Prostaglandin E2 Concentrations Research Articles

The Essential Role of PGF2α/PTGFR in Molding Endometrial Breakdown and Vascular Dynamics, Regulated by HIF-1α in a Mouse Menstrual-like Model.

In women of childbearing age, extensive decidualization, shedding and remodeling of the endometrium during the menstrual cycle are fundamental for successful pregnancy. The role of prostaglandins (PGs) in menstruation has long been proposed in humans, and the rate-limiting enzyme cyclooxygenase was shown to play a key role in endometrial breakdown and shedding in a mouse menstrual-like model in our previous study. However, the specific types of PGs involved and their respective roles remain unclear. Therefore, our objective was to investigate the mechanism through which PGs regulate endometrial disintegration. In this study, the microscopy was observed by HE; the protein levels of prostaglandins E1 (PGE1), prostaglandins E2 (PGE2), prostaglandin F2α (PGF2α) and Prostaglandin I2 (PGI2) were detected by ELISA; the mRNA level of Pfgfr2, Vascular Endothelial Growth Factor(Vegf), Angiostatin and Hypoxia inducible factor-1α (Hif1α) were examined by real-time PCR; PTGFR Receptor (PTGFR), VEGF, Angiostatin and HIF-1α protein levels were investigated by western blotting; the locations of protein were observed by Immunohistochemistry; HIF-1α binding PTGFR promoter was detected by Chromatin Immunoprecipitation (ChIP) and real-time PCR. We found that the concentrations of PGE1, PGE2, and PGF2α all increased significantly during this process. Furthermore, Ptgfr mRNA increased soon after Progesterone (P4) withdrawal, and PTGFR protein levels increased significantly during abundant endometrial breakdown and shedding processes. PTGFR inhibitors AL8810 significantly suppressed endometrial breakdown and shedding, promoted Angiostatin expression, and reduced VEGF-A expressions and vascular permeability. And HIF-1α and PTGFR were mainly located in the luminal/gland epithelium, vascular endothelium, and pre-decidual zone. Interestingly, HIF-1α directly bound to Ptgfr promoter. Moreover, a HIF-1α inhibitor 2-methoxyestradiol (2ME) significantly reduced PTGFR expression and suppressed endometrial breakdown which was in accord with PTGFR inhibitor's effect. Similar changes occurred in human stromal cells relevant to menstruation in vitro. Our study provides evidence that PGF2α/PTGFR plays a vital role in endometrial breakdown via vascular changes that are regulated by HIF-1α during menstruation.

Abstract 178: Adipose tissue upregulates expression of RANKL in B-cell acute lymphoblastic leukemia through prostaglandin E2 signaling

Abstract Obesity increases the risk of developing cancer and worsens treatment outcome. Our lab has shown that adipose tissue contributes to B-cell acute lymphoblastic leukemia (B-ALL) resistance to chemotherapy both in vitro and in vivo. However, the mechanisms by which adipose tissue causes ALL cell resistance are complex and not completely understood. We therefore implanted obese mice (n=3) with syngeneic BCR/ABL B-ALL cells and performed single-cell RNA sequencing (scRNAseq) of ALL collected from epididymal adipose tissue vs. bone marrow. Our data uncovered a strong adipose microenvironment effect on the ALL transcriptome, identifying 577 differentially expressed genes using a log2 fold threshold of 0.25 and Bonferroni corrected p-values <0.05. Notably, we found that ALL cells in adipose tissue upregulate expression of Tnfsf11, the gene that encodes receptor activator of nuclear factor kappa beta ligand (RANKL). 93% of ALL cells in adipose tissue express RANKL compared to 33% in marrow, along with a log2 fold expression increase of 2.12 (p<1 × 10−100, Wilcoxon test). Because ALL expression of RANKL has been linked to bone resorption and CNS invasion, both of which have increased incidence in obese patients, we elected to explore this further. Ex vivo culture with human visceral adipose tissue validated ALL cell RANKL upregulation, demonstrating a 1.63 and 17.5-fold increase in gene expression after 48 hours in BV173 and RS4;11 B-ALL cell lines, respectively (n=4-6, both p<0.01, paired t test). Soluble RANKL was not detected in media from ALL cells cultured alone, detectable in media conditioned by human adipose tissue (0.018 pM), but increased in media from adipose-ALL cocultures (RS4;11: 0.065 pM, BV173: 0.317 pM, n=1). One potential candidate signal to stimulate ALL cell RANKL expression is the inflammatory mediator prostaglandin E2 (PGE2), which is known to be released by adipose tissue. Physiological concentrations of PGE2 (200 ng/mL) caused a similar increase in ALL cell RANKL gene expression as adipose tissue in both cell lines (n=6). PGE2 treatment also caused a mild resistance of ALL cells to daunorubicin chemotherapy in vitro, increasing the EC50 from 27±4 to 53±12 nM in RS4;11 (n=6, p<0.05), and from 39±11 to 62±19 nM in BV173 (n=6, p=0.06). Adipose tissue induction of RANKL expression was partially blocked by the addition of PGE2 receptor EP2 and EP4 blockers. Taken together, our results suggest that PGE2 produced by the adipose microenvironment activates ALL cell RANKL expression, which could potentially contribute to CNS invasion, bone breakdown, and chemoresistance. Citation Format: Michael Cohen, Jia Tan, Tyler Kuk, Blesscamal Nadeak, In Sook Ahn, Xia Yang, Etan Orgel, Steven D. Mittelman. Adipose tissue upregulates expression of RANKL in B-cell acute lymphoblastic leukemia through prostaglandin E2 signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 178.

