There have been some proposals that stem cells exist in the ovarian surface epithelium (OSE) of the adult human ovary; however, no direct evidence of such cells has been given until now. The aim of ...this study was to isolate the putative ovarian stem cells (OSCs) from the OSE layer in women with no naturally present oocytes and follicles—20 postmenopausal women and five women with premature ovarian failure. Small round cells with a bubble-like structure and diameters from 2 to 4 μm were isolated from the material obtained by OSE scraping. They expressed early embryonic developmental markers such as stage-specific embryonic antigen-4 and Oct-4, Nanog, Sox-2, and c-kit transcription markers, and they displayed prominent c-kit immunohistochemical staining. These cells were separated by density gradient centrifugation and grown
in vitro, where they proliferated. Some of them grew intensively and reached a diameter of approximately 20 μm after 5–7 days. In the OSE cell culture, oocyte-like cells developed, which reached a diameter of up to 95 μm and expressed Oct-4A, Oct-4B, c-kit, VASA, and ZP2 transcription markers, corresponding to early oocytes. They did not express SCP3 meiotic marker. In conclusion, the discovered cells are proposed to represent the adult OSCs with the expression of embryonic stem cell markers. The expression of germ lineage marker c-kit points toward their primordial germ cell ancestry. A new term “embryonic-like stem cells of the adult” is proposed for embryonic-like stem cells that might persist in various tissues and organs of adults. These findings could be used for further studies aimed at the autologous treatment of ovarian infertility and degenerative diseases.
To evaluate the involvement of immune abnormality in patients with idiopathic premature ovarian insufficiency (POI). In addition to the known etiology, autoimmune disorders may be a pathologic ...mechanism for POI.
Our study was a prospective controlled trial. Twenty women with POI, reasons other than autoimmune excluded, were enrolled in this study. The control group consisted of 17 healthy women. In both groups, family and personal history were taken and the levels of follicle stimulating hormone, luteinizing hormone, thyroid-stimulating hormone, prolactin, anti-Müllerian hormone, inhibin B, antithyroglobulin and antithyroid peroxidase antibodies were determined. Antiovarian antibodies and subpopulations of peripheral blood T-lymhocytes were also determined.
Participants in the study group exhibited hypergonadotropichypogonadism, while high levels of follicle stimulating hormone and low levels of inhibin B and anti-Müllerian hormone were observed. In 16 (80%) patients, POI was associated in their personal and familial history with another autoimmune disease. Fifty percent of patients presented highly elevated antithyroid antibodies. The lymphocyte subset, especially B cells, was significantly higher (p=0.014), and peripheral regulatory lymphocytes CD25+ high were significantly lower (p=0.015) in the study group than in the control group. Anti- ovarian antibodies were detected in 20% of patients with POI.
We presume that the presence of anti-ovarian antibodies together with abnormalities of cellular immunity may in some cases potentially represent the involvement of an autoimmune mechanism in idiopathic POI.
Summary
High seminal reactive oxygen species (ROS) are related to poor semen quality and impaired fertilization. We aimed at finding whether there is an association between ROS and fertilization, ...embryo quality and pregnancy rates after conventional in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). In prepared semen of 147 male partners of infertile couples, ROS were assessed with luminol chemiluminescence. Spermiogram was assessed in native semen. ROS were negatively correlated with standard sperm characteristics and testicular volume, and positively with abnormal sperm head morphology. Fertilization rate and embryo morphology on day 2 and on day 4 were assessed in 41 IVF and 106 ICSI cycles. The influence of maternal (female age and number of oocytes) and paternal (sperm motility, morphology and ROS) factors on fertilization and embryo quality were assessed by means of regression analyses. After IVF, fertilization and pregnancy rates were negatively associated with ROS level (p = 0.031 and 0.041, respectively). In case of higher ROS, significantly fewer ICSI‐derived embryos (p = 0.036) reached the morula‐blastocyst stage on day 4. High seminal ROS levels are associated with impaired sperm fertilizing ability and lower pregnancy rates after IVF. In ICSI, a negative association of ROS with embryo development to the blastocyst stage has been observed.
A fast and selective analytical method, used to determine the different lysophosphatidic acid (LPA) species in serum, has been developed and validated. LPA species were quantitatively extracted from ...serum using methanol–chloroform (2:1, v/v). The proteins were precipitated by this solvent mixture and separated by centrifugation in one step. LPA levels were determined in clear extracts using the HPLC-MS/MS method. The linearity of this method was established in the concentration range between 0.1 and 16
μM for all LPA species with a correlation coefficient greater than 0.99. Recovery of all LPA species determined by the serum, fortified at approximately 1
μM and 2–3
μM, was between 93% and 111% with an average R.S.D. of less than 8%. This method was used to determine LPA in numerous sera of healthy controls, patients with benign ovarian tumours and ovarian cancer at different stages. Significantly higher total LPA levels were determined in the sera of patients with different types of tumours (benign and malignant).
