MnCl2 Treatment Research Articles

Transcriptional analysis of the molecular response of Arabidopsis to manganese stress and recovery

Manganese (Mn) is an essential element for plants and plays a role in various metabolic processes. However, excess manganese can be toxic to plants. This study aimed to analyze the changes in various physiological activities and the transcriptome of Arabidopsis under different treatments: 1 mmol/L MnCl2 treatment for 1 day or 3 days, and 1 day of recovery on MS medium after 3 days of MnCl2 treatment. During the recovery phase, minor yellowing symptoms appeared on the leaves of Arabidopsis, and the content of chlorophyll and carotenoid decreased significantly, but the content of malondialdehyde and soluble sugar increased rapidly. Transcriptome sequencing data shows that the expression patterns of differentially expressed genes exhibit three major models: initial response model, later response model, recovery response model. Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis identified several affected metabolic pathways, including plant hormone signal transduction mitosolysis activates protein kinase (MAPK) phytohormone signaling, phenylpropanoid biosynthesis, ATP binding cassette transporters (ABC transporter), and glycosphingolipid biosynthesis. Differential expressed genes (DEGs) involved in phenylpropanoid biosynthesis, ABC transporter, and glycosphingolipid biosynthesis, were identified. Sixteen randomly selected DEGs were validated through qRT-PCR and showed consistent results with RNA-seq data. Our findings suggest that the phenylpropanoid metabolic pathway is activated to scavenge reactive oxygen species, the regulation of ABC transporter improves Mn transport, and the adjustment of cell membrane lipid composition occurs through glycerophospholipid metabolism to adapt to Mn stress in plants. This study provides new insights into the molecular response of plants to Mn stress and recovery, as well as theoretical cues for cultivating Mn-resistant plant varieties.

Trinuclear manganese and zinc clusters and a dinuclear manganese complex with multiple oxo-bridges: Syntheses, structures, and magnetic properties of the MnII compounds and luminescent property of the ZnII complex

Two trinuclear clusters [Mn3(L1)2] (1) and [Zn3(L1)2] (2) have been prepared from the reactions of Mn(CH3COO)2·4H2O and Zn(CH3COO)2·4H2O, respectively, with a hydroxy-rich Schiff-base ligand H3L1 (H3L1 = 2-{[bis(2-hydroxy-3-methoxybenzyl)(amino-ethyl)amino]methyl}phenol). One dinuclear compound [Mn2(HL2)Cl2] (3) was synthesized from the treatment of MnCl2·4H2O with another hydroxy-rich Schiff base ligand H3L2 (H3L2 = 2-{[bis(2-hydroxy-3-methoxybenzyl)(amino-ethyl)amino]ethyl}phenol). Complexes 1–3 were characterized by single-crystal X-ray diffraction analysis. The structural analyses indicated that complexes 1 and 2 are isostructural and contain linear MnII–MnII–MnII and ZnII–ZnII–ZnII cores. In compounds 1–3, the metal centers are doubly bridged by phenoxido groups of the ligands. Variable temperature magnetic measurements of 1 and 3 revealed the presence of antiferromagnetic intramolecular couplings, and the magnetic susceptibility data are consistent with S = 5/2 of the MnII ions. The luminescent property of compound 2 was investigated. The 3d10 zinc compound 2 exhibited strong luminescence with λmax = 491 nm (for λex = 418 nm).

Possible Involvement of Cellular Pathway and Cytokines in Manganese Induced Neurotoxicity in Neuroblastoma Cells via KH–Type Splicing Regulatory Protein

