To examine the contribution of both SNPs on predicting utilization, we used the bias-reduced binary logistic regression model to simultaneously analyze Val158Met and Ser9Gly genotypes and then the binary logistic regression model for the Val158Met and Ser9Gly alleles (Table 6). Comparing the odds of having zero utilization between patients with Gly/Ser genotype and those with Gly/Gly, we found an OR of 4.37 (95% CI, 1.39–22.89; P = 0.020). Comparing the odds of having zero utilization between Ser/Ser and those with Gly/Gly, the OR was 2.81 (95% CI, 0.11–21.66; P = 0.338). Analysis of alleles showed an OR of 2.36 (95% CI, 1.09–5.12; P = 0.030) between each additional DRD3 Ser allele and having zero utilization. For COMT Val158Met, we found an OR of 0.32 (95% CI, 0.12–0.83) between having zero utilization and each additional Met allele (P = 0.020).
We further examined models that included age, sex, SC type, and CPI as covariates to exclude potential bias on utilization counts by these variables (Table 7). Consistent with the previous analysis (Table 6), DRD3 heterozygote genotype showed an increased odds of having zero utilization over the homozygotes with an OR of 4.53 (95% CI, 1.41–28.58; P = 0.016). Similarly, this analysis showed an OR of 0.36 (95% CI, 0.13–0.98; P = 0.044) between having zero utilization and each additional COMT Met allele, consistent with our previous analysis.
To examine the influence of the SNPs on the number of utilization counts, we further simultaneously analyzed DRD3 Ser9Gly and COMT Val158Met in a negative binomial regression model (Table 8). This model was chosen to account for overdispersion in the utilization data. There was minimal difference in utilization rates between subjects of DRD3 Ser9Gly genotype and those of Gly/Gly genotype (IRR = 1.03 [95% CI, 0.71–1.48]; P = 0.879) or between Ser/Ser genotype and Gly/Gly (IRR = 0.98 [95% CI, 0.50–1.93]; P = 0.959). COMT rs4680 Met/Met genotype, however, showed a 2.20 times the utilization incident rate than the Val/Val genotype (IRR = 2.20 [95% CI, 1.21–3.99]; P = 0.010). The association between each additional COMT rs4680 Met allele and utilization counts was illustrated by the IRR of 1.33 (95% CI, 1.01–1.75; P = 0.043).
Covariate adjusted models for negative binomial regression analyses confirmed that DRD3 Ser9Gly genotype did not associate with utilization counts (IRR = 0.90 [95% CI, 0.64–1.28]; P = 0.558; Table 9). The IRR between COMT Met/Met genotype and Val/Val genotype was 1.85 (95% CI, 1.03–3.30; P = 0.038).
This is the first genetic study that was specifically designed to examine the influence of monoamine candidate gene polymorphisms on pain in SCD, although a genome-wide association study of SCD disease severity was reported.31 In this study, we examined 2 SNPs that are relevant to the endogenous monoamine system and have been associated with pain phenotypes in non-SC patients. We used an innovative pain-scoring method CPI, and utilization data, to characterize pain phenotypes in SCD. In this study, utilization serves as a surrogate marker for acute pain due to crises. CPI (taken during routine clinic visits) is a quantitative and qualitative measurement of chronic pain on noncrisis days in SCD.
The Ser9Gly polymorphism results in an amino acid substitution at codon 9 in the N-terminal domain of the DRD3 in which the Gly variant has been observed to trigger a greater cellular response to dopamine.32 The Gly variant also displays greater binding affinity to dopamine33 and increased mitogen-associated protein kinase signal duration.32 The Gly variant has also been reported to have greater reward-related dopamine release in the ventral tegmental area.34 It was proposed that the Gly allele would decrease tonic release and increase phasic release of dopamine,34 leading to decreased pain sensitivity since analgesic activity is proposed to be related to the phasic release of mesolimbic dopamine.9,35 Although the exact mechanisms are yet to be elucidated, these studies have implications for the role of DRD3 in pain and analgesia.
