RU-2861434-C1 - METHOD FOR SIMULTANEOUS DETECTION OF RENAL CORPUSCLES AND ARTERIOLES IN KIDNEY

Abstract

FIELD: biotechnology. SUBSTANCE: invention relates to a method for the simultaneous detection of renal corpuscles and arterioles in the kidney. The method includes sequentially performing on paraffin sections of the kidney an immunohistochemical reaction with monoclonal antibodies of clone 1A4 against alpha-actin of smooth muscle cells, visualised using diaminobenzidine chromogen, followed by staining the preparations with a solution of alcian blue 8GX prepared in a 2% aqueous solution of acetic acid, and analysing the preparation using transmitted light microscopy. EFFECT: creating an effective, well-reproducible method for detecting renal corpuscles and arterioles on histological sections of the kidney using transmitted light microscopy. 1 cl, 1 dwg, 1 ex

Inventors

• Korzhevskii Dmitrii Eduardovich
• Podsumkova Iuliia Maksimovna
• Kirik Olga Viktorovna
• ALEKSEEVA OLGA SERGEEVNA

Dates

Publication Date

20260505

Application Date

20250730

Claims (1)

1. A method for the simultaneous detection of renal corpuscles and arterioles in the kidney, consisting of sequentially performing an immunohistochemical reaction on paraffin sections of the kidney with monoclonal antibodies of clone 1A4 at a concentration of 20±1 mg/l against alpha-actin of smooth muscle cells, visualized using a diaminobenzidine chromogen, followed by staining the preparations with a 1% solution of alcian blue 8GX, prepared in a 2% aqueous solution of acetic acid, and analyzing the preparation using microscopy in transmitted light, whereby the smooth muscle elements in the arteriole wall are stained brown, the basal membranes are stained bright blue-green, and the cells of the renal tubules and renal corpuscles are stained pale blue-green.

Description

FIELD OF TECHNOLOGY TO WHICH THE INVENTION RELATES The invention relates to medicine, namely to diagnostic histological studies, conducting histochemical reactions, during which sections of biological material are studied under a microscope on glass slides. LEVEL OF TECHNOLOGY The human kidney is a vital organ responsible for blood filtration, regulation of water-electrolyte balance and acid-base balance, hormone production, and more. The kidney is divided into a cortex and medulla, the structural and functional unit of which is the nephron. The nephron consists of the renal corpuscle (glomerulus) and a system of convoluted tubules consisting of the proximal section, the loop of Henle, and the distal section, which opens into the collecting duct. The renal corpuscle is responsible for blood filtration, followed by the reabsorption and secretion of ions, metabolic products, and other substances within the tubular system. The renal parenchyma is quite sensitive to changes and quickly responds to changes in blood supply, oxygen levels, toxic concentrations, pressure surges in the renal vessels, the deposition of immune complexes, etc. When these parameters change, as well as when there is a lack of blood or oxygen, the renal parenchyma is damaged, which can lead to deterioration of kidney function and the development of diseases associated with structural damage to the renal tissue. Such kidney diseases include infectious glomerulonephritis, glomerular fibrosis, acute and chronic pyelonephritis, diabetic nephropathy, tumors, cysts, and renal sclerosis of various origins. In focal segmental glomerulosclerosis (FSGS), a type of glomerulonephritis, the glomeruli of the kidneys are affected [Shabaka A, Tato Ribera A, Fernández-Juárez G. Focal Segmental Glomerulosclerosis: State-of-the-Art and Clinical Perspective. Nephron.2020;144(9):413-427. doi: 10.1159/000508099], leading to impaired blood filtration and systemic effects such as hypertension, edema, proteinuria, and an increased risk of renal failure. Arterial hypertension has a strong impact on many factors that determine the quality of kidney function, which in turn can lead to structural damage to the kidneys and the development of serious diseases (glomerulosclerosis, interstitial nephritis, etc.). The risk of developing such diseases increases in the presence of concomitant metabolic disorders [M.E. Statsenko, M.V. Derevyanchenko. Features of the structural and functional state of the kidneys and endothelium in patients with arterial hypertension combined with metabolic disorders, Nephrology. Vol. 21, No. 5 (2017) 14-21 https://doi.org/10.24884/1561-6274-2017-21-5-5-18; H.-J. Greene, A.K. Walli, E.F. Gröne. Arterial hypertension and hyperlipidemia as determinants of glomerulosclerosis. The Clinical Investigator. Clin Investig (1993) 71: 834-839]. To identify various pathologies in the kidneys, methods are needed that can be used to accurately and reliably visualize Changes in renal structure [L.V. Melnikova, E.V. Osipova. Kidney damage in essential arterial hypertension: pathogenetic foundations of early diagnosis. Arterial hypertension. 2019;25(1):6–13; Sweat F, Puchtler H, Sesta JJ. PAS--phosphomolybdic acid--Sirius supra blue FGL-CF. Stain for demonstration of glomerular fibrosis. Arch Pathol. 1968;86(1):33–39]. In order to study the cellular and molecular mechanisms of dysfunctions developing in the kidneys with arterial hypertension, it is important to simultaneously assess the state of the renal corpuscles, their components and blood vessels, primarily the arterial type, in the wall of which there are smooth muscle cells containing a characteristic form of actin - alpha-actin of smooth muscle cells. The use of alpha-actin as a marker is explained by its direct involvement in the development of some renal pathologies [Feng D, DuMontier C, Pollak MR. The role of alphaactinin-4 in human kidney disease. Cell Biosci. 2015 Aug 18;5:44. doi:10.1186/s13578-015-0036-8; Badid C, Desmoulière A, McGregor B, Costa AM, Fouque D, Hadj Aïssa A, Laville M. Interstitial alpha-smooth muscle actin: a prognostic marker in membranous nephropathy. Clin Nephrol. 1999 Oct;52(4):210–217; Batyushin M. M., Pasechnik D. G. Detection of vimentin, pancytokeratin, smooth muscle actin, E-catherin and antibodies to CD-10 markers of epithelial-mesenchymal transformation in chronic glomerulonephritis // Nephrology. 2014. , /Vol. 18, No. 5. P. 52-58]. In scientific practice, there are two methods for the simultaneous detection of renal glomeruli and blood vessels: the double immunofluorescence reaction and the PAS reaction [G.L. Snigur, A.V. Smirnov, M.V. Schmidt. The Importance of Modern Methods of Pathohistological Research in the Diagnosis of Kidney Diseases. Volgograd Scientific Medical Journal, 2014, Vol. 4, pp. 43-47]. However, the first method is very labor-intensive and expensive, and the PAS reaction only visualizes renal corpuscles well, while detection of arterioles is unreli