Exploring the Impact of Endometrioma Aspiration and Dienogest Combination Therapy on Cyst Size, Inflammatory Cytokines in Follicular Fluid and Fertility Outcomes.

Endometriomas (chocolate cysts) are cystic lesions that can develop on ovaries, and are characterized by the presence of ectopic endometrial tissue or similar tissue. Such lesions can cause a decline in the number and quality of oocytes, and lead to implantation failure. In this study, we retrospectively assessed the efficacy of repeated endometrioma aspiration and dienogest combination therapy in patients suffering endometriosis-associated infertility with endometriomas. A comparison was made between a treated group that underwent combination therapy followed by controlled ovarian hyperstimulation (COH) (n = 30) and a control group that did not undergo treatment (n = 40), at the IVF Osaka Clinic from September 2019 to September 2021. There were no differences in patient background between the two groups. A reduction in endometrioma size continued for 12 months after treatment. The numbers of follicles that developed to 15 mm or greater in size following COH and mature oocytes were significantly lower in the treated group compared to those in the control group. The levels of inflammatory cytokines in the follicular fluid significantly decreased in the treated group (p < 0.05). In patients in the treated group who underwent a second ova retrieval, the results were compared between those in the first ova retrieval (immediately after the end of treatment) and those in the second ova retrieval (four months after the first retrieval). The numbers of follicles following COH, retrieved, mature and fertilized ova were significantly increased in the second ova retrieval.

Relation between semen oxidative reduction potential in initial semen examination and IVF outcomes.

Purpose: The MiOXSYS system is a new technique to analyze the semen oxidative reduction potential (ORP) that may use to classify the level of sperm DNA integrity. It does not clearly explain how the semen ORP values could help to change the IVF outcomes. We have analyzed correlations between semen ORP value and the IVF results. Methods: Four hundred and thirty couples were enrolled. The male counterparts were divided into two groups according to their semen ORP values and compared the fertilization rate, cell cleavage rate, and embryo quality, following the IVF procedures. The relations between ORP values and the clinical pregnancy, live birth, and abortion rates were analyzed. Results: The ORP values show negative and positive correlations with some conventional semen parameters. The fertilization and the cleavage rate did not show any differences in those two groups, but the transferable embryo rate was significantly high in patients with high semen ORP. However, the patients with high ORP show a tendency to lower clinical pregnancy with a low abortion rate compared to the low ORP group. Conclusion: The main purpose of measuring the ORP value in semen is still questionable and shows controversial results.

Mitochondrial Transfer into Human Oocytes Improved Embryo Quality and Clinical Outcomes in Recurrent Pregnancy Failure Cases.

One of the most critical issues to be solved in reproductive medicine is the treatment of patients with multiple failures of assisted reproductive treatment caused by low-quality embryos. This study investigated whether mitochondrial transfer to human oocytes improves embryo quality and provides subsequent acceptable clinical results and normality to children born due to the use of this technology. We transferred autologous mitochondria extracted from oogonia stem cells to mature oocytes with sperm at the time of intracytoplasmic sperm injection in 52 patients with recurrent failures (average 5.3 times). We assessed embryo quality using the following three methods: good-quality embryo rates, transferable embryo rates, and a novel embryo-scoring system (embryo quality score; EQS) in 33 patients who meet the preset inclusion criteria for analysis. We also evaluated the clinical outcomes of the in vitro fertilization and development of children born using this technology and compared the mtDNA sequences of the children and their mothers. The good-quality embryo rates, transferable embryo rates, and EQS significantly increased after mitochondrial transfer and resulted in 13 babies born in normal conditions. The mtDNA sequences were almost identical to the respective maternal sequences at the 83 major sites examined. Mitochondrial transfer into human oocytes is an effective clinical option to enhance embryo quality in recurrent in vitro fertilization-failure cases.

Real-World Evidence on the Diagnostic and Clinical Characteristics of Asthma in Japanese Patients with COPD: The ACO Japan Cohort Study.

