Role of combinations of polymorphisms of cytokine genes in the diagnosis of Legg–Calve–Perthes disease in children

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Abstract

Background. Legg–Calve–Perthes disease (LCPD) is an idiopathic avascular femoral head osteonecrosis. The early disease stage is associated with the development of synovitis of the hip joint linked to the overproduction of factors induced by hypoxia as well as of interleukin (IL)-6. Associations of individual polymorphic variants of cytokine genes with LCPD have been shown. Moreover, alterations in the cytokine regulatory cascade are considered an important link in the pathogenesis of synovial inflammation in the early stages of LCPD. Accordingly, this process may be associated with a certain combination of polymorphic variants of the genes for pro-inflammatory and anti-inflammatory cytokines.

Aim. This study aimed to study the associations of polymorphic variants of the genes for pro-inflammatory and anti-inflammatory IL with LCPD.

Materials and methods. In this case–control study, the main and control groups were composed of 26 children with LCPD and 40 healthy children (all aged 3–11 years), respectively. Genotyping of IL10 (rs1800896), IL13 (rs20541), IL18 (rs187238), IL18 (rs5744292), IL1a (rs1800587), IL1RA (POL_GF_58), IL-1Ra (rs4251961), IL1B (rs16944), IL1B (rs1143634), IL4 (POL_GF_59), IL4 (rs2243250), IL6 (rs1800796), IL6 (rs1800795), INFγ (rs2430561), TGFβ (rs1800469), and TNF (rs1800629) was performed by polymerase chain reaction (PCR) using TaqMan probes to the corresponding polymorphic variants of genes produced by Thermo Fisher Scientific (USA) on an amplifier ViiATM 7 RealTime PCR System (Life Technologies, USA). Statistical processing of the results was carried out using the SNPstats program and multifactor dimensionality reduction.

Results. The study revealed three separate LCPD-potentiating genotypes of polymorphic variants of cytokine genes: IL10 (rs1800896; T>C)*T/C (OR 6.50), IL4 (POL_GF_49, VNTR, Intron4)*2R/2R (OR 12.32), and IL-6 (rs1800796; G>C)*G/C (OR 4.08). Two polymorphic variants of the IL4 gene (POL_GF_49, VNTR, Intron4, and rs2243250; C>T) had a pronounced synergism with respect to the diagnosis of LCPD. Moderate synergy with respect to the diagnosis of LCPD demonstrated the intergenic interaction of IL6 (rs1800796, G>C) with tumor necrosis factor-α (rs1800629, G>A). Moderate antagonism between LCPD and intergenic interactions was obtained for polymorphic variants of IL18 (rs5744292, T>C) and transforming growth factor-β (rs1800469, A>G) genes.

Conclusions. The pathogenesis of synovitis and subsequent osteonecrosis in LCPD is associated with a combination of polymorphic variants of the genes of pro-inflammatory and anti-inflammatory cytokines, as well as of DNA variants of the pro-allergic IL4 gene.

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About the authors

Nikita A. Shabaldin

Kemerovo State Medical University of the Ministry of Health of the Russian Federation; Kuzbass Regional Children’s Clinical Hospital

Author for correspondence.
Email: Shabaldin.nk@yandex.ru
ORCID iD: 0000-0001-8628-5649

MD, PhD, Associate Professor of the Department of Child Surgical Diseases

Russian Federation, Kemerovo

Andrey V. Shabaldin

Kemerovo State Medical University of the Ministry of Health of the Russian Federation; Institute for Complex Issues of Cardiovascular Diseases

Email: weit2007@yandex.ru
ORCID iD: 0000-0002-8785-7896

MD, PhD, D.Sc., Associate Professor, Professor of the Department of Microbiology, Immunology and Virology

Russian Federation, Kemerovo

Svetlana V. Apalko

City Hospital No. 40 of the Kurortny District

Email: svetlana.apalko@gmail.com
ORCID iD: 0000-0002-3853-4185

PhD in biology, Head of the Biobanking and Translational Medicine Sector

Russian Federation, Saint Petersburg

Anna V. Tsepokina

Institute for Complex Issues of Cardiovascular Diseases

Email: cepoav1991@gmail.com
ORCID iD: 0000-0002-4467-8732

MD, Junior Researcher of the Laboratory of Genomic Medicine

Russian Federation, Kemerovo

Yuri I. Rovda

Kemerovo State Medical University of the Ministry of Health of the Russian Federation

Email: y.i.rovda@rambler.ru
ORCID iD: 0000-0001-8310-5868

MD, PhD, D.Sc., Professor, Professor of the Department of Pediatrics and Neonatology

Russian Federation, Kemerovo

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2. Diagram (a) and diagram (b) Fryuchterman–Reingold for intergenic interactions of polymorphic variants of cytokine genes using MDR. The direction of interactions between candidate genes in the formation of the phenotype is indicated by lines of different colors: red = pronounced synergism, orange = moderate synergism, blue = pronounced antagonism, green = moderate antagonism, light brown = additive interaction. The strength and direction of the interactions are presented as a percentage of entropy

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Copyright (c) 2021 Shabaldin N.A., Shabaldin A.V., Apalko S.V., Tsepokina A.V., Rovda Y.I.

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