Human mesenchymal stromal cells inhibit Mycobacterium avium replication in clinically relevant models of lung infection

IntroductionNovel therapeutic strategies are urgently needed for Mycobacterium avium complex pulmonary disease (MAC-PD). Human mesenchymal stromal cells (MSCs) can directly inhibit MAC growth, but their effect on intracellular bacilli is...

Does Scheduled Low-Dose Short-Term NSAID (Ketorolac) Modulate Cytokine Levels Following Orthopaedic Polytrauma? A Secondary Analysis of a Randomized Clinical Trial.

To determine whether scheduled low-dose, short-term ketorolac modulates cytokine concentrations in orthopaedic polytrauma patients. Secondary analysis of a double-blinded, randomized controlled trial. Single Level I trauma center from August 2018 to October 2022. Orthopaedic polytrauma patients between 18-75 years with a New Injury Severity Score greater than 9 were enrolled. Participants were randomized to receive 15 mg of intravenous (IV) ketorolac every 6 hours for up to 5 inpatient days or 2 mL of IV saline similarly. Daily concentrations of prostaglandin E2 (PGE2), interleukin (IL)-1a, IL-1b, IL-6, and IL-10. Clinical outcomes included hospital and intensive care unit (ICU) length of stay (LOS), pulmonary complications, and acute kidney injury (AKI). Seventy orthopaedic polytrauma patients were enrolled, with 35 participants randomized to the ketorolac group and 35 to the placebo group. The overall IL-10 trend over time was significantly different in the ketorolac group (p = 0.043). IL-6 was 65.8% higher at enrollment compared to Day 3 (p < 0.001) when aggregated over both groups. There was no significant treatment effect for PGE2, IL-1a, or IL-1b (p > 0.05). There were no significant differences in clinical outcomes between groups (p > 0.05). Scheduled low-dose, short-term, IV ketorolac was associated with significantly different mean trends in IL-10 concentration in orthopaedic polytrauma patients with no significant differences in PGE2, IL-1a, IL-1b, or IL-6 levels between groups. The treatment did not have an impact on clinical outcomes of hospital or ICU LOS, pulmonary complications, or AKI. Therapeutic Level I. See Instructions for Authors for a complete description of levels of evidence.

Reproducible Synthesis of Biocompatible Albumin Nanoparticles Designed for Intra-articular Administration of Celecoxib to Treat Osteoarthritis.

Osteoarthritis (OA) is the most common form of arthritis, with intra-articular (IA) delivery of therapeutics being the current best option to treat pain and inflammation. However, IA delivery is challenging due to the rapid clearance of therapeutics from the joint and the need for repeated injections. Thus, there is a need for long-acting delivery systems that increase the drug retention time in joints with the capacity to penetrate OA cartilage. As pharmaceutical utility also demands that this is achieved using biocompatible materials that provide colloidal stability, our aim was to develop a nanoparticle (NP) delivery system loaded with the COX-2 inhibitor celecoxib that can meet these criteria. We devised a reproducible and economical method to synthesize the colloidally stable albumin NPs loaded with celecoxib without the use of any of the following conditions: high temperatures at which albumin denaturation occurs, polymer coatings, oils, Class 1/2 solvents, and chemical protein cross-linkers. The spherical NP suspensions were biocompatible, monodisperse with average diameters of 72 nm (ideal for OA cartilage penetration), and they were stable over 6 months at 4 °C. Moreover, the NPs loaded celecoxib at higher levels than those required for the therapeutic response in arthritic joints. For these reasons, they are the first of their kind. Labeled NPs were internalized by primary human articular chondrocytes cultured from the knee joints of OA patients. The NPs reduced the concentration of inflammatory mediator prostaglandin E2 released by the primaries, an indication of retained bioactivity following NP synthesis. Similar results were observed in lipopolysaccharide-stimulated human THP-1 monocytes. The IA administration of these NPs is expected to avoid side-effects associated with oral administration of celecoxib and to maintain a high local concentration in the knee joint over a sustained period. They are now ready for evaluation by IA administration in animal models of OA.

