Peptide Regulation of Chondrogenic Stem Cell Differentiation.

The search for innovative ways to treat osteoarthritis (OA) is an urgent task for molecular medicine and biogerontology. OA leads to disability in persons of middle and older age, while safe and effective methods of treating OA have not yet been discovered. The directed differentiation of mesenchymal stem cells (MSCs) into chondrocytes is considered one of the possible methods to treat OA. This review describes the main molecules involved in the chondrogenic differentiation of MSCs. The peptides synthesized on the basis of growth factors' structures (SK2.1, BMP, B2A, and SSPEPS) and components of the extracellular matrix of cartilage tissue (LPP, CFOGER, CMP, RDG, and N-cadherin mimetic peptide) offer the greatest promise for the regulation of the chondrogenic differentiation of MSCs. These peptides regulate the WNT, ERK-p38, and Smad 1/5/8 signaling pathways, gene expression, and the synthesis of chondrogenic differentiation proteins such as COL2, SOX9, ACAN, etc.

The Influence of KE and EW Dipeptides in the Composition of the Thymalin Drug on Gene Expression and Protein Synthesis Involved in the Pathogenesis of COVID-19.

Thymalin is an immunomodulatory drug containing a polypeptide extract of thymus that has demonstrated efficacy in the therapy of acute respiratory distress syndrome and chronic obstructive pulmonary disease, as well as in complex therapy related to severe COVID-19 in middle-aged and elderly patients.. KE and EW dipeptides are active substances of Thymalin. There is evidence that KE stimulates cellular immunity and nonspecific resistance in organisms, exerting an activating effect on macrophages, blood lymphocytes, thymocytes, and neutrophils, while EW reduces angiotensin-induced vasoconstriction and preserves endothelium-dependent vascular relaxation by inhibiting ACE2, the target protein of SARS-CoV-2. However, the mechanism of the immunomodulatory action of Thymalin, KE, and EW during COVID-19 remains unclear. To identify the potential mechanism of action underlying the immunomodulatory activity of Thymalin and its active components, EW and KE dipeptides, we assessed inflammatory response in the context of COVID-19. Interactions between EW and KE dipeptides and double-stranded DNA (dsDNA) were investigated by molecular modeling and docking using ICM-Pro. Analysis of the possible effect of EW and KE dipeptides on gene expression and protein synthesis involved in the pathogenesis of COVID-19 was conducted through the use of bioinformatics methods, including a search for promoter sequences in the Eukaryotic Promoter Database, the determination of genes associated with the development of COVID-19 using the PathCards database of human biological pathways (pathway unification database), identification of the relationship between proteins through cluster analysis in the STRING database ('Search Tool for Retrieval of Interacting Genes/Proteins'), and assessment of the functional enrichment of protein-protein interaction (PPI) using the terms of gene ontology (GO) and the Markov cluster algorithm (MCL). After that, in vitro studying of a lipopolysaccharide (LPS)-induced model of inflammation using human peripheral blood mononuclear cells was performed. ELISA was applied to assess the level of cytokines (IL-1ß, IL-6, TNFα) in the supernatant of cells with or without the impact of EW and KE peptides. Blood samples were obtained from four donors; for each cytokine, ELISA was performed 2-4 times, with two parallel experimental or control samples for each experiment (experiments to assess the effects of peptides on LPS-stimulated cells were repeated four times, while additional experiments with unstimulated cells were performed two times). Using molecular docking, GGAG was found to be the best dsDNA sequence in the classical B-form for binding the EW dipeptide, while GCGC is the preferred dsDNA sequence in the curved nucleosomal form for the KE dipeptide. Cluster analysis revealed that potential target genes for the EW and KE peptides encode the AKT1 and AKT2 proteins involved in the development of the cytokine storm. The specific targets for the EW peptide are the ACE2 and CYSLTR1 genes, and specific target for the KE peptide is the CHUK gene. Protein products of the ACE2, CYSLTR1, and CHUK genes are functionally associated with IL-1ß, IL-6, TNF-α, IL-4, and IL-10 cytokines. An in vitro model of an inflammatory reaction demonstrated that Thymalin and EW and KE dipeptides reduced the synthesis of IL-1ß, IL-6, and TNF-α cytokines in human peripheral blood mononuclear cells by 1.4-6.0 times. The immunomodulatory effect of Thymalin under the inflammatory response conditions in COVID-19 is based on the potential ability of its active components, EW and KE dipeptides, to regulate protein synthesis involved in the development of the cytokine storm.