Introduction: The ACO Japan Cohort Study, a multicenter observational study, investigated the proportion of patients with chronic obstructive pulmonary disease (COPD) who met the Japanese Respiratory Society (JRS) asthma-COPD overlap (ACO) diagnostic criteria, characteristics of ACO and non-ACO patients, and the patient transitions between ACO/non-ACO diagnosis over 2 years. Patients and Methods: Patients with COPD were consecutively enrolled between June and December 2018 and followed up continuously for 2 years. All participating study sites were medical institutions where respiratory specialists routinely conducted medical examinations/tests required for ACO diagnosis. Results: Among 708 patients with COPD, 101 (14.3%), 118 (16.7%), and 125 (17.7%) were diagnosed with ACO at registration, 1 year, and 2 years, respectively. In total, 22.6% of patients lacked the data necessary for ACO diagnosis throughout the 2 years. Among patients who had the necessary data for ACO diagnosis, 24.7% were diagnosed with ACO at 2 years. More ACO patients had moderate or severe exacerbations in the past year than non-ACO patients at registration (15.8% vs 6.3%, p = 0.049) and 1 year (19.4% vs 7.6%, p = 0.025). ACO patients had a greater decrease in mean forced expiratory volume in one second over 2 years than non-ACO patients (-92.0 vs 43.4 mL). Among patients diagnosed with ACO at registration, 21.4% transitioned to non-ACO after 1 year. Conversely, almost all non-ACO patients at registration remained non-ACO after 1 year. Conclusion: COPD patients with ACO determined by the JRS criteria had a high risk of exacerbations and a rapid decline in respiratory function, indicating that the JRS criteria for ACO are useful for identifying high-risk COPD patients. Testing necessary for ACO diagnosis is insufficiently performed even in real-world clinical practice of COPD specialists.

Mitochondrial function of human embryo: Decline in their quality with maternal aging.

Background: Female fertility declines with age, due to increased chromosomal aneuploidy and possible reduced mitochondrial function in the embryo. Methods: This review outlines how mitochondrial function in human embryos, as predicted from oxygen consumption rate (OCR) measurements, changes in preimplantation stage, and what factors, particularly maternal age, affect mitochondrial function in embryos. Main findings: The structure of the mitochondrial inner membrane and its respiratory function developed with embryo development, while the copy number of mitochondrial DNA per specimen was transiently reduced compared with that of the oocyte. The undifferentiated state of the inner cell mass cells appears to be associated with a low OCR. In contrast, the copy number of mitochondrial DNA increased in trophoblast cells and mitochondrial aerobic metabolism increased.The OCRs at morulae stage decreased with maternal age, but there was no relationship between maternal age and the copy number of mitochondrial DNA at any stages. The higher oxygen spent at the morula stage; the shorter time was needed for development to the mid-stage blastocyst. Conclusions: The mitochondrial respiratory function of human embryos developed along with embryonic growth. Mitochondrial function at morula stage declined with their maternal age and reduced mitochondrial function decreased the rate of development from morula to blastocyst.

Influence of climatic conditions in the mesothermal climate area on pregnancies following elective fresh single blastocyst transfer.

PURPOSE: To investigate the effect of climatic parameters in the mesothermal climate area on clinical pregnancy and live birth following fresh single blastocyst transfer. METHODS: This study investigated clinical pregnancies and live births that resulted from 555 ovarian stimulation cycles followed by fresh single blastocyst transfer. The samples were stratified according to climatic conditions (low T, temperature < 12.9 °C; middle T, 12.9 °C ≤ temperature < 22.6 °C; high T, temperature ≥ 22.6 °C; low H, relative humidity < 62.1%; middle H, 62.1% ≤ relative humidity < 66.5%; high H, relative humidity ≥ 66.5%; short S, sunlight duration < 5.2 h; middle S, 5.2 h ≤ sunlight duration < 6.7 h; long S, sunlight duration ≥ 6.7 h). Clinical pregnancy and live birth rates among three groups from each climatic parameter were compared. Multivariable analyses were performed to investigate the effects of climatic conditions on blastocyst development, endometrial thickness, clinical pregnancy, and live birth. RESULTS: A statistically significant difference was found in pregnancy rates among low T (48.8%), middle T (37.3%), and high T (36.6%) groups. Multivariable analyses revealed that temperature was associated with clinical pregnancy and live birth rates with adjustment for patient age, BMI, type of ovarian stimulation, endometrial thickness, and expansion grade of the transferred blastocyst. The association between climatic parameters and blastocyst development and endometrial thickness was not confirmed. CONCLUSIONS: This study suggests that lower temperatures in the mesothermal climate area could favorably affect the rates of clinical pregnancy and live birth achieved by fresh single blastocyst transfer.

