Poly-Arginine Peptide-18 (R18) Reduces Brain Injury and Improves Functional Outcomes in a Nonhuman Primate Stroke Model.
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ABSTRACT:
SUBMITTER: Meloni BP
PROVIDER: S-EPMC7283416 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Poly-arginine peptide-18 (R18) has recently emerged as a highly effective neuroprotective agent in experimental stroke models, and is particularly efficacious in protecting cortical neurons against glutamic acid excitotoxicity. While we have previously demonstrated that R18 can reduce excitotoxicity-induced neuronal calcium influx, other molecular events associated with R18 neuroprotection are yet to investigated. Therefore, in this study we were particularly interested in protein expression changes in R18 treated neurons subjected to excitotoxicity. Proteomic analysis was used to compare protein expression patterns in primary cortical neuronal cultures subjected to: (i) R18-treatment alone (R18); (ii) glutamic acid excitotoxic injury (Glut); (iii) R18-treatment and glutamic acid injury (R18 + Glut); (iv) no treatment (Cont). Whole cell lysates were harvested 24 h post-injury and subjected to quantitative proteomic analysis (iTRAQ), coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) and subsequent bioinformatic analysis of differentially expressed proteins (DEPs). Relative to control cultures, R18, Glut, and R18 + Glut treatment resulted in the detection of 5, 95 and 14 DEPs respectively. Compared to Glut alone, R18 + Glut revealed 98 DEPs, including 73 proteins whose expression was also altered by treatment with Glut and/or R18 alone, as well as 25 other uniquely regulated proteins. R18 treatment reversed the up- or down-regulation of all 73 Glut-associated DEPs, which included proteins involved in mitochondrial integrity, ATP generation, mRNA processing and protein translation. Analysis of protein-protein interactions of the 73 DEPs showed they were primarily associated with mitochondrial respiration, proteasome activity and protein synthesis, transmembrane trafficking, axonal growth and neuronal differentiation, and carbohydrate metabolism. Identified protein pathways associated with proteostasis and energy metabolism, and with pathways involved in neurodegeneration. Collectively, the findings indicate that R18 neuroprotection following excitotoxicity is associated with preservation of neuronal protein profiles, and differential protein expression that assists in maintaining mitochondrial function and energy production, protein homeostasis, and membrane trafficking.
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Project description:We examined the neuroprotective efficacy of the poly-arginine peptide R18 and its D-enantiomer R18D in a perinatal hypoxic-ischaemic (HI) model in P7 Sprague-Dawley rats. R18 and R18D peptides were administered intraperitoneally at doses of 30, 100, 300 or 1000 nmol/kg immediately after HI (8% O2/92%N2 for 2.5 h). The previously characterised neuroprotective JNKI-1-TATD peptide at a dose of 1000 nmol/kg was used as a control. Infarct volume and behavioural outcomes were measured 48 h after HI. For the R18 and R18D doses examined, total infarct volume was reduced by 25.93% to 43.80% (P?=?0.038 to <?0.001). By comparison, the JNKI-1-TATD reduced lesion volume by 25.27% (P?=?0.073). Moreover, R18 and R18D treatment resulted in significant improvements in behavioural outcomes, while with JNKI-1-TATD there was a trend towards improvement. As an insight into the likely mechanism underlying the effects of R18, R18D and JNKI-1-TATD, the peptides were added to cortical neuronal cultures exposed to glutamic acid excitotoxicity, resulting in up to 89, 100 and 71% neuroprotection, respectively, and a dose dependent inhibition of neuronal calcium influx. The study further confirms the neuroprotective properties of poly-arginine peptides, and suggests a potential therapeutic role for R18 and R18D in the treatment of HIE.
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Project description:Despite the discovery of several promising neuroprotective therapies in rodent models of stroke, no therapy other than the fibrinolytics has been found to be effective in human clinical trials. To address potential discrepancies between rodent and human studies, the Stroke Therapy Academic Industry Roundtable (STAIR) committee suggested that nonhuman primates (NHPs) be used for preclinical, translational stroke studies. Due to the paucity of stroke studies in NHPs, few experimental models have been described. Critical factors in designing NHP stroke models include the choice of species, the method of inducing the stroke and the choice of outcome measures. In this review, we describe established NHP models of stroke and discuss factors that may influence model development with a focus on models that may be useful in preclinical studies for neuroprotective drug screening prior to clinical trials.
