Understanding luteal maintenance during early pregnancy is of substantial biological and practical importance. Characterizing effects of early pregnancy, however, has historically been confounded by ...use of controls with potential exposure to early Prostaglandin F2-alpha (PGF) pulses or differences in Corpus Luteum (CL) age. To avoid this, the present study utilized bihourly blood sampling to ensure control CL (n = 6) were of a similar age to CL from pregnant animals (n = 5), yet without exposure to PGF pulses. Additionally, CL from second month of pregnancy (n = 4) were analyzed to track fate of altered genes after cessation of embryonic interferon tau (IFNT) secretion. The major alteration in gene expression in first month of pregnancy occurred in interferon-stimulated genes (ISGs), with immune/interferon signaling pathways enriched in three independent over-representation analyses. Most ISGs decreased during second month of pregnancy, though, surprisingly, some ISGs remained elevated in the second month even after cessation of IFNT secretion. Investigation of luteolytic genes found few altered transcripts, in contrast to previous reports, likely due to removal of controls exposed to PGF pulses. An exception to this trend was decreased expression of transcription factor NR4A1. Beyond luteolytic genes and ISGs, over representation analyses highlighted the prevalence of altered genes within the extracellular matrix and regulation of Insulin-like growth factor (IGF) availability, confirming results of other studies independent of luteolytic genes. These results support the idea that CL maintenance in early pregnancy is related to lack of PGF exposure, although potential roles for CL expression of diverse ISGs and other pathways activated during early pregnancy remain undefined. Summary sentence Comparison of Day 20 CL from pregnant and cycling animals without initiation of luteolysis found 94 mRNA transcripts with differential expression, including many ISGs, some acutely regulated and others with elevated expression into second month of pregnancy, although no major alterations in luteolytic or steroidogenic genes indicate luteal maintenance during pregnancy is not transcriptomically opposite to CL regression.
The pulsatile pattern of prostaglandin F2alpha (PGF) secretion during spontaneous luteolysis is well documented, with multiple pulses of exogenous PGF necessary to induce regression using physiologic ...concentrations of PGF. However, during spontaneous regression, the earliest pulses of PGF are small and not associated with detectable changes in circulating progesterone (P4), bringing into question what, if any, role these early, subluteolytic PGF pulses have during physiologic regression. To investigate the effect of small PGF pulses, luteal biopsies were collected throughout natural luteolysis in conjunction with bihourly blood samples to determine circulating P4 and PGF metabolite to retrospectively assign biopsies to early and later regression. Whole transcriptome analysis was conducted on CL biopsies. Early PGF pulses altered the luteal transcriptome, inducing differential expression of 210 genes (Q < 0.05) during early regression, compared with 4615 differentially expressed genes during later regression. In early regression, few of these differentially expressed genes were directly associated with luteolysis, rather there were changes in local steroid and glutathione metabolism. Most (94%) differentially expressed genes from early regression were also differentially expressed during later regression, with 98% of these continuing to be altered in the same direction compared with CL at a similar stage of the cycle that had not yet been exposed to PGF. Thus, early, subluteolytic PGF pulses impact the luteal transcriptome, though not by altering steroidogenesis or causing direct inhibition of cellular function. Rather, small pulses alter pathways resulting in the removal of cellular support systems, which may sensitize the CL to later pulses of PGF. Summary sentence Early stages of spontaneous luteolysis involve prostaglandin F2alpha pulses too small to impact steroidogenesis; nonetheless, these pulses are capable of removing cellular support systems such as local progesterone signaling and glutathione-based anti-oxidant metabolism that may sensitize the CL to later pulses of PGF.
