irritable bowel syndrome ( ibs ) is a heterogeneous , idiopathic , gastrointestinal syndrome characterized by abdominal pain or discomfort , diarrhea , and/or constipation . a questionnaire survey conducted in students from a chinese university found that only 5.7% of respondents could be diagnosed with ibs , but epidemiological surveys in india , iran , japan , and other asian countries suggest that ibs is more prevalent in elderly people than in young adults [ 24 ] . questionnaires and related testing of elderly people in hong kong diagnosed 13.1% with ibs , and epidemiological investigation in the songjiang district of shanghai found a prevalence of 9.4% in adults aged 6079 years old and 10.2% in those over 80 years of age . although the etiology of ibs is currently unknown , several hypotheses have been proposed for its pathogenesis . the development of ibs may involve visceral abnormalities , genetic susceptibility , a neuro - immunological disorder , and/or psychological factors . ibs shows a familial tendency , and the mutations involved in genetic susceptibility to ibs have recently received particular attention . however , identifying genetic markers of ibs is complicated by the multi - factorial pathophysiology of this syndrome . it is hypothesized that exposure to certain environmental factors , such as infection or severe psychological stress , may induce ibs in genetically susceptible individuals . polymorphisms in the 5-hydroxytryptamine ( 5-ht ) transporter or receptors , adrenergic system , and mitochondrial genome have been highlighted as potentially contributing to abnormalities in ibs patients , including enhanced perception of gut stimuli , altered gastrointestinal motor function , and psychiatric symptoms . several neurotransmitter receptors involved in the regulation of gastrointestinal motility , such as the 5-ht receptor , adrenergic receptor , and cannabinoid receptor , are g protein - coupled receptors . one g protein subunit , g protein beta polypeptide 3 ( gn3 ) , is a component of several g protein complexes , and thus polymorphisms in the gn3 gene impact signal transduction from several receptors . a common single nucleotide polymorphism substitution of c with t at position 825 within exon 10 of the gn3 gene ( rs5443 ) results in a truncated gn3 splice variant with enhanced g protein activation . this polymorphism has been reported to be overrepresented in patients diagnosed with some forms of ibs [ 1217 ] . sympathetic dysfunction of the adrenergic nerves has also been associated with the pathogenesis of some cases of ibs [ 1820 ] . dysfunction of catechol - o - methyltransferase ( comt ) , an enzyme capable of degrading catecholamines such as dopamine , norepinephrine , and epinephrine , appears to play a role in the pathogenesis of some cases of ibs [ 1820 ] . a common single nucleotide polymorphism substitution at position 1947 in exon 4 of the comt gene results in the substitution of valine 158 with methionine in the comt protein ( a / a genotype ) , which reduces the activity of comt by about 4-fold . however , comt with val 158 ( g / g genotype ) appears to have higher activity . recent studies have demonstrated that comt gene polymorphisms are correlated to chronic pain and depression . although ibs patients often have abdominal pain accompanied by anxiety or depression , few studies have investigated the relationship between comt gene polymorphisms and ibs . preliminary investigations have found that individuals with 2 copies of the gene producing comt with valine at position 158 are over - represented in groups diagnosed with ibs . we sought to determine whether these single nucleotide polymorphisms in gn3 and comt were associated with ibs in elderly chinese patients . due to the likely multi - factorial nature of ibs , we sub - divided patients diagnosed with ibs according to their symptoms and determined the frequency of these snps in these groups and an age - matched control group unaffected by gastrointestinal disease . between march and december 2012 , 66 patients over 60 years old were enrolled at huashan hospital ( shanghai , china ) . all patients were diagnosed with ibs according to the rome iii diagnostic criteria   and were further subdivided into 3 groups based on symptom presentation : 1 ) ibs with constipation ( ibs - c ) , 2 ) ibs with diarrhea ( ibs - d ) , and 3 ) ibs with both symptoms ( ibs - m ) . patients with liver , gallbladder or pancreatic disease , diabetes , hyperthyroidism , cancer , or any other disease were excluded by examination of medical records , physical examination , ultrasound , and blood biochemical analysis . patients with history of abdominal surgery or adverse drug reactions , including constipation and diarrhea , were also excluded from this study . we enrolled 115 age - matched subjects without ibs as a control group during the same period at routine annual physical examination . subjects in the control group defecated once or twice a day , or once every 2 days , producing yellow , soft , solid stools , and did not experience hard bowel movements , abdominal pain , or discomfort . all participants provided written informed consent , and the study was approved by