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Abstract Background Assisted reproductive technology accounts for an increasing proportion of infertility treatments, and assessments to predict clinical pregnancy outcomes are desired. Extracellular vesicles exist in follicular fluid, and small non coding RNAs in extracellular vesicles underline the possibility of reflecting pregnancy potential. Methods Follicular fluid samples are collected from 20 ovarian follicles of 15 infertile patients undergoing assisted reproductive technology. Extracellular vesicles are isolated by serial centrifugation and small RNA sequencing is performed to investigate the profiles of microRNAs and P-element-induced wimpy testis-interacting RNAs. Results Small extracellular vesicles with a size range of approximately 100 nm are successfully isolated, and the small non coding RNA profiles of pregnant samples (n = 8) are different from those of non-pregnant samples (n = 12). Fourteen dysregulated small non coding RNAs are selected to identify the independent candidates [mean read count >100, area under the curve >0.8]. Among them, we find that a specific combination of small non coding RNAs (miR-16-2-3p, miR-378a-3p, and miR-483-5p) can predict the pregnant samples more precisely using a receiver operating characteristics curves analysis (area under the curve: 0.96). Furthermore, even in the same patients, the three microRNAs are differentially expressed between pregnant and non-pregnant samples. Conclusions Our results demonstrate that small non coding RNAs derived from small extracellular vesicles in follicular fluid can be potential non-invasive biomarkers for predicting pregnancy, leading to their probable application in assisted reproductive technology. Further large-scale studies are required to validate the clinical usefulness of these small non coding RNAs.

Objective: Having a single coronary artery (SCA) is a rare congenital anomaly in which a single artery arises from the aorta. Although most cases of SCA are asymptomatic and incidental, its effects during the perinatal period remain unknown. Herein, we report a case of pregnant woman with suspected SCA, based on transthoracic echocardiography (TTE) findings. Case report: A 33-year-old multiparous woman presented with preterm premature rupture of the membrane at 29 weeks gestation. The patient’s preoperative electrocardiogram (ECG) showed slight ST changes. TTE showed dilated right coronary artery and hypoplastic left coronary artery. Cesarean section was performed at 30 weeks of gestation due to non-reassuring fetal status. Although poor oxygenation was observed postoperatively, the patient was managed appropriately. She was diagnosed with SCA based on coronary computed tomographic angiography findings one month after delivery. Conclusion: Pregnant women with SCA require careful perinatal care.

Anti-Müllerian hormone (AMH), an indirect indicator of the number of remaining follicles, is clinically used as a test for ovarian reserve. Typically, a decline suggests a decrease in the number of remaining follicles in relation to ovarian toxicity caused by interventions, which may implicate fertility. In contrast, serum AMH levels are elevated in patients with polycystic ovary syndrome. AMH is produced primarily in the granulosa cells of the preantral and small antral follicles. Thus it varies in association with folliculogenesis and the establishment and shrinking of the follicle cohort. Ovarian activity during the female half-life, from the embryonic period to menopause, is based on folliculogenesis and maintenance of the follicle cohort, which is influenced by developmental processes, life events, and interventions. AMH trends over a woman’s lifetime are associated with in vivo follicular cohort transitions that cannot be observed directly.

