Direct Cleavage Research Articles

P-308 Abnormal Oocytes are more likely to lead to abnormal embryo divisions (Direct Unequal Cleavage,DUC), but do not compromise embryo quality as assessed using CHLOE-EQ score

Abstract Study question Do oocyte dysmorphisms lead to abnormal embryo divisions and compromised embryo quality? Summary answer Oocyte cytoplasmic abnormalities (granularity, Smooth Endoplasmic Reticulum,SER) did not affect CHLOE-EQ score; whilst zona abnormalities (thickness and unevenness) and SER tend to lead to DUCs. What is known already Oocyte dysmorphisms include extracytoplasmic [zona pellucida (ZP) evenness and thickness] and cytoplasmic abnormalities [SERs, inclusions, darkness, granularity]. The impact of these abnormalities on embryo development and viability as reported in the literature is contradictory. CHLOE-EQ score is an Artificial Intelligence (AI) based algorithm designed to support embryologists in assessing embryo viability, and has previously been demonstrated to automatically detect embryo development anomalies (such as DUCs), to be predictive of blastulation, utilisation, selection for transfer, ploidy, implantation and live birth. Therefore, CHLOE-EQ is a metric of embryo viability. The impact of oocyte dysmorphisms on CHLOE-EQ and DUCs is poorly understood. Study design, size, duration Retrospective cohort analysis of 742 embryo time-lapse videos, cultured at a private fertility clinic between June and July 2022. Participants/materials, setting, methods The clinic provided annotations on extracytoplasmic abnormalities (ZP thickness and uniformity) and cytoplasmic abnormalities [SERs, inclusions, darkness, granularity]. CHLOE-EQ (Fairtility) automatically annotated morphokinetics and DUCs and further quantified embryo viability scores (CHLOE-EQ and Blast Score). Main results and the role of chance CHLOE-EQ score was not affected by the oocyte having cytoplasmic abnormalities (no vs yes: 4.2 ± 4, n = 122vs4.1 ± 4, n = 359, NS): dark (4.1 ± 4, n = 476vs3.1 ± 4, n = 5, NS), granular (4.1 ± 4, n = 179vs4.1 ± 4, n = 302, NS), SER (4.1 ± 4, n = 455 vs 3.8 ± 4, n = 26, NS), inclusion (4.1 ± 4, n = 430vs4.1 ± 4, n = 51, NS); or ZP abnormalities [overall (4.2 ± 4, n = 379vs3.7 ± 4, n = 102, NS), non-uniformity (4.1 ± 4, n = 409vs4.2 ± 4, n = 72, NS), thick ZP (4.2 ± 4, n = 457vs2.8 ± 4, n = 24, NS), thin ZP(4.1 ± 4, n = 475vs2.1 ± 4, n = 6, NS]. DUC embryos were two times more likely to be derived from oocytes with thick ZP (9/98, 9%oocytes) than oocytes without thick ZP (15/383, 3.9%, p = 0.03). DUCs were more likely to have a non-uniform ZP compared to non-DUCs [DUCs: 7%(7/98) vs Non-DUCs: 17%(65/383), p = 0.015]. DUCs were not associated with the following oocyte cytoplasmic dysmorphias: SER (DUC vs Non-DUCs: 6/98,6% vs 20/388,5%,NS), dark (0/98 vs 5/383, NS), granular (61/98 vs 241vs383, NS), inclusions (11/98 vs 40/383, NS). DUCs had lower blastulation rate than non-DUCs [DUC:1.8%(2/113) vs Non-DUCs:77%(298/389), p < 0.001]. DUCs were 4-fold less likely to be multinucleated at the 2 cell stage than non-DUCs [DUC: 7%(2/29)vsNon-DUCs: 30%(91/305), p = 0.03]. DUCs were 7-fold more likely to be multinucleated at the 4 cell stage than non-DUCs [DUC: 7%(2/29)vs Non-DUCs:1%(3/302), p = 0.06]. Patient age was not associated with DUCs (DUCs: 36.9 ± 4 vs non-DUCs:37.1 ± 4, NS). Limitations, reasons for caution This was a retrospective-single clinic study. Causality is not determined. Wider implications of the findings Given the growing evidence that DUCs have compromised viability, it is important to understand the biology of how DUCs are connected to oocyte quality. Using AI to detect DUCs to avoid critical information being missed during embryo assessment can assist embryologists in maximising their efficacy of embryo selection. Trial registration number NA

P-146 Larger time intervals between pronuclear fading and the first mitosis are strongly associated with trichotomous mitoses and cleavage arrest in human embryos