the ethics committee of huashan hospital , fudan university [ 2012(229 ) ] . a professionally trained physician evaluated all participants with the geriatric depression scale ( gds ) , a set of 30 polar questions used to quantify participant emotional state during the preceding week . for each question , a positive response is assigned 1 point , and a negative response is assigned no points . genomic dna was extracted using qiaamp mini - spin columns ( qiagen , usa ) according to the manufacturer s instructions and separated by agarose gel electrophoresis for snp determination . the snp sequences of gnb c825 t ( rs 5443 ) and comt val158met ( rs 4680 ) were based on the gene bank databases . primers and probes were designed using abi primer express software v2.0 and synthesized by invitrogen , as detailed in table 1 . the pcr reaction was carried out in a steponeplus real - time pcr system ( applied biosystems , foster city , ca , usa ) and subjected to 95c for 2 min , then 40 cycles of 95c for 10 s , followed by 53c for 45 s. fluorescence was measured by the real - time pcr system and data was converted by sds 2.0 software . the chi - square test and hardy - weinberg equilibrium theory were used to compare the genotype and allele frequency between the ibs and control groups , and between ibs subgroups . chi - square test and fisher s exact test were used to assess the relationship between clinical factors and genotype . unconditional logistic regression analysis and crossover analysis statistical analysis was performed with spss 13.0 statistical software and p - values < 0.05 was considered statistically significant . between march and december 2012 , 66 patients over 60 years old were enrolled at huashan hospital ( shanghai , china ) . all patients were diagnosed with ibs according to the rome iii diagnostic criteria   and were further subdivided into 3 groups based on symptom presentation : 1 ) ibs with constipation ( ibs - c ) , 2 ) ibs with diarrhea ( ibs - d ) , and 3 ) ibs with both symptoms ( ibs - m ) . patients with liver , gallbladder or pancreatic disease , diabetes , hyperthyroidism , cancer , or any other disease were excluded by examination of medical records , physical examination , ultrasound , and blood biochemical analysis . patients with history of abdominal surgery or adverse drug reactions , including constipation and diarrhea , were also excluded from this study . we enrolled 115 age - matched subjects without ibs as a control group during the same period at routine annual physical examination . subjects in the control group defecated once or twice a day , or once every 2 days , producing yellow , soft , solid stools , and did not experience hard bowel movements , abdominal pain , or discomfort . all participants provided written informed consent , and the study was approved by the ethics committee of huashan hospital , fudan university [ 2012(229 ) ] . a professionally trained physician evaluated all participants with the geriatric depression scale ( gds ) , a set of 30 polar questions used to quantify participant emotional state during the preceding week . for each question , a positive response is assigned 1 point , and a negative response is assigned no points . genomic dna was extracted using qiaamp mini - spin columns ( qiagen , usa ) according to the manufacturer s instructions and separated by agarose gel electrophoresis for snp determination . the snp sequences of gnb c825 t ( rs 5443 ) and comt val158met ( rs 4680 ) were based on the gene bank databases . primers and probes were designed using abi primer express software v2.0 and synthesized by invitrogen , as detailed in table 1 . the pcr reaction was carried out in a steponeplus real - time pcr system ( applied biosystems , foster city , ca , usa ) and subjected to 95c for 2 min , then 40 cycles of 95c for 10 s , followed by 53c for 45 s. fluorescence was measured by the real - time pcr system and data was converted by sds 2.0 software . the chi - square test and hardy - weinberg equilibrium theory were used to compare the genotype and allele frequency between the ibs and control groups , and between ibs subgroups . chi - square test and fisher s exact test were used to assess the relationship between clinical factors and genotype . unconditional logistic regression analysis and crossover analysis were used to analyze the risk factors for ibs disease by gene polymorphism . statistical analysis was performed with spss 13.0 statistical software and p - values < 0.05 was considered statistically significant . participant demographics are shown in table 2 . the genomic distribution of the gn3 c825 t and comt val158met polymorphisms in the control group did not significantly deviate from the hardy - weinberg equilibrium ( p > 0.05 ) . gn3 genotyping determined that the c allele frequency was 85.6% in the ibs group and 89.1% in the control group , and the t allele frequency was 14.4% in the ibs group and 10.9% in the control group ( table 3 ) . the cc genotype of gn3 was found in 75.8% of those enrolled in the ibs group ( 50 instances ) in comparison to 80% of the control group ( 92 instances ) . the ct genotype was present in 19.7% ( 13 instances ) of the ibs group and 