Activation of trimethylation of histone 3 lysine 27 (H3K27me3) by EZH2, a component of the Polycomb repressive complex 2 (PRC2), is suggested to play a role in endometriosis. However, the mechanism ...by which this complex is dysregulated in endometriosis is not completely understood. Here, using eutopic and ectopic tissues, as well as peritoneal fluid (PF) from IRB-approved and consented patients with and without endometriosis, the expression of PRC2 complex components, JARID2, miR-155 (known regulators of EZH2), and a key inflammatory modulator, FOXP3, was measured. A higher expression of EZH2, H3K27me3, JARID2, and FOXP3 as well as miR-155 was noted in both the patient tissues and in endometrial PF treated cells. Gain-or-loss of function of miR-155 showed an effect on the PRC2 complex but had little effect on JARID2 expression, suggesting alternate pathways. Chromatin immunoprecipitation followed by qPCR showed differential expression of PRC2 complex proteins and its associated binding partners in JARID2 vs. EZH2 pull down assays. In particular, endometriotic PF treatment increased the expression of
(
= 0.0474), a gene silencer and co-factor that promotes PRC2 interaction with its targets. Thus, these studies have identified the potential novel crosstalk between miR-155-PRC2 complex-JARID2 and PHF19 in endometriosis, providing an opportunity to test other epigenetic targets in endometriosis.
Endometriosis is a chronic, painful condition with unknown etiology. A differential expression of microRNAs in the endometriotic tissues from women with endometriosis with pain compared to those ...without suggested a plausible role for miRNA or epigenetic mechanisms in the etiology of endometriotic pain. The peritoneal milieu is involved in maintenance of endometriotic lesion and nociception. We recently showed the mechanistic role for oxidized-lipoproteins (ox-LDLs) present in peritoneal fluid (PF) in endometriosis and pain. We explored the possibility of ox-LDLs modulating the expression of miRNAs in a manner similar to PF from women with endometriosis. Expression levels of miRNAs and their predicted nociceptive and inflammatory targets were determined in PF and ox-LDL treated human endometrial cell-lines. Samples from IRB-approved and consented patients with and without endometriosis or pain were used. These were compared to endometrial cell-lines treated with various forms of oxidized-lipoproteins. RNA (including miRNAs) were isolated from treated endometrial cells and expression levels were determined using commercial miRNome arrays. Cell lysates were used in immunoblotting for inflammatory proteins using a protein array. Twenty miRNAs including isoforms of miR-29, miR-181 and let-7 were mutually differentially expressed in cells treated with PF from endometriosis patients with pain and those treated with ox-LDL components. The ox-LDLs and endo-PF treatment also produced significant overexpression of microRNA predicted target genes nerve growth factor, interleukin-6 and prostaglandin E synthase and overexpression of their downstream protein targets Mip1α and MCP1. This study showed similarities between miRNA regulation in PF from endometriotic women and ox-LDLs present in abundance in the PF of these women. Key miRNAs responsible for targeting nociceptive and inflammatory molecules were downregulated in the presence of ox-LDLs and endo-PF, thus playing a role in the etiology of endometriotic pain. These redox-sensitive miRNAs can be of potential use as targets in the treatment of endometriosis-associated pain.
Perennial biofuel crops help to reduce both dependence on fossil fuels and greenhouse gas emissions while utilizing nutrients more efficiently compared to annual crops. In addition, perennial crops ...grown for biofuels have the potential to produce high biomass yields, are capable of increased carbon sequestration, and are beneficial for reducing soil erosion. Various monocultures and mixtures of perennial grasses and forbs can be established to achieve these benefits. The objective of this study was to quantify the effects of feedstock mixture and cutting height on yields. The base feedstock treatments included a monoculture of switchgrass (SG) and a switchgrass:big bluestem 1:1 mixture (SGBBS). Other treatments included mixtures of the base feedstock with ratios of base to native forbs plus legumes of 100:0, 80:20, 60:40, and 20:80. The study was established in 2008. Biomass crops typically require 2 to 3 years to produce a uniform stand. Therefore, harvest data were collected from July 2010 to July 2013. Three harvest times were selected to represent (1) biomass for biofuel (March), (2) forage (July), and (3) forage and biomass (October). Annual mean yields varied between 4.97 Mg ha−1 in 2010 to 5.56 Mg ha−1 in 2011. However, the lowest yield of 2.82 Mg ha−1 in March and the highest yield of 7.18 Mg ha−1 in July were harvested in 2013. The mean yield was 5.21 Mg ha−1 during the 4 year study. The effect of species mixture was not significant on yield. The cutting height was significant (p < 0.001), with greater yield for the 15 cm compared to the 30 cm cutting height. Yield differences were larger between harvest times during the early phase of the study. Yield difference within a harvest time was not significant for 3 of the 10 harvests. Future studies should examine changes in biomass production for mixture composition with time for selection of optimal regional specific species mixtures.