Brain regulates weight bearing bone through PGE2 skeletal interoception: implication of ankle osteoarthritis and pain

Bone is a mechanosensitive tissue and undergoes constant remodeling to adapt to the mechanical loading environment. However, it is unclear whether the signals of bone cells in response to mechanical stress are processed and interpreted in the brain. In this study, we found that the hypothalamus of the brain regulates bone remodeling and structure by perceiving bone prostaglandin E2 (PGE2) concentration in response to mechanical loading. Bone PGE2 levels are in proportion to their weight bearing. When weight bearing changes in the tail-suspension mice, the PGE2 concentrations in bones change in line with their weight bearing changes. Deletion of cyclooxygenase-2 (COX2) in the osteoblast lineage cells or knockout of receptor 4 (EP4) in sensory nerve blunts bone formation in response to mechanical loading. Moreover, knockout of TrkA in sensory nerve also significantly reduces mechanical load-induced bone formation. Moreover, mechanical loading induces cAMP-response element binding protein (CREB) phosphorylation in the hypothalamic arcuate nucleus (ARC) to inhibit sympathetic tyrosine hydroxylase (TH) expression in the paraventricular nucleus (PVN) for osteogenesis. Finally, we show that elevated PGE2 is associated with ankle osteoarthritis (AOA) and pain. Together, our data demonstrate that in response to mechanical loading, skeletal interoception occurs in the form of hypothalamic processing of PGE2-driven peripheral signaling to maintain physiologic bone homeostasis, while chronically elevated PGE2 can be sensed as pain during AOA and implication of potential treatment.

BRIDGING GAPS: ADDRESSING TREATMENT CHALLENGES WITH ORAL PGE2 IN DUCTUS-DEPENDENT CONGENITAL HEART DEFECTS

circulation. During fetal development, it diverts approximately 90% of the right ventricular output from the high-resistance pulmonary circulation to the umbilical-placental circulation for gas exchange with maternal blood. After birth, the ductus arteriosus usually shuts within 72 hours to help with lung circulation. Failure to close leads to a persistent left-to-right shunt known as patent ductus arteriosus (PDA). PDA affects up to 80% of premature infants and constitutes 5–10% of all congenital cardiac abnormalities in term infants. Hemodynamically significant PDA is associated with various morbidities, including neurodevelopmental impairment and respiratory distress syndrome (Slaughter et al., 2019). Conversely, in ductus-dependent heart lesions, which account for 20–25% of all congenital heart defects, it is crucial to maintain patency of the neonatal DA to uphold life-saving pulmonary or systemic circulation before corrective surgery (Subramaniam and Solomon, 2013). Treatment for closing patent ductus arteriosus (PDA) mostly involves nonsteroidal anti-inflammatory medicines (NSAIDs) such as indomethacin and ibuprofen, which block cyclooxygenase enzymes necessary for prostaglandin production. Although effective, NSAIDs carry dangers such as renal impairment and intestinal problems. Acetaminophen is considered a safer option, although its effectiveness in premature infants is unclear because it may not fully inhibit prostaglandin synthesis. However, few medications exist for preserving ductus arteriosus (DA) patency, with prostaglandin E1 (PGE1) being the sole pharmacological option despite its associated side effects, such as peripheral vasodilation, apnea, fever, and physiological disturbances (Sear, 2019). Furthermore, the shortage of PGE1 in certain countries like Pakistan highlights the urgent need for alternate solutions. In such situations, adopting other available choices like PGE2, which is accessible in Pakistan as an oral drug, becomes crucial for the efficient management of ductus-dependent congenital heart anomalies. Prostaglandin E2 (PGE2) has been successfully utilized orally in newborns with ductus-dependent cyanotic congenital heart disease. In research by Silove et al., PGE2 was supplied orally in doses ranging from 12 to 65 micrograms/kg at 1 to 4 hours intervals. This treatment regimen resulted in consistent increases in oxygen saturation (SaO2) and plasma PGE2 concentrations within 15-30 minutes after administration, equivalent to those achieved with intravenous infusions. Treatment lasted for durations ranging from 5 days to 4 months, during which the ductus remained open for extended periods, allowing for delayed surgical intervention to support the growth of the infants and their pulmonary arteries. While adverse effects during oral medication were similar to those found with intravenous administration, they were generally less severe in this series, underlining the effectiveness and simplicity of oral PGE2 administration for long-term treatment (Nabavi et al., 2019). Furthermore, a 1987 study by B.D. Thanopoulos et al. supported the effectiveness of oral PGE2 in the treatment of neonates with cyanotic congenital heart disease, which primarily depends on the ductus. The data from this investigation revealed that oral PGE2 treatment, while less productive than intravenous infusions, nonetheless proved useful for long-term therapy, affording more convenience and lower morbidity (Fernández-Francos et al., 2021). It is crucial to highlight that side effects such as apnea and hypotension may occur in newborns following PGE2 therapy and require close monitoring during treatment. In conclusion, oral PGE2 offers a viable alternative for addressing congenital heart defects in resource-limited countries like Pakistan. Its effectiveness and convenience make it a viable choice, especially if access to injectable drugs is limited. This underlines the necessity to investigate alternative treatments for appropriate care in such circumstances.

