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British Journal of Anaesthesia
BJA

CHRM3 rs2165870 polymorphism is independently associated with postoperative nausea and vomiting, but combined prophylaxis is effective

Open ArchivePublished:April 03, 2018DOI:https://doi.org/10.1016/j.bja.2018.02.025

      Abstract

      Background

      Clinical risk factors for postoperative nausea and vomiting (PONV) are evaluated with the Apfel score, however patients with low Apfel scores still experience PONV suggesting a genetic predisposition. PONV risk associates with specific M3 muscarinic acetylcholine receptor (CHRM3) rs 2165870 polymorphism. We investigated whether the Apfel score and this genetic variation independently contribute to PONV risk and whether prophylaxis reduces PONV in patients with low Apfel score but at high genetic risk.

      Methods

      In a prospective, controlled study, 454 subjects undergoing elective surgery were genotyped for rs2165870 and its association with PONV was investigated with log-binomial regression analysis. Subjects were randomised to receive acustimulation/dexamethasone, acustimulation/vehicle, sham acustimulation/dexamethasone, or sham acustimulation/vehicle to investigate their effects on PONV risk.

      Results

      Early PONV occurred in 37% of subjects. The rs2165870 genotype distribution was GG in 191, GA in 207, and AA in 56 subjects. The CHRM3 polymorphism was associated with a relative risk (RR) of 1.5 for GA vs GG [95% confidence interval (CI): 1.1–1.9; P=0.003] and 1.6 for AA vs GG (95% CI: 1.1–2.2; P=0.009) genotypes to develop PONV, and this was independent from the Apfel score (RR per Apfel point: 1.3, 95% CI: 1.2–1.5; P<0.0001). While dexamethasone and acustimulation each reduced the PONV risk by 30% in AA genotype carriers with low Apfel score, combined therapy reduced the risk by 86% (P=0.015).

      Conclusions

      The CHRM3 polymorphism and the Apfel score independently predict PONV susceptibility. Dexamethasone/acustimulation should be considered in patients with low Apfel score but at high genetic risk.

      Clinical trial registration

      DRKS00005664.

