ABSTRACT
Objective To explore the effects of pelvic nerves denervation (PND) on the expression of transit potential receptor vanilloid 1 (TRPV1) in distal colonic mucosa of rats.Methods The experimental study was conducted.One hundred and eight adult male rats were randomly divided into the control group,sham operation group and PND group:(1) 36 rats in the control group remained untreated and were fed regularly;(2) 36 in the sham operation group received open exclusion for 15 minutes,and then sew up the incision;(3) 36 in the PND group received laparotomy with pelvic nerve transection before abdominal closure.The expression of TRPV1 protein in distal colonic mucosa was respectively detected by Western blot at postoperative day 1,3 and 7.Reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect the mRNA level of TRPV1 in the distal colonic mucosa.Measurement data with normal distribution were represented as (x)±s.Repeated measurement data were analyzed by repeated measures ANOVA.Comparisons at the same time intervals among the 3 groups were analyzed using one-way ANOVA.Pairwise comparison was done by the independent samples t test.Results (1) The results of immunohistochemical staining:the average density of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 was respectively 0.180±0.016,0.179±0.015 and 0.183±0.026 in the control group,with no statistically significant difference (F=0.088,P＞0.05).The average density of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 was respectively 0.132±0.017,0.160±0.023 and 0.173±0.020 in the sham operation group,with a statistically significant difference (F=8.699,P＜0.05).The average density of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 was respectively 0.057± 0.009,0.122±0.016 and 0.180± 0.016 in the PND group,with a statistically significant difference (F =113.315,P ＜ 0.05).There were statistically significant differences in the average density of TRPV1 at postoperative day 1 and 3 among the 3 groups (F =108.960,15.218,P＜ 0.05),while significant differences were respectively detected between the control group and the sham operation group or the PND group at postoperative day 1 (t =5.025,15.979,P＜0.05),and a significant difference was also detected between the sham operation group and the PND group (t =9.590,P＜0.05).There was no statistically significant difference in the average density of TRPV1 between the control group and the sham operation group at postoperative day 3 (t =1.670,P＞0.05),while significant differences were respectively detected between the control group and the PND group and between the sham operation group and the PND group (t=6.543,3.361,P＜0.05).There was no statistically significant difference in the average density of TRPV1 at postoperative day 7 among the 3 groups (F=0.518,P＞0.05).(2) The results of Western blot:the relative expressions of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 were respectively 1.02±0.13,1.00±0.15 and 1.00±0.10 in the control group,with no statistically significant difference (F=0.084,P＞0.05).The relative expressions of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 were respectively 0.51±0.13,0.93±0.14 and 1.01±0.16 in the sham operation group,with a statistically significant difference (F =20.930,P＜0.05).The relative expressions of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 were respectively 0.30±0.10,0.70±0.10 and 1.07±0.16 in the PND group,with a statistically significant difference (F=61.441,P＜0.05).There were statistically significant differences in the relative expressions of TRPV1 at postoperative day 1 and 3 among the 3 group (F=58.014,8.841,P＜0.05),while significant differences were respectively detected between the control group and the sham operation group or the PND group at postoperative day 1 (t =6.677,11.145,P＜0.05),and significant difference was also detected between the sham operation group and the PND group (t =3.287,P＜ 0.05).There was no statistically significant difference in the relative expressions of TRPV1 between the control group and the sham operation group at postoperative day 3 (t =0.798,P＞0.05),while significant differences were respectively detected between the control group and the PND group and between the sham operation group and the PND group (t=4.127,3.398,P＜0.05).There was no statistically significant difference in the relative expressions of TRPV1 at postoperative day 7 among the 3 group (F=0.428,P＞0.05).(3) The results of RTqPCR:the mRNA levels of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 were respectively 1.00±0.15,1.10±0.21 and 1.09±0.18 in the control group,with no statistically significant difference (F=0.489,P＞0.05).The mRNA levels of TRPV1 in distal colonic mueosa at postoperative day 1,3 and 7 were respectively 0.58±0.12,0.99±0.19 and 1.13±0.23 in the shan operation group,with a statistically significant difference (F=13.964,P＜0.05).The mRNA levels of TRPV1 in distal colonic mucosa at postoperative day 1,3 and 7 were respectively 0.31±0.10,0.67±0.12 and 1.09±0.19 in the PND group,with a statistically significant difference (F=44.642,P＜0.05).There were statistically significant differences in the mRNA levels of TRPV1 at postoperative day 1 and 3 among the 3 group (F=44.653,9.700,P＜0.05),while significant differences were respectively detected between the control group and the sham operation group or the PND group at postoperative day 1 (t=5.233,9.264,P＜0.05),and significant difference was also detected between the sham operation group and the PND group (t=4.127,P＜0.05).There was no significant difference in the mRNA levels of TRPV1 between the control group and the sham operation group at postoperative day 3 (t =0.995,P＞0.05),while significant differences were respectively detected between the control group and the PND group and between the sham operation group and the PND group (t =4.411,3.505,P＜0.05).There was no statistically significant difference in the mRNA levels of TRPV1 at postoperative day 7 among the 3 group (F=0.099,P＞0.05).Conclusion The expression of TRPV 1 in distal colonic mucosa of rats is significantly down-regulated after pelvic nerves denervation,however,it is gradually recovered with passage of time,which is consistent with the trend of gradual recovery of colonic transit function after pelvic nerve injury.
