Objectives: The relationship between chronic kidney disease and cardiovascular disease is complex and bidirectional. This relationship may be partially linked to thrombophilic genetic anomalies that may pre-dispose to the progression of both diseases.
Materials and Methods: We analyzed blood samples from 102 Lebanese patients with end-stage renal disease and undergoing hemodialysis and 20 randomly selected healthy volunteers for frequencies of 12 cardiovascular disease gene mutations and traditional risk factors. The frequencies of these mutations were calculated and compared in both groups. We stratified patients by quartiles according to their mean score of genetic mutations and traditional risk factors, as well as their mean age at dialysis initiation. Correlation analyses were performed on the various patient groups.
Results: We observed a high frequency of mutations in patients on dialysis. Homozygous mutations (> 10% of patients) were observed in the PAI-1 (11%), MTHFR A1298C sequence variant (12.7%), and ACE genes (12%); in addition, the FXIII V34L and PAI-1 4G/5G genotypes were significantly associated with early dialysis initiation (P < .001 and P = .004, respectively). We observed a strong linear relationship between the different scores and age at dialysis initiation, with older patients exhibiting the highest genetic, traditional, and total scores versus those shown in the youngest patients (R2 = 0.72 and P < .001; R2 = 0.98 and P < .001; and R2 =0.96 and P < .001, respectively).
Conclusions: Our results revealed a polygenic throm-bophilic profile in our population of Lebanese patients with end-stage renal disease. This profile showed a strong association between early dialysis initiation and specific homozygous cardiovascular disease gene mutations. The cumulative load of these genetic and traditional risk factors may be partly responsible for the increased risk of cardiovascular disease and risk of progression to end-stage renal disease in patients with chronic kidney disease.
Key words : Cardiovascular disease, Chronic kidney disease, Genetic risk factors, Traditional risk factors
The relationship between chronic kidney disease (CKD) and cardiovascular disease (CVD) is complex, reciprocal, and bidirectional, and their combination leads to a vicious circle that worsens patient prognosis and leads to premature patient death.1-4 A large body of evidence has shown considerable increased risk of cardiovascular morbidity and mortality in patients with CKD. This increased risk can be partly explained by an increase in commonly shared traditional risk factors by both entities, including hypertension, diabetes mellitus, and metabolic syndrome.5 Furthermore, CKD alone is sometimes considered as an independent,6 if not the most important, risk factor for CVD.2 Similarly, coronary artery disease (CAD), mainly when severe, has been recently shown to be the most important independent risk factor for advanced CKD.1 This strong association between CKD and CVD has been lately described as the CKD-CVD continuum.7
The mechanisms explaining the association between kidney disease and cardiovascular events have been widely studied. The most important pathophysiologic factor is probably endothelial damage, which is considered to be one of the first steps in the development of atherosclerosis. Furthermore, as renal functions decline, proinflam-matory molecules accumulate, leading to production of oxidative stress and inflammatory cytokines responsible for endothelial damage, arterial stiffness, smooth muscle cell proliferation, and consequent cardiovascular events.5,7 Several studies have focused on nontraditional cardiac risk factors that can explain the pathogenesis of CAD in patients with CKD. These factors have included decreased hemoglobin levels, increased inflammation, oxidative stress, abnor-malities in bone and mineral metabolism, sympathetic overactivation, protein-energy wasting, calciphylaxis, and volume overload.3,8 These uremic environmental risk factors are highly prevalent and are strongly linked to the initiation and progression of vascular disease in patients with CKD.5 Reduced bioa-vailability of nitric oxide has also been shown to be a important factor involved in endothelial dysfunction.9
The relationship between genetics and renal disease development is evident. Genetic linkage analyses have identified chromosomal regions that are associated with renal disease. Genes located in such chromosomal regions may predispose a patient to CKD progression. Identification of these genes will lead to a better understanding of the pathogenesis of end-stage renal disease (ESRD).10 The CVD panel comprises an association of genes in which mutations resulted in increased cardiovascular morbidity and mortality via different mechanisms. Given the bidirectional association between CVD and CKD, these same genes may also be responsible for susceptibility to CKD.