Neuroepigenetic Mechanisms of Action of Ultrashort Peptides in Alzheimer's Disease.

Epigenetic regulation of gene expression is necessary for maintaining higher-order cognitive functions (learning and memory). The current understanding of the role of epigenetics in the mechanism of Alzheimer's disease (AD) is focused on DNA methylation, chromatin remodeling, histone modifications, and regulation of non-coding RNAs. The pathogenetic links of this disease are the misfolding and aggregation of tau protein and amyloid peptides, mitochondrial dysfunction, oxidative stress, impaired energy metabolism, destruction of the blood-brain barrier, and neuroinflammation, all of which lead to impaired synaptic plasticity and memory loss. Ultrashort peptides are promising neuroprotective compounds with a broad spectrum of activity and without reported side effects. The main aim of this review is to analyze the possible epigenetic mechanisms of the neuroprotective action of ultrashort peptides in AD. The review highlights the role of short peptides in the AD pathophysiology. We formulate the hypothesis that peptide regulation of gene expression can be mediated by the interaction of short peptides with histone proteins, cis- and transregulatory DNA elements and effector molecules (DNA/RNA-binding proteins and non-coding RNA). The development of therapeutic agents based on ultrashort peptides may offer a promising addition to the multifunctional treatment of AD.

Transport of Biologically Active Ultrashort Peptides Using POT and LAT Carriers.

Ultrashort peptides (USPs), consisting of 2-7 amino-acid residues, are a group of signaling molecules that regulate gene expression and protein synthesis under normal conditions in various diseases and ageing. USPs serve as a basis for the development of drugs with a targeted mechanism of action. The purpose of this review is to systematize the available data on USP transport involving POT and LAT transporters in various organs and tissues under normal, pathological and ageing conditions. The carriers of the POT family (PEPT1, PEPT2, PHT1, PHT2) transport predominantly di- and tripeptides into the cell. Methods of molecular modeling and physicochemistry have demonstrated the ability of LAT1 to transfer not only amino acids but also some di- and tripeptides into the cell and out of it. LAT1 and 2 are involved in the regulation of the antioxidant, endocrine, immune and nervous systems' functions. Analysis of the above data allows us to conclude that, depending on their structure, di- and tripeptides can be transported into the cells of various tissues by POT and LAT transporters. This mechanism is likely to underlie the tissue specificity of peptides, their geroprotective action and effectiveness in the case of neuroimmunoendocrine system disorders.

Peptides Regulating Proliferative Activity and Inflammatory Pathways in the Monocyte/Macrophage THP-1 Cell Line.

This study evaluates the effects of five different peptides, the Epitalon® tetrapeptide, the Vilon® dipeptide, the Thymogen® dipeptide, the Thymalin® peptide complex, and the Chonluten® tripeptide, as regulators of inflammatory and proliferative processes in the human monocytic THP-1, which is a human leukemia monocytic cell line capable of differentiating into macrophages by PMA in vitro. These peptides (Khavinson Peptides®), characterized by Prof. Khavinson from 1973 onwards, were initially isolated from animal tissues and found to be organ specific. We tested the capacity of the five peptides to influence cell cultures in vitro by incubating THP-1 cells with peptides at certain concentrations known for being effective on recipient cells in culture. We found that all five peptides can modulate key proliferative patterns, increasing tyrosine phosphorylation of mitogen-activated cytoplasmic kinases. In addition, the Chonluten tripeptide, derived from bronchial epithelial cells, inhibited in vitro tumor necrosis factor (TNF) production of monocytes exposed to pro-inflammatory bacterial lipopolysaccharide (LPS). The low TNF release by monocytes is linked to a documented mechanism of TNF tolerance, promoting attenuation of inflammatory action. Therefore, all peptides inhibited the expression of TNF and pro-inflammatory IL-6 cytokine stimulated by LPS on terminally differentiated THP-1 cells. Lastly, by incubating the THP1 cells, treated with the peptides, on a layer of activated endothelial cells (HUVECs activated by LPS), we observed a reduction in cell adhesion, a typical pro-inflammatory mechanism. Overall, the results suggest that the Khavinson Peptides® cooperate as natural inducers of TNF tolerance in monocyte, and act on macrophages as anti-inflammatory molecules during inflammatory and microbial-mediated activity.

