||Comparison of follicle growth rates after IVA (in vitro activation) treatment with those of base line numbers.
|Key secondary outcomes
||To evaluate the presence of residual follicles, we measure the levels of serum biomarker for early follicles before ovariectomy. Using part of the ovarian tissue, we evaluate the conditions of the follicles and oocytes after thawing structurally and histologically. To evaluate the efficacy and presence of residual follicles, part of the ovarian tissues is used for cultivation, ultrasound/ Far infrared ray scanning and histological examination (follicle count, fibrotic evaluation, antithyroid antibody and inflammatory change) to evaluate the effect of IVA and the presence of residual follicles.
To judge the effect of hCG on induction of angiogenetic factors production after autografting, we collect part of uterine endometrium before autografting and determine the expression levels of related genes and proteins in neovascularization. We further measure serum VEGF levels after hCG administration.
In the case of very rare type of POI, Resistant Ovary Syndrome (ROS), we detect anti-FSH and FSHR antibodies, analyze their responsible genes and ovarian tissue structures, and measure serum levels of CCN growth factors after linear cutting of ovarian cortex.
After IVA, we perform ovarian stimulation by hormonal therapy. Under evaluation of estradiol, progesterone, FSH, LH levels, we stimulate follicle growth for subsequent IVF. We compare the proportions of oocyte retrieval, oocyte maturation, fertilization, high-quality embryos, pregnancy and abortion with those of base line levels.
|No. of arms
|Purpose of intervention
|Type of intervention
1.We measure serum molecular markers before surgeries to evaluate the effect of hippo signal inhibition and presence of residual follicles.
2.Whole or part of one ovary is removed under laparoscopic surgery. ovarian cortex is isolated and cut into small tissues (
1x1cm,1-2mm thickness )
before vitrification.to induce secondary follicle growth, in situ incision of ovarian cortex is performed on the residual ovary.
3.To determine follicle numbers in ovarian tissues, we count follicles under histological analyses. part of ovarian tissues is used for cultivation, ultrasound, far infrared ray scanning and histological examination.
4.After thawing, ovarian strips are fragmented into 1-2mm cubes and cultured with IVA
medium containing a PTEN inhibitor (bpV)
and PI3K activators (740YP,phospharidic acid, and /or propranolol) for 48h.
5.After culture, the activated ovarian cubes are washed with medium without IVA drugs. The ovarian cubes are transplanted beneath serosa of Fallopian tubes and to the residual ovary and Douglas' pouch.
6.We treat patients with estrogen for 2 weeks before autografting. After grafting, hCG is administered to induce production of
angiogenesis factor to help vascularization and survival of grafts. To judge the effect of hCG, part of endometrium is collected to measure related gene and protein levels before autografting. Angiogenesis factor levels are measured after hCG.
7.We measure molecular markers to evaluate the effect of Hippo signal inhibition after grafting. We treat the patients with
gonadotropin for ovarian stimulation for IVF-ET. Ovarian stimulation continues for one to two years. If ovarian stimulation is not effective, we perform another autografting.
For patients who have high possibility to have residual follicles, we perform fresh IVA. In fresh IVA, we remove their ovaries and culture the ovarian tissues with IVA medium for 48 hours without cryopreservation. After culture, we perform autograft of ovarian tissues under the same method of IVA. Remaining ovarian tissues are vitrified using the same method. Other methods and outcome measures are same as IVA.
For patients who are predicted to become
POI but still have irregular menstruation,
we fragmented the ovarian tissue into
small cubes and autograft them immediately without culture to stimulate secondary follicle growth leading to increase in the number of retrieved oocytes.
1.Excise one side of whole ovary or partial ovarian tissues under laparoscopic surgery. After removal of medulla tissues to prepare ovarian cortex, the ovarian cortex was dissected into 3x3cm with 1-2mm thickness of tissue stripes and further fragmented into 1-2mm cubes. To measure the number of secondary follicles in the dissected ovarian tissue,10% of volume of the tissue is fixed and then each developmental stage of follicles are counted under histological analyses.
2.The ovarian cubes are auto-transplanted beneath of the serosa of both Fallopian
tubes, remaining ovaries and/or Douglus' pouch. We also directly cut ovarian cortex of the contra lateral ovary to induce physical stimulation.
3.Remaining ovarian tissues are vitrified using the same method. Other methods and outcome measures are same as IVA.
||Resistant Ovary Syndrome(ROS) is a very rare type of POI. In contrast to regular POI, ROS patients still have higher number of residual follicles in their ovaries. However, the follicles do not respond to endogenous and exogenous FSH stimulation, resulting in failure to grow. Based on our previous studies, physical stimulation to ovaries induces the production of CCN growth factors, leading to ovarian follicle stimulation. Thus, we perform in situ incision of ovarian cortex as a less invasive approach for ROS patients. When in situ incision of ovarian cortex is not effective, we perform Drug-free IVA. In some cases of ROS patients, auto-antibodies such as anti-FSH/FSHR antibody are found as causes of the disease, we use steroid hormone treatment to suppress the production of auto-antibodies. Using blood tests, we measure the levels of auto-antibodies. We analyze responsible genes for ROS, and perform histological analysis of ovarian structure. We also measure the levels of CCN growth factors after the surgery.
Correct serum from the patients before surgery to measure molecular markers to ensure suppression of Hippo signaling.
We directly cut ovarian cortex or scratch ovarian cortex to induce physical stimulation to the ovary. To measure the number of secondary follicles in the dissected ovarian tissue, 10% of volume of the tissue is fixed and then each developmental stage of follicles are counted under histological analyses.
Correct serum from the patients after surgery to measure molecular markers to ensure suppression of Hippo signaling. After grafting, patients receive ovarian stimulation using gonadotropin drugs to stimulate follicle growth and retrieve oocytes. Mature oocytes are fertilized by in vitro fertilization and preimplantation embryos are transplant into uterus. The treatment continues for 1-2 years. If first auto-transplantation is not successful and patients have cryopreserved ovarian tissues, patients can repeat these procedures.