Pharmacognosy Research,2023,15,2,307-314.DOI:10.5530/pres.15.2.033Published:February 2023Type:Original Article Authors:Sharad Singh, Sunil More, G. S. Latha, Himanshu Agrawal, Veena S M, and Francois Niyonzima Author(s) affiliations:Sharad Singh1, Sunil More1,*, G. S. Latha2, Himanshu Agrawal3, Veena S M4, Francois Niyonzima5 1School of Basic and Applied Sciences, Dayananada Sagar University, Bangalore, Karnataka, INDIA. 2Department of Physiology, Rajarajeswari Medical College and Hospital, Bengaluru, Karnataka, A Constituent College under Dr. M.G.R. Educational and Research Institute, Chennai, Tamil Nadu, INDIA. 3Jubilant Biosys Limited, Bangalore, Karnataka, INDIA. 4Department of Biotechnology, Sapthagiri College of Engineering, Bengaluru, Karnataka, INDIA. 5Department of Mathematics, Sciences and Physical Education, University of Rwanda-College of Education (UR-CE), Kayonza Rukara Campus, RWANDA. Abstract:Background: Manganese is a toxic essential trace element and too high concentration instigates the neurodegenerative disease known as parkinsonism. Effects of manganese may lead to apoptosis. However, a detailed mechanism of manganese toxicity has not been fully elucidated. Previous published articles have highlighted the augmentation of KHSRP expression following Mn exposure. Objectives: In this work, the importance of KHSRP in Mn–induced toxicity was checked along with the impact of other known neurotoxicity inhibitors on KHSRP. Materials and Methods: KHSRP expression, pro and anti-inflammatory cytokines, chemokines, and pharmacological inhibitors (SAHA, Quercetin, and MCC950) were determined by exposing N2a cells to various MnCl2 concentrations. ANOVA and Dunnett’s test were used to decide on the significance. Results: MnCl2 treatment led to the augmentation of the KHSPR mRNA expression and protein increase in N2a cell line. The MnCl2 treatment of N2a cells also showed an elevated liberation of IL-6, TNF-α, MCP–1, and IL-1β. Pharmacological agents like quercetin inhibiting PI3K, MAPK, and WNT pathways, MCC950 blocking NLRP3 pathways, and SAHA showed a decrease in KHSRP expression post Mn treatment. With the inhibition of KHSRP, a decline in the release of IL-1β, IL-6, MCP–1, and TNF-α was also observed. Conclusion: These results suggested that MnCl2 treatment of N2a cells induce the expression of KHSRP via the PI3K–or NLRP3 pathway. Furthermore, this elevated expression of KHSRP is responsible for an increment in the liberation of pro-inflammatory markers in N2a cells. More exploration is needed to throw light on the pathway driving the KHSRP. Keywords:KH–type Splicing Regulatory Protein, Manganese Neurotoxicity, Neuroblastoma, Neurodegeneration, Neuroinflammation.View:PDF (1.26 MB)

Screening the toxicity profile and genotoxicity mechanism of excess manganese confirmed by spectral shift

In this study, the toxicity induced by excessive doses of manganese (MnCl2), which is one of the essential trace elements for the continuation of the metabolic activities of the organisms, was investigated with the help of the Allium test. Toxicity was investigated by using physiological (percent germination, root length, weight gain), cytogenetic [mitotic index (MI), micronucleus (MN), chromosomal abnormalities (CAs)], biochemical [malondialdehyde (MDA), superoxide dismutase (SOD) catalase (CAT)] and anatomical (root tip meristematic cell damage) parameters. Allium cepa L. bulbs were divided into four groups as one control and three treatments. The control group was germinated with tap water, and the treatment groups were germinated with 250, 500 and 1000 µM doses of MnCl2. The germination process was continued for 72 h without interruption. At the end of the period, the root tips were collected, washed in distilled water and made ready for microscopic and spectrophotometric analyzes with the help of routine preparation techniques. As a result, the highest germination percentage, root length, weight gain and MI, and the lowest MN frequency, CAs numbers, MDA level, SOD and CAT enzyme activities were determined in the control group (group I). MnCl2 exposure caused a decrease in physiological parameter values and an increase in cytogenetic (except MI) and biochemical parameter values, depending on the dose. MnCl2 exposure induced MN and CAs such as fragment, sticky chromosome, vagrant chromosome, unequal distribution of chromatin and bridge. This genotoxic effect of MnCl2 was associated with DNA–MnCl2 interaction, and this interaction was also confirmed by bathochromic and hypochromic shifts in spectral analysis. Anatomical damages such as epidermis cell damage, flattened cell nucleus, cortex cell damage and cortex cell wall thickening were observed after MnCl2 treatment. As a result, it has been determined that excessive doses of the trace element Mn cause physiological, cytogenetic, biochemical and anatomical toxicity and A. cepa test material is a reliable bio-indicator in determining this toxicity.