Systematic search for structural motifs of peptide binding to double-stranded DNA.

A large variety of short biologically active peptides possesses antioxidant, antibacterial, antitumour, anti-ageing and anti-inflammatory activity, involved in the regulation of neuro-immuno-endocrine system functions, cell apoptosis, proliferation and differentiation. Therefore, the mechanisms of their biological activity are attracting increasing attention not only in modern molecular biology, biochemistry and biophysics, but also in pharmacology and medicine. In this work, we systematically analysed the ability of dipeptides (all possible combinations of the 20 standard amino acids) to bind all possible combinations of tetra-nucleotides in the central part of dsDNA in the classic B-form using molecular docking and molecular dynamics. The vast majority of the dipeptides were found to be unable to bind dsDNA. However, we were able to identify 57 low-energy dipeptide complexes with peptide-dsDNA possessing high selectivity for DNA binding. The analysis of the dsDNA complexes with dipeptides with free and blocked N- and C-terminus showed that selective peptide binding to dsDNA can increase dramatically with the peptide length.

Peptide Regulation of Gene Expression: A Systematic Review.

Peptides are characterized by their wide range of biological activity: they regulate functions of the endocrine, nervous, and immune systems. The mechanism of such action of peptides involves their ability to regulate gene expression and protein synthesis in plants, microorganisms, insects, birds, rodents, primates, and humans. Short peptides, consisting of 2-7 amino acid residues, can penetrate into the nuclei and nucleoli of cells and interact with the nucleosome, the histone proteins, and both single- and double-stranded DNA. DNA-peptide interactions, including sequence recognition in gene promoters, are important for template-directed synthetic reactions, replication, transcription, and reparation. Peptides can regulate the status of DNA methylation, which is an epigenetic mechanism for the activation or repression of genes in both the normal condition, as well as in cases of pathology and senescence. In this context, one can assume that short peptides were evolutionarily among the first signaling molecules that regulated the reactions of template-directed syntheses. This situation enhances the prospects of developing effective and safe immunoregulatory, neuroprotective, antimicrobial, antiviral, and other drugs based on short peptides.

Gene expression in human mesenchymal stem cell aging cultures: modulation by short peptides.

Effects of the short peptides Ala-Glu-Asp (AED), Lys-Glu-Asp (KED) and Lys-Glu (KE) on the expression of IGF1, FOXO1, TERT, TNKS2, and NFκB genes were studied in human embryo bone marrow mesenchymal stem cells (line FetMSCs) variously aged in "passages" or "stationary" cultures. Both cell aging models were similar in gene expression. The main difference was in the TERT gene expression level, which showed an eightfold increase at the "stationary" aging. IGF1 gene expression levels were very similar in both cell culture aging models, being enhanced by 3.5-5.6 fold upon the addition of the peptides. The FOXO1 gene was expressed twice more actively in the "stationary" than in the "passages" aging model. KED peptide inhibited FOXO1 gene expression by 1.6-2.3 fold. KE peptide increased FOXO1 gene expression by about two-fold in the "stationary" aging model but did not affect it in the "passage" aging model. The most striking difference in the peptide effect on cell aging between "passages" and "stationary" aging models was in the KED effects on TNKS2 gene expression; this expression was inhibited by KED in the "passages" model, while stimulation was observed in the "stationary" model. AED, KED, and KE stimulated expression of the NFκB gene in both models. Thus, the peptides studied at nanomolar concentrations modulate the expression of some genes known to be involved in cell aging.

EDR Peptide: Possible Mechanism of Gene Expression and Protein Synthesis Regulation Involved in the Pathogenesis of Alzheimer's Disease.

