RIP-Seq analysis of non-PPR chloroplast editing factors reveals broad RNA interactions and enrichment of less efficiently translated RNAs by OZ1 and ORRM1 complexes.

C-to-U RNA editing in angiosperm chloroplasts requires a large suite of proteins bound together in the editosome. The editosome is comprised of PPR proteins, RIP/MORFs, OZ proteins, and ORRM proteins that physically interact in high molecular weight complexes. The specific functions of non-PPR editing factors in the editosome are unclear, however, specific subsets of editing sites are affected by absence of non-PPR editing factors. Unlike the PPR components of editosomes that have predictable nucleotide specificities, domains present in non-PPR editing factors make RNA associations difficult to predict. In this study, chloroplast extracts were isolated from juvenile maize seedlings. RNAs were immunoprecipitated using polyclonal antibodies targeting non-PPR editing factors RIP9, OZ1, and ORRM1. RNA libraries from duplicate experiments were compared. RIP9 was associated with most of the non-ribosomal RNA content of chloroplasts, consistent with a general binding function to PPR L-motifs and tethering of large ribonucleoprotein complexes. The breadth of RNA associations was greater than predicted and include mRNAs without predicted editing sites, tRNA sequences, and introns. OZ1 and ORRM1 were associated with a highly similar pool of RNAs that have a bias toward lower translational efficiency values in mature chloroplasts. Lower translational efficiency was also associated with the pool of edited RNAs compared to RNAs without editing sites. The unexpected breadth of interactions by non-PPR editing factors suggests the editosome is large, diverse, and associated with RNAs with lower relative translational efficiency in mature chloroplasts.

Prediction Model Assessing Risk of Delivery after Diagnosis of Abnormal Umbilical Artery Doppler.

OBJECTIVE: Umbilical artery Doppler (UAD) velocimetry abnormalities are associated with increased neonatal morbidity and mortality. Currently, there are no risk stratification methods to assist in antepartum management such as timing of antenatal corticosteroids (ACS). Therefore, we sought to develop a model to predict risk of delivery within 7 days following diagnosis of abnormal UAD velocimetry in patients with fetal growth restriction (FGR). STUDY DESIGN: Retrospective single referral center study of liveborn singleton pregnancies complicated by FGR and ≥1 abnormal UAD velocimetry value (≥95th percentile for gestational age [GA]). We considered 17 variables and used backward stepwise logistic regression to create a multivariable model for the prediction of delivery within 7 days. We assessed model fit with calibration, discrimination, likelihood ratios, and area under the curve. Internal validation of the model was assessed by using the bootstrap method. RESULTS: Between 2008 and 2015, a total of 176 patients were eligible and included for model development. Median (range) GA at initial eligibility was 32.1 weeks (28.1-36.1 weeks) and from initial eligibility until delivery was 21 days (0-104 days). Fifty-two patients (30%) were delivered in the 7 days following inclusion. GA at first abnormal UAD, severity of first abnormal UAD, oligohydramnios, preeclampsia, and pre-pregnancy BMI were included in the model. The model had an area under the ROC curve of 0.94 (95% confidence interval [CI]: 0.90-0.98), sensitivity of 85%, and specificity of 91%. If the model alone were used for ACS timing, 85% of the cohort who delivered in the following week would have received ACS, and ACS would not have been given to 91% who delivered later. Internal validation yielded similar results with a mean area under the curve (95% CI) of 0.94 (0.88-0.98). CONCLUSION: If validated externally, our model can be used to predict risk of delivery in patients with FGR and abnormal UAD velocimetry, potentially improving timing of ACS. KEY POINTS: · Risk of delivery in seven days can be predicted.. · Risk of delivery can inform corticosteroid timing.. · External validation can further develop a clinical aid..

External Validation of a Prediction Model Assessing Risk of Delivery in Fetuses with Growth Restriction after Diagnosis of Abnormal Umbilical Artery Doppler.

