BACKGROUNDVitrification is a prospective technology in ovarian tissue cryopreservation, but it is still in an initial stage. This study was conducted to investigate a modified vitrification protocol for human ovarian tissue, which can be used as an alternative to preserve fertility for young women with cancer who have to undergo cytotoxic therapy and sterilization.

METHODSOvarian tissue samples were collected from 15 patients and randomly allocated to groups of fresh, vitrification, and conventional slow freezing. A modified carrierless vitrification method was applied. The proportion of morphologically intact follicles in fresh ovarian tissues was compared with that in warmed/thawed tissues. The initial growth of the follicles and the concentrations of estradiol and progesterone were detected to determine the viability and endocrine function of the cryopreserved tissues.

RESULTSThe proportion of morphologically intact primordial follicles in the fresh group (97.6%) was significantly higher than that in the other two groups (vitrification group 80.3% and slow-freezing group 72.6%, P < 0.001). In both the vitrification and slow-freezing groups, estradiol and progesterone were secreted continuously during 2-week culture in vitro, the proportion of primary follicles were both significantly increased compared to the fresh group. No statistically significant differences existed between the two groups after cryopreservation in the proportion of both primordial and primary follicles, and the concentrations of estradiol and progesterone (P > 0.05).

CONCLUSIONThe modified vitrification method for cryopreservation of human ovarian tissues is effective, simple, and inexpensive.

Adult ; Cryopreservation ; methods ; Estradiol ; biosynthesis ; Female ; Humans ; Ovary ; cytology ; metabolism ; Progesterone ; biosynthesis ; Tissue Culture Techniques

OBJECTIVETo investigate the expression of 17 beta-hydroxysteroid dehydrogenase type 1 (17β-HSD1) in the kidney of rats and explore the capacity of the kidney for synthesizing sex hormones.

METHODSThe expressions of 17-HSD1 and sex hormones were detected by Western blotting and radioimmunoassay in rat renal cells in primary cultured for 24 and 48 h in the presence or absence of follicle-stimulating hormone (FSH) and luteinizing hormone (LH).

RESULTSAfter cell culture for 24 h, the primary rat renal cells expressed a low level of 17β-HSD1 (0.1843±0.076), which increased to 1.6651±0.044 (P<0.01) in response to co-stimulation by FSH and LH. Low levels of estradiol, progesterone and testosterone were also detected in rat renal cells (3.30±3.78, 62.60±12.33, and 22.12±3.36, respectively), and co-stimulation of FSH and LH significantly increased their levels to 8.50±2.64, 117.80±9.79, and 45.04±4.39, respectively (P<0.05). The levels of these hormones showed no significant differences between cells cultured for 24 h and 48 h (P>0.05).

CONCLUSIONThe rat renal cells express 17β-HSD1 and are capable of stably secreting sex hormones in response to co-stimulation with FSH and LH, suggesting the capacity of the rat kidneys for synthesizing sex hormones. These findings enrich the understanding of the endocrine function of the kidney.

17-Hydroxysteroid Dehydrogenases ; metabolism ; Animals ; Cells, Cultured ; Estradiol ; biosynthesis ; Follicle Stimulating Hormone ; pharmacology ; Kidney ; enzymology ; Luteinizing Hormone ; pharmacology ; Progesterone ; biosynthesis ; Rats ; Testosterone ; biosynthesis

A rapid decline in egg production of laying hens begins after 480 d of age. Such a rapid decrease results predominantly from the ovarian aging, accompanied by endocrine changes, decreased yolk synthesis and accumulation, and the reduction in follicles selected into the preovulatory hierarchy. In this study, hens at 90, 150, 280, and 580 d old (D90, D150, D280, and D580, respectively) were compared for yolk precursor formation in the liver to elucidate effects of aging on laying performance. The results showed that liver lipid synthesis increased remarkably in hens from D90 to D150, but decreased sharply at D580 as indicated by the changes in triglyceride (TG) levels. This result was consistent with the age-related changes of the laying performance. The levels of liver antioxidants and total antioxidant capacity decreased significantly in D580 hens and the methane dicarboxylic aldehyde in D580 hens was much higher than that at other stages. The serum 17β-estradiol level increased from D90 to D280, but decreased at D580 (P<0.05). The expression of estrogen receptor α and β mRNAs in the liver displayed similar changes to the serum 17β-estradiol in D580 hens. Expressions of the genes related to yolk precursor formation and enzymes responsible for fat acid synthesis were all decreased in D580 hens. These results indicated that decreased yolk precursor formation in the liver of the aged hens resulted from concomitant decreases of serum 17β-estradiol level, transcription levels of estrogen receptors and critical genes involved in yolk precursor synthesis, and liver antioxidant status.
Age Factors ; Animals ; Antioxidants ; metabolism ; Chickens ; Egg Yolk ; metabolism ; Estradiol ; blood ; Female ; Lipids ; biosynthesis ; Liver ; metabolism ; Oviposition ; Receptors, Estrogen ; genetics