      Keywords

      • The rs2165870 polymorphism in the M3 muscarinic acetylcholine receptor gene is associated with an increased risk of postoperative nausea and vomiting (PONV).
      • The GA and AA genotypes and the Apfel score were independently associated with PONV in a cohort of 454 patients undergoing elective surgery.
      • In AA genotype subjects with low Apfel score, combined prophylaxis with dexamethasone and acustimulation significantly reduced PONV.
      The overall incidence of postoperative nausea and vomiting (PONV) in the general surgical population is up to 30%,
      • Cohen M.M.
      • Duncan P.G.
      • DeBoer D.P.
      • Tweed W.A.
      The postoperative interview: assessing risk factors for nausea and vomiting.
      making it the most frequent side effect after general anaesthesia.
      • Eberhart L.H.
      • Hogel J.
      • Seeling W.
      • Staack A.M.
      • Geldner G.
      • Georgieff M.
      Evaluation of three risk scores to predict postoperative nausea and vomiting.
      • Gan T.J.
      Postoperative nausea and vomiting—can it be eliminated?.
      Predispositions for PONV are multifactorial and can be categorised as patient risk factors, anaesthetic techniques, and surgical procedures.
      • Chandrakantan A.
      • Glass P.S.
      Multimodal therapies for postoperative nausea and vomiting, and pain.
      Apfel and colleagues
      • Apfel C.C.
      • Laara E.
      • Koivuranta M.
      • Greim C.A.
      • Roewer N.
      A simplified risk score for predicting postoperative nausea and vomiting: conclusions from cross-validations between two centers.
      defined four risk factors: female gender, history of PONV/motion sickness, non-smoking status, and use of postoperative opioids, which resulted in the Apfel score, a simple and often used tool for PONV risk stratification. If none, one, two, three, or four of these risk factors are present, the incidence of PONV is 10, 21, 39, 61, and 79%, respectively.
      • Apfel C.C.
      • Laara E.
      • Koivuranta M.
      • Greim C.A.
      • Roewer N.
      A simplified risk score for predicting postoperative nausea and vomiting: conclusions from cross-validations between two centers.
      However, even patients at low PONV risk by the Apfel score can experience PONV, suggesting a genetic predisposition. Interestingly, a genome wide association study (GWAS) provided evidence for genetic susceptibility to PONV. A single nucleotide polymorphism (SNP; rs2165870) located upstream from the promoter region of the M3 muscarinic acetylcholine receptor (CHRM3) gene is associated with PONV, and was validated in an independent verification cohort where logistic regression analysis in a dominant model revealed an odds ratio (OR) of 3.1 for the minor A allele in the pooled group and of 2.3 in the verification cohort.
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      This makes likely a true association of this SNP with PONV.
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      Prophylactic strategies, such as the administration of antiemetic drugs
      • Apfel C.C.
      • Korttila K.
      • Abdalla M.
      • et al.
      A factorial trial of six interventions for the prevention of postoperative nausea and vomiting.
      • Gomez-Hernandez J.
      • Orozco-Alatorre A.L.
      • Dominguez-Contreras M.
      • et al.
      Preoperative dexamethasone reduces postoperative pain, nausea and vomiting following mastectomy for breast cancer.
      or stimulation of the P6 acupuncture point,
      • Lee A.
      • Chan S.K.
      • Fan L.T.
      Stimulation of the wrist acupuncture point PC6 for preventing postoperative nausea and vomiting.
      • Frey U.H.
      • Funk M.
      • Lohlein C.
      • Peters J.
      Effect of P6 acustimulation on post-operative nausea and vomiting in patients undergoing a laparoscopic cholecystectomy.
      • Frey U.H.
      • Scharmann P.
      • Lohlein C.
      • Peters J.
      P6 acustimulation effectively decreases postoperative nausea and vomiting in high-risk patients.
      reduce the risk for PONV by ∼30% and are often applied in high-risk patients based on Apfel score. In the IMPACT trial, ondansetron, droperidol, and dexamethasone were equally effective in PONV prophylaxis,
      • Apfel C.C.
      • Korttila K.
      • Abdalla M.
      • et al.
      A factorial trial of six interventions for the prevention of postoperative nausea and vomiting.
      however studies evaluating established PONV treatments only examined 5-hydroxytryptamine type 3 receptor antagonists.
      • Kazemi-Kjellberg F.
      • Henzi I.
      • Tramer M.R.
      Treatment of established postoperative nausea and vomiting: a quantitative systematic review.
      • Meyer T.A.
      • Roberson C.R.
      • Rajab M.H.
      • Davis J.
      • McLeskey C.H.
      Dolasetron versus ondansetron for the treatment of postoperative nausea and vomiting.
      .
      There is a debate about combinations of pharmacological prophylaxis and acustimulation,
      • Cooke M.
      • Rickard C.
      • Rapchuk I.
      • et al.
      PC6 acupoint stimulation for the prevention of postcardiac surgery nausea and vomiting: a protocol for a two-group, parallel, superiority randomised clinical trial.
      • Kim K.H.
      • Kim D.H.
      • Bae J.M.
      • et al.
      Acupuncture and PC6 stimulation for the prevention of postoperative nausea and vomiting in patients undergoing elective laparoscopic resection of colorectal cancer: a study protocol for a three-arm randomised pilot trial.
      and one recent study showed that a combination of dexamethasone and acustimulation was more effective in preventing PONV in gynaecological patients undergoing laparoscopic surgery than dexamethasone alone.
      • Yang X.Y.
      • Xiao J.
      • Chen Y.H.
      • et al.
      Dexamethasone alone vs in combination with transcutaneous electrical acupoint stimulation or tropisetron for prevention of postoperative nausea and vomiting in gynaecological patients undergoing laparoscopic surgery.
      Our primary aim was to test the hypothesis that both genetic factors and the Apfel score contribute independently to the risk for PONV. Secondly, we investigated in a randomised controlled trial whether PONV prophylaxis with dexamethasone, P6 acustimulation, or both mitigates the risk for PONV among patients carrying a genetic variant previously associated with PONV.

      Methods

       Study design

      This prospective, randomised, controlled, single centre study investigating association of the CHRM3 SNP (rs2165870) with PONV and effects of combined prophylaxis on PONV was approved by the University of Duisburg-Essen Medical Faculty Ethics Committee (number 12-5065-BO). Patients' written informed consent was obtained before participation. The study was part of a study investigating effects of multimodal prophylaxis on PONV risk (DRKS00005664). The primary aim of the study was to test the hypothesis that CHRM3 SNP (rs2165870) and Apfel score independently contribute to the risk of PONV (observational study). Secondly, we investigated whether PONV prophylaxis with P6 acustimulation, dexamethasone, or both mitigates the risk for PONV among patients carrying a genetic variant previously associated with the PONV phenotype (randomised controlled trial).

