NFκB and Its Inhibitor IκB in Relation to Type 2 Diabetes and Its Microvascular and Atherosclerotic Complications - ScienceDirect

Human Immunology

Volume 67, Issue 9, September 2006, Pages 706-713

Human Immunology

https://doi.org/10.1016/j.humimm.2006.05.006 Get rights and content

Abstract

Nuclear factor κ B (NFκB) is an important transcription factor that together with its inhibitor (IκB) participates in the activation of genes involved in immune responses. We examined the CA repeat polymorphism of the NFKB1 gene (encoding for NFκB) and A/G point variation in the 3′UTR region of the nuclear factor kappa B inhibitor alpha (NFKBIA) gene (encoding for IκB) in Czech and German patients with type 2 diabetes. The sample consisted of 211 patients, both with and without kidney complications, and 159 controls. Additionally, 152 patients with systemic lupus erythematosus (SLE) were genotyped for NFKBIA polymorphism. We observed a significant increase in the homozygous AA genotype of the NFKBIA gene when compared with the control group (the highest value was in diabetics without diabetic nephropathy [p_c* = 0.0015, odds ratio = 3.59]). No differences were seen between the SLE and control groups. With regard to the polymorphism of the NFKB1 gene, we did not observe any significant differences between the groups. Since the AA genotype of the NFKBIA gene presents a risk for type 2 diabetes development but not for diabetic nephropathy alone, we believe that the NFκB gene polymorphism can influence the pathogenesis of diabetes mellitus and affect its complications. Negative findings relative to other inflammatory autoimmune diseases, such as SLE, suggest a specific relationship between NFκB and type 2 diabetes mellitus.

Introduction

Diabetic nephropathy (DN) is the major cause of chronic renal failure in patients with diabetes mellitus. Nearly 30% of both type 1 and type 2 diabetic patients develop DN independently of glycemic control. The fact that DN manifests in only a subset of diabetics, together with racial/ethnic differences in the prevalence and family clustering, demonstrates its genetic independence from diabetes mellitus [1]. The risk factors that have been identified for the development of DN in longitudinal and cross-sectional studies include: race, genetic susceptibility, hypertension, hyperglycemia, hyperfiltration, smoking, advanced age, male sex, and a high-protein diet [2].

Nuclear factor κ B (NFκB) is a transcription factor that has been shown to be involved in the regulation of many genes that encode mediators of the immune response, embryo and cell lineage development, cell apoptosis, inflammation, cell cycle, oncogenesis, viral replication, and a variety of autoimmune diseases. Because it is activated by a variety of stimuli, the activation of NFκB is thought to be part of the stress response. These activating stimuli include reactive oxygen species and advanced glycation end products, which are toxic products of nonenzymatic glycation caused by long-term hyperglycemia and oxidative stress. At the cellular level, NFκB is activated through phosphorylation of an inhibitor of NFκB (IκB). Phosphorylated IκB is released from NFκB/IκB complex, allowing the translocation of NFκB molecules into the nucleus. Once in the nucleus, they bind to the consensus sequence (5′-GGGACTTTCC-3′) of various genes, thereby activating their transcription [3, 4]. Recent studies have investigated the role of NFκB in the pathogenesis of various human diseases including neurologic disorders, immune deficiency, carcinogenesis, and atherogenesis. In addition, the possible link between NFκB and the development of insulin resistance and type 2 diabetes has also been suggested [3, 5, 6, 7, 8].

The NFκB transcription factor complex has two alternative DNA binding subunits, nuclear factor kappa B p 105 subunit (NFKB1) and NFKB2. The gene coding for NFKB1 is located on chromosome 4q23-q24 [9]. A polymorphic dinucleotide CA repeat, with 18 described alleles, has been identified close to the coding region of the human NFKB1 gene [10]. This polymorphism has recently been investigated for its role relative to increased susceptibility to type 1 diabetes mellitus (Kolostova et al., article in press) [11]. Encouraged by other studies that also suggest that an increased activation of NFκB is associated with the development of diabetic microvascular complications [12, 13], we examined the CA repeat polymorphism of the NFKB1 gene in relation to diabetic nephropathy.