The CVD panel includes the following genes: methylenetetrahydrofolate reductase (MTHFR A1298C and MTHFR C677T sequence variants), factor V Leiden, prothrombin II (PT2), apolipoprotein A1 (APOA1), lipoprotein(a), apolipoprotein B (APOB), β-fibrinogen, plasminogen activator inhibitor 1 (PAI1), angiotensin-converting enzyme (ACE), factor XIII (F8), apolipoprotein E (APOE), and membrane glycoprotein IIb and IIIa (GP2B/3A). Studies have linked anomalies affecting each of these genetic factors with increased risk of CVD.11-28
In this study, we investigated and compared the frequency of the CVD-associated genetic mutations in a sample of Lebanese patients with ESRD versus results shown in a healthy cohort. The frequency of each mutation was determined in the ESRD population as a whole and according to the underlying native kidney disease. We also assessed the potential link between the cumulative load of different CVD genetic anomalies and traditional risk factors (isolated or combined) and age of dialysis initiation, a marker of CKD progression, in the population as a whole and in the population stratified according to presence or absence of consanguinity status.
Materials and Methods
This multicenter cross-sectional study included 102 Lebanese patients with ESRD who were undergoing dialysis at 5 Lebanese dialysis centers. We also included 20 randomly selected healthy control volunteers. Many of the dialysis patients were undergoing work-up in preparation for kidney transplant. The institutional review board at each center approved the study protocol, and all patients gave informed consent before enrollment. Each participant answered and signed a specifically developed questionnaire. Demographic data, infor-mation about the disease (traditional risk factors, diagnosis of the underlying kidney disease, associated cardiovascular morbidities, and age of dialysis initiation), and medical laboratory tests were recorded. Blood samples were collected from patients and sent for genotyping.
The single nucleotide polymorphism variation names and chromosomes of each gene of the CVD panel are summarized in Table 1. Blood samples were used for gene mutation analyses. For DNA isolation, gene amplification, and hybridization of amplification products, we used strip arrays containing allele-specific oligonucleotide probes. The assay covered 12 mutations using their respective primers. For the factor V Leiden G1691A polymorphic variant, we detected the G to A substitution at nucleotide 1691 located in exon 10 of factor V. A region with 241 base pairs (bp) of exon 10 was amplified by using the following primers: 5’-TCA GGC AGG AAC AAC ACC AT-3’ (forward) and 5’-GGT TAC TTC AAG GAC AAA ATA CCT GTA AAG CT-3’ (reverse). For detection of the factor V H1299R polymorphic variant (4070A> G), a 1568-bp fragment was amplified with the following primers: 5’-TGC TCC TTT ATC TCC GAG GAC C-3’ (forward) and 5’-CTC TGG AGG AGT TGA TGT TTG TCC-3’ (reverse).
For the PT gene (FG20210A polymorphic variant), the G to A substitution at nucleotide 20210, located in exon 14, was examined by the polymerase chain reaction-restriction fragment length polymorphism method. A region with 345 bp of exon 14 was amplified using the following primers: 5’-TCT AGA AAC AGT TGC CTG GC-3’ (forward) and 5’-ATA GCA CTG GGA GCA TTG AAG C-3’ (reverse).
Primers for the F8 V34L polymorphic variant were 5’-GTA AAG TCA AAAATG TCA GAA AC-3’ (forward) and 5’-GTT GAC GCC CAG GGG CAC CG-3’ (reverse). The β-fibrinogen genotyping for the 455G/A polymorphic variant was performed using the forward primer 5’-GAA CAT TTT ACC TTA TGT GAA TTA AGG-3’ and reverse primer 5’-GAA GCT CCA AGA AAC CAT CC-3’. The PAI1 4 G/5 G genotype was analyzed with alternative forward primers (5’-GTC TGG ACA CGT GGG GG-3’ for the 5G allele or 5’-GTC TGG ACA CGT GGG GA-3’ for the 4G allele) and a common reverse primer (5’-GAA GCT CCA AGA AAC CAT CC-3’). The forward primer (5’-NA to 5’-PR) 5’-CAG GAG GTA GAG AGT CGC CAT AGT-3’ and reverse primer 5’-AGT TAT CCT TCA GCA GAT TCT CCT TC-3’ were selected to amplify a DNA fragment from the GP3A gene harboring the polymorphic HPA1a/b site.
