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Poly(ADP-ribose) polymerase (PARP) inhibitors have been proposed as pharmacological agents in the treatment of various diseases. Recently, factors and mechanisms responsible for regulating PARP catalytic activity have been identified, some of which can significantly influence the effectiveness of inhibitors of this enzyme. In this regard, it is important to develop new models and methods that would reflect the cellular context in which PARP functions. We proposed to use digitonin-permeabilized adherent cells to study poly(ADP-ribosyl)ation reaction (PARylation) in order to maintain the nuclear localization of PARP and to control the concentrations of its substrate (NAD + ) and tested compounds in the cell. A specific feature of the approach is that before permeabilization, cellular PARP is converted to the DNA-bound state under conditions preventing premature initiation of the PARylation reaction. Experiments were carried out in rat H9c2 cardiomyoblasts. The activity of PARP in permeabilized cells was analyzed by measuring the immunofluorescence of the reaction product poly(ADP-ribose). The method was verified in the studies of PARP inhibition by the classic inhibitor 3-aminobenzamide and a number of new 7-methylguanine derivatives. One of them, 7,8-dimethylguanine, was found to be a stronger inhibitor compared to 7-methylguanine, due to a formation of additional hydrophobic contact with the protein. The proposed approach opens up new prospects for studying the mechanisms of PARP activity regulation in cells and can be used in high-throughput screening of PARP inhibitors.

Synthetic peptides have a wide range of clinical effects. Of particular interest are peptides based on adrenocorticotropic hormone (ACTH) both as already used and as potential drugs for preventing consequences of cerebral ischemia. However, it is necessary to study influence of the peptide on the brain cells under normal physiological conditions, including understanding the risks of their use. Here, we used high-throughput RNA sequencing (RNA-Seq) to identify differentially expressed genes (DEGs) in the brain frontal cortex of rat receiving intraperitoneal administration of ACTH-like peptides ACTH(4-7)PGP (Semax) and ACTH(6-9)PGP, or saline. We identified 258 and 228 DEGs, respectively, with the fold change > 1.5 and Padj  < 0.05 at 22.5 h after the first administration of Semax and ACTH(6-9)PGP. Metabolic pathways, characterizing both common and specific effects of the peptides on the transcriptome were identified. Both peptides predominantly caused decrease in expression of the genes associated with the immune system. At the same time, when comparing the effects of ACTH(6-9)PGP relative to Semax, DEGs were identified that characterized the main differences in the effects of the peptides. These genes were mostly downregulated and associated with neurosignaling systems and regulation of ion channels, thus characterizing differences in the effects of the peptides. Our data show how differences in the structure of ACTH derivatives are associated with the changes in the brain cell transcriptome following exposure to these related peptides. Furthermore, our results demonstrate that when studying influence of regulatory peptides on transcriptome under pathological conditions, it is necessary to take into account their actions under normal physiological conditions.

Current antidepressant therapy shows substantial limitations, and there is an urgent need for the development of new treatment strategies for depression. Stressful events and hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis play an important role in the pathogenesis of depression. HPA axis activity is self-regulated by negative feedback at several levels including adrenocorticotropic hormone (ACTH)-mediated feedback. Here, we investigated whether noncorticotropic synthetic analogs of the ACTH(4-10) fragment, ACTH(4-7)-Pro-Gly-Pro (Semax) and Ac-Nle4-cyclo[Asp5-His6-D-Phe7-Arg8-Trp9-Lys10]ACTH(4-10)-NH2 (Melanotan II (MTII), a potent agonist of melanocortin receptors), have potential antidepressant activity in a chronic unpredictable stress (CUS) rat model of depression. Stressed and control male adult Sprague-Dawley rats received daily intraperitoneal injections of saline or a low dose (60 nmol/kg of body weight (BW)) of Semax or MTII. Rats were monitored for BW and hedonic status, as measured in the sucrose preference test. We found that chronic treatment with Semax and MTII reversed or substantially attenuated CUS-induced anhedonia, BW gain suppression, adrenal hypertrophy and a decrease in the hippocampal levels of BDNF. In the forced swim test, no effects of the CUS procedure or peptides on the duration of rat immobility were detected. Our findings show that in the CUS paradigm, systemically administered ACTH(4-10) analogs Semax and MTII exert antidepressant-like effects on anhedonia and hippocampal BDNF levels, and attenuate markers of chronic stress load, at least in male rats. The results support the argument that ACTH(4-10) analogs and other noncorticotropic melanocortins may have promising therapeutic potential for the treatment and prevention of depression and other stress-related pathologies., 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 manuscript., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Stroke remains the second leading cause of death worldwide. The development of new therapeutic agents focused on restoring vascular function and neuroprotection of viable tissues is required. In this study the neuroprotective activity of melanocortin-like ACTH(4-7)PGP and ACTH(6-9)PGP peptides was investigated in rat brain at 24 h after transient middle cerebral artery occlusion (tMCAO). The severity of ischemic damage, changes in the proliferative activity of neuroglial cells and vascularization of rat brain tissue were analyzed. The administration of peptides resulted in a significant increase in the volume density of neurons in the perifocal zone of infarction compared to rats subjected to ischemia and receiving saline. Immunohistochemical analysis of the proliferative activity of neuroglia cells using PCNA antibodies showed a significant increase in the number of proliferating cells in the penumbra and in the intact cerebral cortex of rats receiving peptide treatment. The effect of peptides on vascularization was examined using CD31 antibodies under tMCAO conditions, revealing a significant increase in the volume density of vessels and their sizes in the penumbra after administration of ACTH(4-7)PGP and ACTH(6-9)PGP. These findings confirm the neuroprotective effect of peptides due to the activation of neuroglia proliferation and the enhancement of collateral blood flow.

Ischemic stroke is an acute local decrease in cerebral blood flow due to a thrombus or embolus. Of particular importance is the study of the genetic systems that determine the mechanisms underlying the formation and maintenance of a therapeutic window (a time interval of up to 6 h after a stroke) when effective treatment can be provided. Here, we used a transient middle cerebral artery occlusion (tMCAO) model in rats to study two synthetic derivatives of adrenocorticotropic hormone (ACTH). The first was ACTH(4-7)PGP, which is known as Semax. It is actively used as a neuroprotective drug. The second was the ACTH(6-9)PGP peptide, which is elucidated as a prospective agent only. Using RNA-Seq analysis, we revealed hundreds of ischemia-related differentially expressed genes (DEGs), as well as 131 and 322 DEGs related to the first and second peptide at 4.5 h after tMCAO, respectively, in dorsolateral areas of the frontal cortex of rats. Furthermore, we showed that both Semax and ACTH(6-9)PGP can partially prevent changes in the immune- and neurosignaling-related gene expression profiles disturbed by the action of ischemia at 4.5 h after tMCAO. However, their different actions with regard to predominantly immune-related genes were also revealed. This study gives insight into how the transcriptome depends on the variation in the structure of the related peptides, and it is valuable from the standpoint of the development of measures for early post-stroke therapy.

