| ObjectiveHelicobacter pylori-related gastric disease(HPGD)includes chronic gastritis(non-atrophic and atrophic),chronic gastritis with erosion,chronic gastritis with intestinal metaplasia and/or dysplasia,gastric ulcer and gastric cancer,which basically summarizes the whole process of benign and malignant pathological evolution of gastric mucosa.This study was conducted from the perspectives of interleukin-10(IL-10)gene polymorphism,mRNA transcription level and protein qualitative location expression.Based on the background of traditional Chinese medicine(TCM)syndrome differentiation,it is intended to explore the essence of individual differences in gastric mucosa histopathology between different HPGD subjects,so as to provide certain basis for early diagnosis and prevention of gastric mucosa malignancy.MethodsThe samples in this study(including routine physical examination subjects)were collected from 287 gastric mucosa specimens(total)in the digestive endoscopy Department of the First Affiliated Hospital of Guangzhou University of Chinese medicine from Jun 2013 to Aug 2021.All recruiters included in this study were numbered and blinded by corresponding gastric mucosa samples after the completion of medical history collection.The patients were divided into syndrome type group and pathological group according to syndrome classification and histological gastric mucosa section under microscope.TCM syndrome groups contain SSDH(syndrome of spleen-stomach dampness-heat),LQIS(syndrome of liver qi invading the stomach),IBSB(syndrome of internal block of static blood),SQD(syndrome of spleen qi deficiency),PTS(syndrome of phlegm turbidness and stagnation),and NON(non-syndrome).Pathological groups contain NOR(normal group),INF(gastric inflammation group),ATR(gastric atrophy without intestinal metaplasia and dysplasia group),MIX(gastric mild to moderate intestinal metaplasia or dysplasia accompanied by inflammation or atrophy group),SD(gastric severe dysplasia or intestinal metaplasia accompanied by inflammation or atrophy group)and GC(gastric cancer group).All subjects underwent digestive endoscopy.The study samples of routine physical examination subjects were clamped near the gastric antrum,those with gastric mucosal lesions were clamped near the lesion,but for gastric ulcer patients,clamps should be clamped around the lesion.In this study,the rapid urease method was used to detect Hp infection and the methylene blue staining method was used to detect the severity of Hp infection.The gastric mucosa samples were treated by hematylin Eosin(HE)staining,and the changes were observed under a microscope.Single nucleotide polymorphism of IL-10(rs 1800898)was detected by direct sequencing.The expression of IL-10 gene was detected by fluorescence quantitative PCR(SYBR dye method).Immunohistochemical labeling was used to detect the expression of IL-10 protein.This study was approved by the Ethics Committee of the First Affiliated Hospital of Guangzhou University of Chinese Medicine,and all the participants or their families had signed informed consent.Results1.Basic information.(1)Characteristics of Subjects:a total of 287 volunteers including 152 males and 135 females were included,whose ages were ranged from 18 to 80 with an average age of 48.16±12.70 years old.Results of this study showed that there was no significant difference in age among all groups,but the difference of course of disease was statistically significant,further analysis revealed that compared with the NOR group,the differences between the INF group,ATR group,MIX group and GC group were statistically significant(P=0.018、0.030、0.001、0.000,P<0.05),and the course of disease in GC group was longer than that in INF group(P=0.027,P<0.05).(2)The distribution of different TCM syndromes in each pathological group:statistical analysis was conducted on the distribution of different syndromes in each pathological group,and Fisher’s exact probability test showed that there was no statistical significance in the overall distribution of syndromes among each group(P=0.47,P>0.05).In terms of trend,the proportion of SSDH was higher in all pathological groups.2.Hp infection and histopathological variations in gastric mucosa.(1)Distribution of Hp infection and infection degree in gastric mucosa:the TCM groups:results of Hp infection rate and infection degree in gastric mucosa of subjects with different syndromes showed that there was no statistically significant difference in Hp infection rate and infection degree among different TCM syndromes(P=0.249、0.171,P>0.05).In terms of the trend,the infection rate of Hp in LQIS group was higher than that in other syndrome groups,and the proportion of Hp severe infection in PTS group was higher than that in other syndrome groups.The pathological groups:results of Hp infection rate and infection degree in gastric mucosa of all pathological groups showed no statistical significance(P=0.495、0.095,P>0.05).In terms of trend,the infection rate of Hp in GC group was higher than that in other groups.Compared with other groups,the proportion of Hp severe infection in SD group was the highest.