Abstract Study question Does the duration of the interval between pronuclear fading and first cell division affect preimplantation embryo developmental competence? Summary answer Larger pronuclear-fading to first-division intervals (t2-tPNf) are associated with greater risk of direct unequal cleavage (DUC) and lower blastulation rate, but not of blastocyst aneuploidy. What is known already During the last decade, time lapse technology (TLT) has made possible detailed and dynamic observation of in vitro embryo development, offering a powerful non-invasive tool for improving embryo selection. Recent evidence suggests that the phase between pronuclear fading and the first cleavage is a perilous bridge between the zygote and the cleaving embryo. Following pronuclear breakdown, delayed progression through mitosis caused by incomplete DNA replication may result in chromosome lagging, whole and segmental segregation errors, and breakage. In this analysis, we specifically tested the hypothesis that delays in a short final phase of fertilization have an impact on embryo development. Study design, size, duration Retrospective study of 1316 zygotes cultured until day 5 using TLT and derived from first ICSI cycles (111 no PGT-A and 94 PGT-A) performed between 2018 and 2022. Testicular sperm or cryopreserved gametes cycles were excluded. DUC was defined as the cleavage of one zygote or blastomere into three daughter blastomeres or cell cycle interval shorter than five hours at the first (DUC1), second (DUC2) or third (DUC3) cell division cycle. Participants/materials, setting, methods t2-tPNf intervals were clustered into quartiles (Q1, <2hpi; Q2, 2–2.5hpi; Q3, 2.5–3hpi and Q4, > 3hpi), which were compared based on embryo development and live birth by linear and logistic regression. The Q1 cluster was set as control, while all other quartiles groups were considered as dummy variables. Multivariate analysis was conducted using the generalized estimating equations, to control for variable numbers of zygotes from each patient. The primary outcome was DUC rate. Main results and the role of chance Overall, rates of DUC1, DUC2 and DUC 3 rates were 10.3%, 8% and 2.9%, respectively. After adjusting for major confounders (maternal age, paternal age, maternal body mass index, cause of infertility, anti-Müllerian hormone, follicle stimulating hormone) we observed a higher DUC 1 and DUC 2 rate with increasing times of t2-tPNf. We observed abnormal distribution of DUC1 and DUC2 rate between quartiles groups. Specifically, 89% of DUC1 was shown among Q3 (21%) and Q4 (78%) and 61% of DUC2 among Q3 (22%) and Q4 (39%). The percentage of DUC1 was higher for Q3 (multivariate-OR = 3.29, 95%CI 0.99-10.85, adjusted-p = 0.01) and Q4 (multivariate-OR = 22.79, 95%CI 7.80-66.62, adjusted-p<0.01]. The percentage of DUC2 was higher only in Q4 (multivariate-OR = 2.58, 95%CI 1.13-5-57, adjusted-p = 0.01). Moreover, we observed a reduced blastulation rate in Q3 (multivariate-OR = 0.68, 95%CI 0.49-0.95, adjusted-p = 0.01) and Q4 (multivariate-OR = 0.27, 95%CI 0.19-0.38, adjusted-p<0.01). No significant differences were observed in DUC3 rate, top quality blastocyst, euploidy rate (PGT-A cycles analysis) and live birth rate. Limitations, reasons for caution The retrospective single-centre design and the limited sample size are limitations of the study Wider implications of the findings The study is consistent with the emerging view that the final phases of fertilization - although short - are crucial for development. Delays in progression towards mitosis probably express chromosome integrity loss and perturbances of chromosome segregation. In most embryos, such perturbances appear to cause trichotomous mitoses and cleavage arrest. Trial registration number Not applicable

O-122 The first study to assess the clinical efficacy of CHLOE-EQ on the assessment of embryo viability of embryos cultured in a GERI time-lapse incubator