Abstract Retained products of conception (RPOC) is a common cause of postpartum bleeding, which may be life-threatening; however, no evidence-based guidelines exist to assist in evaluating the risk of massive hemorrhage in women with RPOC. In this prospective study, we aimed to evaluate the predictive factors for massive hemorrhage in women with RPOC. The primary and secondary endpoints were to validate the usefulness of power Doppler color scoring (PDCS) in evaluating hypervascularity and to identify other predictive factors (such as maximum RPOC diameter and serum βhCG and Hb level at first visit), respectively. Among the 51 women with RPOC included in this study, 16 (31.5%) experienced massive hemorrhage during follow-up. None of the women with PDCS 1 or 2 (18) experienced massive hemorrhage, whereas 16 (48.5%) women with PDCS 3 or 4 (33) did. Multiple logistic regression analysis showed that the odds ratio [95% confidence interval] (P value) for PDCS, assisted reproductive technology (ART), and low serum hemoglobin (Hb) levels were 22.39 [2.25 − 3087.92] (P = 0.004), 5.72 [1.28 − 33.29] (P = 0.022), and 4.24 [0.97 − 22.99] (P = 0.056), respectively. Further, the decision tree method identified PDCS, ART, and low serum Hb levels as potential predictive factors for massive hemorrhage. This study identified PDCS as useful predictor of massive hemorrhage in women with RPOC. With additional inclusion of factors such as ART and low serum Hb levels, the risk of massive hemorrhage may be effectively evaluated, leading to better management of women of reproductive age.

Abstract Purpose This study aimed to investigate whether serum leucine‐rich α2‐glycoprotein (LRG) is a useful diagnostic biomarker for endometriosis, including the evaluation of treatment efficacy and exploration of LRG production in endometriotic lesions. Methods Forty‐three women with endometriomas were compared to 22 women with benign ovarian cysts and 30 women who underwent assisted reproduction as controls. Changes in serum LRG levels were assessed before and after surgery, and during dienogest treatment. LRG expression in endometriotic tissue samples was evaluated using immunoblotting. Results Serum LRG levels in the endometrioma group (80.0 ± 36.3 μg/mL) were significantly higher than those in the benign ovarian cyst (65.1 ± 27.0 μg/mL, p = 0.0265) and control (57.8 ± 22.3 μg/mL, p = 0.0028) groups. Serum LRG levels after endometrioma surgery were significantly lower than preoperative levels (p = 0.0484). Serum LRG levels consistently decreased during dienogest treatment. LRG expression levels were significantly higher in endometriotic tissues than in the normal endometrium. Conclusion Serum LRG, possibly derived from local and systemic origins, could be used as a potential biomarker for the diagnosis and treatment of endometriosis.

Abstract Background Primary ovarian insufficiency (POI) is characterized by the development of hypergonadotropic hypogonadism before 40 years of age and leads to intractable infertility. Although in vitro fertilization and embryo transfer with donated eggs enables pregnancy, not a few patients desire pregnancy using their oocytes. However, follicular development is rare and unpredictable in patients with POI. Thus, there is a need for treatments that promote the development of residual follicles and methods to accurately predict infrequent ovulation. Methods This review discusses the effects of various treatments for obtaining eggs from POI patients. Furthermore, this study focused a potential marker for predicting follicular growth in patients with POI. Main Findings Different treatments such as hormone‐replacement therapy, dehydroepiandrosterone supplementation, platelet‐rich plasma injection, and in vitro activation have shown varying degrees of effectiveness in retrieving oocytes from patients with POI. To predict follicle development in the cycle, elevated serum estradiol and reduced follicle‐stimulating hormone (FSH) levels are important. However, these markers are not always reliable under continuous estradiol‐replacement therapy. As a novel marker for predicting follicle growth, serum anti‐Müllerian hormone (AMH) levels, measured using the picoAMH enzyme‐linked immunosorbent assay, were found to predict follicle growth in patients and the cycle. Conclusion This review highlights the challenges and available interventions for achieving pregnancy using a patient's oocytes in cases of POI. We believe that a combination of currently available treatments and prediction methods is the best strategy to enable patients with POI to conceive using their own eggs. Although AMH levels may predict follicle growth, further research is necessary to improve the chances of successful follicular development and conception in patients with POI.