Fast-tracking antibody maturation using a B cell-based display system.

Affinity maturation, an essential component of antibody engineering, is crucial for developing therapeutic antibodies. Cell display system coupled with somatic hypermutation (SHM) initiated by activation-induced cytidine deaminase (AID) is a commonly used technique for affinity maturation. AID introduces targeted DNA lesions into hotspots of immunoglobulin (Ig) gene loci followed by erroneous DNA repair, leading to biased mutations in the complementary determining regions. However, systems that use an in vivo mimicking mechanism often require several rounds of selection to enrich clones possessing accumulated mutations. We previously described the human ADLib® system, which features autonomous, AID-mediated diversification in Ig gene loci of a chicken B cell line DT40 and streamlines human antibody generation and optimization in one integrated platform. In this study, we further engineered DT40 capable of receiving exogenous antibody genes and examined whether the antibody could be affinity matured. The Ig genes of three representative anti-hVEGF-A antibodies originating from the human ADLib® were introduced; the resulting human IgG1 antibodies had up to 76.4-fold improvement in binding affinities (sub-picomolar KD) within just one round of optimization, owing to efficient accumulation of functional mutations. Moreover, we successfully improved the affinity of a mouse hybridoma-derived anti-hCDCP1 antibody using the engineered DT40, and the observed mutations remained effective in the post-humanized antibody as exhibited by an 8.2-fold increase of in vitro cytotoxicity without compromised physical stability. These results demonstrated the versatility of the novel B cell-based affinity maturation system as an easy-to-use antibody optimization tool regardless of the species of origin.Abbreviations: ADLib®: Autonomously diversifying library, ADLib® KI-AMP: ADLib® knock-in affinity maturation platform, AID: activation-induced cytidine deaminase, CDRs: complementary-determining regions, DIVAC: diversification activator, ECD: extracellular domain, FACS: fluorescence-activated cell sorting, FCM: flow cytometry, HC: heavy chainIg: immunoglobulin, LC: light chain, NGS: next-generation sequencing, PBD: pyrrolobenzodiazepine, SHM: somatic hypermutation, SPR: surface plasmon resonance.

Novel tri-specific tribodies induce strong T cell activation and anti-tumor effects in vitro and in vivo.

BACKGROUND: Immunotherapy based on Bi-specific T Cell Engagers (TCE) represents one of the most attractive strategy to treat cancers resistant to conventional therapies. TCE are antibody-like proteins that simultaneously bind with one arm to a Tumor Associated Antigen (TAA) on cancer cells and with the other one to CD3 complex on a T-cell to form a TCR-independent immune synapse and circumvent Human Leucocyte Antigen restriction. Among them, the tribodies, such as Tb535H, a bi-specific molecule, made up of a Fab and a scFv domain both targeting 5T4 and another scFv targeting CD3, have demonstrated anti-tumor efficacy in preclinical studies. METHODS: Here, we generated five novel tri-specific and multi-functional tribodies, called 53X tribodies, composed of a 5T4 binding Fab arm and a CD3 binding scFv, but differently from the parental Tb535H, they contain an additional scFv derived from an antibody specific for an immune checkpoint, such as PD-1, PD-L1 or LAG-3. RESULTS: Compared with the parental Tb535H bi-specific T cell engager targeting 5T4, the novel 53X tribodies retained similar binding properties of Tb535H tribody, but showed enhanced anti-tumor potency due to the incorporation of the checkpoint inhibitory moiety. In particular, one of them, called 53L10, a tri-specific T cell engager targeting 5T4, CD3 and PD-L1, showed the most promising anti-tumor efficacy in vitro and led to complete tumor regression in vivo. CONCLUSIONS: The novel tribodies have the potential to become strong and safe therapeutic drugs, allowing to reduce also the cost of production as one single molecule contains three different specificities including the anti-TAA, anti-CD3 and anti-IC binding arms.

Prevention of intrauterine fetal growth restriction by administrating C1q/TNF-related protein 6, a specific inhibitor of the alternative complement pathway.