       Subjects

      Patients of either sex older than 18 yr were eligible if ASA physical status I–III and undergoing general anaesthesia for gynaecological, ear, nose, and throat, or endocrine surgery (thyroid) at Essen University Hospital. Exclusion criteria were: patients with a cardiac pacemaker or implanted cardioverter/defibrillator, patients with an allergy to nickel or chrome, or ASA classification >III. Subject characteristics, morphometric data, and risk factors that can influence PONV (e.g. sex, PONV history, and smoking history) were collected before surgery.
      • Apfel C.C.
      • Roewer N.
      • Korttila K.
      How to study postoperative nausea and vomiting.

       Randomisation and group allocation

      Subjects were randomly allocated to receive one of four treatments using a computer-generated random number table: acustimulation/dexamethasone, acustimulation/vehicle, sham acustimulation/dexamethasone, or sham acustimulation/vehicle (Fig. 1).

      Study procedures

       Acustimulation and pharmacological prophylaxis

      Acustimulation was provided by a commercially available device, the ReletexTM (Neurowave Medical Technologies, Chicago, IL, USA), which was applied to the P6 acupoint on the dominant upper extremity.
      • Dundee J.W.
      • Ghaly R.G.
      • Bill K.M.
      • Chestnutt W.N.
      • Fitzpatrick K.T.
      • Lynas A.G.
      Effect of stimulation of the P6 antiemetic point on postoperative nausea and vomiting.
      Sham acustimulation involved an inactivated acustimulation device prepared by inactivating the electrodes with a silicone cover. Vehicle was defined as saline instead of dexamethasone. Dexamethasone is approved in Germany for PONV prophylaxis and is frequently used when anaesthesia is maintained with isoflurane and nitrous oxide; it has equal efficacy compared with ondansetron and droperidol.
      • Apfel C.C.
      • Korttila K.
      • Abdalla M.
      • et al.
      A factorial trial of six interventions for the prevention of postoperative nausea and vomiting.
      On the day before surgery, sealed, opaque, individually numbered envelopes were prepared, which contained a data sheet with information on group allocation and the acustimulation device. Envelopes were opened immediately before induction of anaesthesia by the responsible anaesthetist. After induction of anaesthesia, acustimulation or sham acustimulation devices were activated (31 Hz, strength Grade III)
      • Dundee J.W.
      • Ghaly R.G.
      • Bill K.M.
      • Chestnutt W.N.
      • Fitzpatrick K.T.
      • Lynas A.G.
      Effect of stimulation of the P6 antiemetic point on postoperative nausea and vomiting.
      and subjects received either dexamethasone (4 mg ml1, mibe GmbH, Brehna, Germany) or saline 4 ml according to treatment group allocation to maintain subjects blinding. Acustimulation devices remained in situ for 24 h.

       Anaesthesia

      Induction was performed with fentanyl (3–5 μg kg−1) and either thiopental (5 mg kg−1), propofol (1–2 mg kg−1), or etomidate (0.2–0.4 mg kg−1) as hypnotic agent at the discretion of the responsible anaesthetist according to the subjects' comorbidities and personal preference, and rocuronium (0.6 mg kg−1). For rapid sequence inductions, succinylcholine (1 mg kg−1) or rocuronium (1 mg kg−1) were administered. Anaesthesia was maintained with isoflurane, 0.8–1.0% end-tidal in nitrous oxide (60–70%), and repetitive doses of fentanyl.

       Postanaesthetic care unit

      Patients with pain received morphine (0.1 mg kg−1 i.v.). A rescue therapy with granisetron (3 mg i.v.) was administered to patients experiencing an episode of nausea or vomiting, which is effective for existing PONV.
      • Kazemi-Kjellberg F.
      • Henzi I.
      • Tramer M.R.
      Treatment of established postoperative nausea and vomiting: a quantitative systematic review.
      • Meyer T.A.
      • Roberson C.R.
      • Rajab M.H.
      • Davis J.
      • McLeskey C.H.
      Dolasetron versus ondansetron for the treatment of postoperative nausea and vomiting.
      A consort diagram is presented in Fig. 1.

       Clinical outcome measures

      All investigators responsible for data collection were blind as to study group allocations and treatments administered. Occurrence of nausea, retching, and vomiting was recorded for 2 h in the postanaesthetic care unit (PACU), and at 6 and 24 h, as was administration of pain medication and antiemetics according to recommendations for PONV trials.
      • Apfel C.C.
      • Roewer N.
      • Korttila K.
      How to study postoperative nausea and vomiting.
      • Lerman J.
      Surgical and patient factors involved in postoperative nausea and vomiting.
      • Korttila K.
      The study of postoperative nausea and vomiting.
      Pain was recorded on a scale of 0–10 (0=no pain, 10=strongest pain), and cumulative pain medication was collected, and its administration categorised as yes/no.

