Background
The underlying causes of vulvar pain in women with vulvodynia remain poorly understood. Catechol- O -methyltransferase, an enzyme that metabolizes catecholamines, is a neuromodulator that is involved with perception and sensitivity to pain. The catechol- O -methyltransferase gene is polymorphic, and a single nucleotide polymorphism is associated with low activity and heightened pain sensitivity. The variant allele that encodes this polymorphism commonly is called the “L allele” because of its low enzyme activity as opposed to the normal H (high activity) allele.
Objective
The methionine-containing catechol- O -methyltransferase protein coded by the L allele results in elevated catecholamine levels, reduced inactivation of the dopaminergic and adrenergic systems, and increased sensitivity to pain. This polymorphism not only may decrease the pain threshold in response to acute pain but also may facilitate the development of chronic pain. Therefore, the objective of our study was to assess whether a variation in the catechol- O -methyltransferase genotype is involved in increased pain sensitivity in women with vulvodynia.
Study Design
We conducted a prospective cohort study.
Methods
Buccal swabs were collected from 167 white women with vulvodynia and 107 control subjects; the DNA was tested for a single nucleotide polymorphism at position 158 (rs4680) in the catechol- O -methyltransferase gene.
Results
Women with vulvodynia had a marginally increased, yet not significant, prevalence of the catechol- O -methyltransferase genotype that is associated with high activity of the coded protein: 32.9% in the women with vulvodynia, as opposed to 21.5% in the control subjects (odds ratio, 1.80; 95% confidence interval, 1.02–3.15). Subgrouping the cases based on pain frequency revealed that the elevated occurrence of this catechol- O -methyltransferase genotype was present in 40.6% of the subset of women who experienced pain only with sexual intercourse vs only 21.5% of control subjects (odds ratio, 2.50; 95% confidence interval, 1.27–4.93). Also, women with primary vulvodynia had a significantly higher prevalence of the H allele than did the control subjects (62.9% vs 48.1%; odds ratio, 1.82; 95% confidence interval, 1.05–3.17).
Conclusion
Increased pain sensitivity in women with vulvodynia is not due to a genetically determined low catechol- O -methyltransferase enzyme activity. Other mechanisms may account for alterations in catechol- O -methyltransferase activity in women with pain that is limited to intercourse or primary vulvodynia that contributes to pain sensitivity.
Approximately 8.3% of women in the United States have a current diagnosis of vulvodynia; the disorder affects approximately 25% of women at some point in their lifetime. Vulvodynia is defined as vaginal or vulvar stabbing or burning pain of at least 3 months duration that can be spontaneous or provoked and either intermittent or constant. The cause of vulvodynia is most likely multifactorial and variable among different women. The mechanisms that are responsible for its development and persistence still are not defined clearly.
Vulvodynia is considered a chronic pain condition and can be associated with other chronic pain disorders such as migraine, irritable bowel syndrome, bladder pain syndrome/interstitial cystitis, and chronic pelvic pain. Recent evidence suggests that genetic variations contribute to pain sensitivity and the risk of the development of chronic pain in other chronic pain syndromes. Multiple gene polymorphisms have been identified in women with vulvodynia. However, their relation to pain sensitivity remains incompletely determined.
Catechol- O -methyltransferase (COMT) is an enzyme that metabolizes and inactivates catechol-containing compounds such as the catecholamines adrenaline (epinephrine), noradrenaline (norepinephrine), and their precursor, dopamine. Each of these 3 compounds is a neurotransmitter that is involved in the transmission and perception of pain sensation. Greater rate and extent of degradation of these neurotransmitters by COMT is associated with decreased perception of painful stimuli.
A single nucleotide polymorphism in the COMT gene at position 158 in exon 3 results in the substitution of methionine for valine in the final protein (rs4680). This val 158 met change results in decreased thermostability in the COMT protein and a concomitant 3- to 4-fold reduction in enzyme function. The variant allele commonly is called the “L allele” because of low enzyme activity as opposed to the normal H (high activity) allele. The methionine that contains COMT protein coded by the L allele results in elevated catecholamine levels, reduced inactivation of the dopaminergic and adrenergic systems, and increased sensitivity to pain. Healthy individuals who were L,L homozygous exhibited the greatest degree of pain sensitivity after the application of painful stimuli. This polymorphism not only may decrease the pain threshold in response to acute pain but also may facilitate the development of chronic pain. The val 158 met polymorphism was observed to be more prevalent in some, but not all, studies of individuals who experience fibromyalgia, tension headaches, and temporomandibular disorder.
