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Carbon monoxide poisoning is one of the most common forms of poisoning in the world. Although the primary mode of treatment, oxygen therapy, is highly effective in many cases, there are instances in which it is inadequate or inappropriate. Whereas oxygen therapy relies on high levels of a low-affinity ligand (O2) to displace a high-affinity ligand (CO) from metalloproteins, an antidote strategy relies on introducing a molecule with a higher affinity for CO than native proteins (Kantidote,CO > Kprotein,CO). Based on the fundamental chemistry of CO, such an antidote is most likely required to be an inorganic compound featuring an electron-rich transition metal. A review is provided of the protein-, supramolecular complex-, and small molecule-based CO poisoning antidote platforms that are currently under investigation.

Aberrant response to physiological cell stress is part of the mechanisms underlying the development of diverse human diseases, including neuropathologies. Neuroglobin (NGB), an intracellular monomeric globin, has gained attention for its role in endogenous stress response pathways in neuroprotection. To date, evidence supports the concept of NGB as an inducible protein, triggered by physiological and pathological stimuli via transcriptional and/or post-transcriptional mechanisms, offering cell-autonomous neuroprotective functions under various cellular stresses. Notably, recent evidence suggests the extracellular occurrence of NGB. We aimed to explore whether NGB redistribution in the cell microenvironment may serve in transmitting resilience capability in a model with neuronal characteristics. Results obtained in SH-SY5Y demonstrated that intracellular NGB upregulation is associated with the promotion of the extracellular release of the globin. Additionally, cell secretome from NGB-overexpressing cells, characterized by globin accumulation, exhibits protective effects against oxidative stress and mitochondrial toxicity, as evidenced by reduced apoptosis and preserved mitochondrial structure. These findings shed light on the potential significance of extracellular NGB as part of a common cell response to physiological and stress conditions and as a factor promoting cell resilience. Furthermore, the potential for neuroprotection of extracellular NGB paves the way for future therapeutic opportunities.

[Display omitted] • Ngb protects MMP+- induced MN9D cells against apoptosis. • Ngb rescues mitochondrial function in the PD cell model. • Ngb interacts with NDUFA10 subunit to rescue Complex I activity in PD cell model. The study aimed to validate the protective effect of neuroglobin (Ngb) in a cell model of Parkinson's disease (PD) and explore its therapeutic potential. Lentivirus-Ngb (LvNgb) and siRNA-Ngb (siNgb) were used to achieve Ngb overexpression and knockdown, respectively, in a sporadic PD cell model. Apoptosis was evaluated by flow cytometry-based Annexin V/propidium iodide assays. Activation of the pro-apoptotic factor, Caspase-9, was detected by immunoblotting, and Complex I activities were detected by using enzyme-linked immunosorbent assay (ELISA). Mitochondrial dysfunction was examined by measuring the mitochondrial membrane potential (MMP), NAD+/NADH ratios, and reactive oxygen species (ROS) levels. Additionally, coimmunoprecipitation (Co-IP) assays were conducted in mouse neuroblastoma cell line 9D (MN9D) cells to determine the interactions of Ngb with the Complex I subunit NDUFA10. The results showed that Ngb overexpression reduced the percentages of apoptotic cells, total caspase-9 levels and restored Complex I activities in the PD cell model. Conversely, knockdown of Ngb resulted in an increase in apoptotic cells, higher total caspase-9 levels, and decreased Complex I activities. Furthermore, Ngb overexpression restored MMP and NAD+/NADH ratios and alleviated ROS-mediated oxidative stress in MN9D cells. Finally, Co-IP confirmed the interaction between Ngb and NDUFA10 in MN9D cells. In conclusion, Ngb protects MN9D cells against apoptosis by interacting with Complex I subunit NDUFA10, rescuing its activity and inhibiting the mitochondrial pathway of apoptosis in the MPP+-mediated PD model. [ABSTRACT FROM AUTHOR]

Neuroglobin (Ngb) is a cytosolic heme protein that plays an important role in protecting cells from apoptosis through interaction with oxidized cytochrome c (Cyt c) released from mitochondria. The interaction of reduced Ngb and oxidized Cyt c is accompanied by electron transfer between them and the reduction in Cyt c. Despite the growing number of studies on Ngb, the mechanism of interaction between Ngb and Cyt c is still unclear. Using Raman spectroscopy, we studied the effect of charged amino acid substitutions in Ngb and Cyt c on the conformation of their hemes. It has been shown that Ngb mutants E60K, K67E, K95E and E60K/E87K demonstrate changed heme conformations with the lower probability of the heme planar conformation compared to wild-type Ngb. Moreover, oxidized Cyt c mutants K25E, K72E and K25E/K72E demonstrate the decrease in the probability of methyl-radicals vibrations, indicating the higher rigidity of the protein microenvironment. It is possible that these changes can affect electron transfer between Ngb and Cyt c.

Neuroglobin is a hemoprotein expressed in several nervous system cell lineages with yet unknown physiological functions. Neuroglobin presents a very similar structure to that of the related globins hemoglobin and myoglobin, but shows an hexacoordinate heme as compared to the pentacoordinated heme of myoglobin and hemoglobin. While several reactions of neuroglobin have been characterized in vitro, the relative importance of most of those reactions in vivo is yet undefined. Neuroglobin, like other heme proteins, can reduce nitrite to nitric oxide, providing a possible route to generate nitric oxide in vivo in low oxygen conditions. The reaction kinetics are highly dependent on the nature of the distal residue, and replacement of the distal histidine His64(E7) can increase the reaction rate constants by several orders of magnitude. However, mutation of other distal pocket positions such as Phe28(B10) or Val68(E11) has more limited impact on the rates. Computational analysis using myoglobin as template, guided by the structure of dedicated nitrite reductases like cytochrome cd1 nitrite reductase, has pointed out that combined mutations of the residues B10 and CD1 could increase the nitrite reductase activity of myoglobin, by mimicking the environment of the distal heme pocket in cytochrome cd1 nitrite reductase. As neuroglobin shows high sequence and structural homology with myoglobin, we hypothesized that such mutations (F28H and F42Y in neuroglobin) could also modify the nitrite reductase activity of neuroglobin. Here we study the effect of these mutations. Unfortunately, we do not observe in any case an increase in the nitrite reduction rates. Our results provide some further indications of nitrite reductase regulation in neuroglobin and highlight the minor but critical differences between the structure of penta- and hexacoordinate globins.

Neuroglobin (Ngb) is a member of the globin family. Like other globin proteins, it is involved in the maintenance of oxygen homeostasis. Ngb distribution in the normal brain is well known, but its response to pathological injury, such as cerebral ischemia, has not yet been adequately elucidated. One primary reason for this is that ischemic lesions in cerebral tissues have mostly been studied in transgenic organisms. In addition, the available data on Ngb content are limited to a small number of brain structures. This article examines the patterns of changes in Ngb immunoreactivity in neurons from different parts of the rat brain after subtotal cerebral ischemia of varying duration. An immunohistochemical study of Ngb content in 25 brain structures of white male Wistar rats exposed to 30-min and 3-h subtotal cerebral ischemia was performed. A decrease in Ngb content in all structures (especially 3 h after the ischemia onset and in the phylogenetically older parts), temporal mediation, and dependence on the phylogenetic age were revealed. The obtained results further decipher the correlation between the changes in Ngb content and the degree of cerebral ischemic damage, which is necessary to clarify the functions of the studied protein.

