The aim of this study is to investigate the regulatory mechanism of LncRNA(UCA1)on the miRNA-331-3p/bromine domain protein 4(BRD4)axis through acupuncture,in order to ameliorate colonic inflammation in rats with spleen deficiency.Additionally,this study seeks to explore the contemporary mechanism of“strengthening spleen and benefiting Qi”acupuncture therapy in improving symptoms of spleen deficiency and loose stools.
Sixty male SD rats were randomly assigned to four groups:normal group,model group,non-meridian non-acupuncture group and acupuncture group,with 15 rats in each group.With the exception of the normal group,the remaining three groups were utilized to establish a rat model of spleen deficiency through the compound method of overwork and irregular diet.Following successful modeling,the acupuncture group re-ceived electro-acupuncture at“Zusanli(ST36)”and“Pishu(BL20)”,while the non-meridian non-acupuncture group was treated with electro-acupuncture at the tail tip as a control.The treatment was administered once daily for 20 minutes over a period of 14 days.After treatment,serum and colonic tissues were collected from the rats.The expressions of LncRNA(UCA1)and miRNA-331-3p/BRD4 axis-related genes in rat colon tissue were detected by qPCR.The contents of interleukin-1β(IL-1β)and interleukin-6(IL-6)in rat serum were detected by ELISA.The pathological morphological changes of rat colon tissue were observed by HE staining.The staining of BRD4 in colon was observed by immunohistochemistry method.The immunoactivities of IL-1β,IL-6,tumor necrosis factor-α(TNF-α)and NFκB p65 expression levels were detected by immunofluorescence histochemistry.The apoptosis in colon tissues of each group was detected by TUNEL staining.
Compared with those of the normal group,the expression levels of miRNA-331-3p in colon tissues of rats in the model group and non-meridian non-acupuncture group were significantly decreased(P<0.05),and the mRNA expression levels of LncRNA(UCA1),BRD4,nuclear factor kappa-B(NF-κB)p65,IL-1β,IL-6,IL-8 and TNF-αwere increased(P<0.05).Compared with those of the model group,the expression levels of miRNA-331-3p in the colon tissue of rats were significantly increased(P<0.05)in the acupuncture group,and the mRNA expression levels of LncRNA(UCA1),BRD4,NF-κB p65,IL-1β,IL-6,IL-8 and TNF-αwere decreased(P<0.05).Compared with those of the normal group,the expression levels of IL-1βand IL-6 in the serum of rats in the model group were significantly increased(P<0.05),and the expression levels of IL-1βand IL-6 in the colon tissue of rats in the acupuncture group were decreased(P<0.05)compared with those in the model group.Immunohistochemical results showed that the expression of BRD4 in the colon tissue of rats in the model group was increased compared with that in the normal group(P<0.05).Compared with that in the model group,the expression of BRD4 in the acupuncture group was decreased(P<0.05).The immunofluorescence histochemistry results showed that the mean absorbance values of IL-1β,IL-6,TNF-αand NF-κB p65 immunofluorescence in colon tissues of the model group and non-meridian non-acupuncture group were increased compared with those of the normal group(P<0.05).Compared with those of the model group,the mean absorbance values of IL-1β,IL-6,TNF-αand NF-κB p65 immunofluorescence in colon tissues of rats in the acupuncture group were decreased(P<0.05).In addition,HE staining results showed that acupuncture could effectively reduce inflammatory cell infiltration and loss of cupped cells in colonic mucosal tissues.TUNEL staining results showed that acupuncture could improve colonic inflammation and inhibit apoptosis of colonic mucosal cells in spleen deficiency rats.
Electro-acupuncture at“Zusanli(ST36)”and“Pishu(BL20)”can interfere with the LncRNA(UCA1)regulating miRNA-331-3p/BRD4 axis,inhibit the expressions of intestinal inflammation and pro-inflammatory mediators,and then inhibit the apoptosis of colonic mucosal cells and improve the immunity of the body,thus improving the symptoms of spleen deficiency and loose stools.
| 科 Family | 属数 Number of genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) | 属 Genus | 种数 Number of species | 占总种数比例 Percentage of total species (%) |
|---|---|---|---|---|---|---|
| 鹅膏菌科Amanitaceae | 2 | 11 | 5.26 | 鹅膏菌属 Amanita | 10 | 4.78 |
| 小菇科 Mycenaceae | 2 | 12 | 5.74 | 丝盖伞属 Inocybe | 5 | 2.39 |
| 多孔菌科 Polyporaceae | 8 | 14 | 6.70 | 蜡蘑属 Laccaria | 5 | 2.39 |
| 红菇科 Russulaceae | 3 | 23 | 11.00 | 小皮伞属 Marasmius | 6 | 2.87 |
| 小菇属 Mycena | 11 | 5.26 | ||||
| 光柄菇属 Pluteus | 5 | 2.39 | ||||
| 红菇属 Russula | 17 | 8.13 | ||||
| 栓菌属 Trametes | 5 | 2.39 |