Abstract Study question Can an automatic AI scoring system predict ploidy, live birth and utilization? Are there differences in AI scoring between donor and own gametes? Summary answer CHLOE-EQ Score is directly associated with oocyte quality, ploidy, utilization, live birth, embryo quality, direct uneven cleavage (DUC), blastulation, utilization and selection for transfer. What is known already The integration of AI algorithms, such as CHLOE-EQ, into different Time-lapse systems requires clinical and biological validation. Geri Time-lapse videos have given the embryologists more insight into embryo development. Analyzing this information manually requires time and introduces risk of error. To tackle this issue, AI solutions like CHLOE-EQ (Fairtility) can be used to automatically assess video datapoints. CHLOE-EQ provides an embryo quality score that has been shown to predict embryo viability and ploidy, providing clarity on the underlying biological factors. Before introducing AI tools in clinical practice, it is crucial to confirm their efficacy and validate with clinical data. Study design, size, duration A retrospective cohort analysis was conducted at a private clinic in Spain from April 2021 to November 2022, involving the review of 3196 Geri time-lapse videos with a subset of known ploidy and live birth outcomes. The correlation of CHLOE-EQ score with ASEBIR clinic grading was evaluated. As well as with DUCs, oocyte quality, sperm source, blastulation, utilization, selection for transfer, ploidy and live birth. Participants/materials, setting, methods Geri time-lapse videos were automatically analyzed by CHLOE-EQ (Fairtility). CHLOE-EQ score was assessed in relation to laboratory (ploidy, clinic ASEBIR embryo scoring, utilization, selection for transfer) and clinical outcomes (live birth), as well as between own vs donor gametes (own eggs >40y vs donor eggs and testicular sperm vs donor sperm) using descriptive statistics and t-test. The accuracy of prediction was measured using binary logistic regression (AUC). Main results and the role of chance CHLOE-EQ score was positively correlated with ASEBIR embryo quality (A:8.7±1.9, n = 349 >B:6.8±2.9, n = 470 > C: 5.1±3.0, n = 124 > D:1.3±2.1, n=751; p < 0.05). Non-DUCs had higher CHLOE-EQ Score than DUCs [5.3±3.8, n=1798 vs 1.9±0.38, n=643, p<0.001]. CHLOE-EQ Score was unaffected by the quality of the sperm sample, with similar CHLOE-EQ scores between donor sperm and testicular derived sperm (4.1±3.9, n=335 vs 3.4±4.2, n=56, respectively, NS). Embryos that blastulated (yes vs no: 5.4±3.7, n=1996 vs 0.6±2.1, n=309, p < 0.001), were utilized (7.4±0.28, n=911, vs 1.0±2.1, n = 1309, p < 0.001), selected for transfer (8.7±2.4, n=153 vs 3.3±3.7, n = 2067, p < 0.001), were euploid (7.5±2.5, n=72 vs 6.3±3, n=152, p = 0.001) and resulted in live births (4.4±4.1, n = 332 vs 3.8±4, n = 499, p = 0.02) had a higher CHLOE-EQ score than embryos that did not. CHLOE-EQ Score is higher in embryos derived from oocytes from donors than own eggs, suggesting that oocyte quality affects CHLOE-EQ score (4.0±4, n = 1189 vs 2.7±3.4, n = 356). CHLOE-EQ Score is predictive of utilization (AUC=0.95, n=2220, baseline=41%, p<0.001), euploidy (AUC=0.63, n=224, baseline=32.1%, p=0.003), blastulation (AUC=0.94, n = 2305, baseline=86.6%, p < 0.001) and selection for transfer (AUC=0.89, n = 2220, baseline=41%, p<0.001). Limitations, reasons for caution This is a retrospective single-center study in which embryos for transfer were selected by human embryologists, and forms part of program to validate the responsible integration of AI into clinical practice in each individual clinic. Wider implications of the findings This is the first study presenting the efficacy of prediction of CHLOE-EQ with GERI data. AI tools have the potential to improve consistency, efficiency, and accuracy of embryo assessment and selection. CHLOE-EQ predicts through quantitative and qualitative morphological and morphokinetics information, resulting in more personalized care for each individual embryo. Trial registration number not applicable

P-204 The pattern of the zygote cleavage to three cells must be considered before embryo transfer

Abstract Study question What is the clinical outcome of instant irregular cleavage (IDC) from zygote to three cells? Summary answer IDC embryos, did not develop to blastocysts and did not acieve pregnancy; FC embryos that reached blastocyst stage exerted similar pregnancy rates as control embryos What is known already Evaluation of the zygote morphology is one of the earliest embryonic stages investigated. Several zygote grading systems were proposed based on pronuclei size and NPB (nucleolus precursor bodies) distribution, thus is an effective indicator of the embryo’s potential to result in a life birth. The first cleavage of a zygote is a highly coordinated event. Disruptive timing of the first cleavage, named direct cleavage (DC, less than 5 hours), is associated with formation of poor embryo quality. Early identification of pathological embryos is challenging and impacts the timing of embryo fate decision in ART. Study design, size, duration A total of 1978 fresh IVF cycles from single unit were analyzed and a total of 4012 embryos were studied. Only 2pn embryos, cultured exclusively in time-lapse system for 5 days were included. The morphokinetics of embryos and their outcome were recorded, from z-score analysis through day 5. We discriminated between IDC, FC and normal cleavage pattern of embryos and compared clinical outcome of various morphokinetic parameters. We also evaluated possible early predictors for IDC. Participants/materials, setting, methods We generated three study groups according to embryo cleavage pattern: (I) Control, normal cleavage (n = 551); (II) FC, zygote to three cells within 5 hours (n = 1587); (III) IDC, instant cleavage from zygote to three cells (n = 922). The association between z-score of 138 IDC embryos and their sibling random controls was thoroughly investigated. All time lapse annotations were performed by senior embryologists. Statistical analysis was performed by SPSS software. Main results and the role of chance IDC embryos were mainly arrested at day 3 and the number of usable embryos (reached blastocyst stage and were suitable for embryo transfer or cryopreservation) was negligible; 4/922 (0.4%). In comparison, the amount of usable FC embryos was 108/1587 (6.6%) and control embryos usage reached 180/551 (32.7%). While the pregnancy rate of control and FC embryos, which reached embryo transfer were similar (40.35% and 42.55%, respectively); transfer of IDC embryos did not result in pregnancy. Additionally, the timetable as measured by time of PN fading and time from fading to first cleavage, differed significantly between the three groups. Therefore, we performed a thorough analysis of zygote morphology data of IDC embryos compared to control sibling embryos in search of early possible markers for these findings. We have not detected significant differences in z-score analysis, reflected by number and size of nucleoli, as well as pronuclear symmetry. Limitations, reasons for caution IDC embryos number used for z-score analysis was limited since many didn’t meet the inclusion criteria (lack of overlapping of the pronuclei, presence of random of control embryo and good quality image). Wider implications of the findings The decision regarding the fate of IDC and FC embryos should include the pattern of first cycle cleavage. Culture IDC and FC embryos for 5 days up to the blastocyst will spare transfer of embryos that are fated to arrest even when their morphological grade on day 3 is acceptable. Trial registration number 0043-22-MMC

