The Study On Mechanism Of Endometriosis Associated Pain: Does NGF,TrkA And P75 Change?

Pain is one of the most typical symptoms of endometriosis,which affects the life quality of patients of childbearing age.The mechanism of endometirosis pain is not clear. Research has been focused on sensory nerve sensory nerve fibers AS,sensory fibers C in recent years.There are some reports about neural anatomy mechanisms in endometriosis pain.The role of neurotransmitter systems in pathogenesis of endometriosis is still unknown.There are lots of molecules called "pain related substances",such as nerve growth factor(NGF),interleukins and tumor necrosis factor-α.NGF belongs to neurotrophin family,which promotes the survival of neurons.It has been found that the binding of NGF with its receptor causes self-induced phosphorylation and triggers a variety of biological effects,including the release of 5-HT from mast cells,resulting in hyperalgesia,which plays an important role in the regulation of neuropathic and non-neuropathic pain.Different types of lesions related to different pain.Generally speaking,endometrioma is not associated with pain,deep infiltrating endometriosis always involved with dysmenorrhea,CPP,intercourse pain.Literature reports that innervation of ectopic endometrial lesions may related to estrogen sensitive.However,the research about the relationship between different types of endometriosis and its surgery findings have not yet been fully unified.It needs more clinical research.Objective1 To explore NGF and its receptor TrkA,P75 expression in different types of endometriosis lesions,as well as the relationship between the degree of pain;2 To explore the expression of NGF and its receptor TrkA,P75 in eutopic endometrial. In different groups and hope it can enrich the "eutopic endometrial determinism";3 To investigate the normal endometrial and eutopic endometrial in vitro,to quantitative the expression of NGF,TrkA,and P75NTR.And to observe if the expression of NGF,TrkA,and P75NTR can be change by estrogen and progesterone and GnRHa.Materials and Methods1.Immunohistochemistry and double immunofluorescence assay were performed to observe NGF,TrkA and P75 of endometriosis lesions.2.Western Blot and RT-PCR were performed to detect NGF,TrkA and P75 in endometriosis lesions and eutopic endometrium.3.Real-timePCR were performed to detect NGF,and P75 in endometriosis lesions as well as eutopic endometrium.4.Culture endometrium of patients with endometriosis and the control for detection of NGF,TrkA and P75,RT-PCR,cell immunofluorescence,flow cytometry will be used.5.Different concentrations of estrogen and progesterone and GnRHa will be used to stimulate eutopic endometrial and normal endometrial cells in vitro.To detect NGFmRNA,TrkAmRNA and P75mRNA expression in the cell level.6.Use the same concentration of estrogen,RT-PCR assay NGF,TrkA and P75 expression in the cell level at different time points.7.To detect NGF and pain-related peptide(orphanin FQ) expression in serum of patient by ELISA.Results1.The semi-quantity of NGF and its receptor P75 in endometriosis lesions is as follows: uterosacral ligament > retroperitoneal > endometrioma.TrkA expression in the peritoneum is the highest,followed by uterosacral ligament and endometrioma.2.DIE uterosacral ligament lesions with severe dysmenorrhea expressed NGF,TrkA and its receptor P75 and it it higer than the patient with none-mild dysmenorrheal.3.NGF,TrkA and P75 expression in terosacral ligament lesions in patient with endometriosis,which is higer than non-endometriosis patients.Endometrioma does not express more NGF and its receptor TrkA,P75 of than non-endometriosis patients.4.Expression of P75 in patients' endometrium with severe dysmenorrhea is higher than none -mild dysmenorrheal group and the control group.5.Eutopic endometrial stromal cells from severe dysmenorrheal patient in vitro express more NGF and its receptor TrkA than none-mild dysmenorrheal and normal group.6.NGFmRNA and its receptor TrkAmRNA,P75mRNA expressed by eutopic endometrial stromal cells from severe dysmenorrheal patient in vitro is the same with none-mild dysmenorrheal and normal group.7.The level of NGFmRNA,TrkAmRNA and P75mRNA expressed by eutopic endometrial stromal cells and normal endometrial stromal cells showed no significant change before and after intervention by different concentrations of estrogen, progesterone.8.After administratimg 17βestradiol in stromal cellsof patients with endometriosis and normal endometrial eutopic endometrial stromal cells,the NGFmRNA,from 8 to 24 hours showed an upward trend,while the control group has a downward trend.9.Different concentrations of GnRHa significantly decreased the level of NGFmRNA, TrkAmRNA,P75mRNA in ectopic endometrial cells.There are significant differences compared to the control and the normal group.10.Dysmenorrhea endometriosis patients have a higher level of nociceptin.NGF expression in the serum has the same trend with OFQ.11.Serum NGF decreased in the post-drug period as well as the serum OFQ expression in patients with a history of GnRHa treatment.Conclusions:1.This is the first report of the expression of NGF,TrkA and P75 in endometriosis lesions,which is related to different part of endometriosis lesions and the severity of pain.2.NGF and its receptor TrkA is expressed more in eutopic endometrial stromal cells, suggesting that endometriosis eutopic endometrial is abnormal in patients with dysmenorrhea,and further enrich the "eutopic endometrial determinism".GnRHa may have multiple mechanisms in the treatment of pain.3.Pain substances may be elevated in peripheral blood of patients with endometriosis who have severe dysmenorrhea.GnRHa administration can decreased substance, suggesting that it can reduce pain-related factor expression through multiple ways.