The central effectors of the stress system are greatly interconnected and include, among others, a large group of peptides derived from proopiomelanocortin. In addition to natural corticotropins, a number of artificial molecules that contain some ACTH fragments in their structure are also referred to members of this family. Some of them possess a wide range of biological activity. The molecular mechanism underlying the biological activity of such peptides is partly based on allosteric modulation of various receptors. We analyzed the ability of some biologically active synthetic corticotropins (ACTH(4-7)PGP, ACTH(6-9)PGP, ACTH(7-10)PGP), and glyproline PGPL to affect the GABA-receptor system of rat brain. The effects of the peptides were studied in the isolated plasma membranes of brain cells, as well as after systemic peptide administration in the rat model of acute restraint stress. The delayed effect of stress or preadministration of each of the studied peptides on [ 3 H]GABA binding was different for its high- and low-affinity-specific sites. The studied peptides individually affected the binding of [ 3 H]GABA in their own way. Acute restraint stress caused a decrease in [ 3 H]GABA binding at its low-affine site and did not affected the high-affine site. Preliminary peptide administration did not influence this effect of stress., (© 2023 John Wiley & Sons Ltd.)

Ischemic stroke continues to be one of the leading causes of death and disability in the adult population worldwide. The currently used pharmacological methods for the treatment of ischemic stroke are not effective enough and require the search for new tools and approaches to identify therapeutic targets and potential neuroprotectors. Today, in the development of neuroprotective drugs for the treatment of stroke, special attention is paid to peptides. Namely, peptide action is aimed at blocking the cascade of pathological processes caused by a decrease in blood flow to the brain tissues. Different groups of peptides have therapeutic potential in ischemia. Among them are small interfering peptides that block protein-protein interactions, cationic arginine-rich peptides with a combination of various neuroprotective properties, shuttle peptides that ensure the permeability of neuroprotectors through the blood-brain barrier, and synthetic peptides that mimic natural regulatory peptides and hormones. In this review, we consider the latest achievements and trends in the development of new biologically active peptides, as well as the role of transcriptomic analysis in identifying the molecular mechanisms of action of potential drugs aimed at the treatment of ischemic stroke.

Glyprolines are Gly-Pro (GP)- or Pro-Gly (PG)-containing biogenic peptides. These peptides can act as neutrophil chemoattractants, or atheroprotective, anticoagulant, and neuroprotective agents. The Pro-Gly-Pro (PGP) tripeptide is an active factor of resistance to the biodegradation of peptide drugs. The synthetic Semax peptide, which includes Met-Glu-His-Phe (MEHF) fragments of adrenocorticotropic hormone and the C-terminal tripeptide PGP, serves as a neuroprotective drug for the treatment of ischemic stroke. Previously, we revealed that Semax mostly prevented the disruption of the gene expression pattern 24 h after a transient middle cerebral artery occlusion (tMCAO) in a rat brain model. The genes of this pattern were grouped into an inflammatory cluster (IC) and a neurotransmitter cluster (NC). Here, using real-time RT-PCR, the effect of other PGP-containing peptides, PGP and Pro-Gly-Pro-Leu (PGPL), on the expression of a number of genes in the IC and NC was studied 24 h after tMCAO. Both the PGP and PGPL peptides showed Semax-unlike effects, predominantly without changing gene expression 24 h after tMCAO. Moreover, there were IC genes ( iL1b , iL6 , and Socs3 ) for PGP, as well as IC ( iL6 , Ccl3 , Socs3 , and Fos ) and NC genes ( Cplx2 , Neurod6 , and Ptk2b ) for PGPL, that significantly changed in expression levels after peptide administration compared to Semax treatment under tMCAO conditions. Furthermore, gene enrichment analysis was carried out, and a regulatory gene network was constructed. Thus, the spectra of the common and unique effects of the PGP, PGPL, and Semax peptides under ischemia-reperfusion were distinguished.

The effect of temperature on the effectiveness of the incorporation of deuterium into pyrrolylcarnosine (PC) was studied. Deuterium gas and heavy water were used as a source of deuterium. Isotope exchange was carried out using solid-phase and liquid-phase methods. It was found that it is better to use isotope exchange with deuterated water to obtain preparative amounts of labeled pyrrolylcarnosine. When using y solid-phase method, the main label is in pyrrole. The incorporation of deuterium at a higher temperature occurs more evenly. In addition, the use of deuterated water made it possible to reduce the amount of unlabeled isotopomer to almost 0% and to obtain a product with a yield of 70% and a content of more than seven deuterium atoms. It was established that the content of deuterium in the compound can be increased by pretreating the reaction mixture with deuterium gas. This approach opens up additional opportunities for the synthesis of labeled compounds., (© 2022. Pleiades Publishing, Ltd.)

The involvement of nitric oxide in the responses of the hemostasis system to the appearance of proline-containing peptides in the blood was studied in experiments on rats. It was shown that a single intranasal administration of peptides PGP, RPGP, and PGPL to rats led to an increase in fibrinolytic, anticoagulant, and antiplatelet potential of blood. The use of the non-selective NO-synthase blocker L-NAME almost completely inhibited the anticoagulant effects of the glyprolines. It was found that the mechanism of the anticoagulant-fibrinolytic and antiplatelet action of glyproline peptides is determined by the activation of the enzymatic pathway of nitric oxide formation. The obtained results revealed the involvement of nitric oxide in the implementation of hemostatic and vascular-endothelial functions of the organism., (© 2022. Pleiades Publishing, Ltd.)