P-150 Comparison of morphokinetic parameters of embryos generated from Capacitation In-vitro Maturation and Controlled ovarian stimulation-ICSI

Abstract Study question Is there any difference in the embryo morphokinetic parameters between embryos generated from Capacitation – In-vitro maturation (CAPA-IVM) and Controlled ovarian stimulation – ICSI (COS-ICSI)? Summary answer Embryos generated from CAPA-IVM cycles reached tPNa, t2, t4, t8, tSC, tM, tSB, and tB significantly later than COS-ICSI embryos. What is known already IVF with ovarian hyperstimulation has limitations in some subgroups of women at high risk of ovarian stimulation, such as those with polycystic ovary syndrome. IVM may be a feasible alternative to IVF in women with a high AFC. A recent randomized controlled trial produced statistically inconclusive findings regarding the noninferiority of the CAPA-IVM technique compared with COS-ICSI. Moreover, there is a lack of studies investigating the impact of CAPA-IVM on the morphokinetic parameters of embryos. Study design, size, duration This is a retrospective cohort study conducted from March 2021 to March 2022. The study included 115 couples (68 couples used CAPA-IVM, 47 couples used COS-ICSI) who had their embryos cultured to the blastocyst stage using a time-lapse system. Participants/materials, setting, methods The study was conducted at My Duc Phu Nhuan hospital, Ho Chi Minh City, Vietnam. Couples aged ≤ 40 years old, had blastocyst culture using time-lapse system were included in the study. Couples used oocyte or sperm donation, PGT treatment, had severe male factors, and fertilization failure in previous cycles were excluded. We analyzed 392 blastocysts from 115 couples for morphokinetic parameters including tPNa, tPNf, the onset of t2 to t8, tSC, tM, tSB, and tB. Main results and the role of chance There was no difference in tPNf, t3, and t5 between embryos from CAPA-IVM and COS-ICSI. However, embryos generated from CAPA-IVM reached tPNa, t2, t4, t8, tSC, tM, tSB, and tB significantly later than COS-ICSI embryos [tPNa (9.3 2.0 vs 8.0 1.9; P < 0.001), t2 (27.4 3.7 vs. 25.5 3.1; P < 0.001), t4 (38.7 4.5 vs. 37.3 4.3; P = 0.004), t8 (58.9 9.2 vs. 56.3 8.4; P = 0.011), tSC (76.2 11.1 vs. 71.4 10.4; P < 0.001), tM (90.4 8.6 vs. 85.5 10.0; P < 0.001); tSB (101 8.1 vs. 97.1 8.3; P < 0.001) and tB (108 7.6 vs 106 7.7; P < 0.05)]. The frequency of direct cleavage in the first and the second cell division cycle was similar between two groups (58.6% vs. 30.0%; 41.4% vs. 70%, respectively, P > 0.05). Limitations, reasons for caution The retrospective cohort study design could limit the ability to eliminate sources of bias. The characteristics of participants may influence the study findings, i.e. women in the CAPA-IVM group were PCOS while women in the COS-ICSI were not. In addition, the study population's single-ethnicity nature limits the results' external generalizability. Wider implications of the findings This study found that embryos from CAPA-IVM reached tPNa, tPNf, t2, t4, t8, tSC, tM, tSB, and tB significantly later than COS-ICSI. The findings of this study provide evidence for further studies to evaluate CAPA-IVM embryos and to improve the CAPA-IVM culture system. Trial registration number Not Applicable