Mutant forms of human neuroglobin that carry targeted mutations in the putative interface with cytochrome c: single (E60K, K67E, E87K, K95E) and double (E60K\E87K) substitutions were obtained by site-directed mutagenesis. The E60K, K95E, and E60K\E87K mutations cause slight changes in the UV-vis absorption spectra, which can be associated with both a change in the electrostatic field near the heme and a change in the heme iron spin to the high-spin state. The secondary structure of mutant neuroglobins calculated from the CD spectral data almost did not differ from the secondary structure of wild-type neuroglobin, except for the protein with the K67E substitution, whose β-turn is reorganized into an α-helix. The IR spectra provide further evidence for the predominance of α-helices in protein secondary structure for mutant forms of neuroglobin. Thus, the introduction of these mutations did not have a significant effect on the characteristics of the heme-containing protein neuroglobin. The developed mutant forms will be used to study the contribution of individual amino acid residues to the formation of the reaction complex between neuroglobin and cytochrome c, which will allow rational design of drugs for the therapy of various diseases associated with neuronal death in the future. [ABSTRACT FROM AUTHOR]

Globins have been studied as model proteins to elucidate the principles of protein evolution. This was achieved by understanding the relationship between amino acid sequence, three-dimensional structure, physicochemical properties, and physiological function. Previous molecular phylogenies of chordate globin genes revealed the monophyletic evolution of urochordate globins and suggested convergent evolution. However, to provide evidence of convergent evolution, it is necessary to determine the physicochemical and functional similarities between vertebrates and urochordate globins. In this study, we determined the expression patterns of Ciona globin genes using real-time RT-PCR. Two genes (Gb-1 and Gb-2) were predominantly expressed in the branchial sac, heart, and hemocytes and were induced under hypoxia. Combined with the sequence analysis, our findings suggest that Gb-1/-2 correspond to vertebrate hemoglobin-α/-β. However, we did not find a robust similarity between Gb-3, Gb-4, and vertebrate globins. These results suggested that, even though Ciona globins obtained their unique functions differently from vertebrate globins, the two of them shared some physicochemical features and physiological functions. Our findings offer a good example for understanding the molecular mechanisms underlying gene co-option and convergence, which could lead to evolutionary innovations. [ABSTRACT FROM AUTHOR]

The discovery of neuroglobin (Ngb), a brain- or neuron-specific member of the hemoglobin family, has revolutionized our understanding of brain oxygen metabolism. Currently, how Ngb plays such a role remains far from clear. Here, we report a novel mechanism by which Ngb might facilitate neuronal oxygenation upon hypoxia or anemia. We found that Ngb was present in, co-localized to, and co-migrated with mitochondria in the cell body and neurites of neurons. Hypoxia induced a sudden and prominent migration of Ngb towards the cytoplasmic membrane (CM) or cell surface in living neurons, and this was accompanied by the mitochondria. In vivo, hypotonic and anemic hypoxia induced a reversible Ngb migration toward the CM in cerebral cortical neurons in rat brains but did not alter the expression level of Ngb or its cytoplasm/mitochondria ratio. Knock-down of Ngb by RNA interference significantly diminished respiratory succinate dehydrogenase (SDH) and ATPase activity in neuronal N2a cells. Over-expression of Ngb enhanced SDH activity in N2a cells upon hypoxia. Mutation of Ngb at its oxygen-binding site (His64) significantly increased SDH activity and reduced ATPase activity in N2a cells. Taken together, Ngb was physically and functionally linked to mitochondria. In response to an insufficient oxygen supply, Ngb migrated towards the source of oxygen to facilitate neuronal oxygenation. This novel mechanism of neuronal respiration provides new insights into the understanding and treatment of neurological diseases such as stroke and Alzheimer's disease and diseases that cause hypoxia in the brain such as anemia. [ABSTRACT FROM AUTHOR]

Objectives The objective of this article was to explore the effects of neuroglobin (NGB) on ferroptosis in the nerve cells.

Neuroglobin, which is a heme protein from the globin family that is predominantly expressed in nervous tissue, can promote a neuronal survivor. However, the molecular mechanisms underlying the neuroprotective function of Ngb remain poorly understood to this day. The interactions between neuroglobin and mitochondrial cytochrome c may serve as at least one of the mechanisms of neuroglobin-mediated neuroprotection. Interestingly, neuroglobin and cytochrome c possibly can interact with or without electron transfer both in the cytoplasm and within the mitochondria. This review provides a general picture of molecular interactions between neuroglobin and cytochrome c based on the recent experimental and computational work on neuroglobin and cytochrome c interactions. [ABSTRACT FROM AUTHOR]

