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ORIGINAL ARTICLE
Year : 2014  |  Volume : 27  |  Issue : 3  |  Page : 589-593

The potential association of heat shock protein 70-2 + 1267 A/G gene polymorphism with nephropathy in type II diabetic patients


1 Department of Medical Biochemistry, Faculty of Medicine, Menoufia University, Menoufia, Egypt
2 Department of Internal Medicine, Faculty of Medicine, Menoufia University, Menoufia, Egypt

Date of Submission13-Jun-2013
Date of Acceptance13-Oct-2013
Date of Web Publication26-Nov-2014

Correspondence Address:
Mona Salah El-Din Mahmoud Habib
Shebein Elkom, Menoufia
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-2098.145521

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  Abstract 

Objective
The aim of the study was to assess the association between heat shock protein (HSP) 70-2 + 1267 A/G polymorphism and diabetic nephropathy (DN) in type II diabetes mellitus.
Background
DN is a life-threatening microvascular complication of type II diabetes. Oxidative stress plays a main role in its pathogenesis. As a consequence, a cellular adaptive response occurs requiring functional chaperones, so that the induction of HSPs is a maintained response in counteracting this oxidative stress. HSP70-2 + 1267 A/G and HSP70-hom + 2437 T/C polymorphisms may play an important role in susceptibility to and/or progression of DN.
Patients and methods
Sixty type II diabetic patients (30 patients with DN and 30 patients free of DN) and 20 healthy individuals, as the control group, were selected. Participants were genotyped for the HSP70-2 + 1267 A/G and HSP70-hom + 2437 T/C polymorphisms by PCR/restriction fragment length polymorphism.
Results
There was significant difference for HSP70-2 + 1267 A/G polymorphism. GG genotype and G allele were more frequent in the DN group versus other groups, whereas there was no significant difference in genotype and allele distributions among the three studied groups for the HSP70-hom polymorphism. On comparing diabetics with nephropathy versus diabetics without nephropathy, the odds ratio of the risk allele (G) of HSP70-2 + 1267 was 4.60 (95% confidence interval 2.09-10.12) and was 0.45 (95% confidence interval 0.15-1.42) for the risk allele (C) of HSP70-hom + 2437 T/C polymorphism.
Conclusion
HSP70 -2 + 1267 A/G polymorphism is associated with renal complications in type II diabetic patients and may be useful in identifying patients with increased risk for DN.

Keywords: diabetic nephropathy, gene polymorphism, heat shock proteins, oxidative stress


How to cite this article:
Ghanayem NM, Khamis SS, El-Shafie MK, El-Sattar El-Ghobashi YA, El-Shazly RA, El-Din Mahmoud Habib MS. The potential association of heat shock protein 70-2 + 1267 A/G gene polymorphism with nephropathy in type II diabetic patients. Menoufia Med J 2014;27:589-93

How to cite this URL:
Ghanayem NM, Khamis SS, El-Shafie MK, El-Sattar El-Ghobashi YA, El-Shazly RA, El-Din Mahmoud Habib MS. The potential association of heat shock protein 70-2 + 1267 A/G gene polymorphism with nephropathy in type II diabetic patients. Menoufia Med J [serial online] 2014 [cited 2020 Feb 24];27:589-93. Available from: http://www.mmj.eg.net/text.asp?2014/27/3/589/145521


  Introduction Top


Type II diabetes mellitus has become a global health problem [1] . Diabetic nephropathy (DN) is a life-threatening microvascular complication of type II diabetes mellitus [2] . It is the most common cause of end-stage renal disease [3],[4] . Oxidative stress plays a main role in the pathogenesis of DN. Hence, a cellular adaptive response occurs requiring functional chaperones so that the induction of heat shock proteins (HSPs) is a maintained response in counteracting this oxidative stress [5] . HSP70s are the major stress-inducible proteins promoting cell survival under stressful stimuli [3],[6] . In addition, they inhibit apoptosis [7] . Intracellular HSP70 delays the progression of chronic kidney disease through the antiapoptotic activity and cytoprotection [8] . The human inducible HSP70 protein is encoded by HSP70-1 and HSP70-2 located in the major histocompatibility complex class III region on chromosome 6 [9] , with an additional gene HSP70-hom located 4 kb upstream of HSP70-1 whose expression is constitutive [10] . Nucleotide position +1267 in the HSP70-2 gene is polymorphic and, although it lies in the coding region, it is a silent mutation [11] . Carriage of the G allele is associated with increased risks and poor outcomes in different diseases [12],[13],[14] . The T2437C, present in the coding region of HSP70-hom, could affect the stability and activity of this protein [15] . HSP70 gene polymorphisms were found to be risk factors in several human disorders [16] , and they may play an important role in susceptibility to and/or progression of DN [17] .