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Project description:The poly-arginine peptides R18D and R18 represent novel potential neuroprotective treatments for acute ischaemic stroke. Here we examined whether R18D and R18 had any significant effects on the thrombolytic activity of alteplase (tPA) and tenecteplase (TNK) on clots formed from whole blood in an in vitro thrombolysis plate assay. R18D and R18 were examined at concentrations of 0.25, 0.5, 1, 2, 4, 8 and 16 µM during the 1-h thrombolytic assay. We also included the well-characterised neuroprotective NA-1 peptide as a control. R18D, R18 and NA-1 all reduced tPA or TNK percentage clot lysis by 0-9.35%, 0-3.44% and 0-4.8%, respectively. R18D, R18 and NA-1 had a modest and variable effect on the lag time, increasing the time to the commencement of thrombolysis by 0-9.9 min, 0-5.53 min and 0-7.16 min, respectively. Lastly, R18 and NA-1 appeared to increase the maximal activity of the thrombolysis reaction. In addition, the in vitro anti-excitotoxic neuroprotective efficacy of R18D and R18 was not affected by pre-incubation for 1-2 h or overnight with tPA or TNK, whereas only R18D retained high anti-excitotoxic neuroprotective efficacy when pre-incubated in a synthetic trypsin (TrypLE Express). The present in vitro findings suggest that neither R18D or R18 when co-administered with the thrombolytic inducing agents tPA or TNK are likely to have a significant impact when used clinically during clot thrombolysis and confirm the superior proteolytic stability of the R18D peptide.
| S-EPMC9259545 | biostudies-literature
Project description:Using cortical neuronal cultures and glutamic acid excitotoxicity and oxygen-glucose deprivation (OGD) stroke models, we demonstrated that poly-arginine and arginine-rich cell-penetrating peptides (CPPs), are highly neuroprotective, with efficacy increasing with increasing arginine content, have the capacity to reduce glutamic acid-induced neuronal calcium influx and require heparan sulfate preotoglycan-mediated endocytosis to induce a neuroprotective effect. Furthermore, neuroprotection could be induced with immediate peptide treatment or treatment up to 2 to 4 hours before glutamic acid excitotoxicity or OGD, and with poly-arginine-9 (R9) when administered intravenously after stroke onset in a rat model. In contrast, the JNKI-1 peptide when fused to the (non-arginine) kFGF CPP, which does not rely on endocytosis for uptake, was not neuroprotective in the glutamic acid model; the kFGF peptide was also ineffective. Similarly, positively charged poly-lysine-10 (K10) and R9 fused to the negatively charged poly-glutamic acid-9 (E9) peptide (R9/E9) displayed minimal neuroprotection after excitotoxicity. These results indicate that peptide positive charge and arginine residues are critical for neuroprotection, and have led us to hypothesize that peptide-induced endocytic internalization of ion channels is a potential mechanism of action. The findings also question the mode of action of different neuroprotective peptides fused to arginine-rich CPPs.