OBJECTIVE: We have previously described a model using maternal, antenatal, and ultrasonographic characteristics to assess the risk of delivery within 7 days following diagnosis of abnormal umbilical artery Doppler (UAD) in pregnancies affected by fetal growth restriction (FGR). Therefore, we sought to validate this model in an independent cohort. STUDY DESIGN: Retrospective, single referral center study of liveborn singleton pregnancies from 2016 to 2019 complicated by FGR and abnormal UAD (systolic/diastolic ratio ≥95th percentile for gestational age [GA]). Prediction probabilities were calculated by applying the original model (Model 1) to the current cohort (Brigham and Women's Hospital [BWH] cohort). The variables of this model include GA at first abnormal UAD, severity of first abnormal UAD, oligohydramnios, preeclampsia, and prepregnancy body mass index. Model fit was assessed with area under the curve (AUC). Two alternative models (Models 2 and 3) were created to identify a model with better predictive characteristics than Model 1. The receiver operating characteristics curves were compared using the DeLong test. RESULTS: A total of 306 patients were assessed for eligibility, 223 of whom were included in the BWH cohort. Median GA at eligibility was 31.3 weeks, and median interval from eligibility to delivery was 17 days (interquartile range: 3.5-33.5). Eighty-two (37%) patients delivered within 7 days of eligibility. Applying Model 1 to the BWH cohort resulted in an AUC of 0.865. Using the previously determined probability cutoff of 0.493, the model was 62% sensitive and 90% specific in predicting the primary outcome in this independent cohort. Models 2 and 3 did not perform better than Model 1 (p = 0.459). CONCLUSION: A previously described prediction model to predict risk of delivery in patients with FGR and abnormal UAD performed well in an independent cohort. With high specificity, this model could assist in identifying low-risk patients and improve antenatal corticosteroid timing. KEY POINTS: · Risk of delivery in 7 days can be predicted.. · Risk of delivery can inform corticosteroid timing.. · An externally validated clinical aid can be developed..

Association between Respiratory Morbidity and Labor in Pregnancies with Gestational Diabetes Mellitus.

OBJECTIVE: This study aimed to the assess risk of respiratory morbidity in neonates born to women with gestational diabetes mellitus (GDM) delivered after labor compared with those delivered without exposure to labor. STUDY DESIGN: This is a secondary analysis of a prospective single-center cohort study of singleton pregnancies complicated by GDM. Neonates who were liveborn and delivered at ≥34 weeks' gestation were included. The primary outcome was respiratory morbidity defined as respiratory distress syndrome (RDS) or transient tachypnea of the newborn (TTN) resulting in neonatal intensive care unit (NICU) admission. Neonates born after labor (either spontaneous or induced) were compared with those delivered by cesarean delivery without labor. Associations between labor and neonatal morbidities were estimated using logistic regression. Covariates were adjusted for if they differed significantly between neonates exposed to and not exposed to labor (p < 0.05) and there was biologic plausibility that they would affect neonatal respiratory morbidity. RESULTS: Of the 581 neonates meeting study inclusion criteria, 23.2% delivered without exposure to labor. Those who did and did not experience labor delivered at similar gestational ages (38.6 vs. 38.4 weeks). Thirty-six neonates (6.2%) developed RDS or TTN and were admitted to the NICU. Exposure to labor was associated with a lower frequency of respiratory morbidity requiring admission to NICU, 4.9% (22/446) versus 10.4% (14/135) (p = 0.04). After adjusting for parity, body mass index, birth weight, gestational weight gain more than Institute of Medicine guidelines, race, and exposure to labor were associated with an adjusted odds ratio of 0.41 (95% confidence interval: 0.18-0.89). CONCLUSION: Exposure to labor was associated with decreased odds of respiratory morbidity in neonates born to mothers with GDM. Limiting elective cesarean in this population can reduce health care costs and optimize neonatal health. KEY POINTS: · Labor is associated with less respiratory morbidity.. · We should limit elective cesarean delivery with GDM.. · This approach could reduce health care costs..

Myosin-binding protein C displaces tropomyosin to activate cardiac thin filaments and governs their speed by an independent mechanism.