The effects of the peptide ACTH(6-9)-Pro-Gly-Pro at doses of 5; 50; 500 μg/kg on the Wistar rats' behaviour and gut mucosal microbiota composition under conditions of chronic immobilization stress (CRS) were studied. CRS increased anxiety-like and depressive-like behaviour, disturbances in locomotor activity and gut dysbiosis. Administration of ACTH(6-9)-Pro-Gly-Pro showed many phenotypic results. Peptide demonstrated anti-depressant activity at doses of 5 and 500 μg/kg by a decrease in the total immobile time in the FST. ACTH(6-9)-Pro-Gly-Pro administered at a dose of 50 μg/kg resulted in an anxiolytic effect which is shown by an increase in the time in the open arms of EPM (p < 0.05) and a decrease in the time in the closed arms (p < 0.05). Moreover, the peptide led to a decrease in alpha- and beta-diversity of the gut microbiota (p < 0.01). Correlation and linear regression analysis demonstrated central mechanisms of ACTH(6-9)-Pro-Gly-Pro anxiolytic activity and both central and peripheral ones in an anti-depressant effect. In this way, peptide ACTH(6-9)-Pro-Gly-Pro could prevent the development of behavioural disturbances and gut dysbiosis caused by chronic restraint stress., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Neurodegenerative diseases and depression are multifactorial disorders with a complex and poorly understood physiopathology. Astrocytes play a key role in the functioning of neurons in norm and pathology. Stress is an important factor for the development of brain disorders. Here, we review data on the effects of stress on astrocyte function and evidence of the involvement of astrocyte dysfunction in depression and Alzheimer's disease (AD). Stressful life events are an important risk factor for depression; meanwhile, depression is an important risk factor for AD. Clinical data indicate atrophic changes in the same areas of the brain, the hippocampus and prefrontal cortex (PFC), in both pathologies. These brain regions play a key role in regulating the stress response and are most vulnerable to the action of glucocorticoids. PFC astrocytes are critically involved in the development of depression. Stress alters astrocyte function and can result in pyroptotic death of not only neurons, but also astrocytes. BDNF-TrkB system not only plays a key role in depression and in normalizing the stress response, but also appears to be an important factor in the functioning of astrocytes. Astrocytes, being a target for stress and glucocorticoids, are a promising target for the treatment of stress-dependent depression and AD.

Natural melanocortins (MCs) have been used in the successful development of drugs with neuroprotective properties. Here, we studied the behavioral effects and molecular genetic mechanisms of two synthetic MC derivatives-ACTH(4-7)PGP (Semax) and ACTH(6-9)PGP under normal and acute restraint stress (ARS) conditions. Administration of Semax or ACTH(6-9)PGP (100 μg/kg) to rats 30 min before ARS attenuated ARS-induced behavioral alterations. Using high-throughput RNA sequencing (RNA-Seq), we identified 1359 differentially expressed genes (DEGs) in the hippocampus of vehicle-treated rats subjected to ARS, using a cutoff of >1.5 fold change and adjusted p -value ( Padj ) < 0.05, in samples collected 4.5 h after the ARS. Semax administration produced > 1500 DEGs, whereas ACTH(6-9)PGP administration led to <400 DEGs at 4.5 h after ARS. Nevertheless, ~250 overlapping DEGs were identified, and expression of these DEGs was changed unidirectionally by both peptides under ARS conditions. Modulation of the expression of genes associated with biogenesis, translation of RNA, DNA replication, and immune and nervous system function was produced by both peptides. Furthermore, both peptides upregulated the expression levels of many genes that displayed decreased expression after ARS, and vice versa, the MC peptides downregulated the expression levels of genes that were upregulated by ARS. Consequently, the antistress action of MC peptides may be associated with a correction of gene expression patterns that are disrupted during ARS.

The analysis of the literature data and own experimental studies on the effect of glyproline peptides on fat, carbohydrate metabolism, and hemostasis system when modeling metabolic syndrome in animals (rats) was carried out. Violations of fat and carbohydrate metabolism are characterized by hypercoagulation, increased glucose levels, dyslipidemia, and decreased rheological properties of blood. In this condition, the arginine- and lysine-containing glyprolines (PGP, PRP, RPGP, KKRRPGP, RKKRPGP, and KRRKPGP) at multiple (7 days) intranasal introduction had a complex effect on the hemostatic parameters, increasing antiplatelet, anticoagulant, and fibrinolytic activity of blood plasma. All the studied drugs also showed normoglycemic and normolipidemic effects and led to a slowdown or decrease in body weight growth. The analysis of the presented material allows us to speak about the prospects of using hit compounds as protective therapeutic agents. In the case of violations of fat and carbohydrate metabolism, regulatory glyprolines protect the body by displaying antithrombotic, hypolipidemic, and hypoglycemic properties., (© 2020 Wiley Periodicals LLC.)

The Semax (Met-Glu-His-Phe-Pro-Gly-Pro) peptide is a synthetic melanocortin derivative that is used in the treatment of ischemic stroke. Previously, studies of the molecular mechanisms underlying the actions of Semax using models of cerebral ischemia in rats showed that the peptide enhanced the transcription of neurotrophins and their receptors and modulated the expression of genes involved in the immune response. A genome-wide RNA-Seq analysis revealed that, in the rat transient middle cerebral artery occlusion (tMCAO) model, Semax suppressed the expression of inflammatory genes and activated the expression of neurotransmitter genes. Here, we aimed to evaluate the effect of Semax in this model via the brain expression profiling of key proteins involved in inflammation and cell death processes (MMP-9, c-Fos, and JNK), as well as neuroprotection and recovery (CREB) in stroke. At 24 h after tMCAO, we observed the upregulation of active CREB in subcortical structures, including the focus of the ischemic damage; downregulation of MMP-9 and c-Fos in the adjacent frontoparietal cortex; and downregulation of active JNK in both tissues under the action of Semax. Moreover, a regulatory network was constructed. In conclusion, the suppression of inflammatory and cell death processes and the activation of recovery may contribute to the neuroprotective action of Semax at both the transcriptome and protein levels.

Due to its nootropic, neuroprotective, and immunomodulatory effects, the peptide Semax is utilized in the treatment of ischemic stroke. Our earlier RNA-Seq analysis of the transcriptome in an ischemic model of transient occlusion of the middle cerebral artery showed an increase in the mRNA levels of many proinflammatory genes, and the suppression of their induction by Semax. However, for many relevant genes, including Il1a, Il1b, Il6 and Tnfa, the levels of their expression were too low for detailed quantitative evaluation. Here we utilize qRT-PCR to analyze the effects of the Semax peptide on the expression of weakly expressed mRNAs encoding several proinflammatory mediators, and show that exposure to Semax leads to a statistically significant decrease in the Il1a, Il1b, Il6, Ccl3, and Cxcl2 mRNAs, which compensates for the increase in the transcription of these genes induced by ischemia-reperfusion. We conclude that the observed protective effect of Semax in the model of stroke may be due to its anti-inflammatory effects. We also discuss the limitations of the RNA-Seq when applied to quantifying less abundant transcripts as compared to the real-time RT-PCR method.