       Genotyping

      Buccal mouth swabs were obtained before operation from all patients. DNA was extracted using standard techniques (QIAgen, Hilden, Germany). The CHRM3 SNP (rs2165870) was genotyped using a Genotyping Array (Assay ID: C___9774837_30, Life Technologies, Carlsbad, CA, USA) as recommended by the manufacturer using a StepOne Real-Time PCR System (Applied Biosystems, Darmstadt, Germany).

       Sample size and statistical analysis

      With a sample size of 472, we intended to investigate whether the CHRM3 SNP (rs2165870) is independently associated with PONV, which was a primary aim of the study. We thus performed a post hoc power analysis based on data from Janicki and colleagues,
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      who showed an OR for the development of PONV of 3.1 in the pooled group and of 2.3 in the verification cohort for the CHRM3 SNP (rs2165870), a minor allele frequency of 0.34 for the A allele in Caucasian subjects,
      • Auton A.
      • Brooks R.M.
      • Durbin L.D.
      • et al.
      1000 Genomes Project Consortium
      A global reference for human genetic variation.
      and an anticipated PONV incidence of at least 30%
      • Cohen M.M.
      • Duncan P.G.
      • DeBoer D.P.
      • Tweed W.A.
      The postoperative interview: assessing risk factors for nausea and vomiting.
      for our population. With a sample size of 472, the most conservative power calculation revealed a power of 83.5% given an alpha error of 0.05, sufficient for detection of putative association of CHRM3 SNP (rs2165870) with PONV in our cohort.
      From 472 subjects completing the study protocol, for 18 subjects there was insufficient DNA in buccal swabs (n=6) or genotyping for CHRM3 rs2165870 was unsuccessful (n=12) (Fig. 1). Therefore, the following analysis was done with 454 patients. The CHRM3 (rs2165870) SNP was tested with Hardy–Weinberg expectations and no evidence for a significant deviation was detected.

       Observational study

      Descriptive statistics were summarised for categorical variables as frequencies (%) and compared between groups treatments using the χ2 test. Parametric variables are presented as mean (standard deviation) and were compared by analysis of variance in case of normal distribution. Otherwise, non-parametric variables were presented as median (range) and compared using the Kruskal–Wallis test. Correlation of nausea and retching/vomiting (PONV) with postoperative morphine administration and pain in the PACU was calculated with binary logistic regression analysis. Frequency of nausea, vomiting, and PONV in figures is presented as mean [95% confidence interval (CI)]. Log-binomial regression analysis was done using risk factors of the Apfel score (smoking status, history of motion sickness, history of PONV, postoperative opioid usage), with variables known to be associated with PONV (age, hypnotic induction agent, and anaesthesia duration
      • Apfel C.C.
      • Heidrich F.M.
      • Jukar-Rao S.
      • et al.
      Evidence-based analysis of risk factors for postoperative nausea and vomiting.
      ) and the CHRM3 rs2165870 SNP as exploratory variables using an additive genetic model to estimate the relative risk (RR). Log-binomial regression analysis with the Apfel score and the CHRM3 rs2165870 SNP as exploratory variables was performed to identify and compare the impact on PONV risk. An interaction model with main effects ‘Apfel score’ and ‘CHRM3 rs2165870 SNP’ and their interaction term was performed to analyse the interaction between these two variables. Log-binomial regression analysis was also done with the CHRM3 SNP, Apfel score, and group allocation as independent variables.

       Randomised controlled trial

      We investigated whether PONV prophylaxis mitigates risk for PONV among subjects carrying a genetic variant previously associated with PONV. RR reduction (RRR) was calculated by the percent reduction in risk with the various treatment allocations (acustimulation/dexamethasone, acustimulation/vehicle, sham acustimulation/dexamethasone) compared with the control group (sham acustimulation/vehicle). Association of CHRM3 rs2165870 genotypes with PONV was analysed by the χ2 test for trend.
      Differences were regarded statistically significant with an a priori alpha error of <0.05. Statistical analyses were two-sided and performed using SPSS, version 22.0 (SPSS, Chicago, IL, USA) or Graph Pad Prism, version 6.0 (Graph Pad Software, La Jolla, CA, USA). Power analysis was done with PS Power and Sample Size Calculations, Version 3.0 (Department of Biostatistics, Vanderbilt University, TN, USA).
      • Dupont W.D.
      • Plummer Jr., W.D.
      Power and sample size calculations. A review and computer program.
      • Dupont W.D.
      • Plummer Jr., W.D.
      Power and sample size calculations for studies involving linear regression.