For the MTHFR C677T polymorphic variant, primer sequences were as follows 5’-TGA AGG AGA AGG TGT CTG CGG GA-3’ (forward) and 5’-AGG ACG GTG CGG TGA GAG TG-3’ (reverse). The second A1298C mutation was analyzed using the following primer pairs: 5’-CTT TGG GGA GCT GAA GGA CTA CTA C-3’ and 5’-CAC TTT GTG ACC ATT CCG GTT TG-3’. ACE insertion/deletion genotypes were determined with a set of primers designed to encompass the polymorphic region in intron 16 of the ACE gene (sense primer 5’ CTG GAG ACC ACT CCC ATC CTT TCT 3’, antisense primer 5’-GAT GTG GCC ATC ACA TTC GTC AGA T-3’).
For the APOB R3500Q polymorphic variant, we used the following primers: 5’-CTT AGG AGG GGA CAT TTG AGT GG-3’ (forward) and 5’-TCT AAT ACA GCC CTG ACC TCG TGT-3’ (reverse). For the APOE 112 polymorphic variant, we used 5’-ACA TGG AGG ACG TGT GCG GCC GCC TG-P-3’ as the probe sequence. The APOE 158 probe sequence was 5’-GCG GCT CCT CCG CGA TGC CGA TGA CCT GCA GAA GCG CCT GGC-P-3’.
Frequency of cardiovascular disease gene variants in the population
We determined the sequence of the CVD gene variants for all patients and investigated the frequency of homozygous variants according to their age at dialysis initiation divided by 10-year intervals.
Frequency of cardiovascular disease gene variants according to underlying kidney
We stratified patients according to their underlying kidney disease and investigated the frequency of homozygous CVD gene mutations according to these causes.
Frequency of traditional risk factors according to the age at dialysis
We stratified patients according to their age of dialysis initiation into 10-year interval groups. We investigated the frequency of traditional risk factors according to mean patient age at dialysis initiation. Traditional risk factors included sex, hypertension, smoking, diabetes mellitus, obesity defined as body mass index > 29 kg/m2, anemia, dyslipidemia, hyperuricemia, CAD, congestive heart failure, stroke, peripheral vascular disease, amputation, deep vein thrombosis, abortion, consanguinity, and metabolic syndrome.
Comparison of age at dialysis initiation for various genetic variants
Using t tests, we compared the mean age at dialysis initiation for all patients having or not having each of the genetic variant genotypes.
Comparison of age at dialysis initiation for various traditional risk factors
We also used t tests to compare the mean age at dialysis initiation for all patients having or not having each of the traditional risk factors.
Association of genetic variants, traditional risk factors, and age at
For each patient, we allocated 3 scores: a genetic CVD mutation score, a traditional risk factor score, and a total score defined by the total number of thrombophilic homozygous mutations, the total number of traditional risk factors, and the sum of both. Patients were then stratified according to their genetic score divided by quartiles of 2 mutations. Patients were also stratified according to their traditional risk factor score divided by quartiles of 3. Using Pearson test, we investigated the potential association between the 3 scores and age at dialysis initiation. We also performed the same analysis in consanguineous and nonconsanguineous patients.
Statistical analyses were performed with SPSS software (SPSS: An IBM Company, version 24.0, IBM Corporation, Chicago, IL, USA). P values ≤ .05 were considered significant.
This study included 102 patients with ESRD undergoing hemodialysis and 20 healthy volunteers (control group). The mean age of patients was 52.6 ± 10 years versus 44.3 ± 12 years for the control group, with similar male-to-female ratio of 3:2. Demographic characteristics are summarized in Table 2.
Frequency of genetic variants in patients with end-stage renal disease
Frequencies of normal wild-type, homozygous mutant, and heterozygous variants for each of the genes in the CVD panel for both ESRD patients and healthy volunteers are shown in Table 3. We observed a high frequency of mutations (carriers and/or homozygous) in the following genes: F8, β-fibrinogen, PAI1, GP3A HPA1, MTHFR1 (A1298C), MTHFR2 (C677T), ACE (insertion/deletion), and APOE (E2/E3 and E3/E4). We found a relatively high frequency (> one-third of patients) of heterozygous variants for the β-fibrinogen (37.2%), MTHFR1 A1298C (52%), MTHFR2 C677T (44.1%), and ACE (44%) genes, with the highest being for the PAI1 gene (71.5%). The greatest frequencies of homozygous variants (> 10% of patients) were observed in the PAI1 (11%), MTHFR1 (13%), and ACE (12%) genes. More than one-half of the patients exhibited anomalies in the MTHFR2 (52%), MTHFR1 (65%), and ACE (56%) genes, with the highest being in the PAI1 gene (82%). We observed high frequencies of 2 mutations in the control group for the following genes: PAI1 (90%) and MTHFR1 A1298C (75%).