A GATA-type transcription factor SreA affects manganese susceptibility by regulating the expression of iron uptake-related genes

SreA has been identified as a GATA-type transcription factor that represses iron uptake to avoid iron excess during iron sufficiency. However, knowledge about whether SreA also affects the homeostasis of other divalent metal ions is limited. In this study, by screening Aspergillus fumigatus transcription factor deletion mutant libraries, we demonstrate that the sreA deletion mutant shows the greatest tolerance to MnCl2 among the tested divalent metal ions. Fe and Mn stimuli are able to enhance the expression of SreA with the different time-dependent manner, while the expression of SreA contributes to Mn2+ tolerance. Lack of SreA results in abnormally increased expression of a series of siderophore biosynthesis genes and iron transport-related genes, especially under MnCl2 treatment. Further mechanistic exploration indicated that lack of SreA exacerbates abnormal iron uptake, and iron excess inhibits cellular Mn content; thus, deletion of sreA results in Mn tolerance. Thus, findings in this study have demonstrated a new unexplored function for the transcription factor SreA in regulation of the Mn2+ tolerance.

Loss of slc39a14 causes simultaneous manganese hypersensitivity and deficiency in zebrafish.

ABSTRACTManganese neurotoxicity is a hallmark of hypermanganesemia with dystonia 2, an inherited manganese transporter defect caused by mutations in SLC39A14. To identify novel potential targets of manganese neurotoxicity, we performed transcriptome analysis of slc39a14−/− mutant zebrafish that were exposed to MnCl2. Differentially expressed genes mapped to the central nervous system and eye, and pathway analysis suggested that Ca2+ dyshomeostasis and activation of the unfolded protein response are key features of manganese neurotoxicity. Consistent with this interpretation, MnCl2 exposure led to decreased whole-animal Ca2+ levels, locomotor defects and changes in neuronal activity within the telencephalon and optic tectum. In accordance with reduced tectal activity, slc39a14−/− zebrafish showed changes in visual phototransduction gene expression, absence of visual background adaptation and a diminished optokinetic reflex. Finally, numerous differentially expressed genes in mutant larvae normalised upon MnCl2 treatment indicating that, in addition to neurotoxicity, manganese deficiency is present either subcellularly or in specific cells or tissues. Overall, we assembled a comprehensive set of genes that mediate manganese-systemic responses and found a highly correlated and modulated network associated with Ca2+ dyshomeostasis and cellular stress. This article has an associated First Person interview with the first author of the paper.

LA-ICP-MS bioimaging demonstrated disturbance of metal ions in the brain of Parkinson's disease model mouse undergoing manganese-enhanced MRI.

Manganese-enhanced MRI (MEMRI) is a powerful tool to study neuronal activity and microarchitecture in vivo. Yet the influence of exogenous manganese on the brain of the Parkinson's disease (PD) model mouse is poorly understood. Laser ablation connected to inductively coupled plasma mass spectrometry (LA-ICP-MS) imaging for tissue section is an ideal tool to simultaneously analyze the metabolism of endogenous metal ions. In this study, DJ-1 knockout PD model mice were subjected to an MnCl2 saline treatment and the distribution of Mn and several other endogenous metal ions in brain regions was assessed by MEMRI and LA-ICP-MS imaging. The results demonstrated that Mn mainly deposited in subcortical regions, such as ventricles, hippocampus (HC), medial preoptic nucleus (MPO), lateral septal nucleus (LS), and ventromedial hypothalamic nucleus (VMH). The enhanced signal-to-noise ratio (S/N) determined by MEMRI for Mn is closely related to the signal in LA-ICP-MS imaging. Significantly, the treatment of MnCl2 disturbs the homeostasis of iron, zinc, copper, and calcium in the DJ-1 mouse, which could result in more severe symptoms of PD. Therefore, the application of MEMRI in the study of neurological disease must be made with caution.

