Ten years of in vitro fertilization in Indonesia: Access to infertility care in a developing country.

OBJECTIVE: This research was conducted to assess access to assisted reproductive technologies (ART) and the current status of the in vitro fertilization (IVF) program that have been implemented in Indonesia over the last 10 years. METHODS: We established a retrospective cohort study and descriptive analysis of the current state of access to infertility care in Indonesia. The data were collected from all IVF centers, clinics, and hospitals in Indonesia from 2011 to 2020, including the number of IVF clinics, total ART cycles, retrieved fresh and frozen embryos, average age of IVF patients, IVF pregnancy rate, and causes of infertility. RESULTS: The number of reported fertility clinics in Indonesia has increased from 14 clinics in 2011 to 41 clinics by 2020. As many as 69 569 ART cycles were conducted over the past 10 years, of which 51 892 cycles used fresh embryos and 17 677 cycles used frozen embryos. The leading cause of consecutive infertility diagnosis was male infertility. Nearly half of the women who underwent IVF procedures (48.9%) were under 35 years old. The pregnancy rate outcome of women who underwent IVF ranged from 24.6% to 37.3%. CONCLUSION: Developments in ART in Indonesia have led to improvements in the ART cycles performed throughout the 10 year period. The identification of key areas that require improvement can provide an opportunity to enhance access to infertility care.

Down-Regulation of miR-93 Negatively Correlates with Overexpression of VEGFA and MMP3 in Endometriosis: A Cross-Sectional Study.

BACKGROUND: Endometriosis is identified as presence of the endometrium outside the uterine cavity. Retrograde menstruation contributes to the endometrial tissue implantation and the establishment of endometriotic lesions at ectopic sites. It has been suggested that the endometriotic lesions are rich in angiogenic growth factors, while they have an essential role in survival and invasion of these cells. We investigated regulation of microRNA-93 (miR-93) and its involvement with vascular endothelial growth factor A (VEGFA) and matrix metalloproteinase (MMP) 3 expression in women with endometriosis. MATERIALS AND METHODS: This was a cross-sectional study at Central Surgical Installation, Dr. Cipto Mangunkusumo General Hospital, Jakarta, Indonesia, between October 2020 and November 2021. Eutopic and ectopic endometrial tissues were collected from 30 subjects with laparoscopically-confirmed endometriotic women. Normal endometrial cells of non-endometriosis women served as controls. Total RNA was isolated from all samples and a quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was used to analyze the expression of miR-93, VEGFA and MMP. RESULTS: There was no significant difference in the expression levels of VEGFA (2.14 ± 0.50, P=0.719) and MMP3 (2.99 ± 0.42, P=0.583) between endometriotic lesions of endometriosis women and the healthy endometrium. Expression of miR-93 was significantly lower in the eutopic endometrium (16.7 fold) and ectopic endometriotic lesion (20 fold) compared to the normal endometrium (P<0.001). Furthermore, we also observed a significant correlation between miR-93, VEGFA expression in eutopic endometrium obtained from women with endometriosis (r=-0.544, P=0.029). Expression of the miR-93 was also negatively correlated with MMP3 expression in both eutopic (r=-0.412, P=0.01) and ectopic (r=-0.539, P=0.03) endometrial cells of women with endometriosis. CONCLUSION: VEGFA and MMP3 expression levels trended to be increased in both eutopic and ectopic endometrial tissues of endometriosis women, while down-regulation of miR-93 might be involved in the alteration of VEGFA and MMP3 in endometriosis.

Anti-Müllerian hormone independently affect mtDNA copy number in human granulosa cells.

BACKGROUND: Recently, as a delayed childbearing trend is emerging in modern women's adulthood, diminished reproductive potential due to age-related changes is more prevalent. Reduction in the abundance of mitochondrial DNA (mtDNA) copies and circulating anti-Müllerian hormone (AMH) have been separately reported with aging, contributing to the decrease in successful reproduction. However, there are limited reports on the impact of age on mtDNA and AMH in the same individual and whether mtDNA copy numbers are influenced by age and AMH. METHODS: In the present study, we utilized a real-time quantitative PCR (RT-qPCR) to quantify the mtDNA copy number of granulosa cells obtained from 43 women undergoing an in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) program. RESULTS: According to our analysis, a significant correlation was observed between age and mtDNA copy number (r = -0.54, P < 0.001) and between age and AMH level (r = -0.48, P < 0.001) of the same individual. There was also a positive correlation between mtDNA copy number and AMH (r = 0.88, P < 0.001) with AMH level falling as mtDNA decreases. In our regression, age and AMH were shown to have low collinearity (VIF = 1.297) but only AMH was correlated with mtDNA quantity (P < 0.001). CONCLUSION: Our study suggests that both mtDNA and AMH abundance are influenced by age and that AMH levels independently affect mtDNA copy number regardless of age. Further research is required to understand the role of AMH on mitochondria bioenergetics.

The Correlation of Chronological Age and Micro Ribonucleic Acid-135b Expression in Spent Culture Media of In vitro Fertilisation Patient.