Catalytic transfer hydrogenolysis mechanism of benzyl phenyl ether over NiCu/Al2O3 using isopropanol as hydrogen source

Efficient transfer hydrogenolysis of benzyl phenyl ether (BPE) was accomplished over Ni1.5Cu1.5/Al2O3 using isopropanol as hydrogen source. The characterization results demonstrated that the formation of NiCu alloy and electron transfer between Ni and Cu were responsible for the excellent catalytic performance. Isopropanol displayed better hydrogen-donating ability than other low-carbon alcohols because of its unique dehydrogenation pathway and small steric hindrance. The results of isotopic experiments elucidated that active hydrogen species for the CO bond cleavage originated from isopropanol rather than H2, while both isopropanol and H2 were responsible for hydrogenation of phenol to cyclohexanol when reaction was conducted with mixed hydrogen sources. A mechanism study illustrated that transfer hydrogenolysis of BPE proceeded through direct cleavage of CO bond followed by phenol hydrogenation, in which the active hydrogen species were in the form of H+ and H−.

The developmental potential of mature oocytes derived from rescue in vitro maturation

To examine the developmental competence of immature oocytes in stimulated cycles, that matured after rescue invitro maturation (IVM) compared with their sibling invivo matured oocytes. Retrospective cohort study. IVF clinic. A total of 182 patients underwent 200 controlled ovarian stimulation cycles with intracytoplasmic sperm injection cycles in which immature oocytes were retrieved and at least one mature oocyte was obtained through rescue IVM. Invitro culture of immature germinal vesicle (GV) and metaphase I (MI) oocytes, retrieved in stimulated cycles. Fertilization rate, cleavage rate, blastulation rate, ploidy of embryos evaluated using preimplantation genetic testing for aneuploidy, morphokinetic parameters and pregnancy outcomes. In total, 2,288 oocytes were retrieved from 200 cycles. After denudation, 1,056 of the oocytes (46% ± 16%) were classified as metaphase II (MII). A total of 333/375 (89%) of MI oocytes and 292/540 (54%) of GV oocytes matured overnight and underwent intracytoplasmic sperm injection. The fertilization rates of matured oocytes from MI rescue IVM (R-MI) and from GV rescue IVM (R-GV) were comparable with those of their sibling MII oocytes (71% vs. 66%; 66% vs. 63%, respectively). Early cleavage rates (80% ± 35% vs. 92% ± 20%; 80% ± 42% vs. 95% ± 28%, respectively) and blastulation rates (32 ± 40% vs. 62 ± 33%; 24 ± 37% vs. 60 ± 35%, respectively) were significantly decreased in rescue IVM matured oocytes (R-oocytes)-derived zygotes, but the blastocyst (BL) euploidy rate and "good quality" BL rate were comparable with those of MII sibling-derived embryos. In addition, rescue IVM embryos showed significantly higher levels of multinucleation at the 2- and 4-cell stages, as well as higher rates of zygote direct cleavage from one to 3 to 4 cells. Overall, 21 transfers of rescue IVM embryos resulted in 3 healthy live births. For patients with a low maturation rate and/or low numbers of mature oocytes at retrieval, rescue IVM may contribute more competent oocytes and additional viable BLs for transfer from the same stimulation cycle, maximizing the chances for pregnancy and live birth.

Oxidative cleavage and ammoxidation of organosulfur compounds via synergistic Co-Nx sites and Co nanoparticles catalysis

The cleavage and functionalization of C–S bonds have become a rapidly growing field for the design or discovery of new transformations. However, it is usually difficult to achieve in a direct and selective fashion due to the intrinsic inertness and catalyst-poisonous character. Herein, for the first time, we report a novel and efficient protocol that enables direct oxidative cleavage and cyanation of organosulfur compounds by heterogeneous nonprecious-metal Co-N-C catalyst comprising graphene encapsulated Co nanoparticles and Co-Nx sites using oxygen as environmentally benign oxidant and ammonia as nitrogen source. A wide variety of thiols, sulfides, sulfoxides, sulfones, sulfonamides, and sulfonyl chlorides are viable in this reaction, enabling access to diverse nitriles under cyanide-free conditions. Moreover, modifying the reaction conditions also allows for the cleavage and amidation of organosulfur compounds to deliver amides. This protocol features excellent functional group tolerance, facile scalability, cost-effective and recyclable catalyst, and broad substrate scope. Characterization and mechanistic studies reveal that the remarkable effectiveness of the synergistic catalysis of Co nanoparticles and Co-Nx sites is crucial for achieving outstanding catalytic performance.