  Patients and methods Top


This study was carried out in Internal Medicine and Medical Biochemistry Departments, Menoufia University. It was carried out on 80 participants divided into three groups: group I included 30 diabetic patients with DN who had end-stage renal disease (duration of diabetes is more than 10 years); group II included 30 diabetic patients without DN (duration of diabetes is more than 10 years); and group III included 20 age-matched and sex-matched healthy controls.

All studied participants were submitted to measurement of the following: total cholesterol, serum urea and creatinine, ACR, fasting and 2 h postprandial blood glucose, HbA1c, and genotyping of HSP70-2 + 1267 A/G and HSP70-hom + 2437 T/C polymorphisms by PCR/restriction fragment length polymorphism.

Genotyping of HSP70 polymorphism by PCR/restriction fragment length polymorphism

A volume of 5 ml of blood was withdrawn for lymphocyte separation by lymphoflot solution (Biotest AG, Dreieich, Germany) followed by DNA extraction (QIAamp DNA Blood Mini Kit; Qiagen).

Genotyping of HSP70-2 + 1267 A/G polymorphism

A fragment of 1118 bp that includes the polymorphic site on position 1267 was amplified using the primers 5Ͳ-CATCGACTTCTACACGTCCA-3Ͳ (forward) and 5Ͳ-CAAAGTCCTTGAGTCCCAAC-3Ͳ (reverse). The products of PCR were cleaved with PstI and electrophoresed on 3% agarose. The DNA-lacking polymorphic PstI site within the HSP70-2 gene generates a product of 1118 bp after restriction (A allele), whereas the PstI site produced two fragments of 934 and 184 bp (G allele).

Genotyping HSP70-2 + 2437 T/C polymorphism

A fragment of 627 bp that includes the polymorphic site on position 2437 was amplified using the primers 5Ͳ-GGACAAGTCTGAGAAGGTACAG-3Ͳ (forward) and 5Ͳ-GTAACTTAGATTCAGGTCTGG-3Ͳ (reverse) and subjected to restriction with NcoI. The uncut fragment was 627 bp (C allele) and digestion products were 354 and 273 bp (T allele).

Statistical analysis

Results were statistically analyzed by statistical SPSS program version 16. Two types of statistics were performed: descriptive statistics, for example, percentage, mean, and SD; and analytic statistics, for example, the χ2 -test test. A two-tailed Student's t-test was used to compare quantitative data. Statistical significance was considered significant at the P-value less than 0.05 value.


  Results Top


This study showed highly significant increase in number and percentage of AA genotype and A allele distribution in group II when compared with group I and also in group III when compared with group I, whereas there was no significant difference between group II and group III ([Table 1]). There was highly significant increase in number and percentage of AG and GG genotypes and G allele in group I when compared with group II and group III, whereas there was no significant difference in group II when compared with group III ([Table 1]). The study showed that G allele increased the risk for DN by 5.60-fold on comparing group I versus group III ([Table 2]), whereas on comparing group I versus group II, G allele increased the risk for DN by 4.60-fold ([Table 3] and [Table 4]). There were no significant differences in genotype and allele distributions of HSP70-hom + 2437 T/C polymorphism among the studied groups ([Table 4]).
Table 1: Statistical comparison of genotype and allele distribution of the HSP70-2 + 1267 A/G polymorphism among the three studied groups


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Table 2: Odds ratio of the HSP70-2 genotypes and alleles in diabetics with nephropathy (group I) and controls (group III)


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Table 3: Odds ratio of the HSP70-2 genotypes and alleles in diabetics with nephropathy (group I) and diabetics without nephropathy (group II)


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Table 4: Statistical comparison of genotype and allele distribution of the HSP70-hom + 2437 T/C polymorphism among the three studied groups


