[The Peptide Drug ACTH(4-7)PGP (Semax) Suppresses mRNA Transcripts Encoding Proinflammatory Mediators Induced by Reversible Ischemia of the Rat Brain].

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.

[Effects of ischemia on the expression of neurotrophins and their receptors in rat brain structures outside the lesion site, including on the opposite hemisphere].

Neurotrophins stimulate the regeneration of neural tissue after lesions. It is also known that the sources of neurogenesis and cerebral function recovery are predominantly located in subcortical brain structures. The effects of ischemia on the expression of genes that encode neurotrophins (Bdnf, Ngf, Nt-3) and their receptors (TrkB, TrkA, TrkC, p75) in brain structures outside the lesion site were studied 3, 24, and 72 h after irreversible unilateral occlusion of the middle cerebral artery in rats. Changes in the mRNA expression of these genes were assessed by relative quantification using real-time RT-PCR. Sham surgery was found to stimulate the expression of genes that encode neurotrophins (Bdnf, Ngf) and their receptor (p75). It has been shown that ischemia influenced the expression of neurotrophins (Bdnf, Ngf, Nt-3) and their receptors (TrkB, TrkA, TrkC, p75) in brain structures outside the lesion focus, including the contralateral hemisphere. The downregulation of Bdnf and TrkB transcripts and Ngf and TrkA upregulation in the contralateral cortex on the first day of ischemia obviously reflected stress response. On day 3, Nt-3 transcription increased in all investigated structures outside the lesion focus. In the contralateral hemisphere, relative levels of TrkA and TrkC mRNA expression increased, while p75 expression decreased. Presumably, the observed changes in gene transcription serve to facilitate neuroplasticity and neural tissue regeneration.

[Effect of tripeptide Pro-Gly-Pro on rat brain transcriptome in focal ischemia].

Biologically active regulatory peptide, tripeptide Pro-Gly-Pro (PGP) was used as C-terminal fragment for peptide drugs Semax and Selank. In recent years the independent effects of PGP were observed. The question was raised, whether PGP contributes to the effects ofpeptide drugs containing PGP as a fragment. The genome-wide analysis was performed to investigate the influence of PGP on the transcriptome of ischemic rat brain cortex tissues. The gene expression alterations caused by the action of the tripeptide PGP were compared with the gene expression of the control group "ischemia" at 3 and 24 h after permanent occlusion of left middle cerebral artery. The altered expression was detected for 29 genes at 3 h and 57--at 24 h. The proteins encoded by these genes have variety of functions: cytokines, transport proteins, transcription factors, transmembrane receptors, etc. Biological processes, which are related to the genes with altered expression, were distinguished. The influence of PGP on the diversity of biological processes in different systems of the organism is demonstrated for the first time. The process "Immune response" was the most statistically notable at 24 h after occlusion. The expression of the immune system genes was predominately down regulated.

[Oligomeric organization of recombinant human neurotrophins expressed in Escherichia coli cells].

Genes of human neurotrophins NGF, BDNF, NT-3 were cloned, and the corresponding proteins and their fragments were expressed in Escherichia coli BL-21 (DE3lambda) cells. Their intracellular localization was determined. The conditions for isolation and purification of the target recombinant proteins and for folding of BDNF and NT-3 precursors were selected. The recombinant proprecursors of human neurotrophines have been shown to possess complex oligomeric structure.