Experimental Study On The Endostatin Treatment Of Endometriosis

Background and PurposeThe endometriosis (Endometriosis, EM) is one kind of gynecology department common disease. Although belongs to the benign disease, it has the malignant tumor characteristic like hyperplasia, invasion, recrudescence and etc. There are some kinds of methods to treat EM. But the result for most of the treatments is actually unsatisfied .Except radical cure surgery, of which the 5-year recurrence rate was more than 40 percent. At present the exact cause of endometriosis is still not completely clear . the research indicated that the endometrium fragment (gland epidermis and intersititial cell) can survives,grows, and causes the pathological change and the symptom only through adhere, Infiltration and the vascularization.In this process the establishment of the newborn blood vessel plays the key role to the survival of the endometrium planter and the formation of EM . Recent research indicates that the variety of cytokines play an important role in the occurrence of the disease.Especially the vascular endothelial growth factor (VEGF), which stimulates angiogenesis and promoting the development of EM. Therefore, we speculate inhibition of vascular endothelial growth factor in the disease may have therapeutic effects. At present recombinant human endostatin (ENDOSTAR) has been listed in China for the treatment of malignant tumors, which inhibited tumor growth mainly through inhibition of vascular endothelial growth factor. It suggested that the drug may be similar therapeutic effect on EM. Thus we use human endometrial tissue in nude mice by abdominal subcutaneous injection to establish the animal model of endometriosis.Then we may observe the therapeutic effect of ENDOSTAR on the model by local injection of endostatin.Methods1) First step: to establish endometriosis animal model in nude mice Endometria from benign gynecologic disease due to the removal of human uterine endometrium (or induced abortion uterine decidual tissue curettage) were planted into 3-4 week-old female nude mice abdominal by subcutaneous injection. Then giving estradiol benzoate intramuscular injection at dosage of 2 mg/kg immediately and continuous injection in each five days. 4 weeks later, executes the bare mouse and observe the hypodermic infection's size and the blood vessel growth situation, Then we take graft lesions to observe the histopathological examination by HE staining and immunohistochemical staining with the rabbit anti-human polyclonal anti-keratin antibody, monoclonal mouse anti-ki-67 and microvascular density (MVD) in order to undersdand whether animal model successed or not.2) Second step: to observe the the therapeutic effect of ENDOSTAR on EMBased on the above methodology, We established endometriosis model in nude mice, in the sixth day after surgery , nude mice were divided into two groups: normal saline control group and ENDOSTAR treatment group(whichafter called the ES group), then local injection 0.05 ml saline and 2.5mg/kg ENDOSTAR into graft lesions every other day, respectively. 4 weeks later, executes the bare mouse and observe the hypodermic infection's size and the blood vessel growth situation.Then we take the graft lesions to understand therapeutic effect of ENDOSTAR on endometriosis through HE staining and immunohistochemical staining.Results1) modelling result: 12 modelling nude mouse survived completely.the abdomen hypodermic obviouslly appear soybean size's transplant lesion,which fuses with the nude mouse muscular tissue.The transplant lesion appears sticks out translucently small saccate obvious blood vessel on the appearance. Pathology biopsy can find obviously the human endometrial glands and stroma. Gland epithelial cells stained with keratin, glandular epithelial cells staining ki-67 antibody and the gland epithelial cell ki-67 dyeing masculine gender rate is the 17.68%. Microvascular density with CD34 mark is 18.35±4.5.2) Among the saline group, 12 mice survived. Abdomen hypodermic of 11 mouse obviouslly appear 3-5mm size's transplant lesion,which fuses with the nude mouse muscular tissue.The transplant lesion appears sticks out translucently small saccate obvious blood vessel on the appearance. Pathology biopsy can find obviously the structured and tidy human endometrial glands and stroma. Gland epithelial cells stained with keratin, glandular epithelial cells staining ki-67 antibody and the gland epithelial cell ki-67 dyeing masculine gender rate is the 18.48%. Microvascular density with CD34 mark is 19.60±5.8. The results are similar to the model group's.The experimental group (the ES group) survives 11. Abdomen hypodermic of 10 mouse obviouslly appear 3-5mm size's transplant lesion.The size of lesion have not obvious difference to NS group's (P>0.05). The lesion appears tuberculous and the quality of material is harder, whiter, and without obvious blood vessel proliferation. Pathology biopsy can find obviously the human endometrial glands and stroma. but gland appears disorder and partial gland epithelial cell atrophied. Gland epithelial cells stained with keratin, glandular epithelial cells staining ki-67 antibody and the gland epithelial cell ki-67 dyeing masculine gender rate is the 3.6%, which obviously lower than the NS group (P< 0.01). Microvascular density with CD34 mark is 7.28±3.6,which lower than the NS group (P< 0.05).Conclusion1) We successfully established endometriosis animal model in nude mice using human endometrial tissue. Model traits appeared stable, and retained the original characteristics of the organization.2) Injected endostatin(ENDOSTAR),VEGF expressivess and the amount number of MVD decreased, formed wassular and generation were in effect,ectopic endometriosis inhibited, ectopic endometriosis decreased . It is the expected rescults.