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  Discussion Top


Our results showed that the HSP70-2 gene variant at position + 1267, a silent change in the coding region, was associated with nephropathy in type II diabetic patients. This is consistent with other study [17] . Results about the effect of this polymorphism on the HSP70-2 mRNA levels are controversial [18] . Low levels of HSP70-2 mRNA expression are associated with G allele of the HSP70-2 + 1267 polymorphism, suggesting interindividual differences in HSP70 expression [9,19-21]. Tahara et al. [22] found that the AA genotype of HSP70-2 had the highest level of mRNA expression among the three genotypes, whereas the GG genotype showed the lowest, suggesting that the A allele has a more protective potential than the G allele. Buraczynska et al. [17] and Maugeri et al. [10] demonstrated that HSP70-2 polymorphism is not involved in causing DN but is in linkage disequilibrium with some neighboring gene on chromosome 6p. Buraczynska et al. [17] observed that HSP70-1-110 A/C and HSP70-2 + 1276 A/G are associated with DN. HSP70-1-110 A/C polymorphism alters regulation of the HSP70-1 gene, and hence its expression. In patients with GG genotype, a decrease of mRNA expression might result in impaired cell response to stress leading to the intracellular accumulation of denatured proteins that could determine susceptibility to disease. Data from the Environmental Genome Project showed that the HSP70-2+1267 A/G was in linkage disequilibrium with HSP70-2 + 39 C>T. HSP70-2 + 39 T allele reduces the HSP70 synthesis levels of normal-status cultured cells [23] . HSP70-2 + 1267 A/G polymorphism is associated with various kidney diseases including acute renal failure in preterm neonates [24] , increased prevalence of vesicoureteral reflux and recurrent urinary tract infections in children [25] , and worse allograft survival in adult kidney recipients [26] . Rusai et al. [7] speculated that HSP70-2+1267 A/G leading to decreased HSP70 expression might influence development of congenital malformations of the kidney and urinary tract by impairment of antiapoptotic mechanisms needed to regulate organogenesis and concluded that, both in adults and children, decreased expression of HSP70s is associated with a number of kidney diseases. For HSP70-hom + 2437 T/C polymorphism, our study showed that this polymorphism was not a significant risk factor for DN. This is in accordance with other study [17] ([Figure 1] and [Figure 2]).
Figure 1: Genotype distribution of HSP70-2 + 1267 A/G gene polymorphism among the three studied groups.

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Figure 2: Allelic distribution of HSP70-2 + 1267 A/G gene polymorphism among the three studied groups.

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  Conclusion Top


This study supports the association of HSP70-2 + 1267 A/G polymorphism with the susceptibility to DN in type II diabetic patients as evident by significant higher frequency of G allele and GG genotype in DN patients. This may be useful in identifying patients with increased risk for DN. There is no evidence for the association of HSP70-hom + 2437 T/C polymorphism with DN.


  Acknowledgements Top


Conflicts of interest

There are no conflicts of interest.

 
  References Top

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18.Gombos T, Förhécz Z, Pozsonyi Z, et al. Interaction of serum 70-kDa heat shock protein levels and HspA1B (+1267) gene polymorphism with disease severity in patients with chronic heart failure. Cell Stress Chaperones 2008; 13 :199-206.  Back to cited text no. 18
    
19.Pociot F, Ronningen KS, Nerup J. Polymorphic analysis of the human MHC-linked heat-shock protein 70 (HSP70-2) and HSP70-Hom genes in insulin-dependent diabetes mellitus (IDDM). Scand J Immunol 1993; 38 :491-495.  Back to cited text no. 19
    
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22.Tahara T, Shibata T, Arisawa T, et al. The BB genotype of heat-shock protein (HSP) 70-2 gene is associated with gastric pre-malignant condition in H. pylori-infected older patients. Anticancer Res 2009; 29 :3453-3458.  Back to cited text no. 22
    
23.Guo H, Deng Q, Wu C, et al. Variations in HSPA1B at 6p21.3 are associated with lung cancer risk and prognosis in Chinese populations. Cancer Res 2011; 71 :7576-7586.  Back to cited text no. 23
    
24.Fekete A, Treszl A, Tóth-Heyn P. Association between heat shock protein 72 gene polymorphism and acute renal failure in premature neonates. Pediatr Res 2003; 54 :452-455.  Back to cited text no. 24
    
25.Karoly E, Fekete A, Banki NF. Heat shock protein 72 (HSPA1B) gene polymorphism and toll-like receptor (TLR) 4 mutation are associated with increased risk of urinary tract infection in children. Pediatr Res 2007; 61 :371-374.  Back to cited text no. 25
    
26.Fekete A, Viklický O, Hubácek JA. Association between heat shock protein 70s and toll-like receptor polymorphisms with long-term renal allograft survival. Transpl Int 2006; 1:190-196.  Back to cited text no. 26
    


    Figures

  [Figure 1], [Figure 2]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4]


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