The greatest percentage of patients with CVD homozygous mutations (number of mutations ranging between 1 and 3 per patient) was observed between the age of 20 and 40 years at dialysis initiation, with gradual and linear declines thereafter (Figure 1).
Frequency of genetic variants associated with underlying cause of native kidney disease Cardiovascular disease gene mutations in the ESRD patients were analyzed according to cause of underlying native kidney disease. We observed an aggregation of different CVD gene mutations with certain underlying causes, including nephrosclerosis, diabetes mellitus, and most importantly with cause “unknown.” In contrast, CVD gene mutations were not present in patients with cystic diseases (Figure 2).
Frequency of traditional risk factors
We investigated the frequency of traditional risk factors according to patient age at dialysis initiation. We found that the frequency of traditional risk factors increased when patient age at dialysis initiation was between 50 and 80 years.
Comparison of age at dialysis initiation versus various cardiovascular disease
We found a significant association between age of dialysis initiation and homozygous mutations of the F8 V34L and PAI1 4G/5G polymorphic variants. This significant association was also shown between age of dialysis initiation and heterozygous mutations for the GP3A HPA1a/b variant. Carriers of these mutations started dialysis at a young mean age (at 25.7, 37, and 37.4 y, respectively) (Table 4). Interestingly, for most patients with such mutations, the underlying causes for their ESRD were unknown.
Comparison of age at dialysis initiation versus traditional risk factors
Our results showed a significant association between some traditional risk factors and age at dialysis initiation, including body mass index, hypertension, cardiovascular disease, and consanguinity. The latter was associated with dialysis initiation at the youngest mean age (39.8 y; Table 5).
Association of genetic variants, traditional risk factors, and age at dialysis
initiation in patients with end-stage renal disease
Patients were stratified by quartiles according to total scores of genetic mutations and traditional risk factors. We observed a strong linear relationship between the 3 patient scores and the mean age of dialysis initiation, with the highest scores being associated with older age. The association was the strongest when both genetic and traditional risk scores were combined (total score) (Figure 3).
Association of genetic variants, traditional risk factors, and age at dialysis
initiation in consanguineous patients
A similar relationship was found when total risk scores were analyzed according to mean age of dialysis initiation in both the consanguineous and nonconsanguineous ESRD study populations (Figure 4). Scores increased similarly in both consanguineous and nonconsanguineous groups. However, consanguineous patients started dialysis at a much earlier age, with the youngest patients (≤ 20 years old) in both groups exhibiting a signi-ficantly lower number of CVD genetic mutations compared with those over 20 years (3.0 vs 4.3, respectively; P = .008). In fact, the proportion of consanguineous patients in the study population declined with advancing age (Figure 5).
Our study revealed CVD genetic mutation frequencies similar to those reported by other investigators regarding normal wild-type variants of the following genes: factor V H1299R, PT FG20210A, and F8 V34L.29-32 Interestingly, patients with diabetes mellitus in our study exhibited increased expression of 3 important gene mutations: MTHFR C677T, PAI1, and ACE (Figure 2). These results are in agreement with those observed in other populations of different ethnic backgrounds, where a high frequency of mutant allele for the MTHFR gene was reported in patients with diabetes mellitus and nephropathy.33,34 Other Japanese35 and Chinese36 studies found significant associations between individuals with the T/T genotype of the MTHFR C677T polymorphism and elevated risk of developing CKD. Moreover, Jamison and associates37 recently reported findings similar to ours in 677 patients from 21 Veterans Affairs Medical Centers. These observations provide additional support for the hypothesis that the mutant TT genotype of the gene regulating MTHFR activity may increase the risk of cardiovascular mortality in patients with ESRD.37
In contrast, similar allele distributions regarding MTHFR C677T in patients with and without diabetic nephropathy38 and between Saudi hemodialysis patients and a normal control group have also been reported.39 These reported differences in the frequencies of the MTHFR alleles in these studies may be related to differences in expression between Asian, European, and Middle Eastern populations.