Cerebral ischemia/reperfusion (I/R) injury remains a grievous health threat, and herein effective therapy is urgently needed. This study explored the protection of neuroglobin (Ngb) in rats with cerebral I/R injury. The focal cerebral I/R rat models were established by middle cerebral artery occlusion (MCAO) and neuronal injury models were established by oxygen–glucose deprivation/reoxygenation (OGD/R) treatment. The brain injury of rats was assessed. Levels of Ngb, Bcl‐2, Bax, endoplasmic reticulum stress (ERS)‐related markers, and Syt1 were measured by immunofluorescence staining and Western blotting. The cytotoxicity in neurons was assessed by lactate dehydrogenase (LDH) release assay. Levels of intracellular Ca2+ and mitochondrial function‐related indicators were determined. The binding between Ngb and Syt1 was detected by co‐immunoprecipitation. Ngb was upregulated in cerebral I/R rats and its overexpression alleviated brain injury. In OGD/R‐induced neurons, Ngb overexpression decreased LDH level and neuronal apoptosis, decreased Ca2+ content, and mitigated mitochondrial dysfunction and ERS‐related apoptosis. However, Ngb silencing imposed the opposite effects. Importantly, Ngb could bind to Syt1. Syt1 knockdown partially counteracted the alleviation of Ngb on OGD/R‐induced injury in neurons and cerebral I/R injury in rats. Briefly, Ngb extenuated cerebral I/R injury by repressing mitochondrial dysfunction and endoplasmic reticulum stress‐mediated neuronal apoptosis through Syt1. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is an extremely aggressive malignancy with a very disappointing prognosis. Neuroglobin (NGB), a member of the globin family, has been demonstrated to have a significant role in a variety of tumor forms. The possible role of NGB as a tumor suppressor gene in pancreatic cancer was investigated in this work. Information from the public dataset TCGA combined with GTEx was used to analyze the finding that NGB was commonly downregulated in pancreatic cancer cell lines and tissues, correlating with patient age and prognosis. The expression of NGB in pancreatic cancer was investigated via RT-PCR, qRT-PCR, and Western blot experiments. In-vitro and in-vivo assays, NGB elicited cell cycle arrest in the S phase and apoptosis, hindered migration and invasion, reversed the EMT process, and suppressed cell proliferation and development. The mechanism of action of NGB was predicted via bioinformatics analysis and validated using Western blot and co-IP experiments revealed that NGB inhibited the EGFR/AKT/ERK pathway by binding to and reducing expression of GNAI1 and p-EGFR. In addition, pancreatic cancer cells overexpressing NGB showed increased drug sensitivity to gefitinib (EGFR-TKI). In conclusion, NGB inhibits pancreatic cancer progression by specifically targeting the GNAI1/EGFR/AKT/ERK signaling axis. • In this study, the significant tumor-suppressive effect of NGB in pancreatic cancer is described for the first time. • This study is the first to investigate the mechanism of action of NGB in inhibiting the development of pancreatic cancer. • The relationship between the interaction of NGB with GNAI1 (G protein) and p-EGFR was revealed for the first time. • This study provides a basis for the potential application of NGB in pancreatic cancer. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is a debilitating neurodegenerative disease characterised by the accumulation of amyloid-beta and tau in the brain, leading to the progressive loss of memory and cognition. The causes of its pathogenesis are still not fully understood, but some risk factors, such as age, genetics, and hormones, may play a crucial role. Studies show that postmenopausal women have a higher risk of developing AD, possibly due to the decrease in hormone levels, especially oestrogen, which may be directly related to a reduction in the activity of oestrogen receptors, especially beta (ERβ), which favours a more hostile cellular environment, leading to mitochondrial dysfunction, mainly affecting key processes related to transport, metabolism, and oxidative phosphorylation. Given the influence of hormones on biological processes at the mitochondrial level, hormone therapies are of clinical interest to reduce the risk or delay the onset of symptoms associated with AD. One drug with such potential is tibolone, which is used in clinics to treat menopause-related symptoms. It can reduce amyloid burden and have benefits on mitochondrial integrity and dynamics. Many of its protective effects are mediated through steroid receptors and may also be related to neuroglobin, whose elevated levels have been shown to protect against neurological diseases. Its importance has increased exponentially due to its implication in the pathogenesis of AD. In this review, we discuss recent advances in tibolone, focusing on its mitochondrial-protective effects, and highlight how valuable this compound could be as a therapeutic alternative to mitigate the molecular pathways characteristic of AD. [ABSTRACT FROM AUTHOR]

Current management of glaucomatous optic neuropathy is limited to intraocular pressure control. Neuroglobin (Ngb) is an endogenous neuroprotectant expressed in neurons and astrocytes. We recently showed that exogenous intravitreal Ngb reduced inflammatory cytokines and microglial activation in a rodent model of hypoxia. We thus hypothesised that IVT-Ngb may also be neuroprotective in experimental glaucoma (EG) by mitigating optic nerve (ON) astrogliosis and microgliosis as well as structural damage. In this study using a microbead-induced model of EG in six Cynomolgus primates, optical coherence imaging showed that Ngb-treated EG eyes had significantly less thinning of the peripapillary minimum rim width, retinal nerve fibre layer thickness, and ON head cupping than untreated EG eyes. Immunohistochemistry confirmed that ON astrocytes overexpressed Ngb following Ngb treatment. A reduction in complement 3 and cleaved-caspase 3 activated microglia and astrocytes was also noted. Our findings in higher-order primates recapitulate the effects of neuroprotection by Ngb treatment in rodent EG studies and suggest that Ngb may be a potential candidate for glaucoma neuroprotection in humans. [ABSTRACT FROM AUTHOR]

An efficient system for the biosynthesis, isolation and purification of recombinant human neuroglobin has been developed and optimized, which makes it possible to produce protein in quantities sufficient to study its properties. According to UV-visible, IR-, CD-, and NMR spectroscopy data, recombinant neuroglobin is a structured protein in the holoform state. The data of chromato-mass-spectrometric analysis made it possible to conclude that there is a correctly formed disulfide bond in the structure of the oxidized form of the protein. Using Raman and surface-enhanced Raman spectroscopy with laser excitation at 532 nm, it was shown that heme in the reduced and oxidized forms of neuroglobin has vibrational degrees of freedom typical of b-type hemes, and the iron atom is hexacoordinated. Using Raman spectroscopy with laser excitation at 633 nm, it was found that reduced –SH-groups were present in reduced neuroglobin, while in oxidized neuroglobin a disulfide bridge was formed. The results obtained serve as the basis for detailed studies of the functioning of neuroglobin as a neuroprotector, in particular, during its interaction with oxidized cytochrome c, which is released from mitochondria in violation of their functioning and/or morphology. [ABSTRACT FROM AUTHOR]

Sawsan Abuhamdah,1,2 Tahia H Saleem,3 Bakheet EM Elsadek,4 Omyma Ashraf,5 Ali R Hamdan,6 Eslam El Sayed El-Khateeb,6 Saeda M Abd Elwahab,7 Mohammed H Hassan5 1College of Pharmacy, Al Ain University, Abu Dhabi, United Arab Emirates; 2Department of Biopharmaceutics and Clinical Pharmacy, Faculty of Pharmacy, The University of Jordan, Amman, Jordan; 3Department of Medical Biochemistry, Faculty of Medicine, Assiut University, Assiut, Egypt; 4Department of Biochemistry and Molecular Biology, Faculty of Pharmacy, Al-Azhar University, Assiut, 71524, Egypt; 5Department of Medical Biochemistry, Faculty of Medicine, South Valley University, Qena, 83523, Egypt; 6Department of Neurosurgery, Faculty of Medicine, South Valley University, Qena, 83523, Egypt; 7Radio-Diagnosis Department, Faculty of Medicine, South Valley University, Qena, 83523, EgyptCorrespondence: Mohammed H Hassan, Department of Medical Biochemistry, Faculty of Medicine, South Valley University, Qena, 83523, Egypt, Tel +20 1098473605, Email Mohammedhosnyhassaan@yahoo.comIntroduction: Traumatic spinal cord injury (TSCI) is a life-threatening neurological disorder and there is a lack of biomarker research, particularly human studies that could help to categorize the severity and predict the outcome. We aimed to assess the role of serum Ubiquitin C-terminal hydrolase L1 (UCH-L1) and Neuroglobin (NGB) in predicting severity and outcome of TSCI.Methods: This prospective study included 63 participants categorized into 33 patients with various types of TSCI and 30 unrelated healthy volunteers. Neurosurgical [American spinal injury association (ASIA) impairment score (AIS)] and radiological [using spine computed tomography (CT) and magnetic resonance imaging (MRI)] assessments were performed on the included patients to determine the severity and the level of injury with neurological follow-up of patients within 6 months post-injury. Serum UCH-L1 and NGB were measured for all participants using commercially available ELISA assay kits.Results: Of the included patients, 20 (60.60%) had partial SCI and the remaining 13 patients (39.39%) had complete SCI. On follow-up, 19 patients (57.57%) showed improved AIS, while 14 cases (42.42%) did not show any improvement in their AIS scores. There was significantly higher median serum UCHL1 value among cases compared to controls (1723 pg/mL and 657 pg/mL, respectively), p ˂ 0.05. There was an insignificant rise of serum NGB levels among cases in comparison with the controls (15.2pg/mL and 7.52pg/mL, respectively, p ˃ 0.05). Significantly lower initial median serum UCHL1 levels (pg/mL) were observed in patients with improved AIS during the neurological follow-up compared with those who did not show any improvement in their AIS score (1723, and 4700 respectively, p ˂ 0.05), with lack of significant difference in the initial median serum NGB levels, p ˃ 0.05.Conclusion: Initial serum UCHL1 assay could be a helpful marker in reflecting the degree of TSCI and predicting its outcome, though NGB needs further assessment.Keywords: traumatic spinal cord injury, Ubiquitin C-terminal hydrolase L1, neuroglobin, diagnostic and prognostic markers