Endometriosis is a complex disorder that afflicts many women with chronic pelvic pain and infertility. It is believed that polycomb group of proteins, such as enhancer of zeste homolog 2 (EZH2), ...which plays a major role in cancer, also might contribute to the etiology of endometriosis. We and others have shown upregulation of EZH2 in human endometriotic tissues. However, the factors responsible for this upregulation is unknown. This study hypothesizes that the components of peritoneal fluid (PF) plays a dynamic role in epigenetic pathways in endometriosis. To test this hypothesis we measured EZH2 expression and activity (activation of H3K27me3) in endometrial cells. Ishikawa (human endometrial) cells were treated with 1% PF from IRB‐approved and consented women with and without endometriosis (n=6–8/group age and cycle matched). A subset of cells was also treated with various concentrations of GSK126 (5 nM to 10 μM) either before or after PF treatment. Real‐time PCR was used to analyze EZH2 expression in these treated cells. Our results showed bidirectional effects of GSK126 on EZH2 and H3k27me3 expression, depending on the order of the drug treatment (before or after 24 hours of PF treatment) and the source of the PF (control vs endo). When cells were treated with endo PF, there was a 2–3 fold induction in EZH2 expression when cells were treated with endo PF compared to the media control. Surprisingly, there was a dose dependent increase in EZH2 mRNA expression with GSK126 pre‐treatment followed by the PF treatments. Almost a 9‐fold increase in EZH2 expression was observed when cells were pre‐treated with 2.5 μM GSK126 followed by control PF (*p<0.05), whereas pretreatment of GSK followed by endo PF had no such effects on EZH2 expression. However, when cells were pre‐treated with PF followed by the drug the EZH2 expression levels went down. Western blot of EZH2 and H3K27me3 showed similar findings. When MTT assay was performed to examine cell viability under the above experimental conditions, no significant changes in viability was observed at the 48 or 96 hour time points. These results suggest for an intermediate pathway modulated by the PF components that might be at play. The current studies are investigating the possible role for ARID1A mutation in the bidirectional effect of the EZH2 inhibitors in endometriosis.
Support or Funding Information
PhRMA Foundation Pre Doctoral Fellowship in Pharmacology/Toxicology
This is from the Experimental Biology 2019 Meeting. There is no full text article associated with this published in The FASEB Journal.
Objective: To determine if sperm attachment to oviduct epithelial cells (OEC) in vitro is selective for higher quality sperm and if the system requires homologous species OEC.
Design: Controlled ...prospective study with outcomes assayed by a technician blind to sperm treatment groups.
Setting: An academic research laboratory.
Patient(s): Experiment 1: normospermic donors with children (4 donors, 7 ejaculates). Experiment 2: cryopreserved donor samples (4 donors).
Intervention(s): Semen collection by masturbation after 48 hours of abstinence.
Main Outcome Measure(s): Experiment 1: sperm assays of motility, morphology, membrane integrity, and capacitation status. Experiment 2: sperm chromatin (DNA) integrity and condensation.
Result(s): Experiment 1: sperm not attaching to OEC had lower motility, more membrane disruptions, and more acrosome reactions than did control sperm. This selectivity was equivalent for sperm in coculture with all OEC types. Experiment 2: sperm attached to OEC had fewer abnormalities in chromatin structure compared with sperm that were not attached.
Conclusion(s): Selective attachment of functionally superior sperm to OEC is likely important during sperm reservoir formation in vivo and may be exploitable in vitro as a method to isolate high-quality sperm for clinical procedures. Such a system does not require human origin OEC.
Rotating equipment is widespread in the process industry, where pumps, compressors, and turbines are used to drive continuous manufacturing lines. This class of machinery is meant to run without ...interruption, but invariably experiences degradation that can lead to equipment failure. Any break in a continuous manufacturing line can halt production, so there is a pressing need to diagnose rotating equipment health before failure occurs. Existing health modeling and diagnosis strategies require supplemental tests to collect model training data, and ignore time-series behavior in machine signals that can be useful for diagnosing equipment health. This letter presents a general modeling structure to give context to historical data, which can act as a substitute for supplemental test data, and describes a methodology for assessing repair quality based on trends in signal features. A case study that uses the proposed methodology to assess the quality of repair procedures is provided.