Abstract Background Ovarian endometrioma is a common gynecological disease that is often treated with surgery or hormonal treatment. Ovarian cystectomy, a surgical procedure for ovarian endometrioma, can result in impaired ovarian reserve. Methods We conducted a randomized controlled trial to evaluate the efficacy of hormonal treatment [gonadotropin-releasing hormone agonist (GnRHa) or dienogest (DNG)] for preserving ovarian reserve after cystectomy for ovarian endometrioma. The primary endpoint was the level of serum Anti-Müllerian hormone (AMH) as a marker of ovarian reserve. Results Before and after laparoscopic surgery, 22 patients in the GnRHa group and 27 patients in the DNG group were administered hormonal treatment for a total of 4 months. After 1-year follow-up, >60% of the patients in the DNG group retained over 70% of their pretreatment AMH levels, whereas no patient in the GnRHa group retained their AMH levels after cystectomy (P < 0.01). Interleukin-6 (IL-6) is a key cytokine involved in inflammation. Compared with the GnRHa group, patients in the DNG group had lower IL-6 levels at the end of treatment. Conclusions Our data revealed that DNG is more effective than GnRHa in preserving ovarian reserve after cystectomy of ovarian endometrioma. This is achieved through the reduction of the inflammatory response during the perioperative period and other endometriosis-related inflammatory reactions. Trial registration The registration number of this trial is UMIN-CTR, UMIN000018569, registered 6 August 2015, https://upload.umin.ac.jp/cgi-open-bin/ctr_e/ctr_view.cgi?recptno=R000021492 , and Japan Registry of Clinical Trials, jRCTs041180140, registered 29 March 2019, https://jrct.niph.go.jp/en-latest-detail/jRCTs041180140 . This randomized controlled trial was conducted in accordance with the CONSORT guidelines.

Endometriosis-associated ovarian malignancies have been well documented. Although these malignancies also occur as extraovarian lesions, little is known about them. Thus, this literature review aimed to further explore these rarely experienced tumors. A total of 257 published cases between April 1990 and April 2020 were found using PubMed, and 212 cases were included in the analysis considering Sampson’s criteria and the history of endometriosis. We classified these cases as follows: intestine, abdominal scar, vagina and vulva, peritoneum and deep endometriosis, urinary tract, uterine cervix, and others. Age of patients, history of endometriosis, types of past hormonal therapy, symptoms, histological types, and treatment were identified. The most common tumor site was the intestine. Endometrioid carcinoma was the dominant histological type. Contrary to the ovary, clear cell carcinoma was rare in extraovarian sites. On the other hand, clear cell carcinoma represented the largest number of abdominal scars. This difference may help us to understand the development of endometriosis-related malignancies. Hormonal treatment was mentioned in 67 cases and estrogen replacement therapy in 33 cases. Although risks of estrogen therapy are still controversial, the highly differentiated histological types and hormone-dependent characteristics of endometriosis-associated malignancy should be considered. Physicians should be careful about estrogen monotherapy after hysterectomy and long-term hormone replacement therapy in patients with a history of endometriosis.

Abstract Patients with primary ovarian insufficiency (POI) often have a high prevalence of autoimmune disorders. To identify antigenic molecules associated with ovarian autoimmunity, we performed immunoprecipitation (IP) screening using serum from patients with POI and the established human granulosa cell line (HGrC1). POTE ankyrin domain family member E (POTEE) and POTE ankyrin domain family member F (POTEF), proteins specific to primates, were identified as candidate antigens. Using immunohistochemistry (IHC) with human ovarian tissue, POTEE or POTEF was weakly seen in the granulosa cells (GCs) of primordial follicles and primary follicles, and strongly in large antral follicles and luteal cells. Interestingly, no signals were detected in growing GCs in secondary, preantral, and small antral follicles. Thus, to explore the function of POTEE and POTEF in human folliculogenesis, we established HGrC1 cell lines with drug-inducible expression of POTEF. Expression of POTEF significantly suppressed cell proliferation in HGrC1 cells. Furthermore, chaperonin containing TCP-1 complex (CCT) components, which affect folding proteins required for cell proliferation, was bound to the actin domain of POTEF protein. Although CCT is normally localized only around the Golgi apparatus, TCP-1α, a component of CCT, co-migrated closer to the cell membrane when POTEF expression was induced. These data suggest that the interaction between POTEF and CCT components impairs the usual function of CCT during cell growth. In addition, over-accumulation of POTEF in HGrC1 cells leads to autophagic failure. It was recently reported that knockout of an autophagic gene in mice leads to a phenotype similar to human POI. These results suggested that a proper amount of POTEF is required for the maintenance of GCs in follicle pools, whereas POTEF overaccumulation might be involved in follicle atresia and the development of POI. We also showed the possibility that POTEF could be an antigen involved in ovarian autoimmunity.