      Results

       Subject characteristics

      No significant differences were found regarding height, weight, gender, smoking status, history of motion sickness, history of PONV, Apfel score, ASA status, intraoperative fentanyl dosage, and anaesthesia duration according to CHRM3 rs2165870 SNP (Table 1). However, GG genotype carriers were older (median: 47 yr, range: 18–83) compared with GA (median: 44 yr, range: 17–83) and AA (median: 39 yr, range: 18–86) genotype carriers (P=0.041) (Table 1).
      Table 1Subject characteristics and risk factors for postoperative nausea and vomiting (PONV) for whole cohort and stratified for CHRM3 rs2165870 genotypes
      All genotypesGGGAAA
      Number (n)45419120756
      Age (yr)45 (17–86)47 (18–83)44 (17–83)39 (18–86)
      Body height (cm)169 (145–198)169 (150–198)168 (145–196)169 (156–198)
      Body weight (kg)72 (43–140)74 (44–120)71 (43–140)68 (43–118)
      Gender (f/m)317/137132/59145/6240/16
      Smoker (n; %)151 (33.3)55 (28.8)79 (38.3)17 (30.4)
      History of motion sickness (n; %)103 (22.8)43 (22.8)50 (24.3)10 (17.9)
      History of PONV (n; %)94 (20.8)36 (19)49 (23.8)9 (16.1)
      Apfel score (n, %)
       0–1103 (22.7)43 (22.5)48 (23.2)12 (21.4)
       2156 (34.4)66 (34.6)71 (34.3)19 (34)
       3139 (30.6)56 (29.3)63 (30.4)20 (35.7)
       456 (12.3)26 (13.6)25 (12.1)5 (8.9)
      ASA physical status (n; %)
       1112 (24.7)39 (20.4)60 (29)13 (23.2)
       2298 (65.6)133 (69.6)126 (60.9)39 (69.7)
       344 (9.7)19 (10)21 (10.1)4 (7.1)
      Intraoperative fentanyl dose (mg)0.25 (0.1–1.1)0.25 (0.15–0.8)0.25 (0.1–1.1)0.25 (0.15–0.7)
      Anaesthesia duration (min)107 (30–311)108 (36–288)104 (30–311)114.5 (39–194)
      Surgical type (n, %)
       Gynaecology159 (35)71 (37.2)68 (32.9)20 (35.7)
       Ear, nose, and throat280 (61.7)113 (59.1)133 (64.2)34 (60.7)
       Thyroid15 (3.3)7 (3.7)6 (2.9)2 (3.6)
      Study groups (n, %)
       Acustimulation/vehicle113 (24.9)50 (26.2)49 (23.7)14 (25)
       Acustimulation/dexamethasone122 (26.9)48 (25.1)57 (27.5)17 (30.4)
       Sham acustimulation/vehicle110 (24.2)46 (24.1)52 (25.1)12 (21.4)
       Sham acustimulation/dexamethasone109 (24)47 (24.6)49 (23.7)13 (23.2)

       The CHRM3 rs2165870 polymorphism

      The CHRM3 rs2165870 genotype distribution was GG in 191 subjects, GA in 207 subjects, and AA in 56 subjects resulting in a minor allele frequency of 0.35 for the A allele.

       Observational study

      PONV (nausea and retching/vomiting) in the PACU correlated significantly with pain (P=0.001) and tended to increase not significantly with morphine dosage in the PACU (P=0.055). As pain leads to various physiological and pathophysiological changes and can overlay any genetic association, we only regarded PONV after 2 h, which did not show such a correlation.
      We thus focused on early PONV (2–6 h) which had the highest incidence (Fig. 2) and demonstrated a significant association of the CHRM3 rs2165870 SNP with nausea (P=0.021), retching (P=0.001), and PONV (P=0.006) (Table 2).
      Figure thumbnail gr2
      Fig 2Occurrence of nausea, retching/vomiting, and nausea and retching/vomiting combined (PONV) in the PACU, 2-6, and 6-24 hours after anaesthesia and surgery. Data are presented as means ± 95% confidence interval.
      Table 2Association of postoperative nausea and vomiting (PONV) (2–6 h) stratified for CHRM3 rs2165870 genotypes
      All genotypesGGGAAAP-value
      Nausea (n; %)139 (30.6)45 (23.6)74 (35.7)20 (35.7)0.021
      Retching/vomiting (n; %)117 (25.8)32 (16.8)66 (31.9)19 (33.9)0.001
      Nausea and retching/vomiting combined (PONV) (n; %)169 (37.2)55 (28.8)88 (42.5)26 (46.4)0.006