PURPOSE: The latest treatments do not sufficiently prevent miscarriage and fetal growth restriction (FGR) in pregnant women. Here, we assessed the effects of a human protein, CTRP6, that specifically inhibits the activation of the alternative complement pathway on miscarriage, fetal and placental development. METHODS: Pregnant CBA/J mice mated with DBA/2 male mice as a model of spontaneous abortion and FGR were randomly divided into the control and CTRP6 groups. In the CTRP6 group, the mice were intravenously administered CTRP6 on days 4.5 and 6.5 post-conception (dpc). The abortion rate and fetal and placental weights on 14.5 dpc were examined. Remodeling of the spiral artery was also assessed. RESULTS: The abortion rate in the CTRP6 group (13%) was reduced compared to the control group (21%), but there was no statistical difference. The placental and fetal weights in the CTRP6 group were also heavier than those in the control (P < 0.05). Moreover, the thickness of the blood vessel wall in the CTRP6 group was significantly thinner than that in the control (P < 0.05) and comparable to that in the non-abortion model (CBA/J x BALB). The ratio of the inner-per-the-outer diameter of the spiral artery increased more in the CTRP6 group than that in the control (P < 0.05). As well, the Th1/Th2 cytokine ratio was significantly reduced by CTRP6 treatment. CONCLUSIONS: Taken together, the supplementation with a protein that regulates the alternative complement pathway in vivo improves FGR and promotes spiral artery remodeling in a mouse model of miscarriage and FGR.

Mitochondria Transfer from Adipose Stem Cells Improves the Developmental Potential of Cryopreserved Oocytes.

Although it is not a well-established technology, oocyte cryopreservation is becoming prevalent in assisted reproductive technologies in response to the growing demands of patients' sociological and pathological conditions. Oocyte cryopreservation can adversely affect the developmental potential of oocytes by causing an increase in intracellular oxidative stresses and damage to the mitochondrial structure. In this study, we studied whether autologous adipose stem cell (ASC) mitochondria supplementation with vitrified and warmed oocytes could restore post-fertilization development that decreased due to mitochondrial damage following cryopreservation. ASC mitochondria showed similar morphology to oocytes' mitochondria and had a higher ATP production capacity. The vitrified-warmed oocytes from juvenile mice were supplemented with ASC mitochondria at the same time as intracellular sperm injection (ICSI), after which we compared their developmental capacity and the mitochondria quality of 2-cell embryos. We found that, compared to their counterpart, mitochondria supplementation significantly improved development from 2-cell embryos to blastocysts (56.8% vs. 38.2%) and ATP production in 2-cell embryos (905.6 & 561.1 pmol), while reactive oxygen species levels were comparable. With these results, we propose that ASC mitochondria supplementation could restore the quality of cryopreserved oocytes and enhance the embryo developmental capacity, signifying another possible approach for mitochondrial transplantation therapy.

Real-World Status of Medical Care and Treatment of Chronic Obstructive Pulmonary Disease by Respiratory Specialists in Japan.

INTRODUCTION: The ACO Registry Study was a multicenter, prospective, observational cohort study aiming to clarify the situation of asthma-chronic obstructive pulmonary disease (COPD) overlap (ACO) within the COPD population using the Japanese Respiratory Society (JRS) criteria. We reported the proportion of patients who met the ACO criteria among the COPD population at study registration. METHODS: Using data collected at registration, we investigated the implementation of each diagnostic examination/test required for ACO diagnosis in the full analysis set. Among patients with data necessary for ACO diagnosis, ACO/non-ACO patients with/without asthma diagnosed by a physician and proportions of inhaled corticosteroid (ICS) treatments for COPD were calculated. RESULTS: Of 708 patients analyzed, 396 (55.9%) had the data necessary for ACO diagnosis, and 312 (44.1%) did not. The proportions of patients who underwent laboratory and respiratory function tests (peripheral blood eosinophil count [79.8%], fractional exhaled nitric oxide [63.7%], airway reversibility [46.8%], and total immunoglobulin [Ig] E/specific IgE [33.3%]) were lower than those who underwent subjective examinations (perennial allergic rhinitis [100%], asthma before age 40 years [97.2%], and variable/paroxysmal respiratory symptoms [94.5%]). Among patients with the data necessary for ACO diagnosis and without asthma complications according to the physician's diagnosis, 15.1% (33/219) met the ACO criteria. Of patients who met the ACO criteria, 74.3% (75/101) received ICS, and 25.7% (26/101) did not. By comparison, among patients who did not meet the ACO criteria, 35.6% (105/295) were receiving ICS, and 64.4% (190/295) were not. CONCLUSIONS: The proportion of objective laboratory and physiological tests was lower than expected, despite study sites having the clinical resources for objective tests. Most ACO patients were being treated with ICS as recommended in the JRS treatment guidelines. Attempts should be made to further increase the proper use of ICS among these patients in Japan. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03577795.