Regarding the MTHFR A1298C polymorphic variant, more then one-half of the study population (51.9%) was heterozygous, with an important proportion (12.7%) who were homozygous. These results are in agreement with previous data, in which high incidences of the C677T (47.2%) and A1298C (32.5%) polymorphic variants were shown in a dialysis population.40 These findings are similar to those from a more recent cross-sectional study involving 120 maintenance hemodialysis patients that assessed prevalence of MTHFR C677T and A1298C mutations and their relative association with hyperhomocysteinemia and CVD. Both the MTHFR C677T and A1298C polymorphic variants were reported to be associated with CVD in hemodialysis patients.41
For the PAI1 gene, most of our dialysis patients were heterozygous for 4G/5G (71.57%). Our findings are similar to those from a small observational study that included 36 hemodialysis patients and 40 control individuals,42 in which 64% of patients were heterozygous for 4G/5G. A recent Chinese study43 also reported an association between the PAI1 4G/5G genotype and increased risk of type 2 diabetic nephropathy. In contrast, Al-Muhanna and colleagues39 reported lower prevalence of PAI1 4G/4G polymorphism in Saudi ESRD patients compared with a control group. These differences in the PAI1 allelic frequency could be explained by the different ethnicities of the cohort. Moreover, the frequency of PAI1 heterozygous expression in our dialysis population (71.5%) was comparable to that shown in our normal population (85%), as shown in Table 3. Interestingly, PAI1 homozygous expression was more prevalent in the dialysis population than in the normal group and was strongly associated with initiation of dialysis at a relatively young age (Table 4), a finding not previously reported, which suggests an association between PAI1 homozygous mutation and increased risk for rapid progression to ESRD. Unfortunately, many patients who have these mutations had undetermined causes of ESRD. This raises an important question about its role as a genetic factor in increasing the risk for dialysis initiation at an early age. Unfortunately, given the small number of individuals in the study and healthy cohort populations, we could not make a definite conclusion.
More than two-thirds of our dialysis patients expressed normal genotypes for both the F8 V34L and the GP3A (a/a) genes, which were comparable to results for the control group. Those with either homozygous (F8 V34L) or heterozygous (GP3A, a/a) mutations had a greater risk of dialysis being initiated at a younger age (Table 4), a finding not previously reported. The fact that younger age of dialysis initiation in those with the homozygous mutation (GPIIIA [a/a]) did not reach statistical significance is probably related to the relatively small number of these patients versus those with the heterozygous mutational variant. No association regarding any specific renal cause could be identified.
For ACE, we found that approximately half of the population was heterozygous, with an important proportion (11.7%) being homozygous, although with none in the healthy control cohort (Table 3). These findings are in agreement with results shown in a population of 380 Malaysian patients with ESRD. In that study, the expression of insertion/deletion polymorphisms of the ACE gene and the frequency for the D allele were found to be higher (42.4%) among ESRD patients,44 which appears to be associated with the occurrence of severe left ventricular hypertrophy in kidney transplant patients.45
For APOE, we found that 29.1% of the population was heterozygous and only 4.7% was homozygous. These findings were similar to those reported by Kahraman and associates.46 In our study and as shown previously,47 similar phenotype distributions and allele frequencies were shown between hemodialysis patients and healthy controls; in addition, APOE genotypes were not an independent risk factor for atherosclerotic vascular disease. For APOB, we found that all groups showed normal variants.
The greatest frequencies of homozygous mutations (>10% of patients) were observed with the PAI1 (11%), ACE (12%), and MTHFR A1298C (12.7%) genes. More than half of the patients exhibited anomalies in the following genes: MTHFR A1298C, ACE, and MTHFR C677T (52%, 56% and 65%, respectively), with an even higher percentage shown in the PAI1 gene (82%). We observed a high frequency of 2 mutations in the control group for the following genes: PAI1 and MTHFR A1298C (90% and 70%, respectively). The greatest percentage of patients with CVD homozygous mutations (number of mutations ranging between 1 and 3/patient) was observed between those who were between 20 and 40 years old at dialysis initiation, with gradual and linear declines thereafter (Figure 1). Interestingly, more than two-thirds of the study population within this age interval were consanguineous patients (Figure 5).
When patients with and without diabetes mellitus were compared, we observed similar frequencies of genetic and traditional risk factors, with distinctive late onset of dialysis in the diabetic group. These findings highlight the importance of diabetes mellitus as an isolated major risk, and late onset of dialysis may be influenced by genetic or other environmental factors. Our results are in agreement with those reported in a Swedish registry study.48 This is because diabetes is a chronic silent disease, with most patients having late diagnosis. It gradually leads to con-siderable macrovascular and microvascular com-plications with progression to kidney disease and failure.
