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Introduction: While the provision of unapproved regenerative medicine has been problematic worldwide, few studies have examined the implementation status of regenerative medicine (RM) in the specific field. This study aimed to determine the current status of therapy and clinical research in the obstetrics and gynecology (OBGYN) in Japan under the Act on the Safety of Regenerative Medicine (RM Act). Methods: Detailed data were extracted from publicly available websites provided by the Ministry of Health, Labour, and Welfare. We extracted descriptive details, including risk classification of the RM Act, modality, target disease, locality, institution, and administration route. For therapy, the price for each modality was evaluated. Results: The total number of therapeutic provision plans in OBGYN (1.9% of RM in Japan) are classified as Class II (moderate) risk. Most were administered in clinics in urban areas for treating endometrial or ovarian infertility by locally administering platelet-rich plasma (PRP) or autologous mesenchymal stem cells (MSCs). The price using MSCs is approximately eight times more expensive that of those involving PRP (1832.1 ± 1139.8 vs 240.8 ± 106.5 thousand yen, p < 0.0001). Regarding research, four plans (2.2%) were submitted to target implantation failure and advanced gynecological cancer using autologous lymphocytes, dendritic cells, or MSCs. Conclusion: The RM Act permits knowledge of the current status of regenerative medicine even for unapproved uses in a specific clinical field. The study findings shall prompt a worldwide discussion regarding the required regulations for therapy and clinical research of RM.

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OBJECTIVES: This study aimed to evaluate the dentin bond performance of various resin luting cement (RLC) systems combined with universal adhesives in different luting strategies. MATERIALS AND METHODS: Three self-adhesive resin luting cements (SRLCs) were used with universal adhesives as primers. Twelve specimens per group were prepared to measure shear bond strength (SBS) under distinct luting strategies in etch-&-rinse and self-etch modes. Regarding luting strategies, the bonded specimens were categorized into four groups based on tooth primer application and the curing mode of the SRLC paste: (i) with light irradiation of the tooth primer (wL) + dual-cure mode (DC) of the SRLC paste, (ii) wL + self-cure mode (SC) of the SRLC paste, (iii) without light irradiation of the tooth primer (woL) + DC mode of the SRLC paste, and (iv) woL + SC mode of the SRLC paste. Specimens were also subjected to different storage conditions: 24 h in water (baseline condition) and 10,000 cycles of thermal cycling. RESULTS: Luting strategy, storage condition, and SRLC system type significantly influenced dentin SBS values in both etching modes. Notably, certain SRLCs exhibited significantly higher dentin SBS when the primer was light-irradiated compared with no primer irradiation. CONCLUSION: Most SRLCs demonstrated higher dentin bond strength with light-irradiated primers, suggesting potential enhancement of dentin bond performance via primer light irradiation.

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This study aimed to observe and compare the resin luting cement (RLC)/dentin interfaces of different types of RLC systems with scanning electron microscopy (SEM). This study evaluated six self-adhesive resin luting cement systems (SARLCs), four systems combining a SARLC with a tooth primer (SARLCPs), and six conventional resin luting cement systems (CVRLCs). Cured resin composite rods were bonded to bovine dentin using RLCs in different etching modes. The morphological features at the interfaces were observed using SEM. Although all RLCs in etch-&-rinse (ER) mode showed a clear hybrid layer, most SARLCs had a thinner and more porous hybrid layer than the SARLCPs and CVRLCs. All SARLCPs with primer and CVRLCs showed a thin high-density layer below the primer layer in both etching modes. CVRLCs and SARLCPs with primer treatment systems may be more versatile and reliable when compared to simplified RLC systems.