In Vitro Chemical Mapping of G-Quadruplex DNA Structures by Bis-3-Chloropiperidines.

G-quadruplexes (G4s) are biologically relevant, non-canonical DNA structures that play an important role in gene expression and diseases, representing significant therapeutic targets. Accessible methods are required for the in vitro characterization of DNA within potential G-quadruplex-forming sequences (PQSs). B-CePs are a class of alkylating agents that have proven to be useful chemical probes for investigation of the higher-order structure of nucleic acids. This paper describes a new chemical mapping assay exploiting the specific reactivity of B-CePs with the N7 of guanines, followed by direct strand cleavage at the alkylated Gs. Namely, to distinguish G4 folds from unfolded DNA forms, we use B-CeP 1 to probe the thrombin-binding aptamer (TBA), a 15-mer DNA able to assume the G4 arrangement. Reaction of B-CeP-responding guanines with B-CeP 1 yields products that can be resolved by high-resolution polyacrylamide gel electrophoresis (PAGE) at a single-nucleotide level by locating individual alkylation adducts and DNA strand cleavage at the alkylated guanines. Mapping using B-CePs is a simple and powerful tool for the in vitro characterization of G-quadruplex-forming DNA sequences, enabling the precise location of guanines involved in the formation of G-tetrads.

Selective Photocatalytic C–C Bond Cleavage of Lignin Models and Conversion to High-Value Chemical by Polyoxometalate Under a Mild Water-Based Environment

Selective bond cleavage using a photoredox reaction is a powerful technique in the chemical conversion of biomass such as lignin since it enables us to produce value-added aromatic compounds by controlling the activation of certain chemical bonds. Also, a water-based environment would be preferred for the sake of an eco-friendly reaction, additionally having the advantage of utilizing water-originated species such as •OH and H2O2. However, a direct cleavage of the C–C bond is still challenging due to the difficulties in tailoring redox potentials of catalysts relevant to covalent bond decoupling. Herein, we investigate a selective C–C bond cleavage of the β-O-4 lignin model compound using giant Keplerate polyoxometalates as photocatalysts. The redox potentials of photocatalysts are varied by changing metal compositions, and the correlation between redox potential and covalent bond decoupling is elucidated, probing each set of active radicals by HOMO and LUMOs. Especially, Mo132 shows a highly selective C–C bond cleavage with hydroxyl groups in the lignin model and produces phenyl formate─a effective CO surrogate for the synthesis of various organic compounds─as the main product (97.8% of product selectivity) in a water-based condition. Furthermore, Mo132 decomposes other aromatic polymer models, indicating the potential of photocatalytic C–C bond decoupling of polymers under a benign environment.

Role of Coxsackievirus B3-Induced Immune Responses in the Transition from Myocarditis to Dilated Cardiomyopathy and Heart Failure.

Dilated cardiomyopathy (DCM) is a cardiac disease marked by the stretching and thinning of the heart muscle and impaired left ventricular contractile function. While most patients do not develop significant cardiac diseases from myocarditis, disparate immune responses can affect pathological outcomes, including DCM progression. These altered immune responses, which may be caused by genetic variance, can prolong cytotoxicity, induce direct cleavage of host protein, or encourage atypical wound healing responses that result in tissue scarring and impaired mechanical and electrical heart function. However, it is unclear which alterations within host immune profiles are crucial to dictating the outcomes of myocarditis. Coxsackievirus B3 (CVB3) is a well-studied virus that has been identified as a causal agent of myocarditis in various models, along with other viruses such as adenovirus, parvovirus B19, and SARS-CoV-2. This paper takes CVB3 as a pathogenic example to review the recent advances in understanding virus-induced immune responses and differential gene expression that regulates iron, lipid, and glucose metabolic remodeling, the severity of cardiac tissue damage, and the development of DCM and heart failure.

Plasma Kallikrein as a Forgotten Clotting Factor.

For decades, it was considered that plasma kallikrein's (PKa) sole function within the coagulation cascade is the activation of factor (F)XII. Until recently, the two key known activators of FIX within the coagulation cascade were activated FXI(a) and the tissue factor-FVII(a) complex. Simultaneously, and using independent experimental approaches, three groups identified a new branch of the coagulation cascade, whereby PKa can directly activate FIX. These key studies identified that (1) FIX or FIXa can bind with high affinity to either prekallikrein (PK) or PKa; (2) in human plasma, PKa can dose dependently trigger thrombin generation and clot formation independent of FXI; (3) in FXI knockout murine models treated with intrinsic pathway agonists, PKa activity results in increased formation of FIXa:AT complexes, indicating direct activation of FIX by PKa in vivo. These findings suggest that there is both a canonical (FXIa-dependent) and non-canonical (PKa-dependent) pathway of FIX activation. These three recent studies are described within this review, alongside historical data that hinted at the existence of this novel role of PKa as a coagulation clotting factor. The implications of direct PKa cleavage of FIX remain to be determined physiologically, pathophysiologically, and in the context of next-generation anticoagulants in development.