In the present study, human neuroglobin (hNgb) was found to undergo H2O2‐induced breakdown of the heme center at a much slower rate than other globins, namely in the timescale of hours against minutes. We investigated how the rate of the process is affected by the Cys46/Cys55 disulfide bond and the network of non‐covalent interactions in the distal heme side involving Tyr44, Lys67, the His64 heme iron axial ligand and the heme propionate‐7. The rate is increased by the Tyr44 to Ala and Phe mutations; however the rate is lowered by Lys67 to Ala swapping. The absence of the disulfide bridge slows down the reaction further. Therefore, the disulfide bond‐controlled accessibility of the heme site and the residues at position 44 and 67 affect the activation barrier of the reaction. Wild‐type and mutated species form β‐amyloid aggregates in the presence of H2O2 producing globular structures. Furthermore, the C46A/C55A, Y44A, Y44F and Y44F/C46A/C55A variants yield potentially harmful fibrils. Finally, the nucleation and growth kinetics for the aggregation of the amyloid structures can be successfully described by the Finke–Watzky model. [ABSTRACT FROM AUTHOR]

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Background and Purpose: Intracerebral and subarachnoid hemorrhage are critical conditions with a high mortality, and the outcome for the individual patient is notoriously difficult to predict. Biomarkers that reflect disease severity and predict outcome are therefore warranted.Methods: Blood samples from 40 patients with intracerebral, 46 patients with subarachnoid hemorrhage, and 70 healthy individuals were collected. Levels of glial fibrillary acidic protein (GFAP) and neuroglobin were measured by ultra-sensitive single molecule array and enzyme-linked immunosorbent assay, respectively. Clinical information including mortality and functional outcome was recorded.Results: Blood levels of GFAP and neuroglobin in intracerebral and subarachnoid hemorrhage patients were significantly elevated when compared to healthy individuals (all p < 0.0001). GFAP levels were significantly higher in patients dying or with poor functional outcome than in healthy individuals (all p ≤ 0.01). GFAP levels separated survivors from non-survivors with an area under receiver operating characteristics (AUROC) = 0.78 (confidence interval (CI) 0.59-0.98) for intracerebral hemorrhage and 0.82 (CI 0.69-0.94) for subarachnoid hemorrhage patients. The Akaike and Bayesian information criteria (AIC/BIC) for mortality/poor outcome prediction improved when combining GFAP levels with hematoma volume (p = 0.04/p < 0.01), National Institutes of Health Stroke Scale (NIHSS) (p = 0.09/p < 0.01), Hunt-Hess (p < 0.05/p = 0.21), or Fischer score (p < 0.05/p = 0.02).Conclusions: Elevated GFAP levels at admission to hospital predicted mortality and poor outcome in our cohort of intracerebral and subarachnoid hemorrhage patients. Neuroglobin levels did not provide additional information. Combining GFAP measurements with clinical disease severity scores increased outcome prediction precision. This may suggest that GFAP measurement could improve prognostication in patients with intracerebral or subarachnoid hemorrhage.Registration: This sub-trial was not registered. [ABSTRACT FROM AUTHOR]

We present a comprehensive investigation on the different role of CO in carboxyneuroglobin (1) as ligand of the heme group in the active site forming a bond with the heme iron and (2) dissociated from the heme group but still trapped inside the active site, focusing on two specific orientations, one with CO perpendicular to the plane defined by the distal histidine of the enzyme (form A) and one with CO located parallel to that plane (form B). Our study includes wild type carboxy‐neuroglobin and nine known protein mutations. Considering that the distal histidine interacting with the heme group can adapt two different tautomeric forms and the two possible orientations of the dissociated CO, a total of 36 protein systems were analyzed in this study. Fully optimized geometries and vibrational frequencies were calculated at the QM/MM level, followed by the local mode analysis, to decode CO bond properties. The intrinsic bond strengths derived from the local mode analysis, complemented with NBO and QTAIM data, reveal that the strength of the CO bond, in the hexacoordinate (where CO is a ligand of the heme group) and pentacoordinate (where CO is dissociated from the heme group) scenarios, is dominated by through bond and through space charge transfer between CO and Fe, fine‐tuned by electrostatic and dispersion interactions with the side chain amino acids in the distal heme pocket. Suggestions are made as to advise on how protein modifications can influence the molecular properties of the coordinated or dissociated CO, which could serve the fine‐tuning of existing and the design of new neuroglobin models with specific FeC and CO bond strengths. [ABSTRACT FROM AUTHOR]

Background/Aim: The brain uses 20% of the O2 consumed by the body for energy metabolism. In 2000, found a protein that is thought to be a binding O2 in the brain, namely neuroglobin (Ngb). Ngb is a member of the hemoprotein which has a heme group. The iron ion in the haem group can be oxidized, so a reducing enzyme is needed. In this study, the isolation, purification, and characterization of Ngb protein and the reducing enzyme from oxidized neuroglobin (neuroglobin Fe3+) were carried out. Materials and methods: Ngb protein was isolated by fractionation technique using ammonium sulfate 90% saturation, purified by anion exchange chromatography (DEAE Cellulose) and immunoaffinity chromatography, confirmed by SDS-PAGE and Western blot. The metneuroglobin-reducing enzyme was isolated by RIPA lysis buffer, purified by Affi gel blue chromatography, and confirmed by SDS-PAGE. Results: The isolated Ngb obtained has a molecular weight of 17.26 kDa. Spectrum analysis in the wavelength range of 350- 500nm, showed the afternoon peaks of deoxyNgb, oxyNgb, carboxyNgb and metNgb were 415 nm, 405 nm, 405 nm, and 420 nm, respectively. The results of the isolation of the reducing enzymes obtained consisted of 2 parts, namely the matrix-bound eluate (eluate-1) and matrix-bound eluate (eluate-2). SDS-PAGE results of eluate-1, eluate-2 and Ngb-free fraction (byproduct of Ngb purification) showed the same 3 bands at a molecular weight of 72.45; 26.84 and 16.33 kDa were suspected as reducing enzymes. Conclusion: The reduction kinetics was tested by reacting the fraction and metNgb and measuring the deoxyNgb uptake formed per unit time. The results of the measurement of the ratio of NgbFe3+ to NgbFe2+ from the free fractions Ngb, eluate-1 and eluate-2, which has the best reducing activity is eluate-1 because it has the best regression value of 0.8769. [ABSTRACT FROM AUTHOR]

