Laboratory of Nanobiotechnology, Institute of Biomedical Chemistry, Pogodinskaya St. 10/8, 119121 Moscow, Russia.
Laboratory of Shock Wave Impacts, Joint Institute for High Temperatures of the Russian Academy of Sciences, Izhorskaya St. 13 Bd.2, 125412 Moscow, Russia.
Molecules. 2021 Oct 2;26(19):5979. doi: 10.3390/molecules26195979.
MicroRNAs, which circulate in blood, are characterized by high diagnostic value; in biomedical research, they can be considered as candidate markers of various diseases. Mature microRNAs of glial cells and neurons can cross the blood-brain barrier and can be detected in the serum of patients with autism spectrum disorders (ASD) as components of macrovesicles, macromolecular protein and low-density lipoprotein particles. In our present study, we have proposed an approach, in which microRNAs in protein complexes can be concentrated on the surface of AFM chips with oligonucleotide molecular probes, specific against the target microRNAs. MicroRNAs, associated with the development of ASD in children, were selected as targets. The chips with immobilized molecular probes were incubated in serum samples of ASD patients and healthy volunteers. By atomic force microscopy (AFM), objects on the AFM chip surface have been revealed after incubation in the serum samples. The height of these objects amounted to 10 nm and 6 nm in the case of samples of ASD patients and healthy volunteers, respectively. MALDI-TOF-MS analysis of protein components on the chip surface allowed us to identify several cell proteins. These proteins are involved in the binding of nucleic acids (GBG10, RT24, RALYL), in the organization of proteasomes and nucleosomes (PSA4, NP1L4), and participate in the functioning of the channel of active potassium transport (KCNE5, KCNV2).
微小 RNA 存在于血液中，具有很高的诊断价值；在生物医学研究中，它们可以被视为各种疾病的候选标志物。神经胶质细胞和神经元的成熟微小 RNA 可以穿过血脑屏障，并作为大囊泡、大分子蛋白质和低密度脂蛋白颗粒的成分，在自闭症谱系障碍 (ASD) 患者的血清中被检测到。在我们目前的研究中，我们提出了一种方法，即用针对靶微小 RNA 的寡核苷酸分子探针将蛋白质复合物中的微小 RNA 集中在 AFM 芯片的表面上。选择与儿童 ASD 发展相关的微小 RNA 作为靶标。将固定有分子探针的芯片在 ASD 患者和健康志愿者的血清样本中孵育。通过原子力显微镜 (AFM)，孵育后可以在 AFM 芯片表面上显示出物体。在 ASD 患者和健康志愿者的样本中，这些物体的高度分别为 10nm 和 6nm。芯片表面蛋白质成分的 MALDI-TOF-MS 分析使我们能够鉴定出几种细胞蛋白质。这些蛋白质参与核酸结合（GBG10、RT24、RALYL）、蛋白酶体和核小体的组织（PSA4、NP1L4），并参与活性钾转运通道的功能（KCNE5、KCNV2）。
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