In our study, consanguinity was associated with the youngest age at dialysis initiation, confirming earlier results involving 925 Lebanese patients from all dialysis centers. As reported by Barbari and associates, there is a strong association between consanguinity and early initiation of dialysis (before 30 years old). Moreover, the investigators showed an increased risk for family history of kidney disease among the consanguineous population,49 which represents nearly one-quarter of the total Lebanese dialysis population. These results are in agreement with another study involving 103 white dialysis patients from the United States. That study showed a 3-fold increased risk of developing ESRD in the presence of either a first- or second-degree relative.50
Regarding traditional risk factors, our results revealed the expected significant association between age of dialysis initiation and obesity, hypertension, congestive heart failure, peripheral vascular disease, and amputation. Borderline associations were shown for proteinuria and dyslipidemia (Table 5). These observations are in agreement with recent data on the association between these factors and the increased risk for CKD progression in patients with and without diabetes mellitus.51-55 We found that patients with CAD started dialysis at a relatively young age (mean of 50.2 y), although this result did not reach statistical significance given the small sample size. Our results are in agreement with recent reports showing CAD as the strongest independent predictor of CKD1 and early progression to dialysis.56
Our data revealed new important findings regarding a strong linear and positive relationship between patient age and dialysis initiation and load of genetic and traditional CVD and CKD risk factors, individually and combined, as shown in Figure 3. These results were similar in the consanguineous patient subgroup and were maintained when the analysis was performed between consanguineous and nonconsanguineous dialysis patients (Figure 4). Younger patients who started dialysis at an early age were more likely to have a reduced number of different CVD mutations compared with older patients who started dialysis at a later age and had greater individual and total CVD thrombophilic and traditional risk factor scores. These observations highlight the importance of the cumulative load of CVD genetic and traditional risk factors on age of dialysis initiation, mainly in the older dialysis population. It also explains the bidirectional CVD-CKD relationship in increasing the risk of disease.1
The lower number of CVD genetic mutations in our young dialysis patients suggested a nonvascular origin of the underlying kidney diseases. In younger patients, renal diseases appear to be mostly related to congenital malformation, including vesicoureteral reflux and hereditary nephritis such as medullary cystic kidney disease, familial nephrotic syndrome, and oxalosis. However, a superimposed increased load of CVD thrombophilic mutations, such as the ones reported in our study (F8 V34L, PAI1 4G/5G, and GP3A HPA1a/b) and others mutations, including ACE D/D and MTHFR, may exert a cumulative and additive effect on rapid progression to ESRD. This could lead to dialysis initiation at an early age in young patients with the above renal causes or with hemolytic uremic syndrome. In fact, the greatest percentage of patients with the highest number of CVD homozygous mutations (Figure 1) started dialysis between the ages of 20 and 40 years. Interestingly, the greatest percentage of patients within this age interval were consanguineous, and consanguinity was strongly associated with early onset of dialysis (Table 4) with an inverse linear relationship between mean age at dialysis initiation and consanguinity prevalence within each age interval (R2 = 0.98) as shown in Figure 5. Consanguinity-associated kidney diseases may be related to distinct genetic causes that are related either to CVD thrombophilic mutations and/or to non-throm-bophilia-mediated hereditary nephritis.
We postulated that an increase in the cumulative occurrence of homozygous CVD and non-CVD gene anomalies in addition to increased score of traditional factors may explain the younger age at dialysis initiation in the consanguineous compared with the nonconsanguineous patients with similar overall load of CVD mutations (Figure 4). It is interesting to note that, in our study and in another series,49 a sizeable proportion of the underlying renal causes in the consanguineous dialysis population were unde-termined. Our results point to the important utility of the CVD panel as a screening test for progression to dialysis in older patients to assess the throm-bophilic risk score and in younger CKD patients to detect potential carriers of specific CVD mutations (F8 V34L, PAI1 4G/5G, and GP3A HPA1a/b) shown to be strongly associated with early dialysis initiation.
Major limitations of our study were mostly related to its limited population size, especially concerning the control group, and its retrospective design. Missing data may have limited examination of different traditional risk factors and could have been why many patients had unknown causes of CKD. Despite these limitations, our study demonstrated a strong association, never reported before, between a score of CVD genetic and traditional risk factors and age of initiation of dialysis, a marker for CKD progression. Moreover, although other studies mainly analyzed the impact of one of the genes from the CVD panel on CKD progression, our study analyzed the association of 12 genes and also examined potential traditional CVD and CKD risk factors.