Several recent studies have used transcriptomics to investigate luteal changes during the maternal recognition of the pregnancy period in ruminants. Although these studies have contributed to our ...understanding of luteal function during early pregnancy, few attempts have been made to integrate information across these studies and distinguish key luteal transcripts or functions that are repeatably identified across multiple studies. Therefore, in this study, two independent studies of the luteal transcriptome during early pregnancy were combined and compared. In the first study, corpora lutea (CL) from day 20 of pregnancy were compared with CL collected on day 14 of pregnancy, prior to embryonic signaling. The cattle were nonlactating. In the second study, CL from day 20 of pregnancy were compared with CL collected from day 20 cyclic cattle that had been confirmed as not yet undergoing luteal regression. These were lactating cattle. Three methods were used to compare these two datasets, to identify key luteal regulators. In the first method, all transcripts with Benjamini-Hochberg-adjusted P-value (Q value) < 0.05 in both datasets were considered. This yielded 22 transcripts, including several classical interferon-stimulated genes, as well as regulators of transforming growth factor-beta (TGFB) and latent TGFB-binding proteins (LTBP)1 and 2. In the second, less conservative method, all transcripts with P < 0.01 and changed in the same direction in both datasets were considered. This yielded an additional 20 transcripts that were not identified in the first analysis, for a total of 42 common transcripts. These transcripts were regulators of functions such as inflammatory balance and matrix remodeling. In the third method, transcripts with Q < 0.10 were subject to pathway analysis, and common pathways were identified. Retinoic acid signaling and classical interferon signaling pathways were identified with this method. Finally, regulation by interferon tau (IFNT) was investigated. Among the 42 transcripts identified, 32 were regulated by IFNT in cultured luteal cells (Q < 0.05). Among those not regulated by IFNT were LTBP1 and 2, which are TGFB-binding proteins. In summary, common transcripts from two studies of the luteal transcriptome during early pregnancy were combined and shared changes were identified. This not only generated a list of potential key luteal regulators, which were mostly IFNT regulated, but also included transcripts not regulated by IFNT, including LTBP1 and 2.
Mechanisms of bovine corpus luteum (CL) maintenance during the second month of pregnancy have not been adequately investigated, despite significant reproductive losses. In the first month, ...interferon-tau is believed to suppress oxytocin-stimulated prostaglandin F2α (PGF) production, yet there are conflicting reports of circulating PGF metabolite (PGFM). In this study, characterization of PGFM and P4 occurred through continuous bihourly blood sampling in cows undergoing CL regression (day 18–21, n = 5), and during the first (day 18–21, n = 5) and second month (day 47–61; n = 16) of pregnancy. Cattle in the second month were assigned to control (n = 8) or oxytocin treatment (n = 8; three pulses to mimic luteolysis) to evaluate if oxytocin receptors were active. All cows but one (which had elevated PGFM prior to oxytocin treatment) maintained the pregnancy. Basal PGFM concentrations were low (11.6 ± 0.7 pg/mL) in the first month but increased 2.54-fold in the second month. Few (0.26 ± 0.12 pulses/day) PGFM pulses with low peak concentrations (28.8 ± 3.1 pg/mL) were observed during the first month of pregnancy, similar to cows not undergoing regression. However, in the second month, frequency (1.10 ± 0.26 pulses/day) and peak concentration (67.2 ± 5.0 pg/mL) of PGFM pulses increased, displaying similar frequency but lower peak PGFM than seen in regression (1.44 ± 0.14 pulses/day; 134.5 ± 18.9 pg/mL). Oxytocin treatment increased likelihood of PGFM pulses post-treatment and increased peak concentration (89.7 ± 10.1 pg/mL) in cows during the second month. Thus, cows have more PGFM pulses during second than first month of pregnancy, possibly induced by endogenous oxytocin, indicating suppression of PGF production is an important mechanism for CL maintenance during first but not second month of pregnancy. Summary Sentence Low basal PGFM concentration and minimal pulsatile activity are observed in first month of pregnancy before increasing during second month of pregnancy, suggesting different mechanisms maintain the CL in the first vs second month of pregnancy.