Objective: Hereditary sensory and autonomic neuropathies (HSANs) are a clinical heterogenous group of inherited neuropathies featuring prominent sensory and autonomic involvement. We report on the management of pregnancy and delivery in a woman with HSAN type V (HSAN-V) that is a rare inherited disease characterized by pain insensitivity, and partial anhidrosis. Case report: A 25-year-old woman with HSAN-V at six weeks of gestation was referred to our hospital. She decided to continue her pregnancy after the genetic counseling. A multidisciplinary team including her decided to undergo cesarean section due to her short stature and the risk of an emergency in normal delivery. She successfully gave birth at 38 weeks of gestation by cesarean section under general anesthesia following an uneventful pregnancy course. Conclusion: Cesarean section seems favorable to vaginal delivery in women with HSANs.

Key Clinical Message Our case and the literature review suggest that placenta accreta spectrum, with use of uterotonics and manual removal of placenta, could be risk factors for postpartum unscarred uterine rupture.

Abstract Background Evidence suggests that hypothyroidism and thyroid autoimmunity (TAI) are possibly associated with ovarian dysfunction. This meta‐analysis aimed to investigate whether hypothyroidism and/or TAI affect the ovarian reserve and evaluated using the anti‐Mullerian hormone (AMH). Methods PubMed, EMBASE, Web of Science, and Cochrane Controlled Trials Register databases from inception to October 2020 were searched to identify relevant studies. Studies comparing the AMH levels between the control and the affected groups were included in the data synthesis. The primary endpoint in the meta‐analysis was AMH levels compared with the controls. Main findings Nine trials were included in the analysis. The AMH levels were significantly lower in the adults with euthyroid TAI (mean difference −0.12, [95% CI: −0.18 to −0.06]). The AMH levels tended to be lower in subclinical hypothyroidism and overt hypothyroidism than in the control group, although the differences were not significant. The AMH levels were significantly higher in the euthyroid TAI group in the adolescents (mean difference 2.51, [95% CI 1.82 to 3.21]). Conclusion TAI and hypothyroidism may affect the ovarian reserve. The opposite effects on AMH levels depending on age suggest that TAI may be implicated in the depletion of follicles in adults following extensive activation of primordial follicles in adolescence.

Endometriosis is one of the most common diseases in women of reproductive age, and although there are many theories to explain this enigmatic disease, such as reflux theory, metastasis theory, and metaplasia theory, there is still no single theory that can wholly explain the pathogenesis of the disease, and it is considered a mysterious disease until now [...]

Abstract Background Polycystic ovary syndrome (PCOS) is a common endocrine disorder among women of reproductive age and a major cause of infertility; however, the pathophysiology of this syndrome is not fully understood. This can be addressed using appropriate animal models of PCOS. In this review, we describe rodent models of hormone‐induced PCOS that focus on the perturbation of the hypothalamic‐pituitary‐ovary (HPO) axis and abnormalities in neuropeptide levels. Methods Comparison of rodent models of hormone‐induced PCOS. Main findings The main method used to generate rodent models of PCOS was subcutaneous injection or implantation of androgens, estrogens, antiprogestin, or aromatase inhibitor. Androgens were administered to animals pre‐ or postnatally. Alterations in the levels of kisspeptin and related molecules have been reported in these models. Conclusion The most appropriate model for the research objective and hypothesis should be established. Dysregulation of the HPO axis followed by elevated serum luteinizing hormone levels, hyperandrogenism, and metabolic disturbance contribute to the complex etiology of PCOS. These phenotypes of the human disease are recapitulated in hormone‐induced PCOS models. Thus, evidence from animal models can help to clarify the pathophysiology of PCOS.