Oral administration of MnCl2 attenuated hyperlipidemia-related cardiac remodeling in ApoE−/− mice

Manganese chloride (MnCl2) has been shown to inhibit the Yes-associated protein (YAP) in high-fat diet-fed ApoE−/− mice. Although YAP has been implicated in atherogenesis, there are limited data on the effects of MnCl2 on cardiac remodeling. In this study, we discovered, by electrocardiography, that hyperlipidemia led to spontaneous supraventricular arrhythmia (SVA) in ApoE−/− (KO) mice, with 3 of 9 KO + MnCl2 mice (33%) exhibiting lower incidence of spontaneous SVA than KO mice (6 of 10 mice, 60%). Echocardiography revealed that reduced systolic function in KO mice was reversed by MnCl2 treatment. Oil Red O staining of the aortas and biochemical analysis of lipid levels showed that MnCl2 inhibited plaque formation in a lipid metabolism-independent manner. MnCl2 inhibited inflammatory cell infiltration and reduced fibrosis, as evidenced by hematoxylin and eosin, immunohistochemical and Masson's trichrome staining, respectively. Our findings demonstrate that spontaneous SVA and reduced systolic function were blocked by MnCl2. Our findings show that MnCl2 was useful in delaying cardiac remodeling and reducing susceptibility to spontaneous SVA in a mouse model of hyperlipidemia.

VANGL2 protein stability is regulated by integrin αv and the extracellular matrix

Vang-like 2 (VANGL2) is a four-pass transmembrane protein required for a variety of polarized cell behaviors underlying embryonic development. Recent data show human VANGL2 interacts with integrin αv to control cell adhesion to extracellular matrix proteins. The goal of this study was to further define the functional relationship between integrin αv and VANGL2. We demonstrate integrin αv regulates VANGL2 protein levels both in vitro and in the zebrafish embryo. While integrin αv knockdown reduces VANGL2 expression at membrane compartments, it does not affect VANGL2 transcription. Knockdown of integrin β5, but not β1 or β3, also decreases VANGL2 protein levels. Inhibition of protein translation using cycloheximide demonstrates that integrin αv knockdown cells have increased VANGL2 degradation while interference with either proteasome or lysosome function restores VANGL2. We further show integrin activation and stimulation of cell-matrix adhesion using MnCl2 fails to influence VANGL2. However, MnCl2 treatment stabilizes VANGL2 protein expression levels in the presence of cycloheximide. In the converse experiment, blockage of integrin-mediated cell-matrix adhesion using a cyclic RGD peptide causes a reduction in VANGL2 protein levels. Together, our findings support a model where integrin αv and cellular interactions with the extracellular matrix are required to maintain VANGL2 protein levels and thus function at the plasma membrane.

Manganese promotes intracellular accumulation of AQP2 via modulating F-actin polymerization and reduces urinary concentration in mice.