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Project description:Parthenogenesis is the biological phenomenon by which embryonic development is initiated without male contribution. Whereas parthenogenesis is a common mode of reproduction in lower organisms, the mammalian parthenote fails to produce a successful pregnancy. We herein describe in vitro parthenogenetic development of monkey (Macaca fascicularis) eggs to the blastocyst stage, and their use to create a pluripotent line of stem cells. These monkey stem cells (Cyno-1 cells) are positive for telomerase activity and are immunoreactive for alkaline phosphatase, octamer-binding transcription factor 4 (Oct-4), stage-specific embryonic antigen 4 (SSEA-4), tumor rejection antigen 1-60 (TRA 1-60), and tumor rejection antigen 1-81 (TRA 1-81) (traditional markers of human embryonic stem cells). They have a normal chromosome karyotype (40 + 2) and can be maintained in vitro in an undifferentiated state for extended periods of time. Cyno-1 cells can be differentiated in vitro into dopaminergic and serotonergic neurons, contractile cardiomyocyte-like cells, smooth muscle, ciliated epithelia, and adipocytes. When Cyno-1 cells were injected into severe combined immunodeficient mice, teratomas with derivatives from all three embryonic germ layers were obtained. When grown on fibronectin/laminin-coated plates and in neural progenitor medium, Cyno-1 cells assume a neural precursor phenotype (immunoreactive for nestin). However, these cells remain proliferative and express no functional ion channels. When transferred to differentiation conditions, the nestin-positive precursors assume neuronal and epithelial morphologies. Over time, these cells acquire electrophysiological characteristics of functional neurons (appearance of tetrodotoxin-sensitive, voltage-dependent sodium channels). These results suggest that stem cells derived from the parthenogenetically activated nonhuman primate egg provide a potential source for autologous cell therapy in the female and bypass the need for creating a competent embryo.
| S-EPMC304106 | biostudies-other
Project description:Remdesivir (RDV) is a nucleotide analog prodrug with demonstrated clinical benefit in patients with coronavirus disease 2019 (COVID-19). In October 2020, the US FDA approved intravenous (IV) RDV as the first treatment for hospitalized COVID-19 patients. Furthermore, RDV has been approved or authorized for emergency use in more than 50 countries. To make RDV more convenient for non-hospitalized patients earlier in disease, alternative routes of administration are being evaluated. Here, we investigated the pharmacokinetics and efficacy of RDV administered by head dome inhalation in African green monkeys (AGM). Relative to an IV administration of RDV at 10 mg/kg, an approximately 20-fold lower dose administered by inhalation produced comparable concentrations of the pharmacologically active triphosphate in lower respiratory tract tissues. Distribution of the active triphosphate into the upper respiratory tract was also observed following inhaled RDV exposure. Inhalation RDV dosing resulted in lower systemic exposures to RDV and its metabolites as compared with IV RDV dosing. An efficacy study with repeated dosing of inhaled RDV in an AGM model of SARS-CoV-2 infection demonstrated reductions in viral replication in bronchoalveolar lavage fluid and respiratory tract tissues compared with placebo. Efficacy was observed with inhaled RDV administered once daily at a pulmonary deposited dose of 0.35 mg/kg beginning approximately 8 hours post-infection. Moreover, the efficacy of inhaled RDV was similar to that of IV RDV administered once at 10 mg/kg followed by 5 mg/kg daily in the same study. Together, these findings support further clinical development of inhalation RDV.
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Project description:[(18)F]Nifene is an agonist PET radioligand developed to image α4β2* nicotinic acetylcholine receptors (nAChRs). This work aims to quantify the receptor density (Bmax) of α4β2* nAChRs and the in vivo (apparent) dissociation constant (KDapp) of [(18)F]nifene. Multiple-injection [(18)F]nifene experiments with varying cold nifene masses were conducted on four rhesus monkeys with a microPET P4 scanner. Compartment modeling techniques were used to estimate regional Bmax values and a global value of KDapp. The fast kinetic properties of [(18)F]nifene also permitted alternative estimates of Bmax and KDapp at transient equilibrium with the same experimental data using Scatchard-like methodologies. Averaged across subjects, the compartment modeling analysis yielded Bmax values of 4.8±1.4, 4.3±1.0, 1.2±0.4, and 1.2±0.3 pmol/mL in the regions of antereoventral thalamus, lateral geniculate, frontal cortex, and subiculum, respectively. The KDapp of nifene was 2.4±0.3 pmol/mL. The Scatchard analysis based on graphical evaluation of the data after transient equilibrium yielded Bmax estimations comparable to the modeling results with a positive bias of 28%. These findings show the utility of [(18)F]nifene for measuring α4β2* nAChR Bmax in vivo in the rhesus monkey with a single PET experiment.