The present work shows the biochemical and structural fundamentals for the stress induced anxiety and stress adjustment response of the CA3 hippocampus area. Adult male Wistar rats were repeatedly exposed to a 3 h day restraint stress, for either 3 or 6 days. The concentration of corticosterone and testosterone in the CA3 hippocampus area was divergent, while oxidative stress was progressively increased during the stress exposure. The mitochondrial lysis in the CA3 neurons confirmed the oxidative stress events. Immunohistochemical findings showed that oligodendrocytes (OCs) proliferation and neuroglobin (Ngb) expression were stimulated, whereas MeCP2 expression was decreased as a balance reaction in stress exposure under corticosterone signaling. Remarkably, ultrastructural changes such as mitochondrial lysis, endoplasmic reticulum swelling, and perivascular lysis with platelets adherence to endothelium in the CA3 area were seen in the 6th day of restraining. The anxiety-like behavior was noticed 6 days later after stress exposure. These results suggest that the duration of the exposure, but not the intensity of the stress, is the key factor in the stress-buffering function by the CA3 hippocampus area via up-regulation of the Ngb-OCs bionome. The imbalance of the Ngb-OCs communication may be involved in the development of CA3-dependent anxious behavior. [ABSTRACT FROM AUTHOR]

Purpose: Glial fibrillary acidic protein (GFAP) and neuroglobin (NGB) are important biomarkers of cerebral hypoxia. For this reason, an attempt was made to assess their concentrations in various time intervals and their impact on the severity of neurological symptoms and functional prognosis of thrombolytic ischemic stroke patients., Patients and Methods: The study involved 94 patients reporting to the emergency department of the Collegium Medicum University Hospital in Bydgoszcz within < 4.5 hours of the onset of stroke symptoms. GFAP and neuroglobin levels were measured in plasma at indicated times using a commercial ELISA kit., Results: Based on the data gathered, statistically significant differences were found between the concentration of biomarkers in stroke patients and the control group. The concentrations of both biomarkers, GFAP and NGB, were elevated in patients after ischemic stroke and the changes in their concentrations in the subsequent stages of stroke may suggest their prognostic value strictly dependent on time. NGB was determined on the 7th day, and mRS - after a year (0.35). GFAP measured after 24 h and on day 7 could be a promising biomarker of functional outcome after one year (cut-off point ≤ 0.231 ng/mL, sensitivity 75.0%, specificity 61.2%, cut off point ≤ 0.235 ng/mL, sensitivity 75.0%, specificity 73.9%, respectively) and the severity of the patient's neurological condition. At GFAP concentrations above 0.25 ng/mL, measured within 24 hours, a sharp increase in mortality was observed in stroke patients. In the case of NGB, at the time of stroke occurrence (14 ng/mL) and after 24 hours (10-60 ng/mL). Differences in the concentrations of these biomarkers have been demonstrated in different stroke subtypes., Conclusion: NGB and GFAP are important biomarkers of ischemic brain injury and may also participate in predicting neurological outcomes., Competing Interests: The authors report no conflicts of interest in this work., (© 2024 Pawluk et al.)

Neuroglobin, which is a heme protein from the globin family that is predominantly expressed in nervous tissue, can promote a neuronal survivor. However, the molecular mechanisms underlying the neuroprotective function of Ngb remain poorly understood to this day. The interactions between neuroglobin and mitochondrial cytochrome c may serve as at least one of the mechanisms of neuroglobin-mediated neuroprotection. Interestingly, neuroglobin and cytochrome c possibly can interact with or without electron transfer both in the cytoplasm and within the mitochondria. This review provides a general picture of molecular interactions between neuroglobin and cytochrome c based on the recent experimental and computational work on neuroglobin and cytochrome c interactions.

Alzheimer’s disease (AD) is a debilitating neurodegenerative disease characterised by the accumulation of amyloid-beta and tau in the brain, leading to the progressive loss of memory and cognition. The causes of its pathogenesis are still not fully understood, but some risk factors, such as age, genetics, and hormones, may play a crucial role. Studies show that postmenopausal women have a higher risk of developing AD, possibly due to the decrease in hormone levels, especially oestrogen, which may be directly related to a reduction in the activity of oestrogen receptors, especially beta (ERβ), which favours a more hostile cellular environment, leading to mitochondrial dysfunction, mainly affecting key processes related to transport, metabolism, and oxidative phosphorylation. Given the influence of hormones on biological processes at the mitochondrial level, hormone therapies are of clinical interest to reduce the risk or delay the onset of symptoms associated with AD. One drug with such potential is tibolone, which is used in clinics to treat menopause-related symptoms. It can reduce amyloid burden and have benefits on mitochondrial integrity and dynamics. Many of its protective effects are mediated through steroid receptors and may also be related to neuroglobin, whose elevated levels have been shown to protect against neurological diseases. Its importance has increased exponentially due to its implication in the pathogenesis of AD. In this review, we discuss recent advances in tibolone, focusing on its mitochondrial-protective effects, and highlight how valuable this compound could be as a therapeutic alternative to mitigate the molecular pathways characteristic of AD.

Current management of glaucomatous optic neuropathy is limited to intraocular pressure control. Neuroglobin (Ngb) is an endogenous neuroprotectant expressed in neurons and astrocytes. We recently showed that exogenous intravitreal Ngb reduced inflammatory cytokines and microglial activation in a rodent model of hypoxia. We thus hypothesised that IVT-Ngb may also be neuroprotective in experimental glaucoma (EG) by mitigating optic nerve (ON) astrogliosis and microgliosis as well as structural damage. In this study using a microbead-induced model of EG in six Cynomolgus primates, optical coherence imaging showed that Ngb-treated EG eyes had significantly less thinning of the peripapillary minimum rim width, retinal nerve fibre layer thickness, and ON head cupping than untreated EG eyes. Immunohistochemistry confirmed that ON astrocytes overexpressed Ngb following Ngb treatment. A reduction in complement 3 and cleaved-caspase 3 activated microglia and astrocytes was also noted. Our findings in higher-order primates recapitulate the effects of neuroprotection by Ngb treatment in rodent EG studies and suggest that Ngb may be a potential candidate for glaucoma neuroprotection in humans.