Effects of Meloxicam on the Welfare of Holstein Calves from 6 Weeks to 6 Months Old Undergoing Amputation Dehorning

Amputation dehorning (AD) is a common practice performed on calves, causing harmful effects such as pain, distress, anxiety, and fear. These effects extend to behavioral, physiological, and hematological responses, prompting serious ethical concerns regarding animal welfare, even when performed with local anesthesia. Meloxicam, a non-steroidal anti-inflammatory drug, has been widely used to mitigate the side effects of dehorning and disbudding in calves. However, there is a notable gap in research regarding the effects of meloxicam on calves aged 6 weeks to 6 mo undergoing AD procedures. This study was designed to assess the effectiveness of co-administering meloxicam with lidocaine, a cornual nerve anesthetic, in alleviating the adverse effects caused by the AD procedure in calves within this age range, compared with the use of lidocaine alone. Thirty Holstein calves were enrolled and randomly divided into 2 groups. The first group (Placebo) received a subcutaneous injection of 5 mL of lidocaine in the horn area and a subcutaneous injection of 0.9% saline at a dose of 0.025 mL/kg in the neck, administered 10 min before the AD procedure. The second group (MX) received a combination of lidocaine and meloxicam: a subcutaneous injection of 5 mL of lidocaine in the horn area and a subcutaneous injection of 20 mg/mL meloxicam at a dose of 0.025 mL/kg in the neck, also administered 10 min before the AD procedure. To avoid subjective bias, the researchers were blinded to the treatment groups. Pain-related behaviors, including tail flicking, head shaking, ear flicking, head rubbing, head crossing bar, and kicking, were observed, and physiological parameters, including heart rate, rectal temperature, respiration rate, mechanical nociceptive threshold (MNT), daily active steps, and food intake were monitored. Hematological conditions were determined using enzyme-linked immunosorbent assays and routine blood tests. The data were processed using a generalized linear mixed model (GLMM). The outcomes demonstrated that the AD procedure increased the frequencies of ear flicking and resulted in rises in the respiration rate, heart rate, rectal temperature, and daily active steps. It also led to decreases in total food intake, forage intake, hay intake, MNT, and increased concentrations of prostaglandin E2 (PgE2), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), nitric oxide (NO), and malondialdehyde, as well as glutathione peroxidase activity. However, calves that received meloxicam treatment showed significant improvements in response to the AD procedure, including lower respiration rates, heart rates, and rectal temperatures; higher MNTs; and lower intermediate cell ratio. They also had higher red blood counts, hemoglobin levels, hematocrit values; larger mean platelet volumes; and lower concentrations of PgE2, IL-1β, TNF-α, and NO. These results suggest that co-administration of lidocaine and meloxicam may aid in mitigating the adverse impacts induced by the AD procedure on these calves, thereby supporting the use of meloxicam in conjunction with a local anesthetic in AD procedures for calves aged 6 weeks to 6 mo.

Prostaglandin E2 may clinically alleviate dry eye disease by inducing Th17 cell differentiation.