The acquisition of dominance and ovulatory capacity was evaluated in follicles from cows that were carriers or half-sibling noncarriers of the Trio allele. Follicle size at acquisition of follicular ...dominance was determined by evaluating whether follicles ovulate after GnRH challenge (ovulatory capacity—experiment 1) and by determination of intrafollicular concentrations of estradiol and free insulin like growth factor 1 (IGF1) and relative mRNA expression of cytochrome P450 family 19 subfamily A member 1 (CYP19A1), luteinizing hormone/choriogonadotropin receptor (LHCGR), and pappalysin 1 (PAPPA, previously known as pregnancy-associated plasma protein A, pappalysin 1) in granulosa cells from follicles of different sizes (experiment 2). Ovulatory capacity developed in follicles at 8.3 mm (50% ovulatory capacity) in noncarriers but at smaller sizes (5.5 mm) in Trio carriers. Similarly, in experiment 2, follicles of Trio carriers acquired a dominant phenotype, as determined by intrafollicular estradiol and CYP19A1, LHCGR, and PAPPA mRNA expression in granulosa cells, at significantly smaller sizes but at a similar time after wave emergence. Overall, dominance/ovulatory capacity was acquired when follicles of Trio carriers were ∼30% the size (volume basis) of follicles in noncarriers. In addition, follicles in Trio carriers appear to acquire dominance in a hierarchal manner, as demonstrated by the progressively greater number of follicles with a dominant phenotype between days 2 and 4 after wave emergence. Thus, results from this study provide further support for a physiological model in which selection of multiple follicles in Trio allele carriers is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence. Summary Sentence Results support a model for selection of multiple follicles in Trio allele carriers due to acquisition of dominance at a smaller follicle size but similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence.
Maintenance of the corpus luteum (CL) during pregnancy is essential for continuing the elevated circulating progesterone (P4) that is required to maintain pregnancy. The mechanisms that protect the ...CL during early pregnancy when the non-pregnant animal would typically undergo CL regression have been extensively investigated. It is clear uterine prostaglandin F2α (PGF) causes regression of the CL in non-pregnant ruminants and that maintenance of the CL during early pregnancy is dependent upon secretion of interferon-tau (IFNT) from the elongating embryo. A number of specific mechanisms appear to be activated by IFNT. Most studies indicate that there is an inhibition of oxytocin-induced secretion of uterine PGF. There is also evidence for increased resistance to PGF action, perhaps due to secretion of PGE2 and PGE1 or direct endocrine actions of circulating IFNT. These mechanisms occur concurrently and each may help to maintain the CL during the first month of pregnancy. However, during the second month of pregnancy, IFNT is no longer secreted by the embryo. Attachment of the embryo to the uterus and subsequent placentome development have been linked to silencing of expression from the IFNT gene. In addition, there is some evidence that oxytocin responsiveness of the uterus returns during the second month of pregnancy leading to substantial basal secretion of PGF and perhaps PGF pulses. There is also no evidence that the CL during the second month of pregnancy is resistant to the actions of PGF as observed during the first month. Thus, this manuscript attempts to compare the mechanisms that maintain the CL during the first and second months of pregnancy in ruminants and provides a new, speculative, physiological model for maintenance of the CL during month two of pregnancy that is distinct from the previously-described mechanisms that maintain the CL during the first month of pregnancy.
Our purpose was to establish the prevalence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) in asymptomatic patients scheduled to receive radiation therapy and its effect on ...management decisions.
Between April 2020 and July 2020, patients without influenza-like illness symptoms at four radiation oncology departments (two academic university hospitals and two community hospitals) underwent polymerase chain reaction testing for SARS-CoV-2 before the initiation of treatment. Patients were tested either before radiation therapy simulation or after simulation but before treatment initiation. Patients tested for indications of influenza-like illness symptoms were excluded from this analysis. Management of SARS-CoV-2-positive patients was individualized based on disease site and acuity.
Over a 3-month period, a total of 385 tests were performed in 336 asymptomatic patients either before simulation (n = 75), post-simulation, before treatment (n = 230), or on-treatment (n = 49). A total of five patients tested positive for SARS-CoV-2, for a pretreatment prevalence of 1.3% (2.6% in north/central New Jersey and 0.4% in southern New Jersey/southeast Pennsylvania). The median age of positive patients was 58 years (range, 38-78 years). All positive patients were white and were relatively equally distributed with regard to sex (2 male, 3 female) and ethnicity (2 Hispanic and 3 non-Hispanic). The median Charlson comorbidity score among positive patients was five. All five patients were treated for different primary tumor sites, the large majority had advanced disease (80%), and all were treated for curative intent. The majority of positive patients were being treated with either sequential or concurrent immunosuppressive systemic therapy (80%). Initiation of treatment was delayed for 14 days with the addition of retesting for four patients, and one patient was treated without delay but with additional infectious-disease precautions.