Effects of time-restricted feeding and type of food on fertility competence in female mice.

We assessed the effects of feeding regimen (ad libitum vs. time-restricted food access) and type of food (normal chow (NC: 12% fat) vs. moderately high calorie diet (mHCD: 31% fat)) on fertility competence of female mice. Mice fed mHCD had higher number of oocytes than mice fed NC. On the other hand, when mice were fed NC under time-restricted access to food (NT), the developmental rate to the blastocyst per number of normally fertilized ova was significantly decreased compared to others. The reactive oxygen species (ROS) level in oocytes increased in time-restricted food access and NC group. Transcriptome analysis of whole ovarian tissues from these mice showed a change in the cholesterol metabolism among the four groups. Time-restricted food access decreased serum LDL cholesterol level in both NC and mHCD groups. Moreover, the number of atretic follicles increased in NT mice compared to ad libitum food access mice. The present study shows that mHCD feeding increases the number of ovulated oocytes and that time-restricted feeding of NC impairs the developmental competence of oocytes after fertilization, probably due to the changes in serum cholesterol levels and an increase in the ROS content in oocytes.

Novel Bi-Specific Immuno-Modulatory Tribodies Potentiate T Cell Activation and Increase Anti-Tumor Efficacy.

Cancer immunotherapy has already shown significant improvements by combining different antibodies specific for distinct immune checkpoints, such as Ipilimumab and Nivolumab. Here, we tested combinatorial treatments of immunomodulatory antibodies, previously generated in our laboratory, for their effects on hPBMC activation, either upon stimulation with SEB or in co-cultures with tumor cells by cytokine secretion assays. We found that some of them showed additive or synergistic effects, and on the basis of these observations, we constructed, for the first time, four novel bispecific tribodies (TR), made up of a Fab derived from one anti-IC mAb and two scFvs derived from another mAb targeting a different IC. All four TRs cotargeting either programmed cell death protein 1 (PD-1) and Lymphocyte Activating 3 (LAG-3) or programmed death-ligand 1 (PD-L1) and LAG-3 retained binding affinity for their targets and the antagonistic effects of their parental mAbs, but some of them also showed an increased ability to induce lymphocyte activation and increased in vitro cytotoxicity against tumor cells compared to parental antibodies used either alone or in combinatorial treatments. Furthermore, none of the tribodies showed significant increased cytotoxicity on human cardiomyocytes. Considering that the tribody format reduces production costs (as only one construct provides the inhibitory effects of two antibodies), has an intermediate molecular size (100 kDa) which is well suited for both tumor penetration and an acceptable half-life, we think that these novel immunomodulatory TRBs have the potential to become precious tools for therapeutic applications, particularly in monotherapy-resistant cancer patients.

Effect of X-ray exposure during hysterosalpingography on capabilities of female germ cells.

PURPOSE: To elucidate the effect of X-ray exposure during hysterosalpingography (HSG) on subsequent laboratory outcomes in in vitro fertilization (IVF). METHODS: A total of 1458 oocytes, consisting of 990 oocytes retrieved from 70 women (89 cycles) who underwent HSG prior to IVF and 468 oocytes from 45 women (57 cycles) who underwent IVF without HSG, were evaluated for their retrieval number, maturity, fertilization, and development post fertilization. X-ray exposure during HSG was recorded as reference air kerma (RAK) (mGy). Subjects were stratified according to the amount of RAK (Nil: IVF without HSG, L-RAK: RAK < 16.23, mH-RAK: RAK ≥ 16.23). The number of oocytes retrieved, oocyte maturation, fertilization, and embryo development was compared among 3 groups. Further, multivariate analyses were performed to investigate the effect of X-ray exposure on laboratory outcomes in IVF. RESULTS: There was a statistically significant difference in the fertilization rate among 3 groups (Nil: 71.6%, L-RAK: 80.5%, mH-RAK: 78.3%). The good-quality blastocyst rate in mH-RAK (46.2%) was significantly higher than L-RAK (35.3%) and Nil (32.4%). Multivariate analyses revealed that X-ray exposure was associated with higher fertilization, higher blastocyst development, and higher good-quality blastocyst development rates with adjustment for patient age, BMI, ovarian stimulation types, and fertilization methods. Association between X-ray exposure and the number of oocytes retrieved, and oocyte maturation was not confirmed. CONCLUSIONS: The present study suggests that X-ray exposure of the female reproductive organs during HSG could enhance the potential of oocytes rather than adversely.

