| Postoperative ileus(POI)is a common complication after surgery,especially abdominal surgery,which is the major cause of postoperative ileus.It lasts for 3-7 days or even longer.POI greatly aggravates the pain of postoperative patients,prolongs the hospitalization time and increases the cost of hospitalization.A heavy burden on patients and society.The treatment of postoperative ileus is a challenging clinical problem,and the mechanism of its occurrence and development is still not completely clear.Currently available prokinetic drugs have many side effects.Although motilium,cisapride and metoclopramide have certain curative effects,their clinical application is limited because of the adverse effects of motilium and cisapride on the heart system and the extrapyramidal reaction of metoclopramide.The newly developed avimopam in the United States is used for postoperative intestinal paralysis,but it can not improve postoperative gastrointestinal transit function and inflammatory infiltration.Professor Chen Zhiqiang,a famous doctor of traditional Chinese medicine in Guangdong province,first put forward the pathogenesis theory of poi on the basis of many years of clinical experience.He innovatively put forward the theory of qi stagnation and blood stasis,mixed with deficiency and excess,mainly with deficiency innovative.Based on years of clinical experience,Professor Zhiqiang Chen first put forward the pathogenesis theory of POI.He innovatively put forward the theory of qi stagnation and blood stasis,mixed with deficiency and excess,mainly with deficiency innovative.Xiangbin prescription(XBP)is a compound of Fructus amomi(Sharen),Lindera aggregata Kosterm(Wuyao),Panax ginseng(Renshen),Prunus persica Batsch(Taoren)and Areca catechu L.(Binlang).It has the effect of replenishing qi,invigorating spleen and activating fu.In order to summarize and popularize the application of XBP,we performed the clinical and mechanistic study of it.This study includes two parts:clinical study and experimental study.In the aspect of clinical research,we used the world-recognized advanced high-resolution antroduodenojejunal manometry to dynamically monitor the effects of Xiangbin prescription on Migrating Motor complex(MMC).The efficacy of XBP was evaluated in a randomized controlled trial with placebo.In mechanistic study,the mechanism was mainly studied.POI model of SD rats was established.The mechanism of promoting action of XBP was discussed from mesentery.It is the first time to investigate mesentery,and screened and verified the key gene and protein in mesentery involved in POII.Clinical study:Xiangbin prescription promotes gastrointestinal motility recovery in patients undergoing laparoscopic colorectal cancer surgeryPurpose:Colorectal cancer has been a serious threat to human health[1-2],surgery is the most important means of colorectal cancer.Even with minimally invasive surgery,each patient undergoing abdominal surgery suffers from gastrointestinal dyskinesia or postoperative ileus[3],lasting 3-7 days or more,and patients with colorectal cancer undergo a week of recovery from gastrointestinal dysmotility,which greatly aggravates the pain of patients,prolongs hospitalization,hospitalization costs[4],and high mortality rate5,taking great social and economic burden.Traditional Chinese medicine has advantages and characteristics for the recovery of gastrointestinal motility after colorectal cancer surgery,but it lacks scientific and standardized design.At present,there is no report on the study of gastrointestinal motility monitoring in patients after abdominal surgery with high-resolution antroduodenojejunal manometry in China or abroad.Gastrointestinal motility disorders may occur in patients after laparoscopic colorectal cancer surgery,but how to affect the parameters of interdigestive migrating motor complex(MMC)is not clear.This study is the first time to use Solar GI high resolution manometry to evaluate gastrointestinal motility in patients with colorectal cancer after laparoscopic surgery.Methods:According to the design of clinical studies,40 patients with colorectal cancer were randomly divided into XBP group and placebo group.High-resolution antroduodenojejunal manometry was performed in the two groups on preoperative and postoperative anesthesia awake,and 1-3 days postoperatively.In the XBP group,the patients began to take the medicine on the first day after surgery,twice a day until defecation or 6 days after operation.The placebo group was given a placebo after routine basic treatment on the first day after surgery.Drugs administration were recorded every day(including MMC parameters of the antrum,duodenum,and jejunum(MMC number,amplitude,frequency,motor index,duration of each phase),time to first flatus,defecation,fluid and semi-liquid diet),length of hospital stay,hospitalization costsas as well as the adverse react ions/adverse events(whether abdominal distension,nausea,vomiting,dizziness,headache,etc.).Statistical methods of clinical