(2)Distribution of degree and activity of gastric mucosa inflammation:By non-parametric test,gastric mucosa inflammation degree and activity of Hp positive subjects were significantly higher than that of Hp negative subjects(P<0.05).The TCM groups:there was no statistical significance in gastric mucosa inflammation degree of different TCM syndromes(Hp negative:P=0.530;Hp positive:P=0.706;P>0.05)and activity(Hp negative:P=0.500;Hp positive:P=0.406;P>0.05).In terms of trend,the proportion of severe inflammation degree and inflammation activity in gastric mucosa of Hp positive subjects was the highest in PTS group.The pathological groups:compared with NOR group,gastric mucosa inflammation was more severe in ATR group(P=0.047)and MIX group(P=0.026)(P<0.05).Among Hp positive patients,there was significant difference in gastric mucosa inflammation degree among different pathological groups(P=0.000,P<0.05).A further pairwise comparison found,compared with NOR group,gastric mucosa inflammation was more severe in INF group(P=0.000),ATR group(P=0.000),MIX group(P=0.000),SD group(P=0,000)and GC group(P=0.000)(P<0.05),and gastric mucosa inflammation in SD group was more severe than that in INF group(P=0.010<0.05).There was statistically significant difference in gastric mucosa inflammation activity among subjects in different Hp-negative pathological groups(P=0.013,P<0.05).Further pin-pinal comparison showed that gastric mucosa inflammation activity in GC group was more severe than that in INF group(P=0.014)and ATR group(P=0.023).There were statistically significant differences in gastric mucosa inflammation activity among different Hp-positive pathological groups(P=0.000,P<0.05).A further pairwise comparison found,gastric mucosa inflammation activity in ATR group(P=0.017),MIX group(P=0.041)and GC group(P=0.001)was more severe than that in NOR group(P<0.05).The gastric mucosa inflammation activity in GC group was more severe than that in INF group(P=0.002)and SD group(P=0.015).(3)Correlations between the degree of Hp infection and the severity of pathological variations in gastric mucosa:Spearman rank correlation analysis showed that there was a positive linear correlation between Hp infection and the severity of pathological changes.Specific as follows:The correlation coefficient of gastric mucosa inflammation was Spearman’s r=0.295,the correlation coefficient of inflammation activity was 0.169,the correlation coefficient of atrophy was 0.298,and the correlation coefficient of gastric mucosa intestinal metaplasia and dysplasia were Spearman’s r=0.087 and Spearman’s r=0.119.That is,the degree of gastric mucosa inflammation and activity,atrophy,intestinal metaplasia and dysplasia increased significantly with the aggravation of Hp infection(P<0.05).3.Correlations between IL-10(rs1800898)gene polymorphism and pathological variations of gastric mucosa and TCM syndromes.(1)Correlations between IL-10(rs1800898)SNPs and pathological variations:Fisher’s exact probability test showed that in Hp-negative subjects,gastric mucosa of A/A genotype subjects was more likely to undergo pathological histopathological changes such as atrophy,intestinal metaplasia,dysplasia and even canceration than that of A/G genotype subjects(P=0.01,P<0.05).Further pair comparison showed that the gastric mucosa of subjects carrying A allele may be more prone to benign and malignant pathological changes(P=0.014,P<0.05).(2)Correlations between IL-10(rs1800898)SNPs and TCM syndromes:After Fisher’s exact probability test,there were no statistically significant differences in the relationship between different genotypes and alleles and syndromes of IL-10(rs 1800898)locus in gastric mucosa of the subjects regardless of Hp infection(P>0.05).In terms of trend,subjects with Hp positive A/A genotype or A allele may have more easily tendency to develop SQD syndrome.4.Correlations between the quantitative expression of IL-10 mRNA in gastric mucosa and the pathological variations of gastric mucosa and TCM syndromes.(1)Relationship between IL-10 mRNA and pathological variations in gastric mucosa:Kruskal-wallis H(K)test showed that whether Hp was infected or not,there was no significant difference in the relative expression of IL-10 mRNA in gastric mucosa pathological changes(Hp negative:P=0.53>0.05;Hp positive:P=0.16>0.05).However,in terms of trend,IL-10 mRNA expression in HP-positive gastric cancer patients showed a downward trend compared with ATR group.IL-10 mRNA expression in Hp-positive gastric mucosa atrophy patients showed an increased trend compared with other pathological groups.