Pregnancy in a 46-year-old woman after autologous granular cell mitochondria transfer.

Growth of 4 follicles was achieved in a 46-year-old infertile woman and 2 mature oocytes were subsequently obtained after pituitary desensitization with triptorelin (1.875 mg) and hyperstimulation with recombinant human FSH (Gonal-F) and urinary human menopausal gonadotropin (HMG). The granular cells were collected from the cumulus and follicle fluid of the patient with Percol gradient centrifugation. The cells (about 1 x 10(6) /ml) were homogenized for 6 times with cold glass homogenizer, the resultant crude homogenate centrifuged for 20 min at 2000 x g. The supernatant was again centrifuged for 20 min at 9800 x g, after which the pellets of the mitochondria were resuspended in 1 microl modified human tubal fluid (mHTF) medium and maintained at 4 degrees Celsius. A microinjection needle with the inner diameter of approximately 7 microm was used to aspirate the mitochondrion suspension for about 100 microm in length on the scale of the needle for delivering the mitochondria, along with the immobilized sperms, into a single MII oocyte. About 3000 mitochondria were transferred into each oocyte. The two oocytes were both fertilized, but one had 3 pronucleus, and the normal fertilized oocyte developed into 8-blastomere embryo 3 d after its transfer into the patient's uterus. The urine test confirmed clinical pregnancy in the woman 15 d later, and 30 d after the transfer, the fetal body and heart beat could be observed by ultrasound, but unfortunately, spontaneous abortion occurred at the ninth week. According to our knowledge, this woman is the eldest person in mainland China to receive assisted reproduction operations that successfully got clinical pregnancy with her own oocyte.