The gene coding for IκB (NFKBIA) has been mapped to chromosome 14q13, and A/G point variation in the 3′UTR region of NFKBIA has been detected. We also examined single nucleotide polymorphism of the IκB gene, looking for its involvement in the induction or progression of diabetic microvascular complications in the kidney.

In both analyses, we compared the entire group of diabetic patients (both those with and those without renal complications) with healthy controls drawn from Czech and German populations.

In addition, we also tested NFKBIA polymorphism in a second disease, systemic lupus erythematosus (SLE), to confirm or refute a specific association between NFκB and diabetic complications or diabetes itself.

Section snippets

Subjects

The study of polymorphisms in the NFKBIA and NFKB1 genes involved 395 persons consisting of 246 diabetic patients and 159 control subjects. All subjects were of Caucasian descent and lived in either the Czech Republic or Germany.

The group of diabetic patients, most of whom were type 2 diabetics (n = 211), were subdivided into three groups based on their renal status. The first group of patients (n = 50) included persons with nondiabetic renal disease (NDRD). Diseases in this group included

Inhibitor of NFKBIA

Our study used PCR-based genotyping to investigate single-nucleotide polymorphism (A→G) in the 3′-UTR region of the NFKBIA gene in an attempt to access its possible role in the development or progression of DN in diabetic patients.

To determine whether this possible association is related to just nephropathy we compared diabetic patients with renal disease (n = 117) with those diabetic patients without renal disease (n = 78). We found no significant differences in allele or genotype frequencies

Discussion

In this study we performed genetic analyses of two genes encoding NFKB1 and its inhibitor (NFKBIA) in patients with type 2 diabetes mellitus and SLE. The patients came from three central-European Caucasian populations. More than 200 type 2 diabetic patients, having had diabetes for at least 15 years, were tested. The diabetic patients were divided into a group of patients without complications, a group of patients with diabetic microvascular (DN) complications, and a group with macrovascular

Acknowledgments

This study was financially supported by the Research Program of the Czech Ministry of Education and Youth: Identification code: MSM 0021620814: Prevention, diagnostics and therapy of diabetes mellitus, metabolic and endocrine damage of organism. D. Hohenadel received a travel grant from the Federal Ministry of Education and Research, Germany (Project number: CZE 03/014).