18.3% ( 21 instances ) of the control group , and the tt genotype was present in 4.5% ( 3 instances ) of the ibs group and 1.7% ( 2 instances ) of the control group ( table 4 ) . there was no statistically significant difference between allele frequency ( p=0.323 ) or the genotype distribution ( p=0.534 ) between the ibs and control groups . comt genotyping determined that the g allele frequency was 87.1% in the ibs group and 93.5% in the control group , and the a allele frequency was 12.9% in the ibs group and 6.5% in the control group ( table 5 ) . the gg genotype of comt was found in 86.4% of those enrolled in the ibs group ( 57 instances ) in comparison to 87.8% of the control group ( 101 instances ) . the ga genotype was present in 1.5% ( 1 instance ) of the ibs group and 11.3% ( 13 instances ) of the control group , and the aa genotype was present in 12.1% ( 8 instances ) of the ibs group and 0.9% ( 1 instance ) of the control group ( table 6 ) . the a allele was found significantly more frequently in the ibs group than in the control group ( p=0.040 ) , and the genotype distribution also differed significantly between the ibs group and the control group ( p < 0.001 ) . ibs patients were divided into subgroups according to symptom presentation . seven patients ( 10.6% ) had constipation ( ibs - c ) , 46 patients ( 69.7% ) had diarrhea ( ibs - d ) , and 13 patients ( 19.7% ) had both symptoms ( ibs - m ) ( tables 79 ) . no statistically significant differences in gn3 genotype or allele frequency were found between the control group and any of the ibs subgroups ( tables 79 ) . however , the genotype distribution of comt val158met differed significantly between the control and ibs - d groups ( p<0.001 ) ( table 8) , and this subgroup had a higher a allele frequency ( p=0.029 ) when compared with the control group ( table 8) . no significant difference in comt genotype distribution or allele frequency was detected between the ibs - m or ibs - c subgroups and the control group ( tables 7 and 9 ) . as indicated in table 10 , ibs patients were subdivided into 2 subgroups based on the duration of their ibs symptoms ; 39.4% of enrolled ibs patients had symptoms for longer than 5 years , and 60.6% for less than 5 years . although no difference was detected in the genotype distribution or allele frequency of gnb3-c825 t , the allele frequency of comt 158a was elevated in patients with ibs symptoms for longer than 5 years ( p=0.022 , table 10 ) . after applying non - conditional logistic regression analysis and adjusting for sex , age , and gds score , crossover analysis showed that the effect of the gn3 gene or the comt gene was not statistically significant among ibs patients ( p>0.05 , table 11 ) . interaction analysis indicated no interaction between the gn3 gene and the comt gene in ibs patients ( p>0.05 , table 12 ) . participant demographics are shown in table 2 . the genomic distribution of the gn3 c825 t and comt val158met polymorphisms in the control group did not significantly deviate from the hardy - weinberg equilibrium ( p > 0.05 ) . gn3 genotyping determined that the c allele frequency was 85.6% in the ibs group and 89.1% in the control group , and the t allele frequency was 14.4% in the ibs group and 10.9% in the control group ( table 3 ) . the cc genotype of gn3 was found in 75.8% of those enrolled in the ibs group ( 50 instances ) in comparison to 80% of the control group ( 92 instances ) . the ct genotype was present in 19.7% ( 13 instances ) of the ibs group and 18.3% ( 21 instances ) of the control group , and the tt genotype was present in 4.5% ( 3 instances ) of the ibs group and 1.7% ( 2 instances ) of the control group ( table 4 ) . there was no statistically significant difference between allele frequency ( p=0.323 ) or the genotype distribution ( p=0.534 ) between the ibs and control groups . comt genotyping determined that the g allele frequency was 87.1% in the ibs group and 93.5% in the control group , and the a allele frequency was 12.9% in the ibs group and 6.5% in the control group ( table 5 ) . the gg genotype of comt was found in 86.4% of those enrolled in the ibs group ( 57 instances ) in comparison to 87.8% of the control group ( 101 instances ) . the ga genotype was present in 1.5% ( 1 instance ) of the ibs group and 11.3% ( 13 instances ) of the control group , and the aa genotype was present in 12.1% ( 8 instances ) of the ibs group and 0.9% ( 1 instance ) of the control group ( table 6 ) . the a allele was found significantly more frequently in the ibs group than in the control group ( p=0.040 ) , and the genotype distribution also differed significantly between the ibs group and the control group ( p < 0.001 ) . ibs patients were divided into subgroups according to symptom presentation . seven patients ( 10.6% ) had constipation ( ibs - c ) , 46 patients ( 69.7% ) had diarrhea ( ibs - d ) , and 13 patients ( 19.7% ) had both symptoms ( ibs - m ) ( tables 79 ) . no statistically significant differences in gn3 genotype or allele frequency were found between the control group and any of the ibs subgroups ( tables 79 ) . however , the genotype distribution of comt val158met differed significantly between the control