       PONV according to CHRM3 rs2165870 genotype

      Univariate pairwise comparison of subjects revealed a 30% greater risk for PONV in homozygous AA genotypes (RR=1.3, 95% CI: 1–1.7; P=0.016), while heterozygous GA genotype carriers had a 20% increased risk for PONV compared with GG genotypes (RR=1.2, 95% CI: 1.1–1.4; P=0.005), suggesting a gene-dose effect.

       Regression models

      Log-binomial regression analyses including smoking status, history of motion sickness, history of PONV, postoperative opioid usage, age, hypnotic induction agent, anaesthesia duration, and CHRM3 rs2165870 SNP as exploratory variables revealed that age, smoking status, postoperative opioid usage, hypnotic induction agent, and CHRM3 rs2165870 SNP (GA vs GG) were all independent predictors for PONV (Table 3). When only the Apfel score and CHRM3 rs2165870 polymorphisms were used as exploratory variables, the Apfel score [RR per Apfel score point: 1.3 (1.2–1.5), P<0.0001] and CHRM3 SNP (GA vs GG: RR=1.5, 95% CI: 1.1–1.9; P=0.003, and AA vs GG: RR=1.6, 95% CI: 1.1–2.2; P=0.009) independently contributed to PONV risk. There was no interaction between Apfel score and CHRM3 rs2165870 SNP (P=0.6). Stratification according to Apfel score revealed in subjects with a low risk for PONV (Apfel score 0–2) an association of the AA genotype with a PONV risk of 42% compared with 32% in GA and 20% in GG subjects (P=0.008), while overall PONV incidence in the group with low risk for PONV averaged 28%. Log-binomial regression analysis with CHRM3 SNP, Apfel score, and group allocation as independent variables revealed a significant association of PONV with group allocation, SNP, and the Apfel score. RR for the GA and the AA genotype compared with GG genotype was similar to log-binomial regression analysis with the SNP and the Apfel score as independent variables.
      Table 3Log-binomial regression analyses for known risk factors and the CHRM3 rs2165870 single nucleotide polymorphism on development of postoperative nausea and vomiting (PONV) in the whole study group. RR, relative risk
      PONV risk factorRR (lower 95%–upper 95%)P-value
      Age (per decade)0.9 (0.9–1.0)0.003
      Anaesthesia duration (h)1.1 (1–1.2)0.3
      Hypnotic agent
       Propofol1
      Reference group.
       Thiopental or etomidate1.4 (1.1–1.8)0.01
      Sex
       Male1
      Reference group.
       Female1.3 (0.9–1.7)0.13
      Current smoker
       Yes1
      Reference group.
       No1.4 (1.1–1.8)0.017
      History of motion sickness
       No1
      Reference group.
       Yes1 (0.8–1.3)0.89
      History of PONV
       No1
      Reference group.
       Yes1.2 (0.9–1.6)0.17
      Postoperative opioid use
       No1
      Reference group.
       Yes1.3 (1–1.7)0.048
      CHRM3 rs2165870 SNP
       GG1
      Reference group.
       GA1.5 (1.2–2)0.002
       AA1.4 (1–2)0.06
      a Reference group.

       Randomised controlled trial

       Treatment group distribution

      According to group allocation, 110 subjects received no prophylaxis (sham acustimulation and vehicle). Prophylaxis with dexamethasone or acustimulation was used in 109 and 113 subjects, respectively, and 122 subjects received combined therapy of dexamethasone and acustimulation (Fig. 1).