Myosin-binding protein C (MyBP-C) is an accessory protein of striated muscle thick filaments and a modulator of cardiac muscle contraction. Defects in the cardiac isoform, cMyBP-C, cause heart disease. cMyBP-C includes 11 Ig- and fibronectin-like domains and a cMyBP-C-specific motif. In vitro studies show that in addition to binding to the thick filament via its C-terminal region, cMyBP-C can also interact with actin via its N-terminal domains, modulating thin filament motility. Structural observations of F-actin decorated with N-terminal fragments of cMyBP-C suggest that cMyBP-C binds to actin close to the low Ca(2+) binding site of tropomyosin. This suggests that cMyBP-C might modulate thin filament activity by interfering with tropomyosin regulatory movements on actin. To determine directly whether cMyBP-C binding affects tropomyosin position, we have used electron microscopy and in vitro motility assays to study the structural and functional effects of N-terminal fragments binding to thin filaments. 3D reconstructions suggest that under low Ca(2+) conditions, cMyBP-C displaces tropomyosin toward its high Ca(2+) position, and that this movement corresponds to thin filament activation in the motility assay. At high Ca(2+), cMyBP-C had little effect on tropomyosin position and caused slowing of thin filament sliding. Unexpectedly, a shorter N-terminal fragment did not displace tropomyosin or activate the thin filament at low Ca(2+) but slowed thin filament sliding as much as the larger fragments. These results suggest that cMyBP-C may both modulate thin filament activity, by physically displacing tropomyosin from its low Ca(2+) position on actin, and govern contractile speed by an independent molecular mechanism.

microRNA-34c is a novel target to treat dementias.

MicroRNAs are key regulators of transcriptome plasticity and have been implicated with the pathogenesis of brain diseases. Here, we employed massive parallel sequencing and provide, at an unprecedented depth, the complete and quantitative miRNAome of the mouse hippocampus, the prime target of neurodegenerative diseases such as Alzheimer's disease (AD). Using integrative genetics, we identify miR-34c as a negative constraint of memory consolidation and show that miR-34c levels are elevated in the hippocampus of AD patients and corresponding mouse models. In line with this, targeting miR-34 seed rescues learning ability in these mouse models. Our data suggest that miR-34c could be a marker for the onset of cognitive disturbances linked to AD and indicate that targeting miR-34c could be a suitable therapy.

Insulin growth factor binding protein 7 is a novel target to treat dementia.

Alzheimer's disease (AD) is the most common form of dementia in the elderly but effective therapeutic strategies to treat AD are not yet available. This is also due to the fact that the pathological mechanisms that drive the pathogenesis of sporadic AD are still not sufficiently understood and may differ on the individual level. Several risk factors such as altered insulin-like peptide (ILP) signaling have been linked to AD and modulating the ILP system has been discussed as a potential therapeutic avenue. Here we show that insulin-like growth factor binding protein 7 (IGFBP7), a protein that attenuates the function of ILPs, is up-regulated in the brains of AD patients and in a mouse model for AD via a process that involves altered DNA-methylation and coincides with decreased ILP signaling. Mimicking the AD-situation in wild type mice, by increasing hippocampal IGFBP7 levels leads to impaired memory consolidation. Consistently, inhibiting IGFBP7 function in mice that develop AD-like memory impairment reinstates associative learning behavior. These data suggest that IGFBP7 is a critical regulator of memory consolidation and might be used as a biomarker for AD. Targeting IGFBP7 could be a novel therapeutic avenue for the treatment of AD patients.

Placenta Accreta Spectrum: Evaluation of classic and non-classic presentations, pathologic grading, and uterine scar dehiscence features in a modern institutional series.

INTRODUCTION: The aim of this study is to document the distribution of classic versus non-classic presentation of Placenta Accreta Spectrum (PAS) disorders as well as grading categories by the Society for Pediatric Pathology (SPP) and FIGO systems in an institutional cohort of gravid hysterectomies. We also document the prevalence of uterine scar as a histologic correlate for uterine scar dehiscence, a phenomenon raised by some as central to PAS pathogenesis. METHODS: PAS cases were assigned grade and designated as classic (anterior lower uterine segment implantation, prior C-section) or non-classic (implantation away from anterior lower uterine segment and/or no prior C-section). Features of dehiscence (uterine window, histologic evidence of scar) were recorded. RESULTS: Sixty-two patients were included: 76 % had prior C-section; 55 % had other forms of uterine instrumentation. Classic PAS was recorded in 52 % patients; notably, 48 % had non-classic presentation; of these, all but one had prior instrumentation (curettage, myomectomy, laparoscopy). Uterine window was described in 53 % classic and 23 % non-classic PAS. Scar was demonstrated in 31 % classic and 23 % non-classic PAS; trichrome/reticulin stains were confirmatory. 32 % cases were SPP grade 1, 18 % grade 2, 18 % grade 3a and 32 % grade 3d. Grade 3 was significantly more common in classic (72 %) than non-classic (27 %) PAS. DISCUSSION: While most PAS patients have classic presentation, a large subset does not; in addition, scar tissue is not identified histologically in most PAS hysterectomies; in these settings, PAS cannot be fully attributed to scar dehiscence. Uterine instrumentation often precedes non-classic PAS reinforcing the concept of decidual disruption as central to PAS pathogenesis. PAS grading as defined correlates with presentation (classic vs non-classic).