BACKGROUND: We investigated whether the psychological stress related to a short war (26 June–7 July, 1991) in Slovenia induced changes in fertility, sex ratio at birth and semen quality ...characteristics. METHODS: Sex ratios i.e. males/(males + females) for 4966 births in the general population of Slovenia and separately for 1565 births in the Slovenian capital, Ljubljana, from January–March 1992 were compared with the sex ratio calculated for the same time period in 1991 and 1993. Semen analyses for 38 normozoospermic men attending an outpatient infertility clinic from May to September 1991 were also evaluated. RESULTS: In the general population in Slovenia there was a significant fall in the sex ratio at birth in 1992, compared with 1991 (0.504 versus 0.518; P = 0.03). In Ljubljana, the decline in sex ratio in 1992 was even more pronounced: 0.483 versus 0.537 in 1991 (P = 0.0001) and 0.483 versus 0.516 in 1993 (P = 0.005). A decrease in the proportion of sperm that were progressively motile from 56% before the war to 52% after it (P = 0.01) and of those that were rapidly motile from 40 to 36% (P = 0.01) was observed. CONCLUSIONS: Acute psychological stress in relation to a short war in Slovenia resulted 6 to 9 months later in a decrease in the observed sex ratio at birth. Negative changes in sperm motility may be involved in the sex ratio modifications.
BACKGROUND: Fragile X premutation carriers are at increased risk of premature ovarian failure (POF), which is usually defined as menopause before the age of 40 years. METHODS: We evaluated 83 women ...with sporadic premature ovarian failure, treated at the Department of Obstetrics and Gynaecology, University Medical Centre, Ljubljana, between 1991 and 2001. There was no family history of mental retardation in any of the patients. They were phenotypically normal and had normal female karyotype (46,XX), without a past history of pelvic surgery, chemotherapy or autoimmune diseases. RESULTS: The premutation in the FRAXA locus was found in four of the women screened (4.8%; 95% confidence interval 1.9–11.7). This prevalence (1 in 21) was statistically significantly higher than expected in the female Caucasian population. CONCLUSION: In this study we have confirmed an important association between FRAXA premutation and the pathogenesis of POF. This result has practical implications for genetic counselling and fertility treatment.
The aim of this study was to evaluate the effect of sperm single-stranded DNA, detected by acridine orange (AO), and classical sperm parameters on embryonic quality after ICSI.
Before ICSI, the ...spermatozoa of 183 infertile patients with oligo-, astheno-, teratozoospermia (n = 147), or more than one previous unsuccessful conventional IVF attempt (n = 36) were stained by AO to assess the presence of single-stranded DNA. Two days after ICSI, the embryos of 135 patients were scored for morphology, fragmentation included. Embryos of 48 couples were cultured for 4 days to develop to the morula or blastocyst stage. At most 2 embryos were transferred on Day 2 or 4.
When the level of spermatozoa with single-stranded DNA was increased, there was a significantly lower fertilization rate after ICSI. Besides, increased sperm single-stranded DNA resulted in a higher proportion of heavily fragmented embryos on Day 2 (P < 0.05). In patients with an increased level of spermatozoa with single-stranded DNA, a significantly higher number of embryos were arrested in spite of prolonged culturing (P < 0.05). Classical sperm parameters did not affect the quality and developmental potential of ICSI-derived embryos. No correlation was found between the level of spermatozoa with single-stranded DNA, pregnancy rate, and live-birth rate achieved by ICSI, except in patients with 0% of spermatozoa with single-stranded DNA, in whom the pregnancy rate was significantly higher.
Sperm single-stranded DNA provides additional data on sperm functional capacity in terms of fertilization and embryonic quality after ICSI.
The objective of this prospective study was to evaluate the effect of ammonium accumulated in sequential media and determined by enzymatic spectrophotometric method on the blastocyst development in ...281 human embryos from 100 stimulated and natural in vitro fertilization (IVF) cycles. Ammonium concentration was increased in 62% of cycles and was correlated negatively with the blastocyst development after classical IVF, but not after intracytoplasmic sperm injection (ICSI).
Abstract Objective To evaluate the effect of counseling on women's adherence to hormone therapy (HT) in Slovenia following the publication of the results of the Women's Health Initiative prospective ...study. Method In this 24-month prospective, randomized, controlled study with 125 women in early menopause, those in the study group (n = 64) attended oral presentations on menopause and HT whereas those in the control group (n = 63) did not. Data were collected from 2 questionnaires, one completed before starting HT and the other during follow-up visits after 3, 6, 12, and 24 months. The results were analyzed according to the intention-to-treat principle. Results In the second year, an improved quality of life and friends critical of HT negatively affected adherence to HT. After 24 months, 47% of the participants in the study group and 32% in the control group were still using HT. The difference between the 2 groups in the rates of participants who discontinued HT was the same in the first than and in the last 12 months of the study (49 76.6% vs 15 23.4%). Conclusion Since there were no changes in discontinuation rates between the groups from one study period to the other, and a total of 15 women in each group stopped using HT in the second year of the study, the effect of the educational presentations on HT adherence was long-term rather than temporary.
In the present study, differences in leptin levels between different groups of male patients presenting with infertility problems and possible correlations between leptin levels and clinical, ...spermiological, histological and hormonal characteristics were examined. Two hundred and ten male partners from infertile couples were included in the study. Based on clinical examination, spermiogram and testicular histology results, patients were divided into four groups: 42 men with non-obstructive azoospermia, 15 men with obstructive azoospermia, 68 men with oligoasthenoteratozoospermia and 85 men with normozoospermia. Serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), inhibin B, testosterone, sex hormone binding globulin (SHBG) and leptin were measured. After adjustment for body mass index, there was a negative correlation between serum levels of leptin and inhibin B, total testosterone and SHBG (r = -0.189, p = 0.009, r = -0.250, p = 0.001 and r =-0.221, p = 0.003 respectively) but there was no correlation between leptin and classical sperm characteristics. Our results therefore demonstrate a link between leptin and testicular function, independently of FSH and LH, possibly involving testosterone and SHBG through a regulation of Leydig cell function.
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