Effect Of Angiogenesis Inhibitor Nordihydroguaiaretic Acid In Model Of Swines To Endometriosis Function

Nordihydroguaiaretic acid (NDGA) is the major chemical component distracted from a natural shrub named L.tridentata growing widely in the Southwest of America as well as the Northern part of Mexico. It has been used in traditional therapy for wound healing, pain relieving ,eliminating inflammation and driving out parasites. NDGA can also be synthesized: generally, it is made as the food anticorrosive; as a stong lypoxygenase inhibitor, which has all biological effects including anti- inflamation , anti- virus ,anti-tumor ,anti- angiogenesis, etc. Thus, NDGA is widely used in traditional medicine and a variety of studies.Many studies about the biological activities of NDGA have been conducted. One of the most representative activities of NDGA revealed is that it can inhibit the activity of lipoxygenase. Recently ,Some studies have found that NDGA has an anti-tumour activity .It has been demonstrated that the formation of tumor was critical to the new Angiogenesis and proliferative vascular endothelial cell.NDGA as a anti–Angiogenesis inhibiting agent has strong inhibiting function in proliferating,transfering and forming endotheliocyte.NDGA decreases the growth of VEGF and expression of VEGF- acceptor. It has been suggested that NAGA has a potentially important action in inhibiting the Angiogenesis of tumor.A study by BianXiuwu demonstrated that NDGA has the function of suppressed proliferation , cell differentiation and Apoptosis of spongiocytoma in internal . This indicates that Angiogenesis acts as an extreme actor in tumor.It has also been proved that NDGA has an anti–angiogenesis action .A study in the chicken chorioallantoie membrane assary was carried out to detect the effect of NDGA on angiogenesis.NDGA can effectively inhibit the angiogenesis. Anti–Angiogenesis therapy may be acquired in Endometriosis and angiogenesis, bearing a very important link .In the female reproductive system, the phenomena of angiogenesis can be observed apparently in the processes of periodical proliferation of endometrium.Hence,a basic method to study the anti–Angiogenesis in endometriosis .The aim of this research is to form a model study of animal and to observe the effect of NDGA on ectopic endometriosis. We have investigated the morphological changes of ectopic endometriosis detected in the expression of vascular endothelial growth factor(VEGF), and factorⅧrelating to antigen (FⅧRAg) and to measure the serum concentrations of estradiol (E2), progesterone (P), follicle stimulating hormone (FSH) and luteinizing hormone (LH) by methods employing immunohistochemistry (IHC) stain, HE stain, image analysis and chemiluminoimmunoassay analysis(CLIA).The approaches and main results are as follows:Part 1.construct a model of BaMa swine animal in endometriosis Female six swines with sexual maturation were choosen and endometriosis tissue acquired by operation. The establishment of ectopic endometriosis in the animal model is successful.1.difolliculine with the dose of 30ug/kg was injected before operation in three and six days so as to acquire proliferative phase in endometriosis.2.subcutaneouly tissue and abdominal cavity was implanted in ectopic endom- etriosis .There formed many ectopic focus in corresponding locations.3. difolliculine with the dose of 30ug/kg respectively every other day for 5 days, then, injected after operation .It enhanced the growth of implanted endometriosis.4 . implanted subcutaneouly tissue was observed twice in a week. Implanted endometriosis was gradually growing and forming nodus.5. In 3 weeks , implanted tissue was observed by operation, at the same time, implanted location formed a nodus and cyst.6.A few implanted tissues were elected to test and showed glandular organ and stroma in pathology.It indicated that obtained model in endometriosis was successful.Part 2. Effect of NDGA on the model of ectopic endometriosis by subcutaneous administration The successful models of Six female BaMa swines in normal status were randomly divided into experimental group (NDGA) and contrasting (PBS) group. Subcutaneously injected PBS solution and 3%NDGA at the dose of 20mg/kg were used respectively every other day for 20 days . The serum concentrations of E2, P, FSH and LH were detected.Sections were cut at 4～6μm and stained with HE and IHC with FⅧRAg and VEGF.(1) Variation of Hormone In NDGA group, the secretion of E2 decreased(p<0.01)but that of P was found to be a little increased. FSH and LH showed no difference between before and after injection. There was a low level figure(p>0.05).(2)observing surface endometriosis of swines with NDGA decreased in subcutaneous tissues through observation twice in a week.The controlled groups did not change obviously ,but some masses were increased. In three weeks ,Curing endometriosis of swines with NDGA decreased and even a few disappeared against endometrium. The control groups did not change obviously ,but some of masses were increased.(3) observing morphology in three weeks NDGA group was decreased both in the number of endometrial glands and in the number of stroma cells. The Contrasting group was increased both in the number of endometrial glands and in the stroma cells.(4) FⅧRAg stained sections of MVD analysis NDGA group blood vessel endothelium was few and showed light colour, while the amount of blood vessel was decreased. Anti-FⅧRAg expressed weakly positive in IHC stain sections.; PBS group blood vessel endothelium was many in number and showed deep colour, whereas the amount of blood vessel was increased. Anti-FⅧRAg expressed strongly positive in IHC stain sections.; (P<0.05). There was significant difference in the statistics system.(5) VEGF expressiveness NDGA treated blood vessel endothelium was stained in light-brown colour and expressed weakly positive in IHC stain sections. PBS group blood vessel endothelium was stained in deep brown colour and expressed stongly positive in IHC stain sections. (p<0.05). There was a significant difference between the NDGA group and the PBS group.Conclusion : 1. Sexual maturation Bama animals were choosen and endometriosis tissue acquired by operation. successful model of animal in endometriosis was established.2. Injected NDGA Subcutaneously,VEGF expressiveness and the amount number of MVD decreased; formed vassular and generation were in effect; ectopic endometriosis inhibited; ectopic endometriosis decreased. It is the expected results.