The EDR peptide (Glu-Asp-Arg) has been previously established to possess neuroprotective properties. It activates gene expression and synthesis of proteins, involved in maintaining the neuronal functional activity, and reduces the intensity of their apoptosis in in vitro and in vivo studies. The EDR peptide interferes with the elimination of dendritic spines in neuronal cultures obtained from mice with Alzheimer's (AD) and Huntington's diseases. The tripeptide promotes the activation of the antioxidant enzyme synthesis in the culture of cerebellum neurons in rats. The EDR peptide normalizes behavioral responses in animal studies and improves memory issues in elderly patients. The purpose of this review is to analyze the molecular and genetics aspects of the EDR peptide effect on gene expression and synthesis of proteins involved in the pathogenesis of AD. The EDR peptide is assumed to enter cells and bind to histone proteins and/or ribonucleic acids. Thus, the EDR peptide can change the activity of the MAPK/ERK signaling pathway, the synthesis of proapoptotic proteins (caspase-3, p53), proteins of the antioxidant system (SOD2, GPX1), transcription factors PPARA, PPARG, serotonin, calmodulin. The abovementioned signaling pathway and proteins are the components of pathogenesis in AD. The EDR peptide can be AD.

Peptides: Prospects for Use in the Treatment of COVID-19.

There is a vast practice of using antimalarial drugs, RAS inhibitors, serine protease inhibitors, inhibitors of the RNA-dependent RNA polymerase of the virus and immunosuppressants for the treatment of the severe form of COVID-19, which often occurs in patients with chronic diseases and older persons. Currently, the clinical efficacy of these drugs for COVID-19 has not been proven yet. Side effects of antimalarial drugs can worsen the condition of patients and increase the likelihood of death. Peptides, given their physiological mechanism of action, have virtually no side effects. Many of them are geroprotectors and can be used in patients with chronic diseases. Peptides may be able to prevent the development of the pathological process during COVID-19 by inhibiting SARS-CoV-2 virus proteins, thereby having immuno- and bronchoprotective effects on lung cells, and normalizing the state of the hemostasis system. Immunomodulators (RKDVY, EW, KE, AEDG), possessing a physiological mechanism of action at low concentrations, appear to be the most promising group among the peptides. They normalize the cytokines' synthesis and have an anti-inflammatory effect, thereby preventing the development of disseminated intravascular coagulation, acute respiratory distress syndrome and multiple organ failure.

AEDG Peptide (Epitalon) Stimulates Gene Expression and Protein Synthesis during Neurogenesis: Possible Epigenetic Mechanism.

It was shown that AEDG peptide (Ala-Glu-Asp-Gly, Epitalon) regulates the function of the pineal gland, the retina, and the brain. AEDG peptide increases longevity in animals and decreases experimental cancerogenesis. AEDG peptide induces neuronal cell differentiation in retinal and human periodontal ligament stem cells. The aim of the study was to investigate the influence of AEDG peptide on neurogenic differentiation gene expression and protein synthesis in human gingival mesenchymal stem cells, and to suggest the basis for the epigenetic mechanism of this process. AEDG peptide increased the synthesis of neurogenic differentiation markers: Nestin, GAP43, ß Tubulin III, Doublecortin in hGMSCs. AEDG peptide increased Nestin, GAP43, ß Tubulin III and Doublecortin mRNA expression by 1.6-1.8 times in hGMSCs. Molecular modelling method showed, that AEDG peptide preferably binds with H1/6 and H1/3 histones in His-Pro-Ser-Tyr-Met-Ala-His-Pro-Ala-Arg-Lys and Tyr-Arg-Lys-Thr-Gln sites, which interact with DNA. These results correspond to previous experimental data. AEDG peptide and histones H1/3, H1/6 binding may be one of the mechanisms which provides an increase of Nestin, GAP43, ß Tubulin III, and Doublecortin neuronal differentiation gene transcription. AEDG peptide can epigenetically regulate neuronal differentiation gene expression and protein synthesis in human stem cells.

"I think that the small peptides are the best for healthy ageing...", an interview with Vladimir Khavinson. Interview by Suresh I. S. Rattan.

Since its inception in 2000, Biogerontology has published interviews with some of the most renowned and intellectually influential biogerontologists, including Len Hayflick, Robin Holliday, Denham Harman, Vincent Cristofalo, Claudio Franceschi, Leslie Robert, Ken Kitani, Geroge Martin, Zhores Medvedev and John Maynard Smith. These interviews have explored the minds of these scientists in all aspects of their lives combining the private and the professional. Together, this series is a remarkable document providing an insight into the history of ideas in modern biogerontology. Here we present Vladimir Khavinson talking about his life and work in Russia during and after the Soviet times, his ideas on stress and health, his discoveries of the healthy ageing promoting small peptides, and other anti-ageing interventions.