Aquaporin-2 (AQP2) is a water channel protein expressed in principal cells (PCs) of the kidney collecting ducts (CDs) and plays a critical role in mediating water reabsorption and urine concentration. AQP2 undergoes both regulated trafficking mediated by vasopressin (VP) and constitutive recycling, which is independent of VP. For both pathways, actin cytoskeletal dynamics is a key determinant of AQP2 trafficking. We report here that manganese chloride (MnCl2) is a novel and potent regulator of AQP2 trafficking in cultured cells and in the kidney. MnCl2 treatment promoted internalization and intracellular accumulation of AQP2. The effect of MnCl2 on the intracellular accumulation of AQP2 was associated with activation of RhoA and actin polymerization without modification of AQP2 phosphorylation. Although the level of total and phosphorylated AQP2 did not change, MnCl2 treatment impeded VP-induced phosphorylation of AQP2 at its serine-256, -264, and -269 residues and dephosphorylation at serine 261. In addition, MnCl2 significantly promoted F-actin polymerization along with downregulation of RhoA activity and prevented VP-induced membrane accumulation of AQP2. Finally, MnCl2 treatment in mice resulted in significant polyuria and reduced urinary concentration, likely due to intracellular relocation of AQP2 in the PCs of kidney CDs. More importantly, the reduced urinary concentration caused by MnCl2 treatment in animals was not corrected by VP. In summary, our study identified a novel effect of MnCl2 on AQP2 trafficking through modifying RhoA activity and actin polymerization and uncovered its potent impact on water diuresis in vivo.

Comprehensive investigation of aberrant microRNAs expression in cells culture model of MnCl2-induced neurodegenerative disease

Manganese (Mn) is required in various human physiological processes. Excessive Mn exposure causes manganism, a progressive neurodegenerative disorder similar to idiopathic Parkinson's disease (IPD). However, the detailed mechanism of Mn-induced neurotoxicity is not yet fully understood. MicroRNAs (miRNAs) play important roles in gene expression regulation, and miRNA expression profile provides additional biological and prognostic information of diseases. In our study, RNA sequencing was performed to profile miRNAs in the SH-SY5Y cells following MnCl2 treatment. Expressions of 73 miRNAs were altered following excessive Mn treatment. Furthermore, has-miR-4306 was identified to target 3′UTR of ATP13A2 (PARK9) directly. Inhibition of has-miR-4306 efficiently restored Mn-induced cytotoxicity. Thus, for the first time, we revealed the miRNA effects of Mn ions to neuron cells, highlighted the involvement of miRNA regulation in neurodegeneration caused by Mn exposure, and provided a potential application of miRNAs in future therapeutic intervention.

Oral treatment with herbal formula B401 alleviates penile toxicity in aging mice with manganism.

The present study aims to elucidate the roles of nitric oxide synthase activity, oxidative stress, inflammation, and apoptosis in penile toxicity of aging mice associated with excess manganese (Mn) treatment and to investigate the effect of oral treatment with the herbal formula B401 in this respect. ICR strain mice were divided into two groups: the vehicle (sham group) and the B401 (50 mg/kg) group. The mice were orally treated for 5 days; then a high single dose of MnCl2 (100 mg/kg) was given by intraperitoneal injection to the mice. One day after MnCl2 treatment, corpora cavernosal tissues of both Mn-treated mice and their controls were simultaneously sampled to examine their immunohistochemical staining and Western blot analysis. Nitric oxide (NO) production, levels of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS), expression levels of factors governing angiogenesis (vascular endothelial growth factor), oxidative stress (catalase, superoxide dismutase 2,4-hydroxynonenal), inflammation (tumor necrosis factor alpha), apoptosis (B-cell lymphoma 2 [Bcl-2], Bcl-2-associated X protein [Bax], cleaved poly(adenosine diphosphate-ribose) polymerase [c-PARP], cytochrome C, caspase-12, and caspase-3) were evaluated in penile corpus cavernosum of the mice. We found that penile toxicity in the mice was enhanced under excess Mn treatment through reduction of NOS activity and increase in oxidative stress, inflammation, and apoptosis in the penile cavernous tissue. Furthermore, the penile toxicity in mice with manganism was alleviated by oral B401 treatment through enhancement of both nitric oxide synthesis and angiogenesis, with simultaneous reduction of oxidative stress, inflammation, and apoptosis in penile corpus cavernosum. We suggest that the herbal formula B401 may serve as a potential dietotherapeutic supplement for penile toxicity or dysfunction in aging males.