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There are many previous reports on the effects of ethanol on physiological function, including reports of elevated blood estrogen levels in women who drank alcohol. However, the mechanism of ethanol's effects on ovarian functions, such as follicle development and hormone secretion, has not been fully clarified. Therefore, in this study, we investigated the impacts of ethanol on these phenomena and their mechanisms using a primary culture system of rat ovarian granulosa cells (GCs). In the present experiment, groups were created in which follicle-stimulating hormone (FSH) or ethanol was added alone or FSH and ethanol were co-added, and mRNA and protein expression in each group was measured for luteinizing hormone receptor (LHR) and sex steroid hormone synthase, as well as for estradiol (E2) production, cAMP production, and FSH receptor (FSHR) internalization rate. The addition of FSH induced mRNA expression of LHR and aromatase, which led to membrane LHR expression and E2 production. The coexistence of ethanol enhanced all these responses. The action of FSH is exerted via cAMP, and the co-addition of ethanol enhanced this cAMP production. Ethanol alone did not induce cAMP production. The enhancing effect of ethanol was also observed for cAMP induced by cholera toxin. Ethanol had no significant effect on the internalization rate of FSHR. In conclusion, ethanol increased FSH-stimulated cAMP production by increasing the activity of adenylyl cyclase, which enhanced FSH actions in rat GCs. Alcohol is an exacerbating factor in several female hormone-related diseases, and the mechanism of ethanol-induced increase in estrogen secretion revealed in this study may be involved in the pathogenesis of these diseases.

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Purpose: To create and evaluate a machine-learning model for YOLOv3 that can simultaneously perform morphological evaluation and tracking in a short time, which can be adapted to video data under an inverted microscope. Methods: Japanese patients who underwent intracytoplasmic sperm injection at the Jikei University School of Medicine and Keiai Reproductive and Endosurgical Clinic from January 2019 to March 2020 were included. An AI model that simultaneously performs morphological assessment and tracking was created and its performance was evaluated. Results: For morphological assessment, the sensitivity and positive predictive value (PPV) of this model for abnormal sperm were 0.881 and 0.853, respectively. The sensitivity and PPV for normal sperm were 0.794 and 0.689, respectively. For tracking performance, among the 51 objects, 40 (78.4%) were mostly tracked, 11 (21.6%) were partially tracked, and 0 (0%) were mostly lost. Conclusions: This study showed that evaluating sperm morphology while tracking in a single model is possible by training YOLO v3. This model could acquire time-series data of one sperm, which will assist in acquiring and annotating sperm image data.

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We present a case of spontaneous pregnancy after laparoscopic surgery for a broad ligament pregnancy. A 34-year-old nulliparous woman presented with 6 weeks of amenorrhea. Due to the presence of an empty uterus with a 10 mm right adnexal mass on transvaginal ultrasonography and elevated serum human chorionic gonadotropin (hCG), ectopic pregnancy was suspected. Upon diagnostic laparoscopy, the presence of a 2 cm broad ligament ectopic pregnancy was confirmed. Laparoscopic removal of the gestational tissues was performed. Six months after surgery, a spontaneous pregnancy was established. At the 40th week of gestation, a cesarean section was performed due to arrested labor, resulting in live birth. To the best of our knowledge, there have been no reports of a spontaneous pregnancy occurring after laparoscopic surgery for broad ligament pregnancy. Laparoscopic surgery as a treatment option for broad ligament pregnancy may be useful in early gestational age because it can be completed without complications.

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Heterotopic pregnancy (HP) is a rare but life-threatening disease. We report a rare case of HP that occurred after single embryo transfer (SET) with spontaneous natural pregnancy possibly due to sexual intercourse (SI) during assisted reproductive technology treatment and reviewed previous reports. A 39-year-old woman at 7 weeks 5 days' gestation with anti-sperm antibody who underwent a single frozen-thawed embryo transfer in her natural cycle presented with lower abdominal pain and vaginal bleeding. She had several SIs before the day of SET. A viable intrauterine fetus and an extrauterine mass at the right adnexa were detected on transvaginal ultrasonography. An emergent laparoscopic surgery showed a swollen right fallopian tube, and right salpingectomy was performed. Unfortunately, intrauterine fetal death was diagnosed at 19 weeks' gestation. In conclusion, the possibility of HP should be considered in patients with SIs around the day of SET.