The aim of the present study was to test the hypothesis that women with vulvodynia have a higher prevalence of the L,L genotype and a higher frequency of the L allele than women with no history of this disorder.
Materials and Methods
The study population was recruited from clinics at 5 sites: Florida Hospital Division of Pelvic and Vaginal Pain in Orlando, FL; Center for Sexual Health in Akron, OH; UCLA Medical Center Department of Obstetrics and Gynecology in Los Angeles, CA; Weill Cornell Medicine, Department of Obstetrics and Gynecology in New York, NY; and Women’s Integrated Pelvic Health Program, Denver Health, Denver, CO. The institutional review boards of all 5 sites approved this study, and participants provided written informed consent before participation. Primary institutional review board oversight was provided by Florida Hospital, institutional review board study #238051-22.
Women were enrolled with the use of protocols that were developed for the National Vulvodynia Registry or if they presented to the clinics for treatment of vulvodynia. Women were included in the study as cases if they had vulvar or vulvar vestibular pain of at least 3-months duration in the absence of detectable disease (infection, inflammatory changes, dermatoses, or malignancy). Pain had to be severe enough to impair sexual function or lead to avoidance of activities that involved contact with the vestibule or vulva. Women were included as control subjects if they had no vulvar or vestibular pain and had never had an interval of painful intercourse. Exclusion criteria for both cases and control subjects included non-English speaking, age <21 or >65 years, inability to provide informed consent, recent history of psychiatric hospitalization, current illicit drug abuse, current uncontrolled seizure disorder, history of reproductive tract malignancy, currently pregnant or breast feeding, vaginismus that prevented examination, adnexal/uterine pain on examination, pelvic pain on examination, dermatoses present on examination, or the presence of a pelvic mass.
After participants completed the study questionnaire and underwent a gynecologic examination per protocol, cells for DNA analysis were collected from the buccal cavity using Catch-All Sample Collection swabs (Epicentre Biotechnologies, Madison, Wisconsin). Each participant rinsed her mouth with water, then the swab was rolled firmly several times inside both cheeks. The swab was allowed to dry for 15–30 minutes at room temperature before being placed in tubes that were labeled with the patient’s assigned study code and collection (no other identifying information) and were stored at 4°C until shipped to Weill Cornell Medicine for analysis.
Cellular DNA was obtained by lysis of the cells in a Brij 35 (polyoxyethylene glycol dodecyl ether) detergent, proteinase K-containing buffer, and the COMT polymorphisms were identified by a published procedure. A 10-μL aliquot of lysate was diluted with 15-μL of sterile distilled water and added to an equal volume of 10-mmol/L TRIS-(Tris[hydroxymethyl] aminomethane)- hydrochloride buffer that contained 0.2 mmol/L each of deoxyadenosine triphosphate, deoxycytidine triphosphate, deoxyguanosine triphosphate, and thymidine triphosphate, 1.5 mmol/L magnesium chloride, 50 mmol/L potassium chloride, 1.25 U Taq DNA polymerase, and 30 pmol primer pairs that spanned the polymorphic region. Samples were incubated in a thermal cycler for 10 minutes at 94°C followed by 30 cycles of 94°C for 30 seconds, 61°C for 30 seconds, and 72°C for 30 seconds. This was followed by a 5-minute final elongation cycle at 72°C. The products were then treated with NLA III endonuclease overnight; the resulting amplicons were analyzed on 2% agarose gels and visualized by ethidium bromide staining. The COMT H allele was identified as bands at 114, 36, and 35 base pairs; the L allele formed bands at 96, 36, and 35 base pairs.
Genotype and allele frequencies were determined by direct counting and then were divided by the number of chromosomes to obtain allele frequency and by the number of women to obtain genotype frequency. Goodness of fit to Hardy-Weinberg equilibrium was determined by a comparison of the expected genotype frequencies with the observed values with the use of the chi-square test. Associations between genotype or allele and clinical parameters were analyzed by the Fisher’s exact test. Odds ratios and 95% confidence intervals were calculated for each significant comparison. A probability value of <.05 was considered significant. All analyses were done with Microsoft Excel 2013 software (Microsoft Corporation, Redmond, WA) and STATA software (version 10.0; (STATA Corporation, College Station, TX).
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