Synthesis and supramolecular structures of manganese complexes with N-pyrazolylpropanamide-derived ligands

The syntheses and single crystal X-ray structures of the multifunctional acrylamide-derived ligands N-pyrazolyl-2-methylpropanamide (1,=N-ppaMe) and a mixture of 3- and 5-methylpyrazolylpropanamide (2a and 2b,=N-3Meppa/N-5Meppa), and their Mn2+ complexes are reported, as is the Mn2+ complex of N-triazolylpropanamide (3,=N-tpa). The general synthesis involves treatment of MnCl2·4H2O with the appropriate ligand in solution in the presence of triethylorthoformate as dehydrating agent. This way Mn2Cl4(N-ppaMe)2(EtOH)2 (4), [Mn(N-3Meppa)2(N-5Meppa)][MnCl4] (5), and MnCl2(N-tpa)2(EtOH)2 (6) were obtained. In 4 the N-ppaMe ligand acts as N,O-chelating ligand whose oxygens bridge the two Mn2+ ions. The ligands are also chelating in 5. In contrast, monodentate N-coordination via the pyrazolyl ring was found for 6. The molecular and crystal structures of 1, 2a, 4, 5, and 6 have been determined by X-ray diffraction. The extensive hydrogen bonding that occurs in these compounds is discussed.

Local manganese chloride treatment accelerates fracture healing in a rat model.

This study investigated the effects of local delivery of manganese chloride (MnCl2), an insulin-mimetic compound, upon fracture healing using a rat femoral fracture model. Mechanical testing, histomorphometry, and immunohistochemistry were performed to assess early and late parameters of fracture healing. At 4 weeks post-fracture, maximum torque to failure was 70% higher (P<0.05) and maximum torsional rigidity increased 133% (P<0.05) in animals treated with 0.125 mg/kg MnCl2 compared to saline controls. Histological analysis of the fracture callus revealed percent new mineralized tissue was 17% higher (P<0.05) at day 10. Immunohistochemical analysis of the 0.125 mg/kg MnCl2 treated group, compared to saline controls, showed a 379% increase in the density of VEGF-C+ cells. In addition, compared to saline controls, the 0.125 mg/kg MnCl2 treated group showed a 233% and 150% increase in blood vessel density in the subperiosteal region at day 10 post-fracture as assessed by detection of PECAM and smooth muscle α actin, respectively. The results suggest that local MnCl2 treatment accelerates fracture healing by increasing mechanical parameters via a potential mechanism of amplified early angiogenesis leading to increased osteogenesis. Therefore, local administration of MnCl2 is a potential therapeutic adjunct for fracture healing.

Molecular characterization, heterologous expression and resistance analysis of OsLEA3-1 from Oryza sativa

Late embryogenesis abundant (LEA) proteins in organisms are closely associated with resistance to abiotic stresses. Here we characterized a rice LEA protein, OsLEA3-1, by bioinformatics analysis and heterologous expression in Escherichia coli. Bioinformatics analysis showed that OsLEA3-1 contains a 603-bp open reading frame encoding a putative polypeptide of 200 amino acids, which contains a “LEA_4” motif at positions 5–48 and belongs to a typical group 3 LEA. OsLEA3-1 polypeptide is rich in Ala, Lys, and Thr, but depleted in Cys, Pro, and Trp residues; and is strongly hydrophilic. Secondary structure prediction showed that OsLEA3-1 polypeptide contained an α-helical domain in positions 4-195 but not any β-sheet domain. OsLEA3-1 gene can express in shoot and root of germinating seeds, seedling, panicles, mature embryo, seed, and callus; and was also up-regulated by ultraviolet (UV), heat, cold, salt, and emergency drought. OsLEA3-1 gene was introduced into E. coli. A fusion protein of about 28.03 kDa was expressed in recombinant E. coli cells after the induction by isopropylthio-β-D-galactoside. Compared with control E. coli cells harbouring pET30a, the accumulation of the OsLEA3-1 fusion protein increased the tolerance of the E. coli recombinants under diverse abiotic stresses: high salinity, metal ions, hyperosmotic, heat, and UV radiation. The OsLEA3-1 has the ability to protect the lactate dehydrogenase activity under heating, drying, and MnCl2 treatment in vitro. The findings suggested that the OsLEA3-1 gene may contribute to the ability of adapting to stressful environments of plants.