This study was aimed to investigate whether 2-methoxyestradiol (2ME2) could exert effect of inducing differentiation on myeloma cells. A myeloma cell line CZ-1 secreting lambda light chain protein was used as an object of study. The CZ-1 cell morphology was observed by Wright's staining, the CD49e expression on cell surface after treatment with 2ME2 was detected by flow cytometry, the concentration of lambda light chain protein in the supernatant was assayed by immuno-scattering turbidity method. The results showed that treatments with 0.1-0.5 micromol/L 2ME2 for 72 hours resulted in some mature morphological changes of CZ-1 cells, such as the ratio of karyoplasms going down, nucleolus reducing or disappearing, chromatin getting rougher and more compacted; the CD49e positive CZ-1 cells increased by 2ME2 with concentrations of 0.1 micromol/L to 0.5 micromol/L in a concentration-dependent manner. The statistical difference from the control group was significant; the concentration of lambda light chain protein increased from control group 29.3 +/- 2.77 microg/ml to 35.97 +/- 2.6 microg/ml (P < 0.05) after exposure to 0.1 micromol/L 2ME2 for 72 hours, and the treatment of 0.5 micromol/L 2ME2 up-regulated lambda light chain protein to 79.67 +/- 1.88 microg/ml (P < 0.01) continuously. It is concluded that 2ME2 at low-concentration can induce differentiation of the CZ-1 cells to mature, which provides a new, and safe strategy for myeloma therapy.
Cell Line, Tumor ; Cell Transformation, Neoplastic ; drug effects ; Estradiol ; analogs & derivatives ; pharmacology ; Humans ; Immunoglobulin lambda-Chains ; biosynthesis ; Integrin alpha3 ; biosynthesis ; Multiple Myeloma ; metabolism ; pathology

OBJECTIVETo investigate the effect of Yangjing Zhongyu decoction (YZD) on metalloproteinase-9 (MMP-9) and its inhibitor-1 (TIMP-1) expression and sex hormone regulation in mid-luteal phase endometrium of women with cryptogenic infertility.

METHODSIn situ hybridization and reverse transcription-polymerase chain reaction method was used to detect MMP-9 and TIMP-1 mRNA, and radioimmunoassay was used to determine levels of serum estradiol (E2) and progesterone (P) synchronously, of 22 infertile women during mid-luteal phase.

RESULTSAfter treatment, the mid-luteal serum E2 and P level was 451.501 +/- 226.342 pmol/L and 46.502 +/- 19.948 nmol/L respectively, significantly higher than that before treatment (304.656 +/- 135.853 pmol/L and 33.782 +/- 15.459 nmol/L respectively), the difference was significant (P < 0.01). Staining of MMP-9 mRNA positive granules in cytoplasm and nuclei of adeno-epithelial cell mid-luteal phase endometrium deepened significantly, but the change in mesenchym was insignificant. The MMP-9 mRNA expression after treatment was 0.617 +/- 0.186 (grey level), significantly higher than the level before treatment (0.490 +/- 0.370), comparison between them showed significant difference (P < 0.05). Change of TIMP-1 mRNA expression in adeno-epithelial and mesenchym before and after treatment was insignificant (0.588 +/- 0.191 vs 0.621 +/- 0.146, P < 0.05). Correlation analysis showed that the quantitative difference of P value before and after treatment was positively correlated with the difference of MMP-9 mRNA before and after treatment (r = 0.682, P < 0.01).

CONCLUSIONYZD could soothen Gan and nourish Shen, raise the level of mid-luteal phase serum P, and further promote MMP-9 gene expression in endometrium to benefit the degradation of extracellular matrix of endometrium, and facilitate for blastocyst implantation.