Background: The lack of accuracy in embryo viability assessment methods still remains a challenge to increase the in vitro fertilisation (IVF) success rate. The chronological age and micro ribonucleic acid (RNA)-135b influence the quality of the embryo since microRNA-135b expresses stably in the spent culture media. Therefore, microRNA-135b has the potential to become a non-invasive biomarker of IVF embryo quality. Aims: (1) The aim of this study is to determine the chronological age and microRNA-135b expression distribution of IVF patients. (2) to determine the correlation between chronological age and microRNA-135b expression in spent culture media of IVF patients. Study Setting and Design: An observational study was conducted in Yasmin IVF clinic. Materials and Methods: The chronological age data were collected from the medical records and 31 spent culture media samples from 11 IVF patients were taken on day 5 of embryo culture. We also collected the basal media sample as the control group. The microRNA-135b expression was analysed using quantitative real-time polymerase chain reaction (qPCR) analysis. Statistical Analysis Used: The data analysis was performed using IBM SPSS Statistics 25. Results: The chronological age and microRNA-135b expression were distributed abnormally. There was a significant positive correlation with moderate statistical power between chronological age and microRNA-135b expression in spent culture media. MicroRNA-135b expression increased 4.9-fold in spent culture media than basal media of IVF. Conclusions: The increase of chronological age is followed by the rise of microRNA-135b expression in spent culture media of IVF patients. The microRNA-135b is a potential biomarker to predict IVF embryo quality.

Ovarian cortex freezing as a method of fertility preservation in endometriosis: A case report.

BACKGROUND: Endometriosis affects women in many ways from infertility until reducing ovarian reserve. In women who do not want to immediately conceive, ovarium cortex cryopreservation may be an option for preserving fertility. CASE PRESENTATION: Two patients with chief complaints dysmenorrhea and abdominal enlargement, then checked Anti-Mullerian Hormone (AMH) level and Ca-125 level. Patient underwent transrectal ultrasonography, with the result of endometriosis cyst (sized 12 × 9x3 cm and 7 × 10 × 11 cm for first patient, while second patient had 18 × 10 × 14 cm). Then patients underwent cystectomy and ovarian cryopreservation. Histopathology results revealed endometriosis cyst, with different results of follicle density on the healthy cortex. Patient have an AMH level of 1.82 ng/mL before surgery and may decline after surgery. From the AMH normogram, the patient is below the 25th percentile and almost below the 10th percentile, and her biological age is 34. Normal histopathology result of the ovarian cortex suggested that 1.8 to 166 follicles per mm3 cortical tissue. DISCUSSION: We can see from the histopathology examination the density of the follicle was less than normal in this patient. Patients that suffer from endometriosis may have a low ovarian reserve even before surgery. A thorough consultation, followed by ovarian reserve evaluation, disease progression and recurrence of disease are needed to be monitored closely. CONCLUSION: From all the methods of fertility preservation, we concluded that this patient is most suitable for ovarian cortex freezing.

The Frequency of Chromosomal Euploidy Among 3PN Embryos.

BACKGROUND: The evaluation of embryo morphology is one of the most important parameters used to evaluate developmental timing, also providing an indication of chromosomal failure or degeneration. The first step in the evaluation of a fertilization event is determining the number and shape of the pronuclei (PN). Normally fertilized eggs possess two even PN. However, some embryos which develop from abnormally fertilized zygotes may be tri-pronuclear zygotes (3PN). METHODS: Thirty embryos were collected from 12 women who underwent in vitro fertilization (IVF) at Dr. Cipto Mangunkusumo General Hospital in Jakarta, Indonesia. Embryos were cultured until the blastocyst stage on days 5-6. The blastomere biopsy was performed by piercing the zona pellucida with a laser under a microscope. Chromosomal numerical abnormalities were analyzed using Next Generation Sequencing (NGS). RESULTS: Among the 30 embryos with 3PN zygotes, 33.3% had a normal chromosomal array, with 22 pairs of autosomes and 2 pairs of sex chromosomes. While the rest of sample population detected as abnormal chromosome (66.7%), with the highest percentage of abnormality was triploidy 43.3%, followed by mosaicism 13.4% and aneuploidy 10%. CONCLUSION: This was a preliminary study revealed not all morphologically 3PN embryos are genetically abnormal.

Correlation between luteinizing hormone receptor gene expression in human granulosa cells with oocyte quality in poor responder patients undergoing in vitro fertilization: A cross-sectional study.

Background: This study was performed to evaluate the role of luteinizing hormone (LH) and granulosa cell LH receptor (LH-R) in poor responder patients who underwent controlled ovarian stimulation. Expression levels of LH-R mRNA in granulosa cells was investigated and compared with oocyte morphology, oocyte maturity and fertilization rate.  Methods: Granulosa cells were obtained from 30 patients who underwent in vitro fertilization (IVF) at Dr. Cipto Mangunkusumo Hospital, Jakarta. The patients were divided into two groups: group I (n=10) poor responders; and group II (n=20) non-poor responders. After the extraction of total RNA from granulosa cells, semi-quantitative RT-PCR was performed and the amount of LH-R mRNA was quantified. The relative values were calculated as the ratio of LH-R mRNA and actin beta mRNA. Statistical analysis was performed using Mann-Whitney test and Spearman correlation.  Results: The relative value of LH-R mRNA was higher in group I compared with group II (27.37[0.00-28939.37] vs 0.00[0.00-7196.12]). Oocyte maturity (r=0.267) and morphology (r=0.267) in group I consistently showed a positive correlation with LH-R mRNA; in group II a negative correlation with LH-R mRNA was shown for oocyte maturity (r= -0.552) and morphology (r= -0.164). Group I had a positive correlation between LH-R expression with fertilization rate (r=0.430), and group II showed a negative correlation (r=-0.340).  Conclusions: The expression of LH-R mRNA has a positive correlation with oocyte quality in poor responder patients and a negative correlation in non-poor responders. Our study suggests an optimal expression of LH- R mRNA in granulosa cells during controlled ovarian stimulation to obtain good quality oocytes.