Nitric oxide alleviates manganese toxicity by preventing oxidative stress in excised rice leaves

In the present study, we have investigated the effects of nitric oxide (NO) on alleviating manganese (Mn)-induced oxidative stress in rice leaves. Exogenous MnCl2 treatment to excised rice leaves for 24 and 48 h resulted in increased production of H2O2 and lipid peroxides, decline in the levels of antioxidants, glutathione and ascorbic acid, and increased activities of antioxidative enzymes, superoxide dismutase, guaiacol peroxidase, catalase, ascorbate peroxidase, dehydroascorbate reductase, and glutathione reductase. Treatment of rice leaves with 100 μM sodium nitroprusside (SNP), a NO donor, was effective in reducing Mn-induced increased levels of H2O2, lipid peroxides and increased activities of antioxidative enzymes. The levels of reduced ascorbate and glutathione were considerably recovered due to SNP treatment. The effect of SNP was reversed by the addition of NO scavenger, 2-(4-carboxy-2-phenyl)-4,4,5,5-tetramethyl-imidazoline-1-oxyl-3-oxide (c-PTIO) suggesting that ameliorating effect of SNP is due to release of NO. The results indicate that MnCl2 induces oxidative stress in excised rice leaves, lowers the levels of reduced ascorbate and glutathione, and elevates activities of the key antioxidative enzymes. NO appears to provide a protection to the rice leaves against Mn-induced oxidative stress and that exogenous NO application could be advantageous in combating the deleterious effects of Mn-toxicity in rice plants.

Regulation of Intracellular Manganese Homeostasis by Kufor-Rakeb Syndrome-associated ATP13A2 Protein

Mutations in the ATP13A2 gene are associated with Kufor-Rakeb syndrome (KRS) and are found also in patients with various other types of parkinsonism. ATP13A2 encodes a predicted lysosomal P5-type ATPase that plays important roles in regulating cation homeostasis. Disturbance of cation homeostasis in brains is indicated in Parkinson disease pathogenesis. In this study, we explored the biological function of ATP13A2 as well as the pathogenic mechanism of KRS pathogenic ATP13A2 mutants. The results revealed that wild-type ATP13A2, but not the KRS pathogenic ATP13A2 mutants, protected cells from Mn(2+)-induced cell death in mammalian cell lines and primary rat neuronal cultures. In addition, wild-type ATP13A2 reduced intracellular manganese concentrations and prevented cytochrome c release from mitochondria compared with the pathogenic mutants. Furthermore, endogenous ATP13A2 was up-regulated upon Mn(2+) treatment. Our results suggest that ATP13A2 plays important roles in protecting cells against manganese cytotoxicity via regulating intracellular manganese homeostasis. The study provides a potential mechanism of KRS and parkinsonism pathogenesis.

Synthesis, X-ray crystal structure, thermal and solution studies of a centrosymmetric metal–organic polymer based on proton transfer methodology