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Given the biological effects of NFKB1 and NFKBIA genes in the inflammatory and autoimmune processes, a SNP polymorphism (rs969) in the 3′UTR region of the NFKBIA gene may have a genetic effect on the development of various inflammatory disease associated with altered immune response. In studies with Type 2 diabetes and CD, NFKBIA 3′UTR A → G polymorphism has been associated with development of these diseases, resulting in Th1/Th2 imbalance as the pathogenesis (Romzova et al., 2006; Klein et al., 2004). Katarina et al. (2007) have found that there was no difference in A and G allele frequency of NFKBIA gene in Type 1 diabetes mellitus; however AA genotype of 3′UTR A → G polymorphism was a risk factor for latent autoimmune diabetes in adults.
Hashimoto thyroiditis (HT) is a chronic inflammatory autoimmune disease of thyroid gland affected by interaction of multiple genes and various cytokines. Variants in the genes coding for the NFKB and I_KB proteins can be potentially involved in the development of the inflammatory diseases. NFKB, a key transcription factor of the regulation of immune responses, is interesting candidate for association studies about autoimmune disorder.
The aim of the present study was to investigate the relationship between NFKB1 and NFKBIA (NFKB1 inhibitor gene) polymorphisms, and the risk of HT in a Turkish Population in the context of IL-6 serum levels which may contribute to susceptibility to the disease. We analyzed the distribution of NFKB1-94 ins/del ATTG and NFKBIA 3′UTR A → G polymorphisms using PCR-RFLP method and IL-6 serum levels using ELISA method in 120 HT patients and 190 healthy controls in Turkish population.
Although, there was no statistical significant difference in distribution of the genotypes and alleles of NFKB1-94 ins/del ATTG or NFKBIA 3′UTR A → G polymorphisms in patients and control subjects as single, ins/ins/GG combined genotype had protective effect on the disease when compared to ins/ins/AG combined genotype as combined genotypes of both polymorphisms. In addition to this finding, IL-6 serum levels in HT patients with del/del genotype were significantly higher than in patients with del/ins genotype (p < 0.001). According to the combined genotype analysis of NFKB1-94 ins/del ATTG and NFKBIA 3′UTR A → G polymorphisms, IL-6 levels were also higher in patients with del/del genotype when at least one G allele existing (p = 0.007).
Therefore, our findings suggest that the functional promoter NFKB1-94 ins/del ATTG polymorphism was significantly associated with population HT disease through acting by directly modulating IL-6 serum levels.
Emodin attenuates high glucose-induced TGF-β1 and fibronectin expression in mesangial cells through inhibition of NF-κB pathway
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NF-κB activation, known to be involved in development of renal diseases [24], plays a critical role in high glucose-induced TGF-β1 and FN production in mesangial cells [20,21]. Following the phosphorylation and degradation of IκB-α, NF-κB p65 translocates into the nucleus and binds to a specific DNA sequence to activate the transcription of target genes [25]. Our previous study found increased NF-κB staining in cytoplasm and nuclei and decreased protein level of IκB-α in the diabetic kidney, suggesting the activation of NF-κB pathway in diabetic kidney [26].
The activation of nuclear factor-κB (NF-κB) and the subsequent overexpression of its downstream targets transforming growth factor-β1 (TGF-β1) and fibronectin (FN) are among the hallmarks for the progressive diabetic nephropathy. Our previous studies demonstrated that emodin ameliorated renal injury and inhibited extracellular matrix accumulation in kidney and mesangial cells under diabetic condition. However, the molecular mechanism has not been fully elucidated. Here, we showed that emodin significantly attenuated high glucose-induced NF-κB nuclear translocation in mesangial cells. Interestingly, emodin also inhibited the DNA-binding activity and transcriptional activity of NF-κB. Furthermore, NF-κB-mediated TGF-β1 and FN expression was significantly decreased by emodin. These results demonstrated that emodin suppressed TGF-β1 and FN overexpression through inhibition of NF-κB activation, suggesting that emodin-mediated inhibition of the NF-κB pathway could protect against diabetic nephropathy.
-94 ins/del ATTG NFKB1 gene variant is associated with lower susceptibility to myocardial infarction
2011, Nutrition, Metabolism and Cardiovascular Diseases
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Interestingly, a recent study proved that D/D homozygote endothelial cells displayed a lower increase in eNOS protein levels than the I/I homozygote cells under unidirectional laminar shear stress [13]. Moreover, A/G point variation in the 3′UTR region of the NFKBIA gene has recently been implicated in type 2 diabetes and susceptibility to atherosclerosis [16]. Considering the known physiological relevance of NFкB in inflammation, a possible functional role of NFкB1 and NFкBIA gene variants on susceptibility to myocardial infarction (MI) cannot be ruled out.
An imbalance of Nuclear Factor Kappa B (NFкB) and Inhibitor Kappa B (IкB) is involved in various human diseases including atherogenesis. We aimed to evaluate the relationship between NFKB1 and NFKBIA polymorphism and susceptibility to myocardial infarction (MI).