Adult ; Drugs, Chinese Herbal ; therapeutic use ; Endometrium ; metabolism ; Estradiol ; blood ; Female ; Humans ; Infertility, Female ; drug therapy ; metabolism ; Matrix Metalloproteinase 9 ; biosynthesis ; genetics ; Phytotherapy ; Progesterone ; blood ; RNA, Messenger ; biosynthesis ; genetics ; Tissue Inhibitor of Metalloproteinase-1 ; biosynthesis ; genetics

OBJECTIVE: To investigate the effects of long-term estrogen replacement treatment (ERT) on the expression of bcl-2 and H-ras in rat endometrium. METHODS: Thirty 5-month-old SD female rats were randomly divided into 3 groups: Control group ( sham operated and vehicle injected, n 10) , OVX group (OVX operated and vehicle injected, n = 10) , and ERT group (OVX operated and 17 beta-estradiol injected, n = 10). The rats were killed in the 13th week and the uteri were isolated and weighed, pathologically analyzed, and we measured the thickness of the endometrium. Immunochemistry and in situ hybridization analysis were used to examine the changes of bcl-2 and H-ras mRNA and Bcl and H-ras protein expression in the endometrium of the rats. RESULTS: Uterine weight and endometrial thickness of OVX decreased much more than those of the control (P <0.01 ) and ERT rats (P < 0.01). One simple hyperplasia and one squamous metaplasia of endometrium were found in ERT rats. Quantitatively, bcl-2 and H-ras mRNA and Bcl and H-ras protein level of ERT were higher than those of OVX rats (P < 0.01 ), and there were no statistical significances between the ERT group and the control rats. CONCLUSION Long-term estrogen replacement can keep the endometrium from atrophy, and lead to the genesis of simple hyperplasia and squamous metaplasia of the endometriun, which can increase the risk of endometrial carcinomas. Estrogen may inhibit apoptosis and promote the proliferation of endometrial cells through increasing the expression of bcl-2 and H-ras mRNA and Bcl-H-ras proteins.
Animals ; Endometrium ; metabolism ; Estradiol ; pharmacology ; Estrogen Replacement Therapy ; Female ; Genes, bcl-2 ; Genes, ras ; Ovariectomy ; Proto-Oncogene Proteins c-bcl-2 ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Time Factors ; ras Proteins ; biosynthesis ; genetics

The effect of angiotensin II (Ang II) on the follicular development was studied by using an animal model of follicular atresia induced by pregnant mare s serum gonadotropin (PMSG). The results showed that: (1) a large number of atretic follicles were found in the ovary of 24-day-old mouse after 6-day treatment of PMSG. Deoxyribonucleic acid (DNA) extracted from granulosa cells clearly showed a ladder band under agarose gel electrophoresis analysis. (2) the contents of Ang II in the ovary extremely increased with the development of follicular atresia. (3) Ang II significantly antagonized the stimulating effect of the follicle-stimulating hormone (FSH) on estradiol (E(2)) generation of granulosa cells. It is suggested that Ang II may be involved in the regulation of follicular atresia in mouse.
Angiotensin II ; pharmacology ; physiology ; Animals ; Cells, Cultured ; Estradiol ; biosynthesis ; Female ; Follicle Stimulating Hormone ; pharmacology ; Follicular Atresia ; physiology ; Gonadotropins, Equine ; pharmacology ; Granulosa Cells ; drug effects ; metabolism ; Mice

OBJECTIVETo observe the influence of acupuncture on embryo implantation in rat model of embryo implantation dysfunction, and to primarily explore its possible mechanisms.

METHODSPregnant rats were randomly allocated into the control group, the model group and the acupuncture group. The pregnancy rate and average number of blastocyst were observed, the serum levels of estrodiol (E2), progesterone (P4) and prolactin (PRL) were detected by RIA, and the protein and mRNA expression of progesterone receptor (PR) and prolactin receptor (PRLR) in endometrial tissue of implantation site were determined using immunohistochemical assay and RT-PCR respectively.

RESULTSThe pregnancy rate and average number of blastocyst were significantly higher in the acupuncture group than those in the control group respectively (P <0.01). The serum levels of P4 and PRL as well as the protein and mRNA expression levels of PR and PRLR in the model group were significantly lower than those in the other two groups (P<0.05).