A 3-D metal–organic coordination polymer based on pyrazine-2,3-dicarboxylic acid (pzdcH2) formulated as {[Mn(pzdc)(H2O)2] · 2H2O} n (1), was obtained by the treatment of MnCl2 · 6H2O with pzdcH2, 8-hydroxy quinoline (8-HQ), and 2-amino-4-methyl pyridine (ampy). In this study, we describe the synthesis, elemental analysis, IR spectroscopy, TG analysis, and single X-ray diffraction. The X-ray single crystal structure reveals that the chemical environment around each Mn(II) is a distorted octahedral by participating one nitrogen, five oxygens from three (pzdc)2− and two water molecules, O5MnN. The centrosymmetric 1-D ladder-like structure of 1 is bridged by oxygens of (pzdc)2−, and Mn–pzdc–Mn bonds are the rungs for our ladder structure. In the crystal structure, intermolecular O–H ··· O hydrogen bonds result in the formation of a supramolecular structure, effective for the stabilization of the structure. The thermal decomposition of 1 indicates that it is quite thermally stable. The protonation constants of ampy, 8-HQ and pzdcH2 as the building blocks of proton transfer systems including pzdcH2-ampy and pzdcH2-8-HQ and the corresponding stability constants of these systems were determined by potentiometric study. The stoichiometry and stability constants of complexes of pzdcH2-ampy and pzdcH2-8-HQ with Mn2+ were investigated by this method in aqueous solution. The results obtained from solution study are comparable with the solid state results.

Chronic Exposure to Manganese Alters Brain Responses to Amphetamine: A Pharmacological Magnetic Resonance Imaging Study

The parkinsonian symptoms and increased Mn accumulation in dopaminergic (DAergic) neurons of the basal ganglia implicate impaired dopamine signaling in the neurotoxic effects of chronic manganese overexposure. Using blood oxygenation level-dependent (BOLD) pharmacological magnetic resonance imaging (phMRI), we mapped brain responses to acute amphetamine (AMPH; 3 mg/kg, ip), which stimulates midbrain DAergic systems, in male Sprague-Dawley rats following 6 weeks of chronic MnCl(2) (5 mg Mn/kg, one per week, iv) or saline treatment. Plasma Mn content, measured immediately following phMRI, was elevated twofold in Mn-treated animals (p < 0.05), but the twofold increase in mean striatal Mn content did not reach significance. In saline-treated animals, AMPH stimulated robust positive BOLD responses throughout the basal ganglia and their reciprocally innervated connections. In contrast, acute AMPH stimulated a negative BOLD response in many of these structures in the Mn-treated group, resulting in significant differences between saline- and Mn-treated AMPH-evoked BOLD responses within caudate putamen, globus pallidus, substantia nigra, mediodorsal thalamic nucleus, and somatosensory cortex. These results demonstrate the utility of AMPH-evoked phMRI as readout of the DAergic signaling in vivo and confirm the vulnerability of DAergic systems to Mn.

Protective effect of tert-butylhydroquinone on PC12 cells from neurotoxicity induced by manganese in vitro

To investigate the protective effect of the tert-butylhydroquinone (tBHQ) on PC12 cells from neurotoxicity induced by manganese. Cytotoxicity of PC12 cells was measured by MTT assay, following the PC12 cells treatment with different concentrations of MnCl₂ (300, 600, 900 μmol/L) for 24, 48 or 72 h. PC12 cells were pretreated with 40 μmol/L tBHQ for 12 h, followed by the treatment of 600 micromol/L or 300 μmol/L MnCl₂ for 72 h. Cytotoxicity of PC12 cells was measured by MTT assay, and cell apoptosis was examined by the method of Annexin V-FITC/PI in flow cytometry (FCM). The proliferation of PC12 cells treated with 300, 600, 900 μmol/L MnCl2 was suppressed in the dose dependent pattern (P < 0.01). Proliferation of PC12 cells treated with 600 μmol/L MnCl₂ was suppressed to 40% of that in control group (P < 0.01), but the proliferation rate of PC12 cell pretreated with 40 μmol/L tBHQ was 180% of that in control group (P < 0.01). Apoptotic rate of PC12 cells treated with 300 micromol/L MnCl₂ was higher than the vehicle control group (P < 0.01). Apoptotic rate of 40 μmol/L tBHQ pretreatment followed by 300 μmol/L MnCl₂ treatment was lower than that of MnCl2 treatment group (P < 0.01). The inhibition rate of apoptosis was 61%. Manganese may suppress PC12 cells proliferation and induce apoptosis. tBHQ can reduce PC12 cells proliferation suppressed by manganese and attenuate the apoptosis induced by manganese.