Stroke is a global health and socio-economic problem. However, no efficient preventive and/or palliative treatments have yet been found. Neuroglobin (Ngb) is an endogen neuroprotective protein, but it only exerts its beneficial action against stroke after increasing its basal levels. Therefore, its systemic administration appears to be an efficient therapy applicable to stroke and other neurodegenerative pathologies. Unfortunately, Ngb cannot cross the blood-brain barrier (BBB), making its direct pharmacological use unfeasible. Thus, the association of Ngb with a drug delivery system (DDS), such as nanoparticles (NPs), appears to be a good strategy for overcoming this handicap. NPs are a type of DDS which efficiently transport Ngb and increase its bioavailability in the infarcted area. Hence, we previously built hyaluronate NPS linked to Ngb (Ngb-NPs) as a therapeutic tool against stroke. This nanoformulation induced an improvement of the cerebral infarct prognosis. However, this innovative therapy is still in development, and a more in-depth study focusing on its long-lasting neuroprotectant and neuroregenerative capabilities is needed. In short, this review aims to update the state-of-the-art of stroke therapies based on Ngb, paying special attention to the use of nanotechnological drug-delivering tools. [ABSTRACT FROM AUTHOR]

Most neurodegenerative diseases such as Alzheimer's disease are proteinopathies linked to the toxicity of amyloid oligomers. Treatments to delay or cure these diseases are lacking. Using budding yeast, we report that the natural lipid tripentadecanoin induces expression of the nitric oxide oxidoreductase Yhb1 to prevent the formation of protein aggregates during aging and extends replicative lifespan. In mammals, tripentadecanoin induces expression of the Yhb1 orthologue, neuroglobin, to protect neurons against amyloid toxicity. Tripentadecanoin also rescues photoreceptors in a mouse model of retinal degeneration and retinal ganglion cells in a Rhesus monkey model of optic atrophy. Together, we propose that tripentadecanoin affects p‐bodies to induce neuroglobin expression and offers a potential treatment for proteinopathies and retinal neurodegeneration. [ABSTRACT FROM AUTHOR]

Sevoflurane, a commonly used inhaled anesthetic, causes endogenous apoptosis in fetal rats. Microglia polarization is associated with inflammation, and the IL-10/STAT3/SOCS3 pathway is involved in this process. Neuroglobin (Ngb) is a neuroprotective protein which exhibits an anti-inflammatory effect. The purpose of this study was to investigate whether neurotoxicity induced by sevoflurane exposure in prenatal rats correlates with neuroinflammation and microglia polarization and whether Ngb can moderate this response. We found that exposure to sevoflurane on the 20th day of gestation (G20) induced discernable inflammation in postnatal day 0 (P0) rats, promoted M1 polarization of microglia, and inhibited M2 polarization. Hemin-mediated Ngb elevation inhibited sevoflurane-induced neuroinflammation. Additionally, elevated Ngb inhibited M1 polarization and promoted M2 polarization of microglia. We also found that elevated Ngb could alleviate the effect of sevoflurane on the expression of Interleukin-10 (IL-10), phosphorylated-signal transduction and activators of transcription 3 (P-STAT3), and suppressor of cytokine signaling 3 (SOCS3). Furthermore, we found that elevated Ngb ameliorated the effects of sevoflurane on long-term exploratory behavior and learning and memory in the offspring. Our results show that Ngb alleviates the neurotoxicity of sevoflurane to fetal rats by inhibiting neuroinflammation and affecting microglial polarization, a process which may involve the IL-10/STAT3/SOCS3 pathway. • Exposure to sevoflurane in prenatal rats can cause neuroinflammation in offspring rats. • Hemin-mediated Ngb elevation can inhibit neuroinflammation caused by sevoflurane. • Elevated Ngb affects microglia polarization phenotype. • Elevated Ngb can alleviate the long-term neurotoxicity induced by sevoflurane. [ABSTRACT FROM AUTHOR]

Neuroglobin is a metalloprotein expressed predominantly in a nervous system and involved in the functioning of cells in normal and pathological conditions. Despite numerous studies, information on its regional distribution is ambiguous. The purpose of the study is to identify patterns and features of the regional distribution of neuroglobin in neurons of all structures of the rat brain with a quantitative assessment of its content at the cellular level. Material and methods. Five outbred male rats kept under standard conditions were used in the study. Frontal serial sections were made after decapitation, fixation, dehydration and paraffin-embedding. One section of the series was stained according to the Nissl method for identification of structures according to the stereotaxic atlas, and the second section was immunohistochemically stained for neuroglobin. 100 brain structures were studied cytophotometrically, neuroglobin immunoreactivity was expressed in units of optical density ×103 (conventional units). Results and discussion. Four levels of neuroglobin content were identified – low, moderate, high and very high. Structures without neuroglobin were not found. Among the studied brain structures the amount of neuroglobin contents varies from 140–160 to 459–479 relative units. The low and moderate neuroglobin amount was revealed in supreme number of structures. Among the parts of the brain there is an increase in the content of neuroglobin in the direction telencephalon – diencephalon – midbrain – pons and medulla oblongata. The widest range of values is found in the telencephalon, pons and medulla oblongata. In the cerebellum the largest values are demonstrated by the interposed nucleus and the Purkinje cells of the pyramid. Conclusion. Neuroglobin has been found in neurons in all parts of the rat brain and spinal cord. The amount of neuroglobin depends on phylogenetic age: in the brain its content increases in the anteroposterior direction, and structures of paleocortex contain more neuroglobin than the structures of the neocortex. In the cerebellum a greater amount of neuroglobin is found in structures of the paleocerebellum.

Neuroglobin is a representative of the globin group, an oxygen-binding protein which is predominantly expressed in the nervous system. Despite the large amount of conducted research, its role is still poorly understood. It’s assumed that neuroglobin is able to regulate cell activity in health and disease:in health it provides oxygen homeostasis of neurons, and in pathology it binds free radicals and nitrogen monoxide, blocks mitochondrial factors of apoptosis and suppresses oxidative stress. A clearer understanding of the role of neuroglobin in pathology can offer new approaches in the treatment of neurodegenerative diseases. Special attention in the literature is paid to the role of neuroglobin in cerebral ischemia, which among other cerebrovascular diseases is the cause of disability and mortality of the population all over the world. In the present article an overview of the literature data available to date on the study of the neuroprotective effect of neuroglobin in hypoxia/ischemia is provided: models used to study function in vitro and in vivo, the effects of its increased or decreased expression, neuroglobin-mediated neuroprotective action of hemin, estimated pathways of the induction of neuroglobin and views on the mechanisms of neuroprotection.