Mural granulosa cells support to maintain the viability of growing porcine oocytes and its developmental competence after insemination.

PURPOSE: To enhance the in vitro growth of porcine oocytes, we studied the effect of mural granulosa cells (MGCs) on the viability of oocytes attached to granulosa cells (oocyte-granulosa cell complexes, OGCs) that were obtained from early antral follicles. METHODS AND RESULTS: When OGCs were cultured with MGCs for 12 days, there were significant improvement (P < 0.05) in the robustness of gap junctional communication between the oocyte and the granulosa cells (82% vs. 59%), the survival rate of oocytes (57% vs. 39%), and the diameter of survived oocytes (118 µm vs. 112 µm). The rate of oocyte release of OGCs cultured with MGCs on the 12th day (1.9%) was significantly lower than that of OGCs cultured without MGCs (26%). Complete meiotic arrest was maintained in the group with MGCs (100%), while partial resumption of spontaneous meiosis was noticed in the absence of MGCs (10-19%). Furthermore, the presence of MGCs increased the oocyte maturation rate after maturation culture in both 12- and 14-day culture groups (P < 0.05, 85-88%) compared to OGCs cultured without MGCs (48-60%). MGCs also significantly improved the blastocyst formation rate (day 7) after ICSI (P < 0.05). CONCLUSIONS: The data of this study thus shows that the presence of MGCs during in vitro oocyte growth plays a crucial role in supporting the developmental competence of growing porcine oocytes attached to the granulosa cells via enhancement of their viability.

Statement from the Japanese Respiratory Society: Working diagnosis and initial management of COPD during the COVID-19 pandemic.

The Japanese Respiratory Society (JRS) has recommended spirometry for the diagnosis of respiratory diseases. It is indispensable for the confirmation of airflow obstruction by spirometry in chronic obstructive pulmonary disease (COPD) diagnosis. However, the coronavirus disease 2019 (COVID-19) pandemic has made it difficult for many clinics to perform spirometry as it may lead to possible aerosol infections. Thus, the diagnosis of COPD, especially in the early stage, has become difficult. To overcome this situation, JRS issued a "Flowchart of Working Diagnosis and Management of COPD during the COVID-19 Pandemic". This flowchart may help physicians provisionally diagnose COPD patients without performing spirometry, offering them appropriate intervention even in epidemic and pandemic situations.

A novel embryo quality scoring system to compare groups of embryos at different developmental stages.

PURPOSE: To construct a new embryonic quality scoring system to compare groups of embryos at different developmental stages. METHODS: Based on a hypothesis that the implantation potential of any embryo in an ovum pickup (OPU) cycle remains the same at any stage of development, be it day 2, 3, or 5, a new embryo quality scoring (EQS) system was designed. It was based on the analysis of the clinical results of 1610 single embryo transfers. We validated this scoring system in the comparison of embryonic quality between groups by evaluating the mean scores calculated at day 2, day 3, and day 5 for 957 embryos (150 cycles) from 3 different groups. We then compared EQSs of patients with pregnancy favorable factors (group A) such as young age and high AMH levels, with the patients with contra features (group B). RESULTS: We confirmed that each mean EQS assessed at different stages of embryonic development within the same group was similar. The mean EQSs on day 3 and day 5 in group A were significantly higher than the mean EQSs on days 2, 3, and 5 in group B. CONCLUSION: The novel EQS system proposed by us enables embryonic quality comparison between groups of embryos at different developmental stages.