Genotyping was performed for NFKB1 and NFKBIA gene variants in 253 subjects (86 patients affected by myocardial infarction and 167 control subjects). In 40 patients, biopsy specimens were taken from the left ventricle area of presumed ischemia for p50, p65 and IкBα quantification. The allele frequency and genotype distribution of NFKBIA gene polymorphism did not differ between MI and control group while control subjects had a higher D allele frequency of -94 ins/del ATTG NFKB1 polymorphism, compared to the MI group (P < 0.001; OR = 0.304; 95% CI = 0.177–0.522). Subjects carrying the D allele had significantly lower plasma fibrinogen and CRP (C-reactive protein) levels compared to no carriers (P < 0.05). Fibrinogen–genotype interaction was found to have a significant effect on susceptibility to myocardial infarction. Myocardial p50 (r = 0.627; P = 0.012) and p65 (r = 0.683; P = 0.005) levels significantly correlated with plasma fibrinogen levels while subjects carrying the D allele of the NFкB1 gene variant had lower myocardial p50 (P = 0.007) and p65 (P = 0.009) levels compared to no carriers.
-94 ins/del ATTG NFKB1 gene variant may contribute to lower MI susceptibility via the potential reduction of activated NFкB which in turn is related to plasma inflammatory marker reduction.
Association of IKBA gene polymorphisms with the development of asthma
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Studies have primarily shown a negative association of the four SNPs in the IKBA gene (rs2233407, rs2233408, rs696, and rs2273650) with psoriatic arthritis [19], rs1957106 with multiple sclerosis [20] and multiple myeloma [21], and rs1957106 and rs696 with invasive pneumococcal disease [22]. In contrast, the A/G point variation (rs2273650) in the 3′-UTR region has been associated with development of type 2 diabetes and Crohn's disease, resulting in Th1/Th2 imbalance as the pathogenesis [23,24]. Given the biologic effect of NF-κB and its inhibitor IKBA in airway inflammation, and positive linkage analysis [10], the polymorphism in the IKBA gene may have a genetic effect on the development of asthma.
Nuclear factor–κB (NF-κB) orchestrates the expression of genes responsible for airway inflammation and remodeling in asthma. The activity of NF-κB is tightly regulated by IKBA, which may be modulated by genetic polymorphisms of the IKBA gene. We investigated the association between asthma susceptibility and IKBA gene polymorphisms in a Korean population. Genotyping was performed in BA (bronchial asthma) and NC (normal control). We measured reverse transcription–polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay, and luciferase reporter assays, respectively. A −673A>T (rs2233407) was associated with asthma development in subjects with atopic asthma (odds ratio = 0.56, p = 0.004). The IKBA mRNA level was higher in B-cell lines with the rs2233407 TT genotype compared with those with the AA genotype (p = 0.024). The luciferase activity of the rs2233407 T genotype was higher than that of the A (p = 0.002). The cytoplasmic levels of total IKBA and IKBA [p-S32] were higher in B cell lines of the rs2233407 TT genotype than those of the AA (p = 0.016 and p = 0.036, respectively), whereas nuclear NF-κB activity in cells with the IKBA rs2233407 AA genotype was higher than in cells with the AA (p = 0.038). The IKBA rs2233407 A>T polymorphism may predispose individuals to the development of atopic asthma via regulation of IKBA gene expression at the transcriptional level.
The genetics of vascular complications in diabetes mellitus
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Effects of berberine on matrix accumulation and NF-kappa B signal pathway in alloxan-induced diabetic mice with renal injury
2010, European Journal of Pharmacology
One of the main pathological changes in diabetic nephropathy is the renal fibrosis, which includes glomerulosclerosis and tubulointerstitial fibrosis. In vivo and in vitro studies demonstrated that berberine could ameliorate renal dysfunction in diabetic rats with nephropathy and inhibit fibronectin expression in mesangial cells cultured under high glucose. However, the molecular mechanisms have not been fully elucidated. The purpose of the present study was to investigate the effects of berberine on the nuclear factor-kappa B (NF-κB) activation, intercellular adhesion molecule-1, transforming growth factor-beta1 and fibronectin protein expression in renal tissue from alloxan-induced diabetic mice with renal damage. The distribution of NF-κB p65 in glomerulus and the degradation of IκB-α in renal cortex were examined by immunohistochemistry and Western blot, respectively. The protein expression of intercellular adhesion molecule-1, transforming growth factor-beta 1 and fibronectin in renal cortex were also detected by Western blot. Our results revealed that in alloxan-induced diabetic mice, the nuclear staining of NF-κB p65 was increased in glomerulus, whereas renal IκB-α protein was significantly reduced. The protein levels of intercellular adhesion molecule-1, transforming growth factor-beta 1 and fibronectin were upregulated in kidney from diabetic mice. After berberine treatment, the immunostaining of NF-κB was decreased, and the reduced degradation of IκB-α level was partially restored. The protein levels of intercellular adhesion molecule-1, transforming growth factor-beta 1 and fibronectin were all downregulated by berberine compared with diabetic model group. In conclusion, the ameliorative effects of berberine on extracellular matrix accumulation might associate with its inhibitory function on NF-κB signal pathway.

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Copyright © 2006 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.