultured RGCs was correlated with their activity in cell-free assays. Neonatal rat retinas were dissociated and incubated with ROS-generating systems for hydroxyl radical, superoxide anion (O2-), and H2O2. Scavengers tested were catalase, polyethylene glycol-superoxide dismutase (PEG-SOD), manganese (III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), deferoxamine, and U-74389G. Viability of retrogradely labeled RGCs was determined with calcein-AM 24 hours after plating. O2- and H2O2 scavenging in cell-free assays was measured with dihydroethidium and Amplex Red (Invitrogen, Carlsbad, CA), respectively. Systematic differences were found between ROS scavenging in cell-free assays and the ability of scavengers to protect RGCs in cell culture. Furthermore, many ROS scavengers lost specificity and protected against various ROS, whereas others failed to protect against their unique ROS target. These activities stray from commonly recognized specificities of individual ROS scavengers or generating systems and are important in understanding ROS biology. In addition, antioxidant defense mechanisms used by RGCs and other retinal cells interfere with responses expected from ROS scavengers in well-defined systems. Last, H2O2 induced intramitochondrial O2-, whereas paraquat produced O2- outside of the mitochondria, and these areas of generation can mislead interpretations of ROS scavenger activity and effectiveness. There is discordance between ROS effects in cultured RGCs and cell-free assays, with several mechanisms accounting for this divergence. To identify the roles of ROS signaling in cell death accurately, several approaches should be used. These include using a panel of ROS scavengers and generators, testing the panel in primary neuronal cultures, and quantifying ROS with cell-free assays.

Orchiectomy Attenuates Post-ischemic Oxidative Stress and Ischemia/Reperfusion Injury in Mice: A ROLE FOR MANGANESE SUPEROXIDE DISMUTASE

Males are much more susceptible to ischemia/reperfusion (I/R)-induced kidney injury when compared with females. Recently we reported that the presence of testosterone, rather than the absence of estrogen, plays a critical role in gender differences in kidney susceptibility to I/R injury in mice. Although reactive oxygen species and antioxidant defenses have been implicated in I/R injury, their roles remain to be defined. Here we report that the orchiectomized animal had significantly less lipid peroxidation and lower hydrogen peroxide levels in the kidney 4 and 24 h after 30 min of bilateral renal ischemia when compared with intact or dihydrotestosterone-treated orchiectomized males. The post-ischemic kidney expression and activity of manganese superoxide dismutase (MnSOD) in orchiectomized mice was much greater than in intact or dihydrotestosterone-administered orchiectomized mice. Four hours after 30 min of bilateral ischemia, superoxide formation was significantly lower in orchiectomized mice than in intact mice. In Madin-Darby canine kidney cells, a kidney epithelial cell line, 1 mm H(2)O(2) decreased MnSOD activity, an effect that was potentiated by pretreatment with dihydrotestosterone. Orchiectomy prevented the post-ischemic decrease of catalase activity. Treatment of male mice with manganese(III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), a SOD mimetic, reduced the post-ischemic increase of plasma creatinine, lipid peroxidation, and tissue hydrogen peroxide. These results suggest that orchiectomy accelerates the post-ischemic activation of MnSOD and reduces reactive oxygen species and lipid peroxidation, resulting in reduced kidney susceptibility to I/R injury.

Protective properties of tin- and manganese-centered porphyrins against hydrogen peroxide-mediated injury in rat astroglial cells.

Tin-mesoporphyrin (tin-mp), a potent inhibitor of heme oxygenase, and manganese (III) tetrakis(1-methyl-4-pyridyl)porphyrin (MnTMPyP), a potent superoxide dismutase mimetic, reduced H2O2 toxicity in cultures of transformed rat astroglial cells if added 30 min before, or at the same time as, H2O2. Reduced toxicity was not observed if treatment was delayed for 60 min, the time by which H2O2 was essentially eliminated from cultures. Coadministration of tin-mp and MnTMPyP did not increase protection over either compound administered individually. Tin-mp, but not MnTMPyP, was stable in culture. MnCl2 was not protective, suggesting that protection by MnTMPyP was not dependent on manganous ion, a by-product of MnTMPyP breakdown. Protection by tin-mp and MnTMPyP was not associated with metalloporphyrin-mediated induction of heme oxygenase-1 or with changes in heme oxygenase-2 on western blots. Whereas protective concentrations of tin-mp did not have superoxide dismutase-mimetic properties in vitro, protective concentrations of MnTMPyP partially inhibited heme oxygenase. The data support the hypothesis that heme oxygenase inhibition is protective against acute oxidative injury.

Mechanisms of action of transmitters and other substances on smooth muscle.

Mechanisms of action of transmitters and other substances on smooth muscle.