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Ovarian pregnancy is a rare disease, accounting for 0.5%-3% of ectopic pregnancies. Ovarian pregnancy risk factors and preoperative diagnosis have been extensively reported. However, its histopathology and surgical findings have been poorly studied. To examine appropriate surgical procedures, we investigated the clinical features, surgical findings, and histopathological examinations of four ovarian pregnancy cases treated in our hospital. In histopathological examination, most specimens did not contain ovarian tissues; in some cases, villous tissues were buried in a clot. Therefore, evaluating the appropriateness of surgical resection range from histopathological images was difficult. However, the postoperative course was favorable; no cases manifested complications. Considering all these facts, we regarded the surgical procedures of the four cases in this study as appropriate. For the treatment of ovarian pregnancies, especially for the outward development type, a sufficient therapeutic effect may be achieved even without extensive excision of the ovarian tissues by laparoscopic surgery.

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PURPOSE: To provide information about the relationship between follow-up period and follicular development in patients with infertility due to premature ovarian insufficiency (POI) who are undergoing hormone replacement therapy (HRT). It is necessary to detect follicle development for artificial insemination or in vitro fertilization. METHODS: This retrospective cohort study was conducted at a university hospital in Tokyo, Japan, from April 2014 to February 2019 in 20 patients [follicular development group, 11 women (55%); non-follicular development group, 9 women (45%)] with POI; their follicular development was followed up weekly. Background characteristics, including age, follicle-stimulating hormone (FSH) and anti-Mullerian hormone levels (AMH), the period from the last spontaneous menstruation to hormone replacement therapy initiation, and follow-up period during HRT were investigated. The period without follicular development was tabulated, and the subsequent cumulative follicular development detection rate was calculated. RESULTS: At least 1-year follow-up, the cumulative follicular development rate was 70%; follicular development was observed with a probability of 49.1% at 3 months, 33.4% at 6 months, and 8.3% at 12 months in the follow-up period. CONCLUSIONS: The results show that the longer the non-follicle development period, the lower the probability of subsequent follicular development in patients with POI during HRT.

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OBJECTIVES: The aim of this study was to investigate dentin bonding durability of different etch-and-rinse (ER) adhesive systems under fatigue stress and to compare morphological features of resin/dentin interfaces using SEM. METHODS: Two three-step ER adhesives, a two-step ER adhesive, and a universal adhesive in ER mode were evaluated. Before application of either primer or adhesive, phosphoric acid etching of human dentin was completed. Fifteen bonded specimens for each adhesive system were stored in distilled water at 37 °C for 24 h, then subjected to a shear bond strength (SBS) test. Bonding durability was assessed from the perspective of biomechanical stress. 25 bonded specimens for each adhesive system were subjected to shear fatigue strength (SFS) testing with a repeated subcritical load at a frequency of 20 Hz for 50,000 cycles or until failure. RESULTS: Mean SBS and SFS values ranged from 33.3 to 41.2 MPa, and from 18.3 to 20.3 MPa, respectively. Three-step adhesives showed higher SBS and SFS values than the other adhesive systems. Under SEM, resin tags in different adhesive systems showed similar features, but morphology below the hybrid layer was material dependent. The universal adhesive in ER mode showed an obvious thin, high-density reaction layer below the hybrid layer. SIGNIFICANCE: Three-step adhesives showed higher dentin bond durability than the other ER adhesives; no significant differences in SFS were found between the universal adhesive in ER mode and the three-step ER adhesives. The results of this in vitro study indicate that some ER adhesives might establish chemical bonding with intact dentin below the hybrid layer in addition to micromechanical retention.

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OBJECTIVES: The present study aimed to investigate the changes in the enamel bond strengths of universal adhesives in the early phase of specimen creation and evaluate the effect of double-layer application on enamel bond effectiveness using different etching modes. METHOD: Four universal adhesives, namely Clearfil Universal Bond Quick, G-Premio Bond, Scotchbond Universal, and Tokuyama Universal Bond, were used. Clearfil SE Bond two-step self-etch adhesive system was used as a comparison. Fifteen specimens per group were used for determining the shear bond strength (SBS) to bovine enamel in the etch-and-rinse or self-etch mode. The adhesive was applied to specimens in accordance with the manufacturers' instructions (single-layer application), or the adhesive was applied twice (double-layer application). The bonded specimens were stored in distilled water at 37 °C for 5 min or 1, 6, 12, or 24 h before SBS testing. The flexural properties of the resin composite Clearfil AP-X were measured for the same storage periods. RESULTS: All universal adhesives exhibited increased SBS values with prolonged storage periods regardless of the application technique or etching mode used. Double-layer application was associated with higher SBS values than single-layer application for most universal adhesives over the same storage period. CONCLUSION: Phosphoric acid pre-etching and double-layer application of universal adhesives resulted in increased enamel bond strength in the early phase of specimen bonding. CLINICAL RELEVANCE: Care should be taken when using universal adhesives in the self-etch mode for enamel bonding because the enamel adhesion is sensitive to the duration after filling.