Broad-based pretreatment asymptomatic testing of radiation oncology patients for SARS-CoV-2 is of limited value, even in a high-incidence region. Future strategies may include focused risk-stratified asymptomatic testing.
Purpose: Typical doses of 45–50.4 Gy used to treat regional nodes have demonstrated inadequate control of gross nodal disease (GND) in gynecologic cancer, and accelerated repopulation may limit the ...efficacy of a sequential boost. We reviewed outcomes of patients treated with a simultaneous integrated boost (SIB) at 2.25 Gy per fraction to positron emission tomography (PET) avid GND to evaluate toxicity and tumor control using this dose-escalated regimen.Materials and Methods: A total of 83 patients with gynecologic cancer and PET avid inguinal, pelvic, or para-aortic lymphadenopathy were treated using intensity-modulated radiation therapy (IMRT) with SIB. Primary cancers were mostly cervical (51%) and endometrial (34%), and included patients who received concurrent chemotherapy (59%) and/or brachytherapy boost (78%).Results: Median follow-up from radiation completion was 12.6 months (range, 2.7 to 92.9 months). Median dose to elective lymphatics was 50.4 Gy (range, 45 to 50.4 Gy) at 1.8 Gy/fraction. Median SIB dose and volume were 63 Gy (range, 56.3 to 63 Gy) and 72.8 mL (range, 6.8 to 1,134 mL) at 2–2.25 Gy/fraction. Nodal control was 97.6% in the SIB area while 90.4% in the low dose area (p = 0.013). SIB radiotherapy (RT) field failure-free, non-SIB RT field failure-free, and out of RT field failure-free survival at 4 years were 98%, 86%, and 51%, respectively. Acute and late grade ≥3 genitourinary toxicity rates were 0%. Acute and late grade ≥3 gastrointestinal toxicity rates were 7.2% and 12.0%, respectively.Conclusion: Dose escalated SIB to PET avid adenopathy results in excellent local control with acceptable toxicity.
The pulsatile pattern of prostaglandin F2alpha (PGF) secretion during spontaneous luteolysis is well documented, with multiple pulses of exogenous PGF necessary to induce regression using physiologic ...concentrations of PGF. However, during spontaneous regression, the earliest pulses of PGF are small and not associated with detectable changes in circulating progesterone (P4), bringing into question what, if any, role these early, subluteolytic PGF pulses have during physiologic regression. To investigate the effect of small PGF pulses, luteal biopsies were collected throughout natural luteolysis in conjunction with bihourly blood samples to determine circulating P4 and PGF metabolite to retrospectively assign biopsies to early and later regression. Whole transcriptome analysis was conducted on CL biopsies. Early PGF pulses altered the luteal transcriptome, inducing differential expression of 210 genes (Q <0.05) during early regression, compared with 4615 differentially expressed genes during later regression. In early regression, few of these differentially expressed genes were directly associated with luteolysis, rather there were changes in local steroid and glutathione metabolism. Most (94%) differentially expressed genes from early regression were also differentially expressed during later regression, with 98% of these continuing to be altered in the same direction compared with CL at a similar stage of the cycle that had not yet been exposed to PGF. Thus, early, subluteolytic PGF pulses impact the luteal transcriptome, though not by altering steroidogenesis or causing direct inhibition of cellular function. Rather, small pulses alter pathways resulting in the removal of cellular support systems, which may sensitize the CL to later pulses of PGF. Summary sentence Early stages of spontaneous luteolysis involve prostaglandin F2alpha pulses too small to impact steroidogenesis; nonetheless, these pulses are capable of removing cellular support systems such as local progesterone signaling and glutathione-based anti-oxidant metabolism that may sensitize the CL to later pulses of PGF. Key words: corpus luteum, luteolysis, bovine, transcriptomics, RNAseq.
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