Objective To observe the effect of electroacupuncture on corneal Toll-like receptor 4(TLR4)-mediated inflammatory signaling pathway in diabetic dry eye rats and to explore the mechanism of electroacupuncture in the treat-ment of diabetic dry eye.Methods A type 2 diabetic rat model was established in 32 healthy male Sprague-Dawley(SD)rats by intraperitoneal injection of streptozotocin(30 mg·kg-1)for 12 weeks after feeding with high-sugar and high-fat di-et for 4 weeks.Twenty-five successfully modeled diabetic dry eye rats were randomly divided into the model group(non-in-tervention),electroacupuncture group(the"Jingming","Cuanzhu","Sizhukong","Taiyang"and"Tongziliao"acupoints were treated with acupuncture,and then"Cuanzhu"and"Tongzilao"acupoints were treated with electroacupuncture,15 min for each time,once a day),sham acupuncture group(blunt-tip needle pricking was performed at the same acupoints as the electroacupuncture group),and fluorometholone group(1 g·L-1fluorometholone eye drops were used in both eyes at 8 o'clock,13 o'clock,and 18 o'clock,1 drop each time),with 6 rats in each group,lasting for 2 weeks.Another 6 healthy male SD rats were selected as a blank group.Random blood glucose,tear film breakup time(BUT),tear secre-tion,corneal fluorescein staining(FL)score,and corneal touch threshold(CTT)of rats in each group were detected be-fore modeling,after modeling,and after the intervention.Hematoxylin and Eosin(HE)staining was performed to observe the corneal morphologic changes in each group.Immunofluorescence histochemical staining was adopted to detect the cor-neal TLR4-positive expression in each group.The TLR4,phosphorylated nuclear factor-kappa P65(P-NF-KB P65),inter-leukin(IL)-1β,and IL-18 expression levels in the cornea were detected by Western blot.Results After modeling,com-pared with the blank group,BUT,tear secretion and CTT decreased and FL increased in all experimental groups,and the differences were statistically significant(all P＜0.01).After the intervention,compared with the model group,FL de-creased,and BUT,tear secretion and CTT increased in the electroacupuncture group,and the differences were statistically significant(all P＜0.05).Corneal HE staining showed that after the intervention,the corneal surface of rats in the model group and sham acupuncture group was not smooth,and the corneal epithelial cells were thickened and disorganized;the corneal surface of rats in the electroacupuncture group and fluorometholone group was smooth,and the corneal epithelial cells were arranged neatly.After the intervention,compared with the blank group,the corneal TLR4 expression of rats in all other groups was elevated,and the differences were statistically significant(all P＜0.05);compared with the model group,the corneal TLR4 expression of rats in the electroacupuncture group and fluorometholone group was reduced(both P＜0.01).Compared with the blank group,corneal TLR4,P-NF-κBP65,IL-1β and IL-18 protein expressions increased in the model group and sham acupuncture group(all P＜0.05);compared with the model group,these expressions decreased in the electroacupuncture group and the fluorometholone group(all P＜0.05).Conclusion Electroacupuncture can im-prove the ocular surface signs and inhibit the expressions of TLR4,P-NF-κB P65,IL-1 β,and IL-18 in the cornea of type 2 diabetic dry eye rats,and its mechanism of action may be related to the regulation of TLR4-mediated inflammatory signaling pathway,which inhibits ocular surface inflammation in diabetic dry eye rats.

Objective To reveal the potential mechanism of electroacupuncture regulating the neuroinflammatory signaling pathway mediated by triggering receptor expressed on myeloid cells 2(TREM2)in trigeminal ganglion(TG)and caudate nucleus of spinal trigeminal nucleus(SpVc)of rats with type 2 diabetes and dry eyes and alleviating ocular surface sensory abnormalities.Methods Healthy male Sprague-Dawley(SD)rats were induced by intraperitoneal injection of 10 g·L-1 streptozotocin to establish a type 2 diabetes dry eye rat model after 4 weeks of high-glucose and high-fat diets.In the 12th week of the experiment,successfully modeled rats were randomly divided into the model group,electroacupunc-ture group,sham-acupuncture group and fluorometholone group,and healthy male SD rats fed with normal diets were se-lected as the blank group.Rats in all groups were intervened for 2 weeks.The corneal fluorescein sodium staining(FL),tear secretion[detected by phenol red thread(PRT)test],tear film break-up time(BUT),and corneal touch threshold(CTT)in each group were measured before,after modeling,and after the intervention.The changes in tissue morphology of TG and SpVc and TREM2 positive expression sites in each group were observed.The messenger ribonucleic acid(mR-NA)expression of TREM2,interleukin-18(IL-18)and interleukin-1β(IL-1β)of TG and Sp Vc in each group were detec-ted.Results After modeling,compared with the blank group,the FL scores of rats significantly increased and PRT,BUT and CTT significantly decreased in other groups(all P＜0.01).After the intervention,compared with the model group,the electroacupuncture group and fluorometholone group showed significant reductions in FL scores and significant increases in PRT,BUT and CTT(all P＜0.01);there was no statistically significant difference in these indicators in the sham-acupuncture group(all P＞0.05).Compared with the electroacupuncture group,the PRT and CTT of rats in the flu-orometholone group and sham-acupuncture group were significantly reduced,the FL score significantly increased and BUT significantly decreased in the sham-acupuncture group(all P＜0.01),and there was no statistically significant difference in FL score and BUT of rats in the fluorometholone group(both P＞0.05).Compared with the sham-acupuncture group,the fluorometholone group showed a decrease in FL score and an increase in PRT,BUT and CTT in rats,with statistically dif-ferent significances(all P＜0.05).The immunofluorescence double-labeling assay showed a positive expression of TREM2 in activated microglia of TG and SpVc in the model group.The reverse transcription-polymerase chain reaction of TG and SpVc showed that compared with the blank group,the TREM2 mRNA expression in the TG and SpVc of the model group,electroacupuncture group,fluorometholone group and sham-acupuncture group decreased,the IL-18 and IL-1β mRNA in TG and SpVc of the model group,IL-18 and IL-1β mRNA in TG and IL-1β mRNA in SpVc of the sham-acupuncture group increased(all P＜0.05);compared with the model group,the TREM2 mRNA expression increased and IL-18 and IL-1β mR-NA expression decreased in the TG and SpVc of the electroacupuncture group and fluorometholone group(all P＜0.05).Compared with the electroacupuncture group,the TREM2 mRNA expression decreased and IL-18,IL-1β mRNA expression increased in TG and SpVc of the sham-acupuncture group(all P＜0.05),and no significant difference was found in TREM2,IL-18 and IL-1β mRNA expression in TG and SpVc of the fluorometholone group(all P＞0.05).Compared with the sham-acupuncture group,the TREM2 mRNA expression in TG and SpVc increased in the fluorometholone group,while the IL-18 mRNA expression in SpVc decreased(all P＜0.05);the IL-1β mRNA in TG and SpVc and the IL-18 mRNA in TG of the fluorometholone group showed no statistically significant difference(all P＞0.05).Conclusion Electroacupunc-ture can effectively alleviate the ocular surface sensory abnormalities of rats with type 2 diabetes and dry eyes by regulating the inflammatory signaling pathway mediated by TREM2 in TG and SpVc.