Background: Neuroglobin (Ngb) is the fourth member of globin family which was found to be highly expressed in central and peripheral nervous system of humans. Overexpression of neuroglobin is strongly related to a neuroprotective function. Aim of the Study: Primary aim: to detect the levels of neuroglobin in children with Epilepsy Compared to Normal healthy Controls. Secondary aim: To compare the level of neuroglobin in controlled and uncontrolled epilepsy. Subject and Method: The study is controlled cross sectional study which was conducted on 85 children divided into three groups: group (I) included 30 children with controlled epilepsy, group (II) included 30 children with uncontrolled epilepsy, and group (III) included 25 healthy children selected as controls of matched gender and age. Serum Ngb assay, complete blood count (CBC) were performed to all children. Results: The overall results show that serum neuroglobin levels were significantly increased in patients' groups than in controls. Neuroglobin levels showed no significant difference between controlled epilepsy group and uncontrolled epilepsy one. Conclusion: Higher serum neuroglobin levels among children with epilepsy whether controlled or uncontrolled, reflecting its role as a neuroprotective marker. This study may help to discovernewer antiepileptic therapy through acting on neuroglobin levels. [ABSTRACT FROM AUTHOR]

Human neuroglobin (Ngb) contains a heme group and three Cys residues (Cys46, Cys55, and Cys120) in the polypeptide chain. By introducing an additional Cys at position 15, the X‐ray structure of A15C Ngb mutant was solved at a high resolution of 1.35 Å, which reveals the formation of both the native (C46C55) and the engineered (C15C120) disulfide bonds, likely playing a functional and structural role, respectively, according to the geometry analysis. Unexpectedly, 1,4‐dioxane from the crystallization reagents was bound not only to the protein surface, but also to the heme distal pocket, providing insights into protein‐ligand interactions for the globin and guiding the design of functional heme enzymes. [ABSTRACT FROM AUTHOR]

Nitric Oxide (NO) is a free radical with numerous critical signaling roles in vertebrate physiology. Similar to mammals, in the teleost system the generation of sufficient amounts of NO is critical for the physiological function of the cardiovascular system. At the same time, NO amounts are strictly controlled and kept within basal levels to protect cells from NO toxicity. Changes in oxygen tension highly influence NO bioavailability and can modulate the mechanisms involved in maintaining the NO balance. While NO production and signaling appears to have general similarities with mammalian systems, the wide range of environmental adaptations made by fish, particularly with regards to differing oxygen availabilities in aquatic habitats, creates a foundation for a variety of in vivo models characterized by different implications of NO production and signaling. In this review, we present the biology of NO in the teleost cardiovascular system and summarize the mechanisms of NO production and signaling with a special emphasis on the role of globin proteins in NO metabolism. [ABSTRACT FROM AUTHOR]

Objective: Stroke is closely related to dementia, but there are few prospective studies on cognitive decline after stroke in patients with cerebral hemorrhage. Neuroglobin is an oxygen-binding protein mainly expressed in brain neurons. The aim of our current study was to determine whether neuroglobin could serve as a biomarker for cognitive prognosis in patients with intracerebral hemorrhage (ICH). Methods: Three hundred and sixteen patients with ICH were consecutively enrolled in a prospective study. Baseline data such as age and gender of ICH patients on admission were recorded. Serum neuroglobin concentrations were determined by enzyme-linked immunosorbent assay (ELISA). All ICH patients 3 months after onset were divided into post-stroke cognitive impairment group (PSCI) and non-PSCI group according to MoCA assessment results. Results: The PSCI and Non-PSCI groups had serum neuroglobin concentrations of (4.7 ± 0.9) and (7.5 ± 1.1) ng/ml, respectively, with a statistically significant difference between the two groups (p < 0.05). Age, gender, LDL, FBG, SBP, DBP, NHISS, and Hematoma volume were found to be adversely connected with MoCA (p < 0.05), while education, HDL, and serum neuroglobin were found to be positively correlated with MoCA (p < 0.05). After controlling for baseline data, regression analysis revealed that serum neuroglobin was remained an efficient biomarker for predicting cognitive performance in individuals with ICH (p < 0.05). The diagnostic accuracy of blood neuroglobin concentration for PSCI in ICH patients was 72.6%, the sensitivity was 67.4%, and the specificity was 75.5%, according to receiver operating characteristic (ROC) curve analysis. Conclusions: Serum neuroglobin may serve as a potential biomarker to predict cognitive decline after ICH. [ABSTRACT FROM AUTHOR]

Neuroglobin (Ngb) is a 17 kDa monomeric hexa-coordinated heme protein belonging to the globin family. Ngb is mainly expressed in neurons of the central and peripheral nervous system, although moderate levels of Ngb have been detected in non-nervous tissues. In the past decade, Ngb has been studied for its neuroprotective role in a large number of neurological disorders such as Alzheimer’s disease, Huntington’s disease, brain ischemia and hypoxia. This review discusses and summarizes the natural compounds and the small synthetic molecules capable of modulating Ngb expression that exhibits a protective role against various neurodegenerative diseases.

In this article, we present data on the proteome of human neuroblastoma cells stably overexpressing Neuroglobin (NGB). The neuroprotective role of NGB is clearly established, nevertheless the related mechanistic processes, which are dependent on NGB overexpression, are not known. To address this question, we performed shotgun label-free quantification (LFQ) proteomics using an SH-SY5Y cell model of neuroblastoma that overexpresses an NGB-FLAG construct, and wild type cells transfected with an empty vector as control (CTRL). The proteomes from six biological samples per condition were digested using the S-Trap sample preparation followed by LC-MS/MS analysis with a LTQ-Orbitrap XL mass spectrometer. The quantitative analysis was performed using the LFQ algorithm of MaxQuant, leading to 1654 correctly quantified proteins over 2580 identified proteins. Finally, the statistic comparison of the two analyzed groups within Perseus platform identified 178 differential proteins (107 up- and 71 down-regulated). In addition, multivariate statistical analysis was carried out using MetaboAnalyst 5.0 software. MS proteomics data are available via ProteomeXchange with the dataset identifier PXD029012.

ObjectiveStroke is closely related to dementia, but there are few prospective studies on cognitive decline after stroke in patients with cerebral hemorrhage. Neuroglobin is an oxygen-binding protein mainly expressed in brain neurons. The aim of our current study was to determine whether neuroglobin could serve as a biomarker for cognitive prognosis in patients with intracerebral hemorrhage (ICH).MethodsThree hundred and sixteen patients with ICH were consecutively enrolled in a prospective study. Baseline data such as age and gender of ICH patients on admission were recorded. Serum neuroglobin concentrations were determined by enzyme-linked immunosorbent assay (ELISA). All ICH patients 3 months after onset were divided into post-stroke cognitive impairment group (PSCI) and non-PSCI group according to MoCA assessment results.ResultsThe PSCI and Non-PSCI groups had serum neuroglobin concentrations of (4.7 ± 0.9) and (7.5 ± 1.1) ng/ml, respectively, with a statistically significant difference between the two groups (p < 0.05). Age, gender, LDL, FBG, SBP, DBP, NHISS, and Hematoma volume were found to be adversely connected with MoCA (p < 0.05), while education, HDL, and serum neuroglobin were found to be positively correlated with MoCA (p < 0.05). After controlling for baseline data, regression analysis revealed that serum neuroglobin was remained an efficient biomarker for predicting cognitive performance in individuals with ICH (p < 0.05). The diagnostic accuracy of blood neuroglobin concentration for PSCI in ICH patients was 72.6%, the sensitivity was 67.4%, and the specificity was 75.5%, according to receiver operating characteristic (ROC) curve analysis.ConclusionsSerum neuroglobin may serve as a potential biomarker to predict cognitive decline after ICH.