Direct Oxygen-Oxygen Cleavage through Optimizing Interatomic Distances in Dual Single-atom Electrocatalysts for Efficient Oxygen Reduction Reaction.

The oxygen reduction reaction (ORR) on transition single-atom catalysts (SACs) is sustainable in energy-conversion devices. However, the atomically controllable fabrication of single-atom sites and the sluggish kinetics of ORR have remained challenging. Here, we accelerate the kinetics of acid ORR through a direct O-O cleavage pathway through using a bi-functional ligand-assisted strategy to pre-control the distance of hetero-metal atoms. Concretely, the as-synthesized Fe-Zn diatomic pairs on carbon substrates exhibited an outstanding ORR performance with the ultrahigh half-wave potential of 0.86 V vs. RHE in acid electrolyte. Experimental evidence and density functional theory calculations confirmed that the Fe-Zn diatomic pairs with a specific distance range of around 3 Å, which is the key to their ultrahigh activity, average the interaction between hetero-diatomic active sites and oxygen molecules. This work offers new insight into atomically controllable SACs synthesis and addresses the limitations of the ORR dissociative mechanism.

Transition-Metal-Free Unusual Oxidative Cleavage of Homoallylic Alcohol and Its Application in the Upcycling of Terpene to Value-Added Chemicals.

A NaOtBu-O2 -mediated oxidative dehomologation of homoallylic alcohols into structurally different carboxylic acids through direct oxidative cleavage of either the C(sp2 )-C(sp2 ) or C(sp3 )-C(sp3 ) bond utilizing enolate chemistry was demonstrated. Furthermore, under transition-metal-free conditions, this protocol has been applied to convert terpene as biomass feedstock into value-added chemicals.

Terminal Complement Activation Is Induced by Factors Released from Endplate Tissue of Disc Degeneration Patients and Stimulates Expression of Catabolic Enzymes in Annulus Fibrosus Cells.

Terminal complement complex (TCC) deposition was identified in human degenerated discs. To clarify the role of terminal complement activation in disc degeneration (DD), we investigated respective activating mechanisms and cellular effects in annulus fibrosus (AF) cells. Isolated cells from human AF, nucleus pulposus (NP), and endplate (EP) were stimulated with human serum alone or with zymosan and treated with either the C3 inhibitor Cp40 or the C5 antibody eculizumab. Complement activation was determined via anaphylatoxin generation and TCC deposition detection. Thereby, induced catabolic effects were evaluated in cultured AF cells. Moreover, C5 cleavage under degenerative conditions in the presence of AF cells was assessed. Zymosan-induced anaphylatoxin generation and TCC deposition was significantly suppressed by both complement inhibitors. Zymosan induced gene expression of ADAMTS4, MMP1, and COX2. Whereas the C3 blockade attenuated the expression of ADAMTS4, the C5 blockade reduced the expression of ADAMTS4, MMP1, and COX2. Direct C5 cleavage was significantly enhanced by EP conditioned medium from DD patients and CTSD. These results indicate that terminal complement activation might be functionally involved in the progression of DD. Moreover, we found evidence that soluble factors secreted by degenerated EP tissue can mediate direct C5 cleavage, thereby contributing to complement activation in degenerated discs.

The metabolite alpha-ketoglutarate: a novel target of gasdermin C-dependent pyroptosis.

The metabolite alpha-ketoglutarate: a novel target of gasdermin C-dependent pyroptosis.

Activation of CO2 by Direct Cleavage Triggered by Photoelectrons on Rutile TiO2(110)

The initial activation of the inert CO2 is a key step in its photoreduction to valuable chemicals. This process was proposed to proceed mainly by CO2 accepting a photoelectron to form a CO2•- radical or by CO2 accepting two photoelectrons and a proton to form the HCOO- anion on the prototypical rutile TiO2(110) surface. Here, we reveal a new mechanism, in which CO2 is directly cleaved to CO and the adsorbed O2- anion under the trigger of two photoelectrons, by using density functional theory calculations with the HSE06 hybrid functional. The newly revealed mechanism is more favorable than the two previously proposed pathways. Furthermore, our results show that the deficiency of photoelectrons on the catalyst surface is a potential reason for the current low efficiency of CO2 photoreduction.