Feasibility of Transport of 26 Biologically Active Ultrashort Peptides via LAT and PEPT Family Transporters.

The aim of this work is to verify the possibility of transport of 26 biologically active ultrashort peptides (USPs) into cells via LAT and PEPT family transporters. Molecular modeling and computer-assisted docking of peptide ligands revealed that the size and structure of ligand-binding sites of the amino acid transporters LAT1, LAT2, and of the peptide transporter PEPT1 are sufficient for the transport of the 26 biologically active di-, tri-, and tetra-peptides. Comparative analysis of the binding of all possible di- and tri-peptides (8400 compounds) at the binding sites of the LAT and PEPT family transporters has been carried out. The 26 biologically active USPs systematically showed higher binding scores to LAT1, LAT2, and PEPT1, as compared with di- and tri-peptides, for which no biological activity has been established. This indicates an important possible role which LAT and PEPT family transporters may play in a variety of biological activities of the 26 biologically active peptides under investigation in this study. Most of the 26 studied USPs were found to bind to the LAT1, LAT2, and PEPT1 transporters more efficiently than the known substrates or inhibitors of these transporters. Peptides ED, DS, DR, EDR, EDG, AEDR, AEDL, KEDP, and KEDG, and peptoids DS7 and KE17 with negatively charged Asp- or Glu- amino acid residues at the N-terminus and neutral or positively charged residues at the C-terminus of the peptide are found to be the most effective ligands of the transporters under investigation. It can be assumed that the antitumor effect of the KE, EW, EDG, and AEDG peptides could be associated with their ability to inhibit the LAT1, LAT2, and PEPT1 amino acid transporters. The data obtained lead to new prospects for further study of the mechanisms of transport of USP-based drugs into the cell and design of new antitumor drugs.

Senescence-Associated Secretory Phenotype of Cardiovascular System Cells and Inflammaging: Perspectives of Peptide Regulation.

A senescence-associated secretory phenotype (SASP) and a mild inflammatory response characteristic of senescent cells (inflammaging) form the conditions for the development of cardiovascular diseases: atherosclerosis, coronary heart disease, and myocardial infarction. The purpose of the review is to analyze the pool of signaling molecules that form SASP and inflammaging in cells of the cardiovascular system and to search for targets for the action of vasoprotective peptides. The SASP of cells of the cardiovascular system is characterized by a change in the synthesis of anti-proliferative proteins (p16, p19, p21, p38, p53), cytokines characteristic of inflammaging (IL-1α,ß, IL-4, IL-6, IL-8, IL-18, TNFα, TGFß1, NF-κB, MCP), matrix metalloproteinases, adhesion molecules, and sirtuins. It has been established that peptides are physiological regulators of body functions. Vasoprotective polypeptides (liraglutide, atrial natriuretic peptide, mimetics of relaxin, Ucn1, and adropin), KED tripeptide, and AEDR tetrapeptide regulate the synthesis of molecules involved in inflammaging and SASP-forming cells of the cardiovascular system. This indicates the prospects for the development of drugs based on peptides for the treatment of age-associated cardiovascular pathology.

Biogerontology in Russia: from past to future.

The paper presents major steps of gerontology development in Russia. The issues of training in gerontology and geriatrics, institutional infrastructure within the Gerontological Society of the Russian Academy of Sciences and its activities have been considered therein. Some results of Russian researchers obtained during 2005-2010 have been summarized as well. Special attention is given to the prospects of gerontology in Russia.

Homology between SARS CoV-2 and human proteins.