data:the homogeneity of the variance of the measurement data and the conformity with the normal test were tested by independent sample t.If the variance is not uniform,not meeting the normal distribution of indicators,using the rank sum test to compare.The results are recorded with the median(Lower quartile to upper quartile)P50(P25～P75).Result:1.Baseline data:There were no significant differences in baseline data between gender composition,age distribution,and preoperative self-preparation(P>0.05);no differences with operation and anesthesia time,intraoperative blood loss,intraoperative fluid volume.There were no significant differences in the data of drainage tube and catheter indwelling time(P>0.05).There was no significant difference in the parameters of high-resolution antroduodenojejunal manometry between preoperative and postoperative anesthesia(P>0.05).The baseline data of the two groups of patients included in the study before intervention were consistent and comparable.2.Effect of laparoscopic colorectal cancer resection on gastrointestinal motilityThe gastrointestinal motility monitoring during anesthesia recovery showed no MMC cycle,no MMCIII phase,just scattered and irregular low amplitude contractions.The duration of phase I was prolonged and the duration of phase II was significantly shortened.The MMC of patients with colorectal cancer after laparoscopic surgery was disordered and the gastrointestinal motility was obviously decreased.The results show that laparoscopic surgery is a minimally invasive surgery,but it can also cause postoperative gastrointestinal motility disorders and bring pain to patients.Open surgery may result in longer periods of gastrointestinal motility inhibition3.Comparison of gastrointestinal manometry between two groups 1-3 days after operationOn the first day after operation,high-resolution manometry monitoring was performed for 2～3 hours in the fasting state,and then high-resolution manometry was observed for 2～3 hours after the XBP/placebo granules given.Before the XBP/placebo was given on the first postoperative day,there was no regular MMC and no MMCⅢ activity in either group.After the administration of XBP,the duration of MMCI phase was significantly shortened,and the duration of MMCII phase was prolonged compared with the placebo(P＜0.05).The amplitude of antrum,duodenum and jejunum contraction was significantly enhanced(both P<0.05);MI in the duodenum and jejunum also increased significantly(both P<0.05).On the second postoperative day,there was no regular MMC cycle before the administration of XBP or placebo,but 2 patients in the XBP group recorded a high frequency and amplitude MMCⅢ phase contraction after administration.One case of MMCIII was initiated by the antrum,spread to the duodenum,jejunum,and one case was initiated by the duodenum and spread to the jejunum.The second day after the operation before oral adminiatration,the contraction amplitudes of the antrum,duodenum and jejunum in the XBP group were significantly stronger than those in the placebo group(P＜0.05).After adminiatration of XBP,during of phase I was significantly shortened,during of phase II was significantly prolonged,the antrum,duodenum and jejunum contraction amplitude increased significantly(P<0.05),and the MI of the duodenum and jejunum significantly increased(P<0.05).Oral administration XBP can partially stimulate the activity of MMCⅢ,which is very important to improve the gastrointestinal movement of the disorder.XBP may improve postoperative gastrointestinal motility disorder by stimulating phase Ⅲ.On the third postoperative day,the gastrointestinal motility of the XBP group was significantly restored before administration.Five patients had regular MMC before administration,while only one patient in the placebo group had regular MMC and phase Ⅲ.XBP significantly shortened the duration of phase Ⅰ of the MMC cycle,and increased the duration of phase Ⅱ of the MMC cyclethe and number of MMC(P<0.05).Strong amplitude with high frequency of phase Ⅲ was recorded in most patients in XBP group.XBP significantly increased the motility of the antrum,duodenum and jejunum at phase Ⅲ of the MMC cycle,such as increasing the duration,contraction frequency(both P＜0.05),and the amplitude of MMCⅢ in duodenum and jejunum increased significantly.The amplitude and MI of duodenum and jejunum increased significantly(both P＜0.05).The change in gastrointestinal motility was not significant after placebo administration.On the third day after operation,the gastrointestinal motility of the XBP group was significantly increased after administration,and the amplitude and MI in the three parts were similar to those before surgery(P＞0.05).MI of antrum,duodenum and jejunum did not return to preoperative level,even with first flatus or defecation(both P＜0.05).4.Comparison of the time of first flatus and defecation between XBP and placebo groupIn the two groups,time to first flatus and defecation was compared.The data of homogeneity of