Having an acute pain crisis resulting in a visit to the emergency department or acute care center is a significant and devastating problem because pain crises are associated with serious complications.36 Therefore, patients having zero utilization serve as case controls in this analysis. The probability of heterozygotes having zero utilization were nearly 4.5 times of those with Gly/Gly genotypes, and this remained consistent in the covariate adjusted model. The analysis of DRD3 Ser9Gly homozygotes against heterozygotes has been frequently made since it was first proposed 2 decades ago. Heterozygote advantage allows for the accommodation of environmental conditions37 and has been reported for this SNP in schizophrenia,26 although it was not replicated in a subsequent meta-analysis.38 Frequencies of heterozygote genotypes were decreased in subjects who were more vulnerable for cocaine dependence,27,29 Tourette syndrome,28 and substance abuse in schizophrenia patients.30 Indeed, in the cerebrospinal fluids of healthy volunteers, Jönsson et al.39 found that DRD3 homozygous subjects had significantly higher 5-hydroxyindoleacetic acid levels than heterozygote subjects. Since 5-hydroxyindoleacetic acid is one of the major monoamine metabolites, the result suggested that homozygosity was associated with increased monoaminergic activity.27 The exact mechanism of DRD3 Ser9Gly heterozygote advantage in SCD pain will need to be further studied.
In fibromyalgia patients, Gly/Gly genotype was found to be associated with higher thermal pain threshold and greater pain tolerance.15 Subjects with Gly/Gly or Ser/Ser had increased odds of having 1 or more utilization. Also in this study, Ser allele may be associated with elevated odds of having zero utilization (OR of 2.17 and 2.36, with or without covariables, respectively). In addition to unclear mechanisms for heterozygote advantage, these 2 studies were performed in subjects with different pain conditions and ethnicity backgrounds, which may have contributed to the different findings.
The COMT polymorphism Val158Met (rs4680) was first reported in association studies with schizophrenia.42 It has since been associated with a myriad of disorders and behaviors such as fibromyalgia, aggression, and suicide.42–45 The Val/Val genotypes have the highest activity of COMT, while individuals with Met/Met are expected to have the lowest activity.13 In addition to the Met/Met genotype associating with higher sensory and affective ratings of pain,14 it has also been shown to associate with stronger pain-related signals in several brain structures after repeated pain stimuli46 and more pain on days when pain catastrophizing or pain attention was increased in fibromyalgia patients.43 COMT Val158Met is also associated with thermal pain and pressure pain stimuli in fibromyalgia patients.47 Furthermore, healthy subjects with Met/Met genotype reported significantly more pain compared with those with Val/Val when they were subject to pain induced to the hand by a heat probe.48 However, a few studies have been unable to find significant associations of Val158Met with pain. A study observing the susceptibility to neuropathic pain in a Spanish population did not find an association with Val158Met.22 An acute postoperative pain study also did not find association,49 and another study50 did not find an association of Val158Met with nociceptive sensitivity. However, the same study50 and several others14,48,50,51 found that Val158Met was primarily associated with the temporal integration of painful stimuli. In this study, we found the Met allele was associated with a lower probability of zero utilization (i.e., more acute pain) using the binomial logistic regression model that simultaneously analyzed DRD3 Ser9Gly and COMT Val158Met together. Furthermore, the Met/Met genotype showed 2.20 times the incident rate of utilization than the Val/Val genotype in the negative binomial regression model.
DRD3 Ser9Gly heterozygosity had increased odds of having zero utilization, even after factoring other variables including age, sex, SC type, and CPI, though it remains to be elucidated how Ser9Gly heterozygosity produced this advantage in acute pain in SCD. For the COMT Val158Met polymorphism, the Met allele and Met/Met genotype were associated with utilization, indicative of more acute pain rates. Neither SNP, however, appears to alter baseline pain.
Individual pain difference in SCD is expected to be complex and will likely involve a number of genetic polymorphisms in various biological systems and pathways.49,52 We took a candidate gene approach to specifically focus on 2 key members of the monoamine neurotransmitter systems based on findings in other pain types with these SNPs. We found that 2 COMT and DRD3 polymorphisms appear to influence acute pain in subjects with SCD, as indicated by the OR or IRR of having utilization due to acute pain crisis. We found no relationship of these polymorphisms with CPI scores (an indicator of chronic pain). In addition to candidate gene studies, approaches such as genome-wide association studies can aid the search for additional polymorphisms influencing pain in SCD.31 Ultimately, elucidating genetic contributions made by a full set of relevant polymorphisms will help to guide the individualized management of pain in patients with SCD.
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