| S-EPMC3824181 | biostudies-other
Project description:Group A Streptococcus (GAS) infections account for an estimated 500,000 deaths every year. This bacterial pathogen is responsible for a variety of mild and life-threatening infections and the triggering of chronic autoimmune sequelae. Pharyngitis caused by group A Streptococcus (GAS), but not asymptomatic GAS carriage, is a prerequisite for acute rheumatic fever (ARF). Repeated bouts of ARF may trigger rheumatic heart disease (RHD), a major cause of heart failure and stroke accounting for 275,000 deaths annually. A vaccine that prevents pharyngitis would markedly reduce morbidity and mortality from ARF and RHD. Nonhuman primates (NHPs) have been utilized to model GAS diseases, and experimentally infected rhesus macaques develop pharyngitis. Here we use an NHP model of GAS pharyngitis to evaluate the efficacy of an experimental vaccine, Combo5 (arginine deiminase [ADI], C5a peptidase [SCPA], streptolysin O [SLO], interleukin-8 [IL-8] protease [SpyCEP], and trigger factor [TF]), specifically designed to exclude GAS components potentially linked to autoimmune complications. Antibody responses against all Combo5 antigens were detected in NHP serum, and immunized NHPs showed a reduction in pharyngitis and tonsillitis compared to controls. Our work establishes the NHP model as a gold standard for the assessment of GAS vaccines.IMPORTANCE GAS-related diseases disproportionally affect disadvantaged populations (e.g., indigenous populations), and development of a vaccine has been neglected. A recent strong advocacy campaign driven by the World Health Organization and the International Vaccine Institute has highlighted the urgent need for a GAS vaccine. One significant obstacle in GAS vaccine development is the lack of a widely used animal model to assess vaccine efficacy. Researchers in the field use a wide range of murine models of infection and in vitro assays, sometimes yielding conflicting results. Here we present the nonhuman primate pharyngeal infection model as a tool to assess vaccine-induced protection against colonization and clinical symptoms of pharyngitis and tonsillitis. We have tested the efficacy of an experimental vaccine candidate with promising results. We believe that the utilization of this valuable tool by the GAS vaccine research community could significantly accelerate the realization of a safe and effective GAS vaccine for humans.
| S-EPMC6495378 | biostudies-literature
Project description:Background[11C]Cimbi-36 is a serotonin 2A receptor agonist positron emission tomography radioligand that has recently been examined in humans. The binding of agonist radioligand is expected to be more sensitive to endogenous neurotransmitter concentrations than antagonist radioligands. In the current study, we compared the effect of serotonin releaser fenfluramine on the binding of [11C]Cimbi-36, [11C]MDL 100907 (a serotonin 2A receptor antagonist radioligand), and [11C]AZ10419369 (a serotonin 1B receptor partial agonist radioligand with established serotonin sensitivity) in the monkey brain.MethodsEighteen positron emission tomography measurements, 6 for each radioligand, were performed in 3 rhesus monkeys before or after administration of 5.0 mg/kg fenfluramine. Binding potential values were determined with the simplified reference tissue model using cerebellum as the reference region.ResultsFenfluramine significantly decreased [11C]Cimbi-36 (26-62%) and [11C]AZ10419369 (35-58%) binding potential values in most regions (P < 0.05). Fenfluramine-induced decreases in [11C]MDL 100907 binding potential were 8% to 30% and statistically significant in 3 regions. Decreases in [11C]Cimbi-36 binding potential were larger than for [11C]AZ10419369 in neocortical and limbic regions (~35%) but smaller in striatum and thalamus (~40%). Decreases in [11C]Cimbi-36 binding potential were 0.9 to 2.8 times larger than for [11C]MDL 100907, and the fraction of serotonin 2A receptor in the high-affinity state was estimated as 54% in the neocortex.ConclusionsThe serotonin sensitivity of serotonin 2A receptor agonist radioligand [11C]Cimbi-36 was higher than for antagonist radioligand [11C]MDL 100907. The serotonin sensitivity of [11C]Cimbi-36 was similar to [11C]AZ10419369, which is one of the most sensitive radioligands. [11C]Cimbi-36 is a promising radioligand to examine serotonin release in the primate brain.
| S-EPMC5581490 | biostudies-literature