variance and normal distribution were tested by independent sample t test.If the variance was not consistent with normal distribution,rank sum test was used:the results were expressed by median(Lower quartile to upper quartile)M(P25～P75)For normal distribution,independent sample t-test was used for comparison between groups.Statistical analysis showed that tiem to first flatus and defecation in the XBP group was significantly earlier than that in the placebo group(P＜0.05).5.Comparison of early oral dietary time between XBP and placebo groupThrough the normality test,the time to liquid and semi-liquid diet in the two groups was normally distributed.Therefore,the independent sample t test was used.The statistical results are as follows:time to liquid and semi-liquid diet was shorter than that of the placebo group(P＜0.05).6.Comparison of length of hospital stay and hospitalization costs between between XBP and placebo groupThe length of hospital stays(including total length of hospital stay and postoperative hospital stay)and hospitalization costs were all normal distribution,so the independent sample t-test was used.The statistical results showed that total length of hospital stay and postoperative hospital stay were not significantly different(P>0.05).There were significant differences in total hospitalization costs(P＞0.05).7.Comparison of gastrointestinal hormones and inflammation factors between XBP and placebo groupBlood samples were analyzed before and 1,2,3 and 7 days after operation to compare the levels of gastrointestinal hormones and inflammatory factors in different time.The results showed that Ghrelin decreased significantly on the first day after operation in both groups and increased gradually on the second day.The level of ghrelin in XBP group was significantly higher than that in placebo group on the 7th day after operation(P＜0.05).The plasma MTL decreased significantly on the first day after surgery but gradually increased from the second day.MTL level in the XBP group was significantly different from that in the placebo group on day 2,3,7 day after surgery(P＜0.05).XBP could significantly promote the release of MTL.XBP can significantly reduce plasma level of IL-4,IL-6.IL-4,IL-6 did not fall to preoperative levels in the placebo group on the seventh day after surgery.According to the statistical analysis,the levels of IL-4 and Il-6 increased significantly on the first day after operation,which was consistent with the previous study that the peak of inflammatory infiltration was 24 hours after operation.The level of IL-4 in the XBP group was significantly lower than that in the placebo group at 2,3,and 7 days after surgery(P＜0.05).The IL-6 level was significantly lower than the placebo group on the 7th day after surgery(P<0.05).The level of TNF-α was significantly increased on the first day after colorectal cancer in both groups,which was consistent with the previous study for the peak of inflammatory infiltration.XBP can significantly reduce plasma levels of TNF-α and inhibit postoperative inflammatory response;TNF-α levels in the placebo group did not fall to preoperative levels on the seventh day after surgery.There were significant differences in level of TNF-α between the two groups on 2,3,and 7 days after surgery(P＜0.05).Conclusions:The traditional Chinese medicine XBP can obviously shorten the during of phase I of MMC,prolong the during of phase II,stimulate the activity of phase Ⅲ by enhancing the amplitude of contraction and MI,correct the disorder of MMC after operation,recover the earlier regular MMC,and promote the movement of gastrointestinal coordination.XBP can also effectively shorten time to first flatus and defecation,shorten the postoperative liquid and semi-liquid diet,and reduce hospitalization costs;XBP can also promote the release of MTL and Ghrelin,and reduce the plasma level of IL-4,IL-6,TNF-α.In all XBP promotes rapid recovery of postoperative gastrointestinal motility for patients with colorectal cancer by coordination of MMC and inhibition the inflammatory reaction.Ⅱ.Study on the mechanism of XBP promoting gastrointestinal motility recovery after abdominal surgeryPurpose:Recent studies have confirmed that the mesentery is a single and continuous organ in the digestive system.A deeper understanding of how the mesentery interacts with the digestive system may lead to better disease management and development strategies.Scientists have also come to realize that mesenteric dysfunction can stop blood flow and cause intestinal adhesion and necrosis.In this study.It is the first time to observe the effects of XBP on the recovery of gastrointestinal motility in POI rats from the mesentery.The mechanism of the effect of XBP on POI was elucidated from mesentery-inflammatory factor pathway.Methods:Forty male Sprague-Dawley(SD)rats weighing 200±20 g were purchased from Guangdong Medical Laboratory Animal Company.The animals were allowed free access to standard laboratory chow and water,housed at a