Ovarian endometriosis (OE) provides women of reproductive age with not only severe menstrual pain but also infertility and an increased risk for ovarian carcinogenesis. Whereas peritoneal endometriosis models have been developed with syngeneic implantation of minced uterine tissue and oncogenic K-ras allele with conditional Pten deletion within ovarian surface epithelium generated preneoplastic endometrial glandular morphology, followed by endometrioid adenocarcinoma, there has been no mouse model of OE similar to human counterparts, applicable to preclinical studies. Here we for the first time established a murine OE model that reveals infertility, and evaluated the involvement of iron catalyzed oxidative stress in the pathogenesis. Minced uterine tissue from female mice was implanted on ovarian surface of syngeneic mice after bursectomy to induce OE. Ectopic growth of endometrium was observed in association with ovary 4 weeks after implantation in 85.7% (12/14) of the operated mice with our protocol. Endometriotic lesions involved intestine, pancreas and peritoneal wall. Fibrosis around the ovary was prominent and increased time-dependently in the OE group. Iron accumulation was significantly increased in the OE group, leading to oxidative stress in each stage of the follicles as evaluated by 4-hydroxy-2-nonenal-modified proteins and 8-hydroxy-2′-deoxyguanosine. Expression of follicle stimulating hormone receptor in the follicles revealed a significant decrease during pre-antral, antral and pre-ovulatory phases in the OE group. Finally, the number of pups was significantly reduced in the OE group in comparison to the controls. This model affords an opportunity to evaluate agents or procedures to counteract ovarian endometriosis in the preclinical settings.

Most plants are exquisitely sensitive to their environment and adapt by reprogramming post-embryonic development. The systematic understanding of molecular mechanisms regulating developmental reprogramming has been underexplored because abiotic and biotic stimuli that lead to reprogramming of post-embryonic development vary and the outcomes are highly species-specific. In this review, we discuss the diversity and similarities of developmental reprogramming processes by summarizing recent key findings in reprogrammed development: plant regeneration, nodule organogenesis in symbiosis, and haustorial formation in parasitism. We highlight the potentially shared molecular mechanisms across the different developmental programs, especially a core network module mediated by the AUXIN RESPONSIVE FACTOR (ARF) and the LATERAL ORGAN BOUNDARIES DOMAIN (LBD) family of transcription factors. This allows us to propose a new holistic concept that will provide insights into the nature of plant development, catalyzing the fusion of subdisciplines in plant developmental biology.