Selective serotonin reuptake inhibitors (SSRI) are commonly used to treat depression during pregnancy. SSRIs cross the placenta and may influence the maturation of the foetal brain. Clinical and preclinical findings suggest long-term consequences of SSRI perinatal exposure for the offspring. The mechanisms of SSRI effects on developing brain remain largely unknown and there are no directional approaches for prevention of the consequences of maternal SSRI treatment during pregnancy. The heptapeptide Semax (MEHFPGP) is a synthetic analogue of ACTH(4-10) which exerts marked nootropic and neuroprotective activities. The aim of the present study was to investigate the long-term effects of neonatal exposure to the SSRI fluvoxamine (FA) in white rats. Additionally, the study examined the potential for Semax to prevent the negative consequences of neonatal FA exposure. Rat pups received FA or vehicle injections on postnatal days 1-14, a time period equivalent to 27-40 weeks of human foetal age. After FA treatment, rats were administered with Semax or vehicle on postnatal days 15-28. During the 2nd month of life, the rats underwent behavioural testing, and monoamine levels in brain structures were measured. It was shown that neonatal FA exposure leads to the impaired emotional response to stress and novelty and delayed acquisition of food-motivated maze task in adolescent and young adult rats. Furthermore, FA exposure induced alterations in the monoamine levels in brains of 1- and 2- month-old rats. Semax administration reduced the anxiety-like behaviour, improved learning abilities and normalized the levels of brain biogenic amines impaired by the FA exposure. The results demonstrate that early-life FA exposure in rat pups produces long-term disturbances in their anxiety-related behaviour, learning abilities, and brain monoamines content. Semax exerts a favourable effect on behaviour and biogenic amine system of rats exposed to the antidepressant. Thus, peptide Semax can prevent behavioural deficits caused by altered 5-HT levels during development., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The cytotoxicity of doxorubicin (Dox) and its peptide modifications Z-Gly-Pro-Dox and Boc-Gly-Pro-Dox were studied. Tetrahymena pyriformis was used as a test system, which made it possible, due to the short life cycle and high reproduction rate of ciliates, to trace their response to the effects of toxicants over several generations. It was found that peptide modification of the Dox molecule markedly reduces its cytotoxic and cytostatic effect. The Z-Gly-Pro-Dox modification has less cytotoxic and cytostatic effect compared to Boc-Gly-Pro-Dox. When determining the ability of drugs (at a concentration of 100 μM) to prevent bacterial contamination of samples, it was shown that the smallest degree of overgrowth was recorded in the presence of Dox (OD 600nm 81.1). Boc-Gly-Pro-Dox also had a bacteriostatic effect, though less pronounced (OD 600nm 93.8). The degree of overgrowth in the presence of Z-Gly-Pro-Dox was close to that of distilled water. The results obtained on ciliates did not contradict the data obtained in similar studies on mice.

Here we present a review of studies on the effects of peptides with neuroprotective properties on gene transcription in nerve cells. The few published works in this area clearly demonstrate massive changes in cell transcriptomes induced by peptides under normal conditions and under conditions of experimental brain ischemia. These changes significantly affect signaling and metabolic pathways, affecting various body systems and confirming the multiple target actions of peptides. The importance of noncoding RNAs in the regulation of these processes is shown, and we discuss the prospects of research for determining the main mechanisms of peptide regulation, which is necessary for the further development of drugs with targeted neuroprotective effects., (© 2020 Wiley Periodicals LLC.)

The phytohormone abscisic acid (ABA) plays an important role in plant growth and in response to abiotic stress factors. At the same time, its accumulation in soil can negatively affect seed germination, inhibit root growth and increase plant sensitivity to pathogens. ABA is an inert compound resistant to spontaneous hydrolysis and its biological transformation is scarcely understood. Recently, the strain Rhodococcus sp. P1Y was described as a rhizosphere bacterium assimilating ABA as a sole carbon source in batch culture and affecting ABA concentrations in plant roots. In this work, the intermediate product of ABA decomposition by this bacterium was isolated and purified by preparative HPLC techniques. Proof that this compound belongs to ABA derivatives was carried out by measuring the molar radioactivity of the conversion products of this phytohormone labeled with tritium. The chemical structure of this compound was determined by instrumental techniques including high-resolution mass spectrometry, NMR spectrometry, FTIR and UV spectroscopies. As a result, the metabolite was identified as ( 4RS )-4-hydroxy-3,5,5-trimethyl-4-[( E )-3-oxobut-1-enyl]cyclohex-2-en-1-one (dehydrovomifoliol). Based on the data obtained, it was concluded that the pathway of bacterial degradation and assimilation of ABA begins with a gradual shortening of the acyl part of the molecule.

We studied the effect of intraperitoneal administration ACTH (4-7) -PGP in doses of 5, 50, 150, and 450 μg/kg to Wistar male rats 12-15 min before modeling restraint stress on the morphofunctional state of the colon. In rats exposed to restraint stress, signs of atrophy and inflammatory reaction in the colon wall, changes in functional activity and number of mast cells, and increased serum level of corticosterone were observed. Administration of the peptide led to a decrease in corticosterone concentration, alleviated stress-induced pathomorphological changes, and promoted adaptation of the intestinal wall to stress. The positive effects of ACTH (4-7) -PGP can be determined by multifunctional nature of the physiological and pharmacological effects of the neuropeptide.

Anticoagulant effects of 12 short peptides of the glyproline series - Arg-Glu-Arg-Pro-Gly-Pro (RERPGP), Arg-Glu-Arg-Val-Gly-Pro (RERVGP), Arg-Glu-Arg-Gly-Pro (RERGP), Arg-Pro-Gly-Pro (RPGP), Pro-Leu-Pro (PLP), Pro-Leu-Pro-Ala (PLPA), Pro-Gly-Pro-Leu (PGPL), Phe-Pro-Leu-Pro-Ala (FPLPA), Pyr-Arg-Pro (PyrRP), Lys-Lys-Arg-Arg-Pro-Gly-Pro (KKRRPGP), Arg-Lys-Lys-Arg-Pro-Gly-Pro (RKKRPGP), and Lys-Arg-Lys-Pro-Gly-Pro (KRKPGP) in concentrations of 10-3 and 10-2 mg/ml in vitro was demonstrated by the thromboelastographic method. The effects of 6 peptides ((RERPGP, RPGP, PLP, PLPA, RKKRPGP, and KKRRPGP) were also observed in vivo after intranasal or oral administration. Changes in the studied thromboelastographic parameters towards hypocoagulation in comparison with the control group were noted. Arginine and leucine glyprolines produced the maximum anticoagulant effect.

We studied the effect of Semax on the state of intestinal microbiota in rats subjected to restraint stress. Semax was injected to Wistar male rats intraperitoneally in doses of 5, 50, 150, 450 μg/kg 12-15 min before modelling chronic restraint stress. It was found that stress exposure reduced the number of obligate bacteria in the colon microbiota, but increased the content of opportunistic microorganisms. Semax in doses of 50 and 150 μg/kg prevented the stress-induced changes in the composition of colon microbiota. The observed effects of Semax might be mediated by the central neurotropic effects as well as by binding to peripheral melanocortin receptors of the intestine.