Dry eye (DE) is a multifactorial ocular surface disease characterised by an imbalance in tear homeostasis. The pathogenesis of DE is complex and related to environmental, immunological (e.g., T helper 17 cells) and other factors. However, the DE disease pathogenesis remains unclear, thereby affecting its clinical treatment. This study aimed to explore the mechanism through which prostaglandin E2 (PGE2) affects DE inflammation by regulating Th17. The DE mouse model was established through subcutaneous injection of scopolamine hydrobromide. The tear secretion test and break-up time (BUT) method were used to detect tear secretion and tear film BUT, respectively. Enzyme-linked immunosorbent assay (ELISA) was used to detect the concentrations of PGE2, interleukin (IL)-17, IL-6 and tumour necrosis factor (TNF-α) in tear fluid and those of PGE2 and IL-17 in the serum. RT-qPCR and western blotting were used to test the mRNA and protein expression levels of IL-17 and retinoid-related orphan receptor-γt (RORγt). PGE2 was highly expressed in the DE mouse model. The mRNA and protein levels of IL-17 and the key Th17 transcription factor RORγt were increased in tissues of the DE mice. Moreover, PGE2 promoted tear secretion, reduced the BUT, increased the IL-17 concentration in tears and increased the Th17 cell proportion in DE, whereas the PGE2 receptor inhibitor AH6809 reversed the effects of PGE2 on tear secretion, BUT, and the Th17 cell proportion in draining lymph node (DLN) cells. Taken together, the study findings indicate that PGE2 could induce DE-related symptoms by promoting Th17 differentiation.

Loss of effective lubricating viscosity is the primary mechanical marker of joint inflammation in equine synovitis.

Inflammation of the synovium, known as synovitis, plays an important role in the pathogenesis of osteoarthritis (OA). Synovitis involves the release of a wide variety of pro-inflammatory mediators in synovial fluid (SF) that damage the articular cartilage extracellular matrix and induce death and apoptosis in chondrocytes. The composition of synovial fluid is dramatically altered by inflammation in OA, with changes to both hyaluronic acid and lubricin, the primary lubricating molecules in SF. However, the relationship between key biochemical markers of joint inflammation and mechanical function of SF is not well understood. Here, we demonstrate the application of a novel analytical framework to measure the effective viscosity for SF lubrication of cartilage, which is distinct from conventional rheological viscosity. Notably, in a well-established equine model of synovitis, this effective lubricating viscosity decreased by up to 10,000-fold for synovitis SF compared to a ~4 fold change in conventional viscosity measurements. Further, the effective lubricating viscosity was strongly inversely correlated (r = -0.6 to -0.8) to multiple established biochemical markers of SF inflammation, including white blood cell count, prostaglandin E2 (PGE2 ), and chemokine ligand (CCLs) concentrations, while conventional measurements of viscosity were poorly correlated to these markers. These findings demonstrate the importance of experimental and analytical approaches to characterize functional lubricating properties of synovial fluid and their relationships to soluble biomarkers to better understand the progression of OA.

Prostaglandin E2 accumulation is closely associated with S. aureus-infected bovine endometritis

S. aureus isolated from bacterial bovine endometritis is common in epidemiological reports, but is often ignored as a subclinical pathogenic microorganism. In a previous study, we showed that live S. aureus (LSA) and heat killed S. aureus (HK–SA) induce different inflammatory responses in bovine endometrial tissue, and possibly being associated with the accumulation of prostaglandin E2 (PGE2). Thus, in this study, we varied PGE2 concentrations using inhibitors or agonists in HK–SA-treated bovine endometrial tissues. The results demonstrated that PGE2 has a positive relationship with IL-6, TNF-α, and damage-associated molecular patterns (DAMPs; e.g., HMGB-1 and HABP-1) expression and tissues damage, and is regulated by the EP4-p38 MAPK pathway. We concluded that lipoproteins of S. aureus are associated with PGE2 generation. To further explore the relationship between LSA and PGE2 accumulation, we used the S. aureus strain SA113 lipoprotein knockout (SA113Δlpl) to infect bovine endometrial epithelial cells (BECs). LSA decreased PGE2, cAMP, EP4, IL-6, IL-8, cAMP secretion, and the MAPK and PKA signaling pathways when infected with SA113Δlpl, as compared with SA113-infected groups. Moreover, the adhesion and invasion of BECs were similarly downregulated when lipoproteins in S. aureus were knocked out. The results of this study show that PGE2 is involved in both HK–SA- and LSA-induced inflammatory responses in the bovine endometrium. We suggest that S. aureus infection is associated with bovine endometritis, and although HK–SA and LSA induce different inflammatory responses, the strategy of decreasing PGE2 