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The aim of this study was to determine the influence of different surface treatments of CAD/CAM composite blocks on bonding effectiveness of resin cements, based on the shear bond strength test and the surface science examination. Specimens were fabricated from two types of CAD/CAM composite blocks (Cerasmart and VITA Enamic), and the specimen surfaces were alumina sandblasted. Specimen surfaces were treated with i) silane, ii) primer, and iii) silane+primer, then two types of cements (Block HC Cem and Super Bond) were bonded. After 24 h storage, bond strengths were measured. Surface free energy measurements were performed of the treated CAD/CAM composite blocks surfaces. The groups ii) and iii) showed significantly higher bond strengths than the group i), apart from VITA Enamic with Block HC Cem. Although the total surface free energy showed different trends in different surface treatments, a similar trend was found in both CAD/CAM composite blocks.

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PURPOSE: This study compared Gavi®, an automated system for the equilibration and dehydration steps of vitrification, and a manual vitrification procedure in terms of effects on clinical outcomes. METHODS: The authors retrospectively compared survival rate, and clinical and perinatal outcomes after vitrified-thawed single blastocyst transfer between Gavi® (G method) in 398 cases and Cryotop® (C method) in 208 cases. RESULTS: With C and G methods, survival rates were 98.6% (208/211) and 99.3% (398/401), total pregnancy rates were 34.3% (72/208) and 33.4% (133/398), and total miscarriage rates were 22.2% (16/72) and 24.8% (33/133), respectively. Among women <35 years old, pregnancy rates were 41.1% (30/73) and 40.5% (62/153) and miscarriage rates were 13.3% (4/30) and 16.1% (10/62) with C and G methods, respectively. Among women ≥35 years old, pregnancy rates were 31.1% (42/135) and 29.0% (71/245) and miscarriage rates were 28.6% (12/42) and 32.4% (23/71) with C and G methods, respectively. C and G methods showed no significant differences in any trials, including gestational age, cesarean section rate, or birthweight (P > .05 each). CONCLUSIONS: Gavi® showed comparable clinical outcomes to the manual vitrification method and can be considered an alternative vitrification procedure in assisted reproductive technology.

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In order for facilitating the synthesis of oligosaccharides, transglycosylation reactions mediated by glycoside hydrolases have been studied in various contexts. In this study, we examined the transglycosylating activity of a Golgi endo-α-mannosidase. We prepared various glycosyl donors and acceptors, and recombinant human Golgi endo-α-mannosidase and its various mutants were expressed. The enzyme was able to mediate transglycosylation from α-glycosyl-fluorides. Systematic screening of various point mutants revealed that the E407D mutant had excellent transglycosylation activity and extremely low hydrolytic activity. Substrate specificity analysis revealed that minimum motif required for glycosyl acceptor is Manα1- 2Man. The synthetic utility of the enzyme was demonstrated by generation of a high-mannose-type undecasaccharide (Glc1 Man9 GlcNAc2 ).

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Endo-α-mannosidase, a GH99-family glycoside hydrolase, cleaves α-mannoside linkages with glucose residues. This enzyme is proposed to play a critical role in N-glycan processing for deglucosylation. To measure endo-α-mannosidase activity, we synthesized a fluorescently labeled tetrasaccharide derivative (Glcα1-3Manα1-2Manα1-2Manα1-O-C3H6-NH-Dansyl) in a stereocontrolled manner. The tetrasaccharide skeleton was prepared by step-wise coupling using mannose donors 4 and 7. The 1,2-cis α-glycosidic linkage on the non-reducing end of the glucose residue was constructed by inversion of the stereochemistry of the C-2 hydroxyl group in the α-mannose residue. Finally, the dansyl group was introduced at the reducing end via an aminopropyl linker. This probe successfully measured endo-α-mannosidase activity.

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