A method for analyzing the concentration of heat shock protein 70 kDa in sections of brain tissue of transgenic mice HSP70[IN] and HSP70[EX] by classifying raman spectra

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Abstract

Introduction. Heat shock proteins (HSPs) are a group of highly prospective compounds that can be used in various branches of clinical medicine, such as neurology, pharmacology and oncology. Studying the content of these proteins in various parts of the brain of transgenic mice that express increased concentrations of exo- and endogenous HSP will make it possible to clarify the knowledge about the content of these proteins and their distribution in the structures of the central nervous system.

The aim. To develop a method for identifying and analyzing the content of heat shock proteins weighing 70 kDa in sections of brain tissue of transgenic mice using the method of Raman spectroscopy.

Material and methods. To conduct the experiment, were selected 9 mice of the C57BL/6 line (n=3 per group), among them the first group consisted of wild-type mice, the second and third were transgenic mice of the line with an integrated genetic cassette containing copies of the human heat shock protein expressed in the cytosol and into the interstitial space. In this study, Raman spectra of brain sections were measured using a Renishaw inVia Qontor confocal Raman microscope (Renishaw, UK). Raman spectra were recorded from the primary motor cortex and striatum of the brain with an accumulation time of 5 s, the laser excitation wavelength was 633 nm.

Results. The obtained data underwent mathematical preprocessing, which included normalization, removal of outlines and baseline, dimensionality reduction, and noise filtering. There were trained 4 models for classifying Raman spectra, 2 of which showed high accuracy in differentiating spectra from brain sections of mice of the intact group and mice with overexpression of the 70 kDa heat shock protein.

Conclusion. The obtained results demonstrate the feasibility of an attitude to data processing and interpretation, and also allow us to judge the prospects of using the method of Raman spectroscopy to analyze the distribution of heat shock proteins in brain tissue.

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About the authors

Sviatoslav Alexeevich Temezhnikov

Sechenov First Moscow State Medical University (Sechenov University)

Author for correspondence.
Email: temezhnikov_s_a@staff.sechenov.ru
ORCID iD: 0009-0006-6681-392X

Laboratory assistant of the Department of Human Anatomy and Histology, N.V. Sklifosovsky institute of clinical medicine

Russian Federation, Trubetskaya str., 8/2, Moscow, 119048

Ekaterina Dmitrievna Belitskaya

Shemyakin-Ovchinnikov Institute of bioorganic chemistry of Russian Academy of Sciences

Email: belitskayakatya@yandex.ru
ORCID iD: 0009-0000-2456-567X

Engineer of the Laboratory of Molecular Biophysics

Russian Federation, Miklukho-Maklaya str., 10/16, Moscow, 117997

Vasilisa Yuryevna Makarenko

Shemyakin-Ovchinnikov Institute of bioorganic chemistry of Russian Academy of Sciences

Email: vasilisa.makarienko0901@gmail.com
ORCID iD: 0009-0001-0265-7714

Junior researcher of the laboratory of quantum oscillators

Russian Federation, Miklukho-Maklaya str., 10/16, Moscow, 117997

Igor Alexandrovich Dubinkin

National Research Nuclear University MEPhI (Moscow Engineering Physics Institute)

Email: konqstorm@gmail.com
ORCID iD: 0009-0001-8852-8330

student

Russian Federation, Kashirskoe shosse, 31, Moscow, 115409

Egor Alexandrovich Kuzmin

Sechenov First Moscow State Medical University (Sechenov University)

Email: kuzmin_e_a@staff.sechenov.ru
ORCID iD: 0000-0003-4098-1125

Trainee-researcher of the Department of Human Anatomy and Histology, N.V. Sklifosovsky institute of clinical medicine

Russian Federation, Trubetskaya str., 8/2, Moscow, 119048

Ksenia Sergeevna Pokidova

Sechenov First Moscow State Medical University (Sechenov University)

Email: pokidova.ks@gmail.com
ORCID iD: 0009-0000-0323-7669
SPIN-code: 4016-5570

Student of the clinical institute of children’s health named after N.F. Filatov

Russian Federation, Trubetskaya str., 8/2, Moscow, 119048

Gennadii Alexandrovich Piavchenko

Sechenov First Moscow State Medical University (Sechenov University)

Email: gennadii.piavchenko@staff.sechenov.ru
ORCID iD: 0000-0001-7782-3468
SPIN-code: 6120-1057

Associate professor of the department of human anatomy and histology N.V. Sklifosovsky institute of clinical medicine, PhD (Medicine); MD (Neurology), MSc (Pedagogics)

Russian Federation, Trubetskaya str., 8/2, Moscow, 119048

Anton Vladlenovich Zalygin

Р.N. Lebedev Physical Institute of Russian Academy of Sciences

Email: zalygin.anton@gmail.com
ORCID iD: 0000-0003-0132-0674
SPIN-code: 1990-1880

Highly qualified senior researcher, PhD (Physics and Mathematics)

Russian Federation, Leninsky av., 53, Moscow, 119991

Sergey Lvovich Kuznetsov

Sechenov First Moscow State Medical University (Sechenov University)

Email: kuznetsov_s_l@staff.sechenov.ru
ORCID iD: 0000-0002-0704-1660
SPIN-code: 3824-2646

Professor of the department of human anatomy and histology N.V. Sklifosovsky institute of clinical medicine, PhD (Medicine), professor, Corresponding member of the RAS

Russian Federation, Trubetskaya str., 8/2, Moscow, 119048

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Supplementary files

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2. Fig. 1. Normal spectra (left) and spectra containing emissions (right)

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3. Fig. 2. Plotting the baseline. Mean spectra (blue), polynominal curve approximating the baseline (orange)

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4. Fig. 3. Spectra after baseline removal

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5. Fig. 4. Spatial data of two main components, selected using PCA method. Data answering spectra of the control group (blue), “endo” group (orange), “exo” group (green)

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6. Fig. 5. Smoothing application. Reference spectra (blue), spectra after Savitsky – Golay filter application (orange)

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