An extremely high contagiousness of SARS CoV-2 indicates that the virus developed the ability to deceive the innate immune system. The virus could have included in its outer protein domains some motifs that are structurally similar to those that the potential victim's immune system has learned to ignore. The similarity of the primary structures of the viral and human proteins can provoke an autoimmune process. Using an open-access protein database Uniprot, we have compared the SARS CoV-2 proteome with those of other organisms. In the SARS CoV-2 spike (S) protein molecule, we have localized more than two dozen hepta- and octamers homologous to human proteins. They are scattered along the entire length of the S protein molecule, while some of them fuse into sequences of considerable length. Except for one, all these n-mers project from the virus particle and therefore can be involved in providing mimicry and misleading the immune system. All hepta- and octamers of the envelope (E) protein, homologous to human proteins, are located in the viral transmembrane domain and form a 28-mer protein E14-41 VNSVLLFLAFVVFLLVTLAILTALRLCA. The involvement of the protein E in provoking an autoimmune response (after the destruction of the virus particle) seems to be highly likely. Some SARS CoV-2 nonstructural proteins may also be involved in this process, namely ORF3a, ORF7a, ORF7b, ORF8, and ORF9b. It is possible that ORF7b is involved in the dysfunction of olfactory receptors, and the S protein in the dysfunction of taste perception.

Neuroprotective Effects of Tripeptides-Epigenetic Regulators in Mouse Model of Alzheimer's Disease.

KED and EDR peptides prevent dendritic spines loss in amyloid synaptotoxicity in in vitro model of Alzheimer's disease (AD). The objective of this paper was to study epigenetic mechanisms of EDR and KED peptides' neuroprotective effects on neuroplasticity and dendritic spine morphology in an AD mouse model. Daily intraperitoneal administration of the KED peptide in 5xFAD mice from 2 to 4 months of age at a concentration of 400 µg/kg tended to increase neuroplasticity. KED and EDR peptides prevented dendritic spine loss in 5xFAD-M mice. Their action's possible molecular mechanisms were investigated by molecular modeling and docking of peptides in dsDNA, containing all possible combinations of hexanucleotide sequences. Similar DNA sequences were found in the lowest-energy complexes of the studied peptides with DNA in the classical B-form. EDR peptide has binding sites in the promoter region of CASP3, NES, GAP43, APOE, SOD2, PPARA, PPARG, GDX1 genes. Protein products of these genes are involved in AD pathogenesis. The neuroprotective effect of EDR and KED peptides in AD can be defined by their ability to prevent dendritic spine elimination and neuroplasticity impairments at the molecular epigenetic level.

Peptide bioregulation of aging: results and prospects.

The review comprises the results of author's long-term investigation in the mechanisms of aging and a role of peptide bioregulators in prevention of age-related pathology. A number of small peptides have been isolated from different organs and tissues and their analogues (di-, tri-, tetrapeptides) were synthesized from the amino acids. It was shown that long-term treatment with some peptide preparations increased mean life span by 20-40%, slow down the age-related changes in the biomarkers of aging and suppressed development of spontaneous and induced by chemical or radiation carcinogens tumorigenesis in rodents. Possible mechanisms of the biological effects of small peptides are discussed in the paper. The results of clinical applications of peptide preparation during the period of 6-12 years are presented as well.

Peptide Regulation of Cell Differentiation.

Short peptides are molecules with small molecular weight, capable of penetrating the cell membrane and nuclear membrane for epigenetic regulation of gene expression, including the genes responsible for cell differentiation. The direction of cell differentiation induction depends on the peptide structure and concentration. AEDG and AEDP peptides induce differentiation of pluripotent cells in the epidermis, mesenchyme and nervous tissue. Peptides KE, AED, KED, AEDG and AAAAEKAAAAEKAAAAEK activate neural differentiation. Peptides AEDL and KEDW induce lung and pancreatic cell differentiation. Differentiation of immune cells is stimulated by KE, DS, (Nα-(γ-E)-E), K(Н-E-OH)-OH, AED, KED, EDA, and KEDG peptides. IRW, GRGDS and YCWSQYLCY peptides activate osteogenic differentiation of stem cells. KE, AEDL, and AEDG peptides also induce plant cells differentiation. Short peptides can take part in activation of the signaling pathways regulating expression of differentiation genes. They can interact with histones changing the availability of genes for transcription, regulate gene methylation and activate or inhibit their expression, as well as directly interact with the DNA. Research in the area of directed stem cell differentiation by peptide regulation is of special importance for developing innovative approaches to molecular medicine and cell therapy.

Towards realization of longer life.

The current demographic situation in the world is characterized by an increase in average life expectancy, low birth rate, as well as an increase in the number of older and senior people, which is why our epoch is referred to as «the age of ageing¼. [...].