Neuroglobin (Ngb) is an oxygen-binding protein with a demonstrated role in endogenous neuroprotective mechanisms. It has been shown to function as a scavenger for reactive oxidizing species thereby assisting in cellular defense against oxidative stress. In the present study, we characterized the presence of Ngb in the human cochlea. Immunohistochemical staining was performed on formalin fixed celloidin human cochlea sections obtained from human temporal bones, using affinity purified polyclonal antibodies against Ngb. Thirty-six temporal bones were analyzed, 15 with normal otologic histories and 21 diagnosed with different inner ear pathologies. Ngb immunoreactivity (Ngb-IR) was consistently expressed in the neurons of spiral ganglia (SG) and supporting cells of the organ of Corti. There was a significant decrease of Ngb-IR in SGNs from specimens with inner ear pathologies when compared to normal specimens. In contrast, Ngb-IR in the organ of Corti did not show significant changes between pathological and normal specimens. The differential pattern of Ngb expression in these cochlear structures suggests that Ngb may participate in defense mechanisms in inner ear pathologies where oxidative stress is involved.

Microglia are the immune competent cell of the central nervous system (CNS), promoting brain homeostasis and regulating inflammatory response against infection and injury. Chronic or exacerbated neuroinflammation is a cause of damage in several brain pathologies. Endogenous carbon monoxide (CO), produced from the degradation of heme, is described as anti-apoptotic and anti-inflammatory in several contexts, including in the CNS. Neuroglobin (Ngb) is a haemoglobin-homologous protein, which upregulation triggers antioxidant defence and prevents neuronal apoptosis. Thus, we hypothesised a crosstalk between CO and Ngb, in particular, that the anti-neuroinflammatory role of CO in microglia depends on Ngb. A novel CO-releasing molecule (ALF826) based on molybdenum was used for delivering CO in microglial culture. BV-2 mouse microglial cell line was challenged with lipopolysaccharide (LPS) for triggering inflammation, and after 6 h ALF826 was added. CO exposure limited inflammation by decreasing inducible nitric oxide synthase (iNOS) expression and the production of nitric oxide (NO) and tumour necrosis factor-α (TNF-α), and by increasing interleukine-10 (IL-10) release. CO-induced Ngb upregulation correlated in time with CO's anti-inflammatory effect. Moreover, knocking down Ngb reversed the anti-inflammatory effect of CO, suggesting that dependents on Ngb expression. CO-induced Ngb upregulation was independent on ROS signalling, but partially dependent on the transcriptional factor SP1. Finally, microglial cell metabolism is also involved in the inflammatory response. In fact, LPS treatment decreased oxygen consumption in microglia, indicating a switch to glycolysis, which is associated with a proinflammatory. While CO treatment increased oxygen consumption, reverting LPS effect and indicating a metabolic shift into a more oxidative metabolism. Moreover, in the absence of Ngb, this phenotype was no longer observed, indicating Ngb is needed for CO's modulation of microglial metabolism. Finally, the metabolic shift induced by CO did not depend on alteration of mitochondrial population. In conclusion, neuroglobin emerges for the first time as a key player for CO signalling against exacerbated inflammation in microglia. [ABSTRACT FROM AUTHOR]

Prunus cerasoides (PC) has been reported to have antimicrobial and anti-inflammatory properties, but its potential as a neuroprotective agent in a mouse model of cerebral ischemia has not been explored. Considering neuroglobin (Ngb), an endogenous neuroprotective factor, as a novel approach to neuroprotection, in this study, Ngb promoter activity, Ngb expression changes, and antioxidant protection by PC extract (PCE) and PC component compounds (PCCs) were analyzed in oxygen– glucose deprivation (OGD)-treated neurons. In vivo analysis involved transient middle cerebral artery occlusion (tMCAO) in mice with pre- and post-treatment exposure to PCE. Following ischemic stroke induction, neurological behavior scores were obtained, and cellular function-related signals were evaluated in the ischemic infarct areas. In addition to PCE, certain component compounds from PCE also significantly increased Ngb levels and attenuated the intracellular ROS production and cytotoxicity seen with OGD in primary neurons. Administration of PCE reduced the infarct volume and improved neurological deficit scores in ischemic stroke mice compared with the vehicle treatment. Increased Ngb levels in infarct penumbra with PCE treatment were also accompanied by decreased markers of apoptosis (activated p38 and cleaved caspase-3). Our findings point to the benefits of Ngb-mediated neuroprotection via PCE and its antioxidant activity in an ischemic stroke model. [ABSTRACT FROM AUTHOR]

Neuroglobin, a member of the globin superfamily, is abundant in the brain, retina, and cerebellum of mammals and localizes to mitochondria. The protein exhibits neuroprotective capacities by participating in electron transfer, oxygen supply, and protecting against oxidative stress. Our objective was to determine whether neuroglobin overexpression can be used to treat neurological disorders. We chose Harlequin mice, which harbor a retroviral insertion in the first intron of the apoptosis-inducing factor gene resulting in the depletion of the corresponding protein essential for mitochondrial biogenesis. Consequently, Harlequin mice display degeneration of the cerebellum and suffer from progressive blindness and ataxia. Cerebellar ataxia begins in Harlequin mice at the age of 4 months and is characterized by neuronal cell disappearance, bioenergetics failure, and motor and cognitive impairments, which aggravated with aging. Mice aged 2 months received adeno-associated viral vectors harboring the coding sequence of neuroglobin or apoptosis-inducing factor in both cerebellar hemispheres. Six months later, Harlequin mice exhibited substantial improvements in motor and cognitive skills; probably linked to the preservation of respiratory chain function, Purkinje cell numbers and connectivity. Thus, without sharing functional properties with apoptosis-inducing factor, neuroglobin was efficient in reducing ataxia in Harlequin mice., Competing Interests: Declaration of interests An application for a patent has been filed on neuroglobin in Europe (EP22306002.1) and the United States (PCTEP2023/068369) in 2022 and 2023 (M.C.-D. and P.G)., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Peptide nucleic acid (PNA) is a DNA mimic that shows good stability against nucleases and proteases, forming strongly recognized complementary strands of DNA and RNA. However, due to its feeble ability to cross the cellular membrane, PNA activity and its targeting gene action is limited. Halloysite nanotubes (HNTs) are a natural and low-cost aluminosilicate clay. Because of their peculiar ability to cross cellular membrane, HNTs represent a valuable candidate for delivering genetic materials into cells. Herein, two differently charged 12-mer PNAs capable of recognizing as molecular target a 12-mer DNA molecule mimicking a purine-rich tract of neuroglobin were synthetized and loaded onto HNTs by electrostatic attraction interactions. After characterization, the kinetic release was also assessed in media mimicking physiological conditions. Resonance light scattering measurements assessed their ability to bind complementary single-stranded DNA. Furthermore, their intracellular delivery was assessed by confocal laser scanning microscopy on living MCF-7 cells incubated with fluorescence isothiocyanate (FITC)-PNA and HNTs labeled with a probe. The nanomaterials were found to cross cellular membrane and cell nuclei efficiently. Finally, it is worth mentioning that the HNTs/PNA can reduce the level of neuroglobin gene expression, as shown by reverse transcription-quantitative polymerase chain reaction and western blotting analysis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)