The influence of beef source and cooking method on neutral (NL) and polar (PL) lipid contents, fatty acid profiles, and cholesterol contents of Wagyu and domestic sources of beef was determined. ...Longissimus dorsi muscle from Japanese Wagyu, American Wagyu, Longhorn, Angus, and U.S. Choice was boiled or roasted. Beef from Wagyu breeds had a significantly higher (P 0.05) NL content than that of domestic sources. The NL from Japanese Wagyu samples was lower in saturated fatty acid content and higher in monounsaturated fatty acid content than samples from other beef sources. The PL from the Japanese Wagyu had the lowest content of saturated fatty acids, while the PL from the American Wagyu had the highest content of monounsaturated fatty acids and the lowest content of polyunsaturated fatty acids. Cholesterol content was highest for the Wagyu breeds. The contents of total NL and individual fatty acids were significantly (P 0.05) higher in the roasted beef than boiled beef. Cooking method did not have a significant effect on the PL content, PL fatty acid profiles, and cholesterol content
The influence of beef source, cooking method, and refrigerated storage time on the contents of volatile lipid oxidation products of Wagyu and domestic sources of beef was determined. Longissimus ...dorsi muscle from Japanese Wagyu, American Wagyu, Longhorn, Angus, and U.S. Choice carcasses was boiled or roasted and stored for 0 or 3 days at refrigerator temperatures. With storage time, the contents of lipid oxidation products increased significantly (P < 0.05). Interactions between beef source and storage time were significant (P < 0.05) for hexanal, 2-pentylfuran, octanal, nonanal, and other major lipid oxidation products. Beef source was shown to have a significant effect on the content of these compounds following 3 days of storage but not immediately following cooking (0 day). The contents of lipid oxidation products were higher in the Japanese and American Wagyu breeds than in other domestic beef sources. In general, cooking method had only minor influences on the content of lipid oxidation products. Keywords: Flavor; beef; breeds; lipid oxidation; Wagyu
Conical diffusers are used in hundreds of engineering applications in various industries. Some of the operating conditions that they operate under cause swirling flow to enter the diffuser. It is ...generally well documented that the addition of swirl to the flow of a diffuser allows for greater divergence angles without wall separation, resulting in better overall performance of the diffuser and the machine it’s attached to. It is also known that as swirl strength is increased, the flow will eventually breakdown, resulting in internal flow recirculation and decreased diffuser performance. However, the relationship between the diffuser geometry and its performance at these higher swirl strengths has not been investigated in detail. This link between diffuser geometry, swirl, and performance is investigated using a hybrid RANS-LES based computational model. A series of simulations are performed with the computational model, varying the swirl strength and diffuser half angle φ. Overall, there was found to be little relationship between adjusting the diffuser geometry and diffuser performance at high swirl numbers.
Endometriosis is an enigmatic disease that is associated with severe symptoms and consequences such as total hysterectomy. Upwards of 10% of women of reproductive age have endometriosis, most of whom ...are afflicted with chronic pelvic pain and/or infertility. Despite many treatment options available to endometriosis patients, there is no cure for this condition and recurrence of symptoms is extremely common. In CHAPTER 1, we describe the mechanisms involved in endometriotic pain and the currently available treatment strategies. Our laboratory has provided evidence for the role of oxidative stress in the etiology of endometriosis and its associated pain. In this dissertation research, we wanted to specifically understand the mechanistic role for the abundance of oxidized lipoproteins (ox-LDLs) in the peritoneal fluid (PF) of women with endometriosis. We hypothesize that the ox-LDLs and the peritoneal milieu play a dynamic role in endometriosis. These components are epigenetic modulators of inflammatory and nociceptive processes. Our IRB-approved study used eutopic and ectopic endometrial tissues and peritoneal fluid from patients with (endo) and without (control) endometriosis (IRB-114954-20(9074)). Tissue sources included eutopic endometrium of control patients (EuNN), eutopic endometrium from patients with endometriosis (EuYY), and ectopic tissue from women with endometriosis (EcYY). Peritoneal fluid (PF), was collected from patients categorized as having neither endometriosis nor pain (NN-PF), endometriosis and pain (YY-PF), or endometriosis without pain (YN-PF). In CHAPTER 2, we provide evidence for the ability of the ox-LDL components of PF to induce inflammation and nociception in Sprague-Dawley rats (IACUC 485397498-7). The ox-LDL nociceptive and inflammatory responses were similar to that observed with PF from women with endometriosis and pain. Antioxidants were able to alleviate these nociceptive responses. In CHAPTER 3, we provide a mechanism by which these ox-LDL components induce inflammatory and nociceptive responses. Global miRNA expression was measured in tissues and PF-treated cells and key miRNAs and their targets were validated by qPCR. Several miRNAs (e.g. isoforms of let-7, miR-98, and miR-374) and their gene targets (e.g. IL-10, Mip1α?, and MCP1) were differentially expressed in both ox-LDL and endo PF-treated cells. In CHAPTER 4, an epigenetic mechanism involving Enhancer of zeste 2 (EZH2) is proposed to contribute to endometriosis and associated pain. RT-qPCR and Western blots were used to measure the expression of key epigenetic factors in patient tissues and PF-treated cells while ChIP-qPCR identified interactions between upstream regulator Jumonji protein 2 and epigenetic genes. This study provides mechanistic evidence for oxidized lipoprotein components playing a role in endometriosis associated pain. We also provide evidence for epigenetic changes in the endometriosis pain. Future studies will test drugs that target oxidation and/or epigenetic pathways in animal models of endometriosis and patients with endometriosis.
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