Laboratory of Neurobiology, Nencki Institute of Experimental Biology PAS, 02-093 Warsaw, Poland.
Department of Electrode Processes, Institute of Physical Chemistry, Polish Academy of Sciences, Kasprzaka 44/52, 01-224 Warsaw, Poland.
Int J Mol Sci. 2021 Jun 4;22(11):6086. doi: 10.3390/ijms22116086.
Ischemic stroke is a disturbance in cerebral blood flow caused by brain tissue ischemia and hypoxia. We optimized a multifactorial in vitro model of acute ischemic stroke using rat primary neural cultures. This model was exploited to investigate the pro-viable activity of cell-penetrating peptides: arginine-rich Tat(49-57)-NH (RKKRRQRRR-amide) and its less basic analogue, PTD4 (YARAAARQARA-amide). Our model included glucose deprivation, oxidative stress, lactic acidosis, and excitotoxicity. Neurotoxicity of these peptides was excluded below a concentration of 50 μm, and PTD4-induced pro-survival was more pronounced. Circular dichroism spectroscopy and molecular dynamics (MD) calculations proved potential contribution of the peptide conformational properties to neuroprotection: in MD, Tat(49-57)-NH adopted a random coil and polyproline type II helical structure, whereas PTD4 adopted a helical structure. In an aqueous environment, the peptides mostly adopted a random coil conformation (PTD4) or a polyproline type II helical (Tat(49-57)-NH) structure. In 30% TFE, PTD4 showed a tendency to adopt a helical structure. Overall, the pro-viable activity of PTD4 was not correlated with the arginine content but rather with the peptide's ability to adopt a helical structure in the membrane-mimicking environment, which enhances its cell membrane permeability. PTD4 may act as a leader sequence in novel drugs for the treatment of acute ischemic stroke.
缺血性中风是由于脑组织缺血和缺氧引起的脑血流紊乱。我们使用大鼠原代神经培养物优化了一种多因素体外急性缺血性中风模型。该模型用于研究穿膜肽的促存活活性：富含精氨酸的 Tat(49-57)-NH(RKKRRQRRR-酰胺)及其碱性较弱的类似物 PTD4(YARAAARQARA-酰胺)。我们的模型包括葡萄糖剥夺、氧化应激、乳酸酸中毒和兴奋性毒性。这些肽的神经毒性在低于 50μm 的浓度下被排除在外，并且 PTD4 诱导的促存活作用更为明显。圆二色性光谱和分子动力学 (MD) 计算证明了肽构象特性对神经保护的潜在贡献：在 MD 中，Tat(49-57)-NH 采用无规卷曲和多脯氨酸 II 型螺旋结构，而 PTD4 采用螺旋结构。在水相环境中，肽大多采用无规卷曲构象 (PTD4) 或多脯氨酸 II 型螺旋 (Tat(49-57)-NH) 结构。在 30%TFE 中，PTD4 表现出采用螺旋结构的趋势。总体而言，PTD4 的促存活活性与精氨酸含量无关，而与肽在模拟膜环境中采用螺旋结构的能力有关，这增强了其细胞膜通透性。PTD4 可能作为治疗急性缺血性中风的新型药物的先导序列。