and ibs - d groups ( p<0.001 ) ( table 8) , and this subgroup had a higher a allele frequency ( p=0.029 ) when compared with the control group ( table 8) . no significant difference in comt genotype distribution or allele frequency was detected between the ibs - m or ibs - c subgroups and the control group ( tables 7 and 9 ) . as indicated in table 10 , ibs patients were subdivided into 2 subgroups based on the duration of their ibs symptoms ; 39.4% of enrolled ibs patients had symptoms for longer than 5 years , and 60.6% for less than 5 years . although no difference was detected in the genotype distribution or allele frequency of gnb3-c825 t , the allele frequency of comt 158a was elevated in patients with ibs symptoms for longer than 5 years ( p=0.022 , table 10 ) . after applying non - conditional logistic regression analysis and adjusting for sex , age , and gds score , crossover analysis showed that the effect of the gn3 gene or the comt gene was not statistically significant among ibs patients ( p>0.05 , table 11 ) . interaction analysis indicated no interaction between the gn3 gene and the comt gene in ibs patients ( p>0.05 , table 12 ) . in this study we found that comt 158met allele , encoding an enzyme with reduced activity , was significantly associated with ibs in elderly chinese patients , and was particularly significantly associated with ibs accompanied by diarrhea and ibs that had persisted for longer than 5 years . we also found that ibs patients were significantly less likely to possess heterozygous comt genes than the control patients . the frequency of comt 158met in the ibs group was significantly higher than that in the control group , suggesting that the met allele predisposes elder people to ibs , and the val allele protects against ibs . our results are consistent with previous reports that found that sympathetic dysfunction of the adrenergic nerves is associated with the pathogenesis of some cases of ibs [ 1820 ] , but inconsistent with another study that found the comt 158val allele to be significantly associated with pathogenesis of ibs and increased frequency of defecation . we found no significant difference in the percentage of patients with a gds score above 10 in the ibs group and the control group , no significant difference in the genotype distribution or allele frequency of gn3-c825 t between the ibs and the control group , and no interaction between the gn3 gene and the comt gene in ibs patients . no difference in the genotype distribution and allele frequency of gn3-c825 t was detected when ibs patients were further subdivided according to symptom presentation and persistence of symptoms . our findings concerning gn3-c825 t further support the results of case - control studies conducted in the united states in 2007   and in korea in 2012 . however , several additional studies found different results in other populations . a 2011 study in a greek population found the gn3 - 825 t allele encoding a g protein that mediates enhances activation   to be closely associated with the occurrence of ibs . a 2001 study conducted in a caucasian population found that gn3 - 825c was associated with increased defecation frequency , and korean studies in 2010 and 2012 found gn3 - 825 t to be associated with ibs with constipation in adults   and children , and gn3 - 825c to be associated with ibs with diarrhea in children . we did not detect a statistically significant association of particular gn3 or comt genotypes with ibs with constipation , but our analysis may have been limited by sample size because we enrolled only a small number of patients ( 7 ) who experienced ibs with constipation . the inconsistent results of these studies may be partially explained to be relatively small sample sizes . for example , both kim   and saito   subdivided small populations ( 60 and 50 ibs patients , respectively ) into cc , ct , and tt genotype subgroups , resulting in a smaller size in each group . the studies that found association between gn3-c825 t and ibs were conducted in ethnic koreans   and greeks . fang et al . performed a correlation study between the gn3c825 t polymorphism and functional dyspepsia ( fd ) in a han chinese population , and found that this polymorphism is not a risk factor for the onset of fd . currently , both fd and ibs are thought to share a similar pathogenesis , and both are categorized as functional gastrointestinal diseases ( fgid ) . therefore , one can speculate that the c825 t polymorphism of gn3 may not be a risk factor for the pathogenesis of ibs in the chinese population . we found that the comt val158met single nucleotide polymorphism was associated with susceptibility to ibs , and the met allele was over represented in patients diagnosed with ibs with diarrhea or ibs that had persistsed for longer than 5 years . however , no evidence for a correlation between gn3 c825 t single nucleotide polymorphisms and the pathogenesis of ibs was found among this population of elderly ibs patients . to further validate genetic markers of ibs , larger cohorts will need to be enrolled , as identifying genetic markers of ibs is complicated by the multi - factorial pathophysiology of this heterogeneous syndrome .