Abstract Background Given the seriousness of chemotherapy-induced ovarian injury in female cancer patients, the preservation of fertility, including through the use of cryopreservation technology and pharmaceuticals, requires investigation. Previous studies have shown that damage to the ovaries is related to oxidative stress caused by anticancer drugs. Therefore, superoxide dismutase (SOD) may represent a key factor in the pharmacological protection of the ovaries. The aim of our study was to identify the effects of mangafodipir, a manganese chelate and SOD-mimetic, on suppression of apoptosis in granulosa cells and primordial follicle activation induced by anticancer drugs. Methods Cell viability assays using methyltrichlorosilane solutions and immunoblotting for cleaved caspase-3 were performed in in vitro experiments with the simultaneous addition of mangafodipir to human non-luteinized granulosa cell line (HGrC) cultures treated with hydrogen peroxide (H2O2), cisplatin, or paclitaxel. Count and morphological analyses of follicles at each developing stage in the ovaries and immunohistochemistry for cleaved caspase-3, Ki67 and 4-hydroxynonenal, a marker for oxidative stress, were also performed using mangafodipir-injected 6-week-old female ICR mice treated with cisplatin or paclitaxel. Further, mangafodipir was injected into 6-week-old female BALB/c mice inoculated with ES-2 to analyze whether mangafodipir inhibits the anti-tumor effects of cisplatin or paclitaxel treatment. Results Mangafodipir attenuated apoptosis induced by H2O2 and anticancer drugs in vitro. Mangafodipir also decreased the expression of 4-hydroxynonenal and reduced cisplatin- and paclitaxel-induced apoptosis in granulosa cells in vivo. In addition, mangafodipir inhibited the loss of primordial follicles. Tumor xenograft studies in mice showed that mangafodipir did not affect anticancer drug antitumor effects. Conclusions Oxidative stress might be one of the mechanisms of cisplatin- and paclitaxel-induced the loss of primordial follicles. Mangafodipir can reduce cisplatin- and paclitaxel-induced apoptosis in granulosa cells and primordial follicle activation partially via its SOD activity. At the same time, mangafodipir might have other potential mechanisms to inhibit the activation of primordial follicles. Further, mangafodipir attenuated the ovarian damage caused by cisplatin and paclitaxel without affecting their antitumor activities. Mangafodipir, therefore, though its efficacy might be limited, may be a new option for the preservation of fertility during anticancer treatment.

The elevation of the serum squamous cell carcinoma (SCC) antigen unrelated to disease progression occurs during the follow-up of patients with cervical cancer treated with radiotherapy. Although known empirically, the incidence and characteristics of this non-cancer specific elevation in SCC remain unclear. Here, we examined the post-treatment kinetics of SCC in 143 consecutive patients with squamous cell carcinoma of the cervix treated with definitive radiotherapy; in all patients, progression-free disease status was confirmed by periodic monitoring for at least 36 months (median, 61 months). We found that the 5-year cumulative incidence of post-treatment SCC elevation was unexpectedly high at 37.3% (59/143 patients), and that 59.3% (35/59) of event-positive patients experienced multiple events. The median peak SCC level for a given event was 2.0 ng/mL (interquartile range, 1.7–2.9 ng/mL). The multivariate analysis showed that renal dysfunction was associated significantly with a greater incidence of SCC elevation (p = 0.046). In addition, the 5-year cumulative incidence of SCC elevation was significantly greater in patients with renal dysfunction than in those without (54.8% vs. 32.9%, respectively; hazard ratio, 2.1 [95% confidence interval, 1.1–4.2]; p = 0.028). These data will be useful for monitoring cervical cancer patients treated with radiotherapy.

Abstract Background Serum anti-Müllerian hormone (AMH) concentration has been used to assess ovarian reserve in patients with endometriosis, especially when endometrioma surgery is involved. Previously, we reported that decreased serum AMH levels after cystectomy for endometriomas can recover to preoperative levels in some cases. In this present study, we assessed the sequential changes in serum AMH levels before and after cystectomy in terms of the state of the mesosalpinx prior to surgery. Methods The retrospective cohort study recruited 53 patients from a series of prospective studies conducted from 2009 to 2015. All patients underwent laparoscopic cystectomy for endometriomas. If either mesosalpinx was involved in the endometrioma or adnexal adhesion before cystectomy, the case was defined as ‘involved mesosalpinx’ (n = 14). If both mesosalpinx remained anatomically correct, the case was classified as ‘intact mesosalpinx’ (n = 39). Blood samples were obtained from the patients 2 weeks before surgery, and at 1 month and 1 year after surgery to assess serum AMH levels. Results The serum AMH levels (the involved group vs. the intact group) were 1.92 vs. 0.98 (P = 0.552) preoperatively, 0.59 vs. 1.99 (P = 0.049) at 1 month postoperatively, and 0.48 vs. 2.37 ng/mL (P = 0.007) at 1 year postoperatively. The involved mesosalpinx group showed a further decrease in serum AMH levels at 1 year postoperatively, while serum AMH levels in the intact mesosalpinx group tended to recover. Conclusion These results suggest that pre-existing mesosalpinx disturbance, in combination with adhesiolysis, may be involved in the medium- and long-term decrease in ovarian reserve after endometrioma surgery. A disturbance in ovarian blood supply via the mesosalpinx may underlie this. Trial registration UMIN-CTR UMIN000019369 . Retrospectively registered October 15, 2015.