CONCLUSIONAcupuncture can promote embryo implantation in rats to a certain degree, and its mechanism might be related with the effects of acupuncture in mediating the sexual hormone levels and the receptor expression of rats.

Acupuncture Therapy ; Animals ; Embryo Implantation ; physiology ; Embryonic Development ; physiology ; Estradiol ; blood ; Female ; Immunohistochemistry ; Male ; Pregnancy ; Progesterone ; blood ; RNA, Messenger ; genetics ; metabolism ; Radioimmunoassay ; Random Allocation ; Rats ; Rats, Wistar ; Receptors, Progesterone ; biosynthesis ; genetics ; Receptors, Prolactin ; biosynthesis ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

To investigate the relationship between the insulin resistance (IR) of polycystic ovarian syndrome (PCOS) rat model induced by dehydroeplandrosterone (DHEA) and hormonal changes in the ovarium and the resistin mRNA levels in adipose tissue, 21-day-old female SD rats were divided into two groups in pairs. The rats in group 1 were injected daily (s.c.) with DHEA for up to 20 days and the rats in group 2 injected with oil at the same time. Ovarian weight, serum insulin levels and sex hormone levels in rat blood of both groups were determined. Oral glucose tolerance tests, light microscopic and electronic microscopic examination were performed. The levels of resistin mRNA in adipose tissue were measured by reverse transcription-polymerase chain reaction (RT-PCR). Our results showed that the ovarian weight in group 1 was greater than that in group 2 (P<0.05). The ovaria in group 1 showed multiple follicular cysts, The serum testeosterone and etrasdiol concentration were significantly higher in group 1 than those in group 2 (P<0.001 and P<0.05 respectively), so as the fasting serum glucose (P<0.001) and fasting serum insulin (P<0.05). The value of 1/FINS x FGC was significantly higher in group 1 than that in group 2 (P<0.001). Moreover, the resistin mRNA level of white adipose tissue in the DHEA-induced group was significantly higher than that in the control rats (P<0.05). It is concluded that the DHEA-induced PCOS rat models were similar to those of the patients with PCOS, and the IR was observed. Resistin secreted by adipose tissue may mediate IR in PCOS, and it is likely involved in the pathogenesis and development of PCOS.
Adipose Tissue ; metabolism ; Animals ; Animals, Newborn ; Dehydroepiandrosterone ; Estradiol ; blood ; Female ; Insulin Resistance ; Polycystic Ovary Syndrome ; chemically induced ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Resistin ; biosynthesis ; genetics ; Testosterone ; blood

OBJECTIVETo study the effect of 17beta-estradiol (17beta-E(2)) and medroxyprogesterone acetate (MPA) on the expression of VEGF mRNA in epithelial ovarian carcinoma cell line in vitro.

METHODSHuman epithelial ovarian carcinoma cell line SKOV3 cells were exposed to 17beta-E(2) (10(-12) approximately 10(-8) mol/L) or MPA (10(-9) approximately 10(-5) mol/L), the expression level of VEGF mRNA was determined by RT-PCR.

RESULTSThe PCR products of VEGF were mainly 606 bp and 474 bp. The expression of VEGF mRNA in the 17beta-E(2)-treated cells was significantly higher than in the untreated control (P < 0.05), the expression level of VEGF mRNA increased with the increase in 17beta-E(2) concentration. The expression of VEGF mRNA in SKOV3 cells treated with MPA (10(-9) approximately 10(-5) mol/L) was significantly lower than in the untreated control (P < 0.05), especially that of 606 bp.

CONCLUSIONEstrogen can up-regulate, while progesterone can down-regulate mRNA expression of VEGF. Further studies are needed to elucidate the mechanism.

Adenocarcinoma ; metabolism ; pathology ; Antineoplastic Agents, Hormonal ; pharmacology ; Cell Line, Tumor ; Dose-Response Relationship, Drug ; Estradiol ; pharmacology ; Female ; Humans ; Medroxyprogesterone Acetate ; pharmacology ; Ovarian Neoplasms ; metabolism ; pathology ; RNA, Messenger ; biosynthesis ; genetics ; Vascular Endothelial Growth Factor A ; biosynthesis ; genetics