       PONV prophylaxis according to treatment group

      In the AA genotype with an Apfel score of 0–2 (n=31), PONV incidence without prophylaxis was 71%, while the incidence with acustimulation or dexamethasone was 50% or 50%, respectively (Fig. 3). Therefore, both acustimulation and dexamethasone were associated with a 30% RRR for PONV. Combined prophylaxis with acustimulation and dexamethasone lead to an RRR of 86%, so PONV incidence in AA genotype carriers was reduced to 10% (P=0.015).
      Figure thumbnail gr3
      Fig 3Occurrence of PONV as stratified by CHRM3 rs2165870 genotypes (AA vs. GG/GA) in patients with an Apfel Score of 0–2, comparing the treatment interventions. Relative risk reduction (RRR) is given for each treatment when compared to the control group (Sham acustimulation/vehicle). Data are presented as means ± 95% confidence interval. Association of CHRM3 rs2165870 genotypes with PONV was analysed by the X2-test for trend.
      PONV prophylaxis also reduced the PONV risk in GG/GA genotype carriers with an Apfel score of 0–2 albeit to a lesser extent (Fig. 3). PONV incidence for the GG/GA genotypes without prophylaxis was 34%, while the incidences with acustimulation, dexamethasone, and combined prophylaxis were 28, 26, and 16%, respectively (P=0.046) (Fig. 3). Analysis of PONV prophylaxis in the whole study group (Apfel score 0–4) revealed a trend to the described effect that is observed in Apfel score 0–2 patients, but this effect was less strong (Supplementary Fig. S1) because of variable results in Apfel score 3–4 patients (Supplementary Fig. S2).

      Discussion

      We show that the rs2165870 SNP, located upstream of the promoter region of the M3 muscarinic acetylcholine receptor (CHRM3), and the Apfel score contribute independently to PONV susceptibility. Furthermore, among subjects found to have a low risk for PONV based on Apfel score, but carrying a genetic variant associated with PONV, combined prophylaxis with acustimulation and dexamethasone was effective in preventing PONV. This highlights both the impact of genetic predisposition to PONV and the efficacy of PONV prophylaxis even in patients with a low Apfel score.
      To our knowledge, the study from Janicki and colleagues
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      is the only GWAS aimed to identify genetic risk variants associated with PONV. Only few SNPs have been hitherto associated with PONV, such as the dopamine D2 receptor Taq IA SNP
      • Nakagawa M.
      • Kuri M.
      • Kambara N.
      • et al.
      Dopamine D2 receptor Taq IA polymorphism is associated with postoperative nausea and vomiting.
      and variations in the HTR3A and HTR3B genes.
      • Rueffert H.
      • Thieme V.
      • Wallenborn J.
      • et al.
      Do variations in the 5-HT3A and 5-HT3B serotonin receptor genes (HTR3A and HTR3B) influence the occurrence of postoperative vomiting?.
      However, a further study focussing on the dopamine D2 receptor Taq IA SNP revealed only an association with a history of PONV.
      • Frey U.H.
      • Schnee C.
      • Achilles M.
      • Silvanus M.T.
      • Esser J.
      • Peters J.
      Postoperative nausea and vomiting: the role of the dopamine D2 receptor TaqIA polymorphism.
      Therefore, the CHRM3 rs2165870 SNP is the only SNP detected in a GWAS associated with PONV in two independent cohorts. Our results further confirm the association of the CHRM3 rs2165870 SNP with PONV.
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      While in the study of Janicki and colleagues,
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      DNA data pooling analysis revealed an OR of 3.1 and an OR of 2.3 for the A/G substitution in the verification cohort,
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      we found an RR of 1.5 for GA vs GG and of 1.6 for AA vs GG genotypes. Irrespective of the difference in reporting RR or ORs, inclusion criteria differed between these two studies. Recruitment criteria in the study from Janicki and colleagues
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      for the PONV group included an extensive history of PONV after at least three separate surgeries. In our study cohort, a history of PONV was not an inclusion criterion, which might explain differences in the observed risk between these studies. Our cohort also represents a more common type of patients. Furthermore, Janicki and colleagues
      • Janicki P.K.
      • Vealey R.
      • Liu J.
      • Escajeda J.
      • Postula M.
      • Welker K.
      Genome-wide Association study using pooled DNA to identify candidate markers mediating susceptibility to postoperative nausea and vomiting.
      used a dominant genetic model to study the effects of the CHRM3 rs2165870 SNP, whereas we observed a gene-dose effect and analysis by an additive model.
      The minor A allele frequency of the CHRM3 rs2165870 SNP in our study cohort of 0.35 is in line with a reported minor A allele frequency of 0.34 in Caucasian subjects.
      • Auton A.
      • Brooks R.M.
      • Durbin L.D.
      • et al.
      1000 Genomes Project Consortium
      A global reference for human genetic variation.
      Age was significantly different in the three genotype groups, GG genotypes being older than GA and AA genotypes. However, log-binomial regression analyses revealed a significant association of PONV with the CHRM3 rs2165870 SNP despite correction for age. Thus, both age and the polymorphism were, among others, independent predictors for PONV.