XRN1 deletion induces PKR-dependent cell lethality in interferon-activated cancer cells.

Emerging data suggest that induction of viral mimicry responses through activation of double-stranded RNA (dsRNA) sensors in cancer cells is a promising therapeutic strategy. One approach to induce viral mimicry is to target molecular regulators of dsRNA sensing pathways. Here, we show that the exoribonuclease XRN1 is a negative regulator of the dsRNA sensor protein kinase R (PKR) in cancer cells with high interferon-stimulated gene expression. XRN1 deletion causes PKR pathway activation and consequent cancer cell lethality. Disruption of interferon signaling with the JAK1/2 inhibitor ruxolitinib can decrease cellular PKR levels and rescue sensitivity to XRN1 deletion. Conversely, interferon-ß stimulation can increase PKR levels and induce sensitivity to XRN1 inactivation. Lastly, XRN1 deletion causes accumulation of endogenous complementary sense/anti-sense RNAs, which may represent candidate PKR ligands. Our data demonstrate how XRN1 regulates PKR and how this interaction creates a vulnerability in cancer cells with an activated interferon cell state.

Follicular fluid hydrogen peroxide and lipid hydroperoxide in bovine antral follicles of various size, atresia, and dominance status.

PURPOSE: To avoid inducing a state of oxidative stress (OS), assisted reproductive technologies (ART) must maintain a balance of reactive oxygen species (ROS) and antioxidants during the in vitro culture of oocytes. However, oocyte requirements and tolerance thresholds for ROS during in vivo development are still unclear. Previous studies have examined ROS levels in follicular fluid (FF) using pooled samples or according to follicle size. This study sought to examine two OS markers, lipid hydroperoxides (LPO) and hydrogen peroxide (H2O2), in FF of individually sampled follicles from bovine ovary pairs according to follicle size, atresia, and dominance status. METHODS: TUNEL and cleaved Caspase-3 labeling were used to identify apoptotic granulosa cells and determine follicle atresia status. LPO were measured directly for the first time in FF. RESULTS: Non-atretic follicles and dominant follicles contained more FF H2O2 than atretic follicles and corresponding subordinate follicles, respectively. FF LPO did not vary in relation to atretic status, and no difference existed between dominant and subordinate follicles. However, FF LPO was significantly lower in first subordinate follicles than in the second subordinate follicles from each ovary pair. Neither H2O2 nor LPO levels correlated with follicle size. CONCLUSIONS: These data provide clear evidence that the events of antral folliculogenesis are relevant to ROS dynamics in vivo. Furthermore, such studies will help to optimize in vitro conditions for oocyte culture protocols, particularly when combined with a comparison of oocyte quality with respect to source follicle characteristics.

XRN1 deletion induces PKR-dependent cell lethality in interferon-activated cancer cells.

Emerging data suggest that induction of viral mimicry responses through activation of double-stranded RNA (dsRNA) sensors in cancer cells is a promising therapeutic strategy. One approach to induce viral mimicry is to target molecular regulators of dsRNA sensing pathways. Here, we show that the exoribonuclease XRN1 is a negative regulator of the dsRNA sensor protein kinase R (PKR) in cancer cells with high interferon-stimulated gene (ISG) expression. XRN1 deletion causes PKR activation and consequent cancer cell lethality. Disruption of interferon signaling with the JAK1/2 inhibitor ruxolitinib can decrease cellular PKR levels and rescue sensitivity to XRN1 deletion. Conversely, interferon-ß stimulation can increase PKR levels and induce sensitivity to XRN1 inactivation. Lastly, XRN1 deletion causes accumulation of endogenous complementary sense/anti-sense RNAs, which may represent candidate PKR ligands. Our data demonstrate how XRN1 regulates PKR and nominate XRN1 as a potential therapeutic target in cancer cells with an activated interferon cell state.