temperature of 23±2℃ with a relative humidity of 55±10%,and a 12:12 h light/dark cycle.Rats were acclimated for 7 days before any experimental procedures.Ethical approval for the study protocol used in this study was obtained from the Institutional Animal Care and Ethics Committee of Guangdong Provincial Hospital of Traditional ChineseRats were randomly assigned into four groups:Control group,no operation with saline(n=10);POI,POI manipulation plus treatment with saline(n=10);Procapride group,POI manipulation plus treatment with Prucapride(2 mg/kg,n=10)dissolved in saline solution;and XBP,POI manipulation plus treatment with XBP(500 mg/kg,n=10).Saline,Prucalopride and XiangBin Prescription were orally administered after surgery at the same points,and 2 mL each time in all treatment groups.The mesentery was taken at 24 hours for histopathology;the levels of TNF-α,IL-4,IL-6 and CRP in mesentery tissue were detected by immunohistochemistry;CD68 and iNOS in mesenteric membrane were detected by immunofluorescence;RNA-seq was used to screen the genes closely related to POI in mesentery.Western blot and RT-qPCR were used to verify the function of the key genes.The results were expressed by X±SE,and the efficacy was compared by repeated measurement of variance analysis(P<0.05).Result:1.XBP improved the mesentery histopathology of POI ratsHistological examination of the mesentery from control rats showed normal architecture and absence of inflammatory cells.In contrast,mesenterium sections from the POI group revealed mild inflammatory response infiltrated with neutrophil and macrophage cells in the mesenteric tissue.Treatment with Pru and XBP reduced the number of infiltrating cells and protected the mesenterium from histological damage induced by POI.2.Anti-inflammatory activity of XBP in POI modelIn POI rats,there was a significant increase in the levels of inflammatory mediator IL-6 in mesentery as compared to the controls.Pru and XBP significantly(P＜0.05)reduced the level of IL-6 compared to respective POI group.The level of chemokine CRP was also greatly reduced in XBP-treated animals as compared with vehicle-treated POI rats.A significant increase in the expression of CD68 and iNOS was observed in POI rats when compared with the control(P＜0.05).The oral treatment with Pru and XBP significantly reversed changes in the levels of CD68 and iNOS induced by POI(P<0.05).3.Identification of differentially expressed genes and regulatory networks of differentially expressed mRNA genesTo gain insights into the molecular mechanisms of POI treated with XBP in rats,we profiled the expression of mRNA genes in the CON,POI and XBP groups using RNA-seq.In total,1862 differentially expressed genes(DEGs)were identified by pairwise comparisons.As expected,there are a large number of DEGs(n=1547)between the POI sample and the CON sample whereas there are fewer DEGs when comparing the XBP-treated sample(n=331)with the POI sample.The distributions of DEGs and their overlapping expression in different pairwise comparisons were illustrated by Venn diagram analysis.16 DEGs were shared by the POI vs CON and XBP vs POI groups,indicating a disease-specific correlation involved in the mechanism mediated by XBP during POI.In order to better understand the regulatory roles of DEGs in POI rats treated with XBP,we used the STRING database,Kyoto Encyclopedia of Genes and Genomes pathway and Cytoscape software to identify their most relevant molecular networks and biological functions.The results show that the 16 DEGs from the comparison of the POI vs CON and XBP vs POI groups are enriched in a network associated with activation of immune system processes such as lymphocyte chemotaxis,leukocyte chemotaxis,neutrophil chemotaxis,granulocyte chemotaxis,response to interleukin-13,etc.Among these 16 genes,CCL11,Hspalb,Cyp7b1,Cypllal,Pik3c2g,and SPP1 are significant hub genes,which are centrally located in their associated processes and may thus be critical components within the network.This implies that the inflammatory responses are important in XBP-mediated protection because the components of this activity are expressed both in the XBP and POI group.4.Verification of the DEGs in rat mesenteryThe important proteins significantly altered by XBP treatment were also evaluated and confirmed at the protein level.Moreover,we also examined CCL11,Cyp7b1,Hspalb,Cyp11a1,and SPP1 mRNA expression in rat mesenterium using semi-quantitative analysis.In the protein level,CCL11,Cyp7b1,Cyp11a1 and sppl of XBP were significantly lower than those of model group(P<0.05),and Hspalb was significantly higher than that of model group(P<0.05);The relative levels of CCL11,Cyp7b1,Cyp11a1 and SPP1 mRNA in XBP group and model group were compared.The difference was statistically significant(P＜0.05).XBP can obviously down-regulate pro-inflammatory factors,up-regulate anti-inflammatory factors,thereby inhibiting the mesentery inflammatory response. |
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