Cerebral ischaemia is the most common cause of impaired brain function. Biologically active peptides represent potential drugs for reducing the damage that occurs after ischaemia. The synthetic melanocortin derivative, ACTH (4-7) PGP (Semax), has been used successfully in the treatment of patients with severe impairment of cerebral blood circulation. However, its molecular mechanisms of action within the brain are not yet fully understood. Previously, we used the transient middle cerebral artery occlusion (tMCAO) model to study the damaging effects of ischaemia-reperfusion on the brain transcriptome in rats. Here, using RNA-Seq analysis, we investigated the protective properties of the Semax peptide at the transcriptome level under tMCAO conditions. We have identified 394 differentially expressed genes (DEGs) (>1.5-fold change) in the brains of rats at 24 h after tMCAO treated with Semax relative to saline. Following tMCAO, we found that Semax suppressed the expression of genes related to inflammatory processes and activated the expression of genes related to neurotransmission. In contrast, ischaemia-reperfusion alone activated the expression of inflammation-related genes and suppressed the expression of neurotransmission-related genes. Therefore, the neuroprotective action of Semax may be associated with a compensation of mRNA expression patterns that are disrupted during ischaemia-reperfusion conditions., Competing Interests: The authors declare no conflict of interest.

We studied the effects of Selank on the condition of the colon wall in Wistar male rats subjected to restraint stress. Selank was injected intraperitoneally in doses of 80, 250, and 750 μg/kg 15 min before stress exposure. In rats subjected to stress, signs of atrophy, inflammatory reaction, and changes in the number and functional activity of mast cells were observed against the background of increased corticosterone level. Selank administration led to a decrease in corticosterone levels, reduced pathomorphological manifestations of stress exposure, and accelerated adaptation. These effects were presumably realized due to multifunctional biological effects of Selank.

The importance of studying the action mechanisms of drugs based on natural regulatory peptides is commonly recognized. Particular attention is paid to the peptide drugs that contribute to the restoration of brain functions after acute cerebrovascular accidents (stroke), which for many years continues to be one of the main problems and threats to human health. However, molecular genetic changes in the brain in response to ischemia, as well as the mechanisms of protective effects of peptides, have not been sufficiently studied. This limits the use of neuroprotective peptides and makes it difficult to develop new, more efficient drugs with targeted action on brain functions. Transcriptome analysis is a promising approach for studying the mechanisms of the damaging effects of cerebral ischemia and neuroprotective action of peptide drugs. Beside investigating the role of mRNAs in protein synthesis, the development of new neuroprotection strategies requires studying the involvement of regulatory RNAs in ischemia. Of greatest interest are microRNAs (miRNAs) and circular RNAs (circRNAs), which are expressed predominantly in the brain. CircRNAs can interact with miRNAs and diminish their activity, thereby inhibiting miRNA-mediated repression of mRNAs. It has become apparent that analysis of the circRNA/miRNA/mRNA system is essential for deciphering the mechanisms of brain damage and repair. Here, we present the results of studies on the ischemia-induced changes in the activity of genes and peptide-mediated alterations in the transcriptome profiles in experimental ischemia and formulate the basic principles of peptide regulation in the ischemia-induced damage.

Glutamate (Glu) excitotoxicity, which accompanies brain ischemia or traumatic brain injury, is the leading mechanism of neuronal death. In the present work, we studied the effects of the peptides HFRWPGP (ACTH 6-9 PGP), KKRRPG, and PyrRP on the survival of cultured cortical neurons on the background of excitotoxic effect of Glu (100 µM). Biochemical (MTT/WST) and morphometric analyzes showed that, depending on the dose, ACTH 6-9 PGP and KKRRPGP protect neurons from the cells death, while PyrRP, conversely, enhances it. The neuroprotective effect of ACTH 6-9 PGP is accompanied by a slowdown in the development of delayed calcium dysregulation and synchronous mitochondrial depolarization. Among the studied peptides, only ACTH 6-9 PGP significantly increased the number of neurons that restored Ca 2+ homeostasis after Glu was abolished. The influence of KKRRPGP was less pronounced, whereas PyrRP, on the contrary, reduced the number of neurons with low [Ca 2+ ] i . Thus, this study revealed the high therapeutic significance of ACTH 6-9 PGP and allowed assessing the prospects for its possible clinical use.

Background: Currently, the most dynamic areas in the glutamate receptor system neurobiology are the identification and development of positive allosteric modulators (PAMs) of glutamate ionotropic receptors. PAM-based drugs are of great interest as promising candidates for the treatment of neurological diseases, such as epilepsy, Alzheimer's disease, schizophrenia, etc. Understanding the molecular mechanisms underlying the biological action of natural and synthetic PAMs is a key point for modifying the original chemical compounds as well as for new drug design., Objective: We are trying to elaborate a system of molecular functional screening of ionotropic glutamate receptor probable PAMs., Methods: The system will be based on the radioligand - receptor method of analysis and will allow rapid quantification of new AMPAR probable PAMs molecular activity. We plan to use a tritiumlabeled analogue of recently elaborated ionotropic GluR probable PAM ([3H]PAM-43) as the main radioligand., Results: Here, we characterized the specific binding of the ligand and its ability to potentiate ionotropic GluR currents. The existence of at least two different sites of [3H]PAM-43 specific binding has been shown. One of the above sites is glutamate-dependent and is characterized by higher affinity. "Patchclamp" technique showed the ability of PAM-43 to potentiate ionotropic GluR currents in rat cerebellar Purkinje neurons in a concentration-dependent manner., Conclusion: The possibility of using PAM-43 as a model compound to study different allosteric effects of potential regulatory drugs (AMPAR allosteric regulators) was shown. [3H]PAM-43 based screening system will allow rapid selection of new AMPAR probable PAM structures and quantification of their molecular activity., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

The present study was aimed at the assessment of effects of anxiolytic Selank and nootropic Semax on the whole-brain resting-state functional connectivity (FC) of each of the predefined regions of interest (ROIs) in 52 healthy participants. The ROIs included amygdala (one of the key regions for the regulation of anxiety) and dorsolateral prefrontal cortex (DLPFC; the key region for executive functions, including working memory) in the right and left hemisphere. Resting-state fMRI was carried out three times, namely before, after 5 and 20 min of the injection of either Semax, or Selank, or placebo. Between-group alongwith between-condition differences were revealed in FC between the right amygdala and a region in fusiform, inferior and middle temporal as well as parahippocampal gyri in the right hemisphere. Post hoc analysis allowed us to define both general and specific effects of Selank and Semax on FC between the right amygdala and the right temporal cortex for the first time.

We studied the effect of chronic intranasal and peroral administration of a new peptide preparation AСTH 15-18 PGP in a dose of 100 mg/kg body weight on the state of vascular-platelet and plasma hemostasis in animals. It was found that this synthetic regulatory peptide administered intranasally can produce antiplatelet, anticoagulant, and antifibrin-stabilizing effects on the blood plasma in healthy rats. In both administration routes, the peptide induced activation of the anticoagulation system of the hemostasis by increasing enzymatic and non-enzymatic fibrinolysis; after intranasal administration, the fibrinolytic effects were more pronounced.