Circulating microparticle proteins predict pregnancies complicated by placenta accreta spectrum.

Placenta accreta spectrum (PAS) is characterized by abnormal attachment of the placenta to the uterus, and attempts at placental delivery can lead to catastrophic maternal hemorrhage and death. Multidisciplinary delivery planning can significantly improve outcomes; however, current diagnostics are lacking as approximately half of pregnancies with PAS are undiagnosed prior to delivery. This is a nested case-control study of 35 cases and 70 controls with the primary objective of identifying circulating microparticle (CMP) protein panels that identify pregnancies complicated by PAS. Size exclusion chromatography and liquid chromatography with tandem mass spectrometry were used for CMP protein isolation and identification, respectively. A two-step iterative workflow was used to establish putative panels. Using plasma sampled at a median of 26 weeks' gestation, five CMP proteins distinguished PAS from controls with a mean area under the curve (AUC) of 0.83. For a separate sample taken at a median of 35 weeks' gestation, the mean AUC was 0.78. In the second trimester, canonical pathway analyses demonstrate over-representation of processes related to iron homeostasis and erythropoietin signaling. In the third trimester, these analyses revealed abnormal immune function. CMP proteins classify PAS well prior to delivery and have potential to significantly reduce maternal morbidity and mortality.

Peak performance and cardiometabolic responses of modern US army soldiers during heavy, fatiguing vest-borne load carriage.

INTRODUCTION: Physiological limits imposed by vest-borne loads must be defined for optimal performance monitoring of the modern dismounted warfighter. PURPOSE: To evaluate how weighted vests affect locomotion economy and relative cardiometabolic strain during military load carriage while identifying key physiological predictors of exhaustion limits. METHODS: Fifteen US Army soldiers (4 women, 11 men; age, 26 ± 8 years; height, 173 ± 10 cm; body mass (BM), 79 ± 16 kg) performed four incremental walking tests with different vest loads (0, 22, 44, or 66% BM). We examined the effects of vest-borne loading on peak walking speed, the physiological costs of transport, and relative work intensity. We then sought to determine which of the cardiometabolic indicators (oxygen uptake, heart rate, respiration rate) was most predictive of task failure. RESULTS: Peak walking speed significantly decreased with successively heavier vest loads (p < 0.01). Physiological costs per kilometer walked were significantly higher with added vest loads for each measure (p < 0.05). Relative oxygen uptake and heart rate were significantly higher during the loaded trials than the 0% BM trial (p < 0.01) yet not different from one another (p > 0.07). Conversely, respiration rate was significantly higher with the heavier load in every comparison (p < 0.01). Probability modeling revealed heart rate as the best predictor of task failure (marginal R2, 0.587, conditional R2, 0.791). CONCLUSION: Heavy vest-borne loads cause exceptional losses in performance capabilities and increased physiological strain during walking. Heart rate provides a useful non-invasive indicator of relative intensity and task failure during military load carriage.

Fluctuations in total antioxidant capacity, catalase activity and hydrogen peroxide levels of follicular fluid during bovine folliculogenesis.

Follicular fluid is an important environment for oocyte development, yet current knowledge regarding its in vivo oxidant and antioxidant levels remains limited. Examining follicular fluid oxidants and antioxidants will improve understanding of their changes in vivo and contribute to optimisation of in vitro maturation conditions. The aim of the present study was to consider selected markers, namely catalase (CAT) enzyme activity, total antioxidant capacity (TAC) and hydrogen peroxide (H(2)O(2)) in follicular fluid samples (n = 503) originating from bovine antral follicles. The dynamic changes in two relevant antioxidant measures and one reactive oxygen species (ROS) were measured through stages of bovine follicular development and the oestrous cycle. CAT activity and H(2)O(2) levels decreased significantly as follicle size increased, whereas TAC increased significantly as follicle size increased. Lower TAC and higher H(2)O(2) in small follicles suggest increased ROS in the initial stages of folliculogenesis. Because CAT levels are highest in the follicular fluid of small follicles in the setting of an overall low TAC, CAT may represent a dominant antioxidant defence in the initial stages of folliculogenesis. Future studies must focus on other reactive oxygen species and their various scavenger types during antral folliculogenesis.