Objective: To evaluate a protocol for selection of placental polyp management, including expectant management and hysteroscopic resection with or without transarterial embolization (TAE), using power Doppler color score (PDCS) as the vascularity parameter. Materials and Methods: This retrospective case–control study included 25 patients who were diagnosed with placental polyps. We evaluated the vascularity of placental polyps with PDCS measured by transvaginal ultrasonography as follows: PDCS 1, no blood flow; PDCS 2, minimal flow; PDCS 3, moderate flow; and PDCS 4, marked blood flow. We then selected expectant management or hysteroscopic resection with or without TAE. Results: Three of 17 patients with PDCS 1 or 2 underwent surgical intervention, and expectant management was successful in 14. Seven of eight patients with PDCS 3 or 4 underwent surgical intervention, while expectant management was successful in only one patient. Conclusion: PDCS is a simple examination for evaluating the vascularity of placental polyps. PDCS might be useful for selecting the optimal treatment for placental polyps, such as expectant management or surgical intervention, according to their vascularity.

Perivascular epithelioid cell tumors (PEComa) are mesenchymal tumors composed of histologically and immunohistochemically distinctive perivascular epithelial cells. Although the uterine corpus seems to be one of the most prevalent sites of involvement, PEComa of the uterine cervix are very rare. Only four cervical PEComa cases have been described, and were treated with hysterectomy and radiotherapy. We report a case of a 24-year-old nulligravida woman who presented with acute abdominal pain and was diagnosed with a rupture of an ovarian chocolate cyst. Subsequent surgery revealed that the tumor arose in the uterus, and the histological diagnosis was uterine PEComa with low potential malignancy. Recurrent PEComa in the uterine cervix were excised twice, and she remains disease free 12 months after the last operation. To the best of our knowledge, this is the first report of recurrent cervical PEComa with fertility-preserving surgery. Estimating the malignant potential and appropriate surgery are essential for young patients with uterine PEComa.

In the final step of cytokinin biosynthesis, the main pathway is the elimination of a ribose-phosphate moiety from the cytokinin nucleotide precursor by phosphoribohydrolase, an enzyme encoded by a gene named LONELY GUY (LOG). This reaction accounts for most of the cytokinin supply needed for regulating plant growth and development. In contrast, the LOG-independent pathway, in which dephosphorylation and deribosylation sequentially occur, is also thought to play a role in cytokinin biosynthesis, but the gene entity and physiological contribution have been elusive. In this study, we profiled the phytohormone content of chromosome segment substitution lines of Oryza sativa and searched for genes affecting the endogenous levels of cytokinin ribosides by quantitative trait loci analysis. Our approach identified a gene encoding an enzyme that catalyzes the deribosylation of cytokinin nucleoside precursors and other purine nucleosides. The cytokinin/purine riboside nucleosidase 1 (CPN1) we identified is a cell wall–localized protein. Loss-of-function mutations (cpn1) were created by inserting a Tos17-retrotransposon that altered the cytokinin composition in seedling shoots and leaf apoplastic fluid. The cpn1 mutation also abolished cytokinin riboside nucleosidase activity in leaf extracts and attenuated the trans-zeatin riboside-responsive expression of cytokinin marker genes. Grain yield of the mutants declined due to altered panicle morphology under field-grown conditions. These results suggest that the cell wall–localized LOG-independent cytokinin activating pathway catalyzed by CPN1 plays a role in cytokinin control of rice growth. Our finding broadens our spatial perspective of the cytokinin metabolic system. [ABSTRACT FROM AUTHOR]