ABSTRACT
Objective To investigate the effects of pelvic nerve transection on the colonic motility and the expression of transit receptor potential ankyrin 1 (TRPA1) in the colon mucosa.Methods Ninety-six Sprague-Dawley rats were divided into 3 groups based on a random number table:(1) 18 rats in the control group remained untreated and were fed regularly;(2) 39 rats in the sham operation group received laparotomy,and the pelvic nerves were stripped;(3) 39 rats in the operation group received laparotomy with pelvic nerve transection before abdominal closure.Colonic transit was assessed respectively at postoperative day 1,3,and 7 by injecting and calculating the geometric center (GC) of the distribution of 51Cr after 3 hours of propagation.The expression of TRPA1 in the colonic mucosa was determined by Western blot at postoperative day 1,3,7.Data with normal distribution were expressed by (x)± s,and were analyzed using the repeated measures ANOVA or LSD test.Results The GC values of the distribution of 51Cr in the sham operation group at postoperative day 1,3,7 were 5.8 ± 0.9,7.5 ± 0.5,7.3 ± 0.5,with a significant difference (F =9.508,P ＜ 0.05).The GC values of the distribution of 51Cr in the operation group at postoperative day 1,3,7 were 4.9 ± 0.4,5.6 ± 0.4,6.4 ± 0.8,with a significant difference (F =11.689,P ＜ 0.05).There were significant differences in the GC values of the distribution of 51 Cr at postoperative day 1 and 3 between the sham operation group and the operation group (t =2.227,7.144,P ＜ 0.05),while no significant difference was detected at postoperative day 7 (t =2.162,P ＞ 0.05).The results of Western blot showed that the relative expressions of TRPA1 in the proximal part of the colon at postoperative day 1,3,7 were 1.00 ± 0.05,1.00 ± 0.07,1.00 ± 0.06 in the control group,with a significant difference (F =0.055,P ＞ 0.05).The relative expressions of TRPA1 in the proximal part of the colon at postoperative day 1,3,7 were 0.78 ± 0.09,0.94 ± 0.08,0.95 ± 0.12 in the sham operation group,with a significant difference (F =5.651,P ＜ 0.05).The relative expressions of TRPA1 in the proximal part of the colon at postoperative day 1,3,7 were 0.37 ± 0.12,0.89 ± 0.10,0.92 ± 0.14 in the operation group,with a significant difference (F =41.005,P ＜0.05).There was significant difference in the relative expressions of TRPA1 in the proximal part of the colon among the 3 groups at postoperative day 1 (F =73.497,P ＜ 0.05),while significant differences were respectively detected between the control group and the sham operation group and the operation group at postoperative day 1 (t =4.224,11.954,P ＜ 0.05),and significant difference between the operation group and the sham operation group was also observed (t =7.730,P ＜ 0.05).There was no significant difference in the relative expression of TRPA1 in the proximal part of the colon among the 3 groups between day 3 and day 7 (F =2.087,0.656,P ＞ 0.05).The relative expressions of TRPA1 in the distal part of the colon at postoperative day 1,3,7 were 1.00 ± 0.05,1.00 ± 0.07,1.00 ± 0.06 in the control group,with no significant difference (F =0.055,P ＞ 0.05).The relative expressions of TRPA1 in the distal part of the colon at postoperative day 1,3,7were 0.68 ±0.11,0.98 ±0.12,1.11 ±0.16 in the sham operation group,with a significant difference (F =16.975,P ＜ 0.05).The relative expressions of TRPA1 in the distal part of the colon at postoperative day 1,3,7 were 0.39 ± 0.12,0.78 ± 0.10,1.06 ± 0.13 in the operation group,with a significant difference (F =50.417,P ＜ 0.05).There were significant differences in the relative expression of TRPA1 in the distal part of the colon among the 3 groups between day 1 and day 3 (F =58.773,8.680,P ＜ 0.05),while significant differences were respectively detected between the control group and the sham operation group and the operation group at postoperative day 1 (t =5.706,10.837,P ＜ 0.05),and significant difference was also detected between the operation group and the sham operation group (t =5.131,P ＜ 0.05).There was no significant difference in the relative expression of TRPA1 in the distal part of the colon between the control group and the sham operation group at postoperative day 3 (t =0.166,P ＞ 0.05),while significant differences were respectively detected between the control group and the operation group and between the sham operation group and the operation group at postoperative day 3 (t =3.694,3.528,P ＜ 0.05).There was no significant difference in the relative expression of the TRPA1 in the distal part of the colon between the 3 groups at postoperative day 7 (F =1.319,P ＞ 0.05).Conclusions Injury to pelvic nerves adversely affects colonic transit and expression of TRPA1 in mucosa.With a compensatory mechanism from the intestinal itself,these alterations in intestinal motility function normalize over time suggesting expression of TRPA1 in mucosa plays a crucial role in the recovery of intestinal motility function.