Our study revealed a higher frequency of hetero-zygous mutations for the PAI1 and MTHFR (A1298C) genes and homozygous mutations for PAI1, MTHFR (C677T), and ACE in the ESRD patients, suggesting a CVD polygenic mechanism. We demonstrated a strong association between early dialysis initiation and specific homozygous CVD gene mutations and consanguinity. We established an ascending linear relationship between cumulative loads of different traditional environmental and CVD genetic risk factors and age of dialysis initiation, with the highest scores being associated with initiation of dialysis at an older age. The cumulative load of these combined CVD genetic and traditional risk factors may be at least partly responsible for the increased risk and prevalence of ESRD in older adults and the considerable magnitude of cardiovascular mortality in the CKD population. The observed low genetic CVD mutation score in young consanguineous and nonconsanguineous ESRD patients suggested a predominantly nonvascular origin of ESRD in this population, with a potential rapid CKD progression in the presence of specific and/or high loads of homozygous CVD thrombophilic mutations. Our results confirmed the strong association between consanguinity and early onset of ESRD and dialysis initiation at a young age. In addition, our results suggest a potential increased load of nonvascular and vascular genetic anomalies in combination with traditional risk factors that may explain the initiation of dialysis at a young age. We propose the CVD panel as a screening test for progression to dialysis in older patients to assess the thrombophilic risk score and in younger CKD patients to detect potential homozygous carriers of specific CVD mutations shown to be associated with early initiation of dialysis. Given the small size of our ESRD population and that of the healthy cohort, further and larger-scale prospective studies are needed to confirm these interesting new observations.
Volume : 16
Issue : 6
Pages : 639 - 650
DOI : 10.6002/ect.2018.0164
From the 1Faculty of Medical Sciences, Lebanese University, Beirut, Lebanon; the
2Rafic Hariri University Hospital, Beirut, Lebanon; the 3School of Pharmacy,
Lebanese American University, Byblos, Lebanon; the 4Department of Nephrology,
Mount-Lebanon Hospital, Beirut, Lebanon; the 5Department of Nephrology and
Hypertension, Al Rassoul Al Aazam Hospital, Beirut, Lebanon; the 6Department of
Nephrology, Abou Jaoudeh Hospital, Beirut, Lebanon; the 7Department of
Nephrology, Bahman Hospital, Beirut, Lebanon; and the 8Transmedical for Life,
Acknowledgements: The authors have no sources of funding for this study and have no conflicts of interest to declare.
Corresponding author: Antoine Barbari, Lebanese University, Faculty of Medical Sciences, Department of Internal Medicine, Nephrology Division/Renal Transplant Unit, Rafik Hariri University Hospital, Bir Hassan, Beirut, Lebanon
Phone: +961 1832040
Table 1. Single Nucleotide Polymorphism Variations, Names, and Chromosome of Each Gene in the Cardiovascular Disease Panel
Table 2. Demographic Characteristics of Patients With End-Stage Renal Disease and Control Group
Table 3. Frequency of Wild-type, Heterozygous Carriers, and Homozygous Mutants of Cardiovascular Disease Genes
Table 4. Mean Age at Initiation of Dialysis for All Genetic Variants
Table 5. Mean Age at Initiation of Dialysis for All Traditional Risk Factors
Figure 1. Percentage of Patients with Homozygous Mutations in Cardiovascular Disease Genes According to Patient Age at Dialysis Initiation
Figure 2. Frequency of Different Homozygous Cardiovascular Disease Genetic Mutations Among the Causes of Chronic Kidney Disease
Figure 3. Relationship Between Mean Genetic, Traditional, and Total Scores and Corresponding Mean Age at Dialysis Initiation in the Whole End-Stage Renal Disease Population
Figure 4. Relationship Between Mean Total Genetic and Traditional Scores and Corresponding Mean Age at Dialysis Initiation in the Consanguineous (+) and Nonconsanguineous (-) End-Stage Renal Disease Population
Figure 5. Proportion of Consanguineous Patients According to Mean Age at Dialysis Initiation by 20-Year Quartiles in the Whole End-Stage Renal Disease Study Population