      Anaesthesia induction was performed with thiopental, propofol, or etomidate at the discretion of the anaesthetist in charge. Beyond that, the anaesthetic regime was standardised. Accordingly, the induction agent was included in the log-binomial regression analysis, which showed that rs2165870 was still significantly associated with PONV despite use of several induction agents.
      The Apfel score and the CHRM3 rs2165870 SNP had independent impacts on PONV susceptibility, highlighting the importance of genetic variation for PONV. This finding might also explain why patients with a low Apfel score can still experience PONV. Whether patients should receive PONV prophylaxis or not is currently based mainly on the Apfel score. As both acustimulation and dexamethasone were able to decrease PONV in patients with a genetic variant associated with PONV phenotype, both pharmacological prophylaxis and acustimulation might be applied to patients with the genetic variant. The combination of dexamethasone and acustimulation had an additive effect and significantly decreased the risk of PONV by 86% in patients with the genetic variant but a low Apfel score. Our results present an option for PONV prophylaxis in patients with a genetic variant associated with PONV, and confirm recent studies emphasising combined prophylaxis as an important strategy against PONV.
      • Yang X.Y.
      • Xiao J.
      • Chen Y.H.
      • et al.
      Dexamethasone alone vs in combination with transcutaneous electrical acupoint stimulation or tropisetron for prevention of postoperative nausea and vomiting in gynaecological patients undergoing laparoscopic surgery.
      As genetic testing before anaesthesia, which might be a reasonable option in the future, is not yet implemented in daily practice, questions about the clinical relevance of our findings arise. Only 12% of patients had a low Apfel score but carried the genetic variant associated with PONV. PONV guidelines
      • Gan T.J.
      • Meyer T.A.
      • Apfel C.C.
      • et al.
      Society for Ambulatory Anesthesia guidelines for the management of postoperative nausea and vomiting.
      suggest a multimodal approach to patients at high risk for PONV. Applying these criteria, patients at a high risk of PONV based only on their genetic predisposition would also deserve such a multimodal approach, although only 12% of patients with a low Apfel score would be the respective target population. In the absence of genetic testing and the presumed low incidence of side effects of antiemetics, acustimulation, or both, an implication might be antiemetic prophylaxis of all patients irrespective of their Apfel score. Clearly, further studies are needed to support such a far reaching implication.
      There are limitations to this study. We included a broad spectrum of patients who underwent anaesthesia in different surgical departments (gynaecology, ear, nose and throat, or endocrine). Although this represents a common patient mix, no conclusions can be drawn regarding other types of more extensive surgery. The ‘blinding’ of our study should be noted. Subjects were blinded to their treatment which started after induction of anaesthesia. Although the responsible anaesthesiologist was not blinded to treatment, the investigators responsible for data collection were blinded to study group allocations and treatments. The study was not powered to answer the secondary aim of the study, analysis of PONV prophylaxis in patients carrying the genetic variant associated with PONV. Therefore, further studies are needed to clarify our findings and to verify recommendations for clinical practice.
      In summary, we demonstrated that a genetic variation (the rs2165870 SNP in CHRM3) and the Apfel score independently contribute to PONV susceptibility. Furthermore, combined PONV prophylaxis markedly decreased the risk of PONV in subjects who were at low risk based on the Apfel score but had a genetic variant associated with PONV.

      Authors' contributions

      Conceived and designed the study, and wrote the manuscript: S.K., J.P., U.H.F.
      Analysed and interpreted the data: S.K., U.H.F.
      Recruitment of patients and collection of data: G.J.d.V., L.S., N.S.
      Genetic analysis and power analysis: H.S.B.

      Acknowledgements

      We thank J. Rekowski from the Institute for Medical Informatics, Biometry and Epidemiology, University of Duisburg-Essen, Essen, Germany for statistical assistance.

      Funding

      The Reletex TM device was provided by Neurowave Medical Technologies (Chicago, IL, USA).

      Declaration of interest

      None declared.

      Appendix A. Supplementary data

      Figure thumbnail figs1
      Supplementary Figure 1Occurrence of PONV as stratified by CHRM3 rs2165870 genotypes (AA vs. GG/GA) in the whole study group (Apfel Score of 0–4), comparing the treatment interventions. Data are presented as means ± 95% confidence interval.
      Figure thumbnail figs2
      Supplementary Figure 2Occurrence of PONV as stratified by CHRM3 rs2165870 genotypes (AA vs. GG/GA) in patients with an Apfel Score of 3–4, comparing the treatment interventions. Data are presented as means ± 95% confidence interval.

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