MiR164–NAC21/22 module regulates the resistance of Malus hupehensis against Alternaria alternata by controlling jasmonic acid signaling

Apple leaf spot disease caused by Alternaria alternata apple pathotype (A. alternata AP) is one of the most severe fungal diseases affecting apple cultivation. Transcription factors are involved in various disease-resistance responses, and many of them are regulated by miRNAs. Here, we performed RNA-Seq to investigate gene expression changes during the defense response of Malus hupehensis against A. alternata AP. NAC21/22 was induced upon A. alternata AP infection and silenced by miR164 via direct mRNA cleavage. Contrasting expression patterns were noted between mature miR164 and NAC21/22 during infection. Contrary to NAC21/22 silencing, transiently overexpressing NAC21/22 in M. hupehensis alleviated disease symptoms on ‘gala’ leaves, impeded A. alternata AP growth, and promoted jasmonic acid (JA) signaling-related gene expression. Importantly, transient miR164f overexpression in ‘gala’ leaves enhanced A. alternata AP sensitivity, due perhaps to NAC21/22 downregulation, whereas miR164 suppression produced an opposite effect. In summary, the miR164–NAC21/22 module plays a pivotal role in apple resistance against A. alternata AP by regulating JA signaling.

Direct cleavage during the first mitosis is a sign of abnormal fertilization in cattle

Direct cleavage, a type of abnormal cleavage in which one zygote divides into three or more blastomeres, has been reported in mammals. The incidence of direct cleavage increases in zygotes with three or more pronuclei (multi-PN) and those showing abnormal pronuclei migration. However, there are few reports on the relationship between pronuclei and direct cleavage, and the effects of these relationships on subsequent embryogenesis have not been clarified. It is difficult to observe pronuclei under visible light, especially in bovine zygotes, because of abundant dark lipid droplets in the cytoplasm. We visualized pronuclei by removing lipid droplets from bovine zygotes and analyzed the relationship between the number of pronuclei and direct cleavage using time-lapse cinematography. The direct cleavage rate of multi-PN zygotes was 78.6%, which was significantly higher than that of zygotes with one pronucleus (1 PN, 0.0%) and two pronuclei (2 PN, 8.2%). Observation of pronuclei migration in 2 PN zygotes showed that 3.1% of 2 PN zygotes had non-apposed pronuclei. The direct cleavage rate of zygotes with non-apposed pronuclei was 66.7%, which was significantly higher than that of zygotes with apposed pronuclei (6.4%). Among multi-PN zygotes, the proportions of zygotes with apposed pronuclei and non-apposed pronuclei were 37.5% and 64.3%, respectively. The direct cleavage rate of multi-PN zygotes with non-apposed pronuclei was 100.0%, which was significantly higher than that of zygotes with apposed pronuclei (40.0%). Three-dimensional live-cell imaging of bovine zygotes injected with the mRNA-encoding histone H2B-mCherry showed that the direct cleavage rates of 2 PN and multi-PN zygotes bypassing syngamy were 63.2% and 75.5%, respectively. These rates were significantly higher than that of 2 PN and multi-PN zygotes that underwent syngamy (5.6% and 20.0%, respectively). Regardless of the number of pronuclei, a high frequency of direct cleavage was observed in zygotes in which the pronuclei did not migrate inward the cytoplasm and bypassed syngamy. These results suggest that abnormal fertilization such as multi-PN and migration error of pronuclei in cattle is the primary reason for direct cleavage during the first mitosis. Assessment of direct cleavage during the first mitosis allows exclusion of embryos with abnormal fertilization and may contribute to in vitro produced embryo transfer success.

Mutant plasminogen in hereditary angioedema is bypassing FXII/kallikrein to generate bradykinin.

Hereditary angioedema (HAE) is characterized by recurrent localized edema in various organs, which can be potentially fatal. There are different types of hereditary angioedema, which include genetic deficiency of C1 inhibitor (C1-INH) and hereditary angioedema with normal C1-INH (HAEnCI). In HAEnCI patients mutations have been identified in the F12, PLG, KNG1, ANGPT1, MYOF, and HS3ST6 genes. The release of bradykinin from kininogen via the kallikrein-kinin system (KKS) has been shown to be the main mediator in HAE-FXII, but for HAE-PLG there are only first indications how the PLG mutations can result in bradykinin release. Here we identified in a multi-generation HAE-PLG family an additional F12 mutation, resulting in the loss of one F12 allele. There were no differences in the clinical presentation between HAE-PLG patients with and without the additional F12 mutation, thus we concluded that the kallikrein-kinin system is bypassed in HAE-PLG. Structural modeling and in vitro assays using purified proteins confirmed the PLG mutation c.988A>G; p.K330E to be a gain of function mutation resulting in an increased bradykinin release by direct cleavage of high molecular weight kininogen (HMWK). Thus, we can provide clinical and experimental evidence that mutant plasminogen in HAE-PLG is bypassing FXII/kallikrein to generate bradykinin.