(2)Relationship between the relative expression level of IL-10 mRNA in gastric mucosa and TCM syndromes:after Kruskal-wallis H(K)test,there was no statistically significant difference between TCM syndromes and the relative expression level of IL-10 mRNA in Hp-negative subjects(P=0.63>0.05).However,the relative expression level of IL-10 mRNA in gastric mucosa of Hp positive subjects with different syndromes showed statistical significance(P=0.000<0.05).Concretely,the expression level of IL-10 mRNA in gastric mucosa of Hp positive subjects with IBSB and LQIS showed an increasing trend compared with other syndromes groups.5.Relationship between IL-10 protein expression and pathological changes of gastric mucosa and TCM syndromes.(1)Qualitative quantification integral distribution of IL-10 protein expression in gastric mucosa of different pathological groups:there was no statistical significance in the expression of semi-quantitative score in Hp negative groups(P=0.057>0.05).Hp positive group semi-quantitative score comparison results showed statistically significant difference(P=0.000<0.05).Compared with NOR group,the expression level of IL-10 protein showed the trend of SD group>GC group>MIX group>NOR group.Compared with INF group,IL-10 protein expression showed a trend of SD group>GC group>MIX group>INF group.Compared with the SD group,the expression intensity of IL-10 protein showed a trend of SD group>MIX group>ATR group.(2)Proportional distribution of IL-10 protein expression in gastric mucosa of different pathological groups:abnormal expression of IL-10 protein appeared in gastric mucosa in the form of cytoplasmic expression,cytoplasmic nuclear expression and co-expression.In this study,the composition ratio of IL-10 protein location and expression in gastric mucosa of all pathological groups showed that there was no statistically significant difference in IL-10 protein expression in gastric mucosa of Hp negative groups(P=0.065>0.05).IL-10 protein expression in gastric mucosa of Hp positive groups showed statistically significant difference(P=0.000<0.05).Compared with the INF group,the probability of positive expression of IL-10 protein showed a trend of SD group>GC group>MIX group>INF group.(3)Qualitative quantification integral distribution of IL-10 protein expression in gastric mucosa of TCM syndromes:Non-parametric test(K-W test)showed that there was no statistically significant difference in the expression of semi-quantitative scores in the Hp-negative and Hp-positive groups in the distribution of syndrome types(P=0.197,0.484>0.05).In terms of trend,regardless of Hp infection,IL-10 expression in gastric mucosa of patients with SQD showed an enhanced trend compared with other syndromes.14.The expression rate distribution of IL-10 protein in gastric mucosa in different syndrome type groups:the composition ratio of IL-10 protein expression in gastric mucosa of different pathological groups was compared and analyzed by Chi-square test.The results showed that there was no significant difference in IL-10 protein expression in gastric mucosa between Hp negative and Hp positive groups(P=0.143,0.260>0.05).From the trend,IL-10 protein in gastric mucosa was mainly expressed in cytoplasm in Hp negative patients.The expression of IL-10 protein in gastric mucosa of IBSB patients with Hp positive was mainly in cytoplasm.However,the expression of IL-10 protein in gastric mucosa of other syndrome type groups was mainly cytoplasmic expression in membrane plasma co-expression.Conclusion1.Hp infection and abnormal expression of IL-10 in gastric mucosa can cause or aggravate the progression of benign and malignant pathological evolution of gastric mucosa in patients with HPGD.On the basis of Hp infection,the increased expression of SNPs and mRNA at the IL-10(rs 1800898)site in gastric mucosa and the strong positive expression of its protein may be involved in the formation of TCM syndromes.2.In light of trend,gastric mucosa IL-10(rs 1800898)locus A/A genotype or carrying A allele and associated with Hp infection,its mRNA was highly expressed and the syndrome was spleen deficiency syndrome and blood stasis syndrome,which may be A high risk factor for gastric mucosal malignant lesions.3.IL-10(rs1800898)locus in gastric mucosa of Hp infected patients carries A/A genotype or A allele,and its IL-10 protein is highly expressed in glandular cytoplasm,which may be involved in the formation of SQD.In this state,the high level of mRNA expression may be involved in the formation of IBSB. |
PDF Full Text Request