Heat shock protein 70 gene polymorphisms in Iranian patients with Multiple sclerosis.

Am J Clin Exp Immunol 2024;13(6):278-284 www.ajcei.us /ISSN:2164-7712/AJCEI0161327 Original Article Heat shock protein 70 gene polymorphisms in Iranian patients with Multiple sclerosis Azam Bakhshandeh, Alireza Kargar Dolatabadi, Touraj Farazmanfar, Majid Shahbazi Medical Cellular and Molecular Research Center, Golestan University of Medical Sciences, Gorgan, Iran Received October 20, 2024; Accepted December 11, 2024; Epub December 25, 2024; Published December 30, 2024 Abstract: Genetic factors are effective reagents in susceptibility to multiple sclerosis (MS). Previous studies have shown the relationship between heat shock protein (HSP) gene polymorphisms. So, HSP70 single nucleotide polymorphisms (SNPs) were evaluated as MS risk factors. Here, DNA genotyping was done for HSP70 gene polymorphisms, including HSP70-1 +190 G>C, HSP70-1 -110 A>C, HSP70-1 +438 A>C, and HSP70-hom +2437 A>G in two groups including Iranian MS patients and controls. A standard phenol/chloroform method isolated DNA samples from peripheral blood. Sequence-specific amplification (SSP) polymerase chain reaction (PCR) was used for genotyping polymorphisms. Overall, 76 (35.80%) MS patients and 136 (65.10%) controls were studied with an age mean of 36.0 ± 8.0 years. Female/male was significantly higher in patients than in controls (4.43 vs. 0.10, P < 0.001). The average age was significantly lower in patients (P < 0.001). The most common clinical feature was relapsingremitting (RR) MS; more than half of the population was Fars. Results showed that genotypes of HSP70-hom +2437 C>T had a significant relation with MS (OR = 2.0, 95% CI = 1.0-5.0, P = 0.03) and the same applies to HSP70-1 -110 A>C (OR = 0.0, 95% CI = 0.0-1.0, P < 0.001). Allele and genotype frequency of two other HSP70 SNPs (HSP701 +190 G>C, HSP70-1 +438 A>C) showed no significant differences between patients and controls. HSP70-hom +2437 C>T and HSP70-1 -110 A>C can be considered as risk factors for MS in our population. However, other HSP SNPs should be studied in a larger population in the future. Keywords: HSP70-1, DNA genotyping, relapsing-remitting multiple sclerosis Introduction Multiple sclerosis (MS) is an autoimmune demyelinating disease with inflammation and myelin sheath damage in the central nervous system [1]. Globally, the prevalence of the disease is about 2.5 million people [2]. Iran is a medium prevalence country (with 5 to 30 MS cases per 100000 population) [3]. The evidence reported a higher prevalence of MS in the northern hemisphere than in the southern [4]. The most common age of this disease is 20-40 years old, and its prevalence in women is twice that of men [5]. The main reason for this disease has yet to be determined. A series of risk factors affect the disease and its occurrences, such as immune system failure, genetic predisposition, family background, geographic region, viral infections, and psychological pressures [6]. Like other chronic inflammatory disorders, manifestations of MS are various, from benign disease to rapidly developing and debili- tating disease. MS may affect many organs and systems of the body and causes disability, reduction of the life quality of patients, and death [7, 8]. Sclerotic plaques are the essential symptom of MS that occurs in an inflammatory process, demyelination, and repair with the loss of different axons [9]. Genetic and environmental factors are effective reagents in MS. HSPs are a group of proteins expressed under stress conditions, such as free radicals [10]. As oxidative stresses affect MS pathogenesis and progression, HSPs are involved in this disease [11]. HSPs act as housekeeping genes in physiological conditions and as molecular chaperones. These proteins play roles in the suitable folding of newly synthesized proteins; the functions of HSPs are essential in preventing protein accumulation, weakening unstable and incompatible proteins, and transferring proteins in cell parts [12-16]. High expression of HSPs in the second autoinduchttps://doi.org/10.62347/CMYA9839 HSP-70 gene polymorphism and multiple sclerosis tion, such as heat shock, promotes the development of cell survival function [15]. These proteins consist of HSP100, HSP90, HSP70, HSP60, and HSP40 families and small families [17]. According to previous studies, HSP plays sone roles in neurodegenerative diseases. Especially, HSP70 expression has been observed in CNS that is a powerful antiapoptotic protein by inhibition of programmed cell death signaling. Functions of HSP70 are as intracellular (acting as a chaperone protein) and extracellular (triggering immunological responses) models. The functions of HSP70 in MS pathogenesis are complex. Some evidences reported HSP70 is overexpressed in the CNS of MS patients that is related to the neuroprotective function of HSP70 in an inflammatory environment. On the other hand, high quantities of HSP70 are released into the milieu, when overexpression of HSP70 fails to prevent cell death and promote the activation of immune system mediated by its cytokinelike property [18]. The cytoprotective and immunomodulatory functions of HSP70 can be affected by different polymorphisms of it that cause quantitative and qualitative changes in HSP70 expression [11]. Some studies have shown the relationship between HSP70 gene polymorphisms and different polymorphisms studied in MS, including HSP70-2 1267 A/G, HSP70-hom 2437 T/C, promotor region, and 190 G/C polymorphism of HSP70-1 [17]. This study survived the relationship between four HSP70 SNP gene polymorphisms and MS using the polymerase chain reaction (PCR) method. Materials and methods Patients Iranian patients with the diagnosis of MS from Golestan and Tehran provinces, Iran, were recruited in this study. A neurologist precisely diagnosed MS. The Control group included healthy blood donor volunteers from Blood Donation Centers of Gorgan and Gonbad, in the north of Iran, aged 23-62 years. Age, sex, and ethnicity matched control subjects who were randomly selected and had no autoimmune or inflammatory diseases. A demographic questionnaire was prepared for MS patients. The demographic data included sex, age, onset, 279 and ethnicity. Informed Consent forms were completed and signed by all individuals. The exclusion criteria were lack of participant satisfaction or underlying disease. The ethics committee of Golestan University of Medical Sciences, Gorgan, Iran (IR.GOUMS.REC.1397.196) approved this study. All individuals completed and signed informed consent forms. DNA isolation Peripheral blood samples were collected in 5-mL coated tubes from MS patients and controls. DNA was isolated by a standard phenol/ chloroform method [19] with some modifications. Red blood cells were briefly lysed three times with a buffer (ammonium chloride, potassium dihydrogen phosphate, and disodium hydrogen phosphates). The pellets were resuspended with SDS (10%) (Merck, Germany), EDTA (Merck, Germany), and 10 μL proteinase K (QIAGEN, Germ (...truncated)