Development of therapeutic preparations involves several steps, starting with the synthesis of chemical compounds and testing them in different models for selecting the most effective and safest ones to clinical trials and introduction into medical practice. Cultured animal cells (both primary and transformed) are commonly used as models for compound screening. However, cell models display a number of disadvantages, including insufficient standardization (primary cells) and disruption of cell genotypes (transformed cells). Generation of human induced pluripotent stem cells (IPSCs) offers new possibilities for the development of high-throughput test systems for screening potential therapeutic preparations with different activity spectra. Due to the capacity to differentiate into all cell types of an adult organism, IPSCs are a unique model that allows examining the activity and potential toxicity of tested compounds during the entire differentiation process in vitro. In this work, we demonstrated the efficiency of IPSCs and their neuronal derivatives for selecting substances with the neuroprotective activity using two classes of compounds - melanocortin family peptides and endocannabinoids. None of the tested compounds displayed cyto- or embryotoxicity. Both melanocortin peptides and endocannabinoids exerted neuroprotective effect in the neuronal precursors and IPSC-derived neurons subjected to hydrogen peroxide. The endocannabinoid N-docosahexaenoyl dopamine exhibited the highest neuroprotective effect (~70%) in the differentiated cultures enriched with dopaminergic neurons; the effect of melanocortin Semax was ~40%. The possibility of using other IPSC derivatives for selecting compounds with the neuroprotective activity is discussed.

An integrated methodological approach to study the molecular aspects of short regulatory neuropeptides biological mechanism is proposed. The complex research is based on radioligand-receptor method of analysis and covers such points of peptides molecular activity as: specific binding of peptides to brain cells plasmatic membranes, formation of tissue specific synacton, influence of peptides (as allosteric modulators) on functionality of different neuroreceptors as well as delayed in time effects of peptides on receptor-binding activity of well-known neuroreceptor systems. Radiolabeled ligands in such complex study are the one of the best and precision instruments to uncover the molecular mechanism of multiple and multitarget biological effects of regulatory peptides. In this issue we used heptapeptide Semax as a model regulatory peptide, [ 3 H]Ach and [ 3 H]GABA as an effector molecules, and the rat model of stress-induced memory and behavior impairment as a morbid state. We showed the ability of Semax to modulate in a dose-dependent manner [ 3 H]Ach and [ 3 H]GABA specific binding to some of its corresponding receptors as well as to affect the number of [ 3 H]GABA specific binding places on rat neurons plasmatic membranes after complex stress exposure., (© 2019 John Wiley & Sons, Ltd.)

Boc-Gly-Pro-DP, Z-Gly-Pro-DP, LA-Gly-Pro-DP, Boc-Gly-Pro-Srt, Z-Gly-Pro-Srt were synthesized for the first time. The stability of these compounds in the presence of leucine aminopeptidase, carboxypeptidase Y, carboxypeptidase B and proline endopeptidase (PEP) was determined. It turned out that the compounds are stable in the presence of aminopeptidases and carboxypeptidases. In the presence of PEP, dopamine (DP) and serotonin (Srt) are cleaved from the synthesized preparations. Thus, new proline-containing Srt and DP derivatives were obtained, Srt and DP could be gradually released from them. This suggest the possibility of a prolonged action of these biologically active compounds on the vital activity of cells and, consequently, of the whole organism.

A mass spectrometric method has been developed for determining the content of dopamine and serotonin derivatives, which allows evaluating the efficiency of their penetration through artificial membranes depending on the structure of their peptide fragment. In this case, the diffusion of dopamine and serotonin derivatives through the membrane occurred as a result of competitive interactions. It was shown which compounds in this mixture more easily penetrate through artificial membranes. It was found that the most promising in terms of overcoming the BBB are Boc-Pro-Srt and Boc-Pro-DOPA.

We studied the effects of Selank on morphological parameters of the liver in Wistar male rats subjected to chronic foot-shock stress. Selank was injected intraperitoneally in doses of 100, 300 and 1000 μg/kg 15 min before each stress session. Morphological and morphometrical analysis showed that chronic foot-shock stress induced hydropic degeneration of hepatocytes, an increase of the nucleus/cytoplasm ratio due to an increase in the area of nuclei and reduction of the cytoplasm area, the appearance of focal necroses, and lymphohistiocyte infiltration. Injection of Selank in all doses reduced the intensity of stress-induced degenerative changes. Administration of Selank in doses of 300 and 1000 μg/kg restored the nucleus/cytoplasm ratio in hepatocytes. The maximum stress-limiting effect was attained after administration of 300 μg/kg Selank.

We studied the effects of Selank on intestinal microbiota in Wistar male rats subjected to chronic restraint stress. Selank was injected intraperitoneally in doses of 80, 250 and 750 μg/kg 15 min before stress exposure. Chronic restraint stress led to a decrease in the content of obligate microflora, while the content of opportunistic microorganisms increased. Selank restored intestinal microbiota presumably via central (neurotropic) and peripheral (immunotropic) mechanisms.

We studied immunocorrecting effects of Semax (Met-Glu-His-Phe-Pro-Gly-Pro) on the model of "social" stress caused by sensory contact and intermale confrontation. Functional activity of the immune system of laboratory animals was evaluated in standard immunopharmacological tests: delayed-type hypersensitivity reaction, direct agglutination test, latex test for studying phagocytic activity of peripheral blood neutrophils, changes in differential leukocyte count, and weight of immunocompetent organs. It was found that changes in the immune response caused by "social" stress are multidirectional, which confirms the theory of stress-induced "immune imbalance". Semax acted as effective immune corrector restoring cellular and humoral immunogenesis reactions and phagocytic activity of neutrophils. This attested to the presence of immunomodulating properties in Semax and necessitates further studies in this field.