Hypotensive and Antihypertensive Properties and Safety for Use of Annona muricata and Persea americana and Their Combination Products.

INTRODUCTION: In the management of hypertension (a cardiovascular disease and the leading metabolic risk factor in noncommunicable diseases) with herbal medicines, efficacy and safety are of uttermost concern. This study sought to establish hypotensive, antihypertensive, drug interaction, and safety for use of the aqueous leaf extracts of Annona muricata (AME), Persea americana (PAE), or their combination products (CAPE). Methodology. Systolic and diastolic blood pressure (SBP and DBP), mean arterial blood pressure (MAP), and heart rate (HR) were measured in normotensive Sprague-Dawley rats treated with 50-150 mg/kg of AME, PAE, or CAPE to establish a hypotensive effect. "Combination index" was calculated to establish interaction between AME and PAE. The antihypertensive effect of CAPE was established by measuring SBP, DBP, MAP, and HR in ethanol-sucrose- and epinephrine-induced hypertension. Full blood count, liver and kidney function tests, and urinalysis were determined in ethanol/sucrose-induced hypertension to establish safety for use. RESULTS: AME, PAE, and CAPE significantly (p ≤ 0.001) decreased BP in both normotensive and hypertensive animals. Effects of CAPE 1, CAPE 2, and CAPE 3 were synergistic (combination indices of 0.65 ± 0.07, 0.76 ± 0.09, and 0.87 ± 0.07, respectively). There was a significant decrease (p ≤ 0.01 - 0.001) in SBP and MAP with 100 mg/kg CAPE 1 and 75 mg/kg CAPE 2 treatment in hypertension as well as with nifedipine (p ≤ 0.001) treatment. Epinephrine-induced hypertension in anesthetized cats was significantly and dose-dependently inhibited (p < 0.05 - 0.001) by 25-100 mg/ml CAPE 1 and 37.5-75 mg/ml CAPE 2. CAPE administration had no deleterious effect (p > 0.05) on full blood count, liver and kidney function, and urine composition in hypertensive rats. CONCLUSION: The aqueous leaf extracts of Annona muricata, Persea americana, and their combination products possess antihypertensive properties, with combination products showing synergism and safety with use.

Feasibility of a Regional Approach to an Obstetrics and Gynecology Transition to Residency Course.

OBJECTIVE: To demonstrate the feasibility of a regional Obstetrics and Gynecology (Ob/Gyn) Transition to Residency Course (TRC) through compliance, satisfaction, and sustainability. METHODS: We implemented a two-week, multi-institutional regional TRC (RTRC) for fourth-year medical students matched in Ob/Gyn or Family Medicine from four New England medical schools. Curriculum was developed to meet Ob/Gyn Milestone One (M1) and Core Entrustable Professional Activity (CEPA) objectives. Compliance, satisfaction, and sustainability were identified as feasibility outcomes. RESULTS: From 2015-2018, a total of 63 fourth-year students have participated. The number of students remained stable each year. All students attended 100% of sessions. There was an average of >9/10 in all satisfaction metrics all four years. The number of faculty members from each institution remained stable over the four years. CONCLUSION: A RTRC is feasible as measured through compliance, satisfaction and sustainability.

The diphenylpyrazole compound anle138b blocks Aß channels and rescues disease phenotypes in a mouse model for amyloid pathology.

Alzheimer's disease is a devastating neurodegenerative disease eventually leading to dementia. An effective treatment does not yet exist. Here we show that oral application of the compound anle138b restores hippocampal synaptic and transcriptional plasticity as well as spatial memory in a mouse model for Alzheimer's disease, when given orally before or after the onset of pathology. At the mechanistic level, we provide evidence that anle138b blocks the activity of conducting Aß pores without changing the membrane embedded Aß-oligomer structure. In conclusion, our data suggest that anle138b is a novel and promising compound to treat AD-related pathology that should be investigated further.