New data on peptide drugs have been summarized; their high stability is due to both the introduction of Pro-Gly-Pro in various amino acid sequences and the modification of the glyproline fragment itself. Pro-Gly-Pro-Leu, ACTH(6-9)Pro-Gly-Pro, 5-oxo-Pro-Arg-Pro and 5-oxo-Pro-His-Pro-NH2 were used as proline-containing peptides. Tritiated peptides were obtained: Pro-Gly-Pro-Leu with specific radioactivity of 135 Ci/mmol, ACTH(6-9)Pro-Gly-Pro - 26 Ci/mmol, 5-oxo-Pro-Arg-Pro - 60 Ci/mmol and 5-oxo-Pro-His-Pro-NH2 - 75 Ci/mmol. The concentration of Pro-Gly-Pro-Leu, ACTH(6-9)Pro-Gly-Pro, 5-oxo-Pro-Arg-Pro and 5-oxo-Pro-His-Pro-NH2 in the blood was found to be about 200 times more than in the brain for intranasal administration, and in average 600 times more for intravenous administration. The stability of proline-containing peptides in vitro experiments was determined using different commercially available peptidases (leucine aminopeptidases, dipeptidases, carboxypeptidases B and Y), and using nasal mucus, microsomal fraction of the rat brain (IMPC) and rat blood plasma. During peptidase hydrolysis of Pro-Gly-Pro-Leu, the main metabolites were Gly-Pro-Leu, Pro-Gly-Pro, Gly-Pro and Pro-Gly. For ACTH(6-9)Pro-Gly-Pro, the main metabolites were Phe-Arg-Trp-Pro-Gly-Pro and Trp-Pro-Gly-Pro. In peptidase hydrolysis of 5-oxo-Pro-His-Pro-NH2, the major metabolite was 5-oxo-Pro-His-Pro. It was shown that with different methods of peptides administration the composition of the metabolites formed is different. Based on the data obtained, resistance to enzymatic cleavage of peptides and their metabolic pathways were evaluated. Thus, these new data have shown that the above approaches can be used to prolong the action of glyprolines in living objects. In this case, the degradation of proline-containing peptides occurs mainly not due to the action of proteases, but due to other ways of degradation. In general, the data presented in the review indicate the promise of intranasal way of introducing biologically active peptides into the brain of living organisms.

It was established that in neurodegeneration models in the human neuron-like cell line SH-SY5Y, amide derivatives of arachidonic and docosahexaenoic acids were inactive in experiments with MPP + and CoCl 2 but protected from H 2 O 2 . The protective activity of neurolipins decreased in the series DHA-DA > AA-SER ≥ AA-GLY > AA-GABA ≥ AA-EA and was manifested starting from a concentration of 0.5 nM.

The effects of nootropic drug Semax on the neuronal network of the brain were studied by the resting state functional magnetic-resonance imaging (resting state fMRI). The study was carried out on two groups of healthy volunteers (11 men and 13 women aged 43.9±9.5 years). Resting state fMRI was carried out 3 times: directly before and 5 and 20 min after intranasal 1% Semax (14 subjects) or placebo (10 subjects). The topography of the resting state default mode network was studied. A greater volume of the default mode network rostral (medial frontal cortex) subcomponent was detected in the Semax group in comparison with controls. Resting state fMRI confirmed Semax effects on the neuronal network of the brain and demonstrated topography of these effects.

The concentration-clamp experiments with neurons isolated from the rat brain showed that nootropic and neuroprotective drug Semax added to perfusion solution at concentration of 1 μM augmented the amplitude of GABA-activated ionic currents in cerebellum Purkinje cells by 147±13%. In addition, Semax in perfusion solution (0.1 and 1 μM) diminished the amplitude of glycine-activated chloride currents in hippocampal pyramidal neurons down to 68 and 43% control level, respectively. Both potentiating and inhibitory effects developed slowly, and they were poorly reversible, which indicated a probable implication of second messengers in the observed phenomena. Semax accelerated the falling edge of glycine-activated current both after a short-term co-application with agonist and after addition of this peptide into perfusion solution.

A comparative study of the influence of regulatory proline peptides Pro-Gly-Pro, Pro-Arg-Pro, Pro-Gly-Pro-Leu, and Arg-Pro-Gly-Pro on the state of the hemostasis system was carried out in an experiment on male rats with metabolic syndrome. Under these conditions, repeated (7-fold) intranasal administration of the peptides in a daily dose of 50 μg/kg resulted in an increase in the anticoagulant potential of the blood, namely, in an increase in the anticoagulant, fibrinolytic, and antiplatelet activity 20 h and 7 days after the last peptide injection. The arginine-containing peptide Arg-Pro-Gly-Pro had the most pronounced and stable effect on haemostasis under these experimental conditions.

One of the most urgent and important tasks of modern biological and medical research is the search and research of pharmacological agents that combine lipid-lowering and antithrombotic effects in the organism. The unique effects of the regulatory peptides of the oxoproline series (5-oхo-Pro-His-Pro-NH2, 5-oxo-Pro-Trp-Pro and 5-oxo-Pro-Arg-Pro or 5-oхo-Pro-His-Pro-NH2, Pyr-Trp-Pro and Pyr-Arg-Pro) have been found in rats with hypercholesterolemia (metabolic syndrome). Multiple intranasal of these peptides to animals with developed hypercholesterolemia increased anticoagulant, fibrinolytic and antiplatelet potential of the blood and simultaneously lowered increased concentrations of total cholesterol, low-density lipoprotein cholesterol and triglycerides. In addition, they contributed to the normalization of blood glucose levels. A week after the last admistration of these peptides, the hypocholesterolemic, normoglycemic and anticoagulant effects persisted. The relationship between the structure of peptides of the oxoproline series and their functional properties is discussed. A conclusion is made about the prospects of further studies of oxoproline peptides as drugs that combine antithrombotic effects with the improvement of fat metabolism in the body.

We studied the effects of peptide drugs (HLDF-6, PGP, RPGP, and PGLP) and peptide pharmaceutical products (Semax, Selank, and thyroliberin) on proliferation and survival of mouse embryonic stem cells and their derivatives. Differentiation of mouse embryonic stem cells into neuronal precursors was evaluated. PGP and PGLP in concentrations of 10 and 0.1 μM, respectively, had little, but significant inhibitory effect on proliferative activity of cells. These peptides in concentrations of 10 and 0.1 μM, respectively, and Semax (10 and 0.1 μM) significantly increased the survival rate of mouse embryonic stem cells (serum deprivation). Moreover, study peptides had little effect on the formation of neuronal precursors from mouse embryonic stem cells. HLDF-6, Selank, and thyroliberin produced an insignificant effect on the differentiation of these cells into mature neurons. Analysis of differentiation of embryonic stem cells into GABA + neurons showed that Selank, thyroliberin (100 μM), and NGF (100 ng/ml) decrease the ratio of these cells by 61, 58, and 87%, respectively, in comparison with the control. Our results indicate that these peptide compounds do not produce toxic effect during the embryonic and fetal period of life.

Acetyl, oleoyl, arachidonoyl, and docosahexaenoyl derivatives of the Pro-Gly-Pro-Leu peptide with a chemical purity of 99.8% were synthesized. The degradation kinetics of the Pro-Gly-Pro-Leu derivatives under the action of leucine aminopeptidase, nasal mucus, and microsomal fraction of the brain and blood of rats was studied. It was shown that the N-acyl derivatives of Pro-Gly-Pro-Leu proved to be more resistant to the action of leucine aminopeptidase and other enzyme systems. The study of the cytotoxic and anti-inflammatory activity of preparations on the mouse macrophage cell line RAW264.7 showed that acylation with oleic and arachidonic acid makes the peptide cytotoxic with LC50 in the range of 70-15 μM and gives it anti-inflammatory properties with EC50 of 32 and 36 μM, respectively.

Emerging evidence implicates impaired self-regulation of the hypothalamic-pituitary-adrenal (HPA) axis and inflammation as important and closely related components of the pathophysiology of major depression. Antidepressants show anti-inflammatory effects and are suggested to enhance glucocorticoid feedback inhibition of the HPA axis. HPA axis activity is also negatively self-regulated by the adrenocorticotropic hormone (ACTH), a potent anti-inflammatory peptide activating five subtypes of melanocortin receptors (MCRs). There are indications that ACTH-mediated feedback can be activated by noncorticotropic N-terminal ACTH fragments such as a potent anti-inflammatory MC1/3/4/5R agonist α-melanocyte-stimulating hormone (α-MSH), corresponding to ACTH(1-13), and a MC3/5R agonist ACTH(4-10). We investigated whether intraperitoneal administration of rats with these peptides affects anhedonia, which is a core symptom of depression. Inflammation-related anhedonia was induced by a single intraperitoneal administration of a low dose (0.025mg/kg) of lipopolysaccharide (LPS). Stress-related anhedonia was induced by the chronic unpredictable stress (CUS) procedure. The sucrose preference test was used to detect anhedonia. We found that ACTH(4-10) pretreatment decreased LPS-induced increase in serum corticosterone and tumor necrosis factor (TNF)-α, and a MC3/4R antagonist SHU9119 blocked this effect. Both α-MSH and ACTH(4-10) alleviated LPS-induced anhedonia. In the CUS model, these peptides reduced anhedonia and normalized body weight gain. The data indicate that systemic α-MSH and ACTH(4-10) produce an antidepressant-like effect on anhedonia induced by stress or inflammation, the stimuli that trigger the release of ACTH and α-MSH into the bloodstream. The results suggest a counterbalancing role of circulating melanocortins in depression and point to a new approach for antidepressant treatment., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

We studied the effect of Selank administered intraperitoneally in doses of 100, 300, and 1000 μg/kg to male Wistar rats 15 min prior to restraint stress on the content of aminotransferases and total protein concentration in blood serum and intensity of free radical oxidation in the liver. Under conditions of acute restraint stress, Selank in doses of 100 and 300 μg/kg decreased catalase and superoxide dismutase activities and malondialdehyde concentration and increased total antioxidant activity in the liver homogenate. Administration of Selank in a dose of 1000 μg/kg reduced the content of aminotransferases in blood serum, decreased superoxide dismutase activity in the liver, and increased total antioxidant activity. Under conditions of chronic stress, Selank in all doses produced similar effects: reduced superoxide dismutase activity and malondialdehyde concentration in the liver tissue and AST activity in the serum. The other parameters remained unchanged.

Brain stroke continues to claim the lives of million people every year. To build the effective strategies for stroke treatment it is necessary to understand the neuroprotective mechanisms that are able to prevent the ischemic injury. Consisting of the ACTH (4-7) fragment and the tripeptide Pro-Gly-Pro (PGP), the synthetic peptide Semax effectively protects brain against ischemic stroke. However, the molecular mechanisms underlying its neuroprotection and participation of PGP in them are still needed to be clarified. To reveal biological processes and signaling pathways, which are affected by Semax and PGP, we performed the transcriptome analysis of cerebral cortex of rats with focal cerebral ischemia treated by these peptides. The genome-wide biochip data analysis detected the differentially expressed genes (DEGs) and bioinformatic web-tool Ingenuity iReport found DEGs associations with several biological processes and signaling pathways. The immune response is the process most markedly affected by the peptide: Semax enhances antigen presentation signaling pathway, intensifies the effect of ischemia on the interferon signaling pathways and affects the processes for synthesizing immunoglobulins. Semax significantly increased expression of the gene encoding the immunoglobulin heavy chain, highly affects on cytokine, stress response and ribosomal protein-encoding genes after occlusion. PGP treatment of rats with ischemia attenuates the immune activity and suppresses neurotransmission in the CNS. We suppose that neuroprotective mechanism of Semax is realized via the neuroimmune crosstalk, and the new properties of PGP were found under ischemia. Our results provided the basis for further proteomic investigations in the field of searching Semax neuroprotection mechanism.

We studied the effect of intraperitoneal administration of peptide ACTG 4-7 -PGP to male Wistar rats in doses of 5, 50, 150, and 450 μg/kg on the morphofunctional state of hepatocytes in chronic emotional and painful stress. A dose-dependent stress-limiting effect of the peptide was observed: it normalized the protein synthesis function of the liver and serum activity of ALT. The anticytolytic effect of the peptide increased with increasing its dose against the background of the increase in the relative number of multinucleated and multinucleolated cells and deceleration of the recovery of serum protein concentration. The decrease of hepatocyte cytolysis against the background of more intense morphological signs of protein synthesis processes attests to activation of reparative processes in the liver parenchyma via enhanced constitutional synthesis of protein.

Short endogenous peptides represent one of the most important constituents of the mammalian body's general regulatory system. Some synthesized analogs and modified natural peptides (eg, corticotropins) also show high biological activity. Nevertheless, the mechanism of action of regulatory peptides remains unclear. To explain the effects of peptides of intermolecular processes, the hypothesis that a synactonal mechanism underlies the action of regulatory peptides, exemplified by the heptapeptide Semax, has been proposed. Thus, in the total pool of Semax metabolites, which includes the cleavage products of the parental molecule, we can distinguish the functional core, represented by the major metabolic products-peptides HFPGP and PGP. These peptides have their own binding sites with similar although differing characteristics. Together with Semax, they constitute a single complex of bioregulators acting in a certain sequence and in interaction, ie, synacton. It can be assumed that the diverse clinically significant effects of the drug Semax are determined by its synacton. Specific interactions between some tritium-labeled peptides (basic constituents of the Semax synacton) and plasma membranes of neurons have been characterized. Only a few peptides of the Semax synacton showed competitive activity for the Semax binding sites. Fragments comprising 5 amino acid residues (EHFPG and HFPGP) showed the highest competitive activity. We also characterized the processes of specific ligand-receptor interactions of some tritium-labeled corticotropins ([ 3 H-Pro]MEHFPGP, [ 3 H-Pro]HFPGP, and [ 3 H-Pro]PGP) by applying mathematical discriminative models (Scatchard, Hill, Bjerrum, and Lineweaver-Burk plots). So the intermolecular interactions of these peptides with plasma membranes of neuronal brain targets are probably not limited by specific binding at orthosteric sites. The effect of peptides that act in the synacton considerably extends the regulatory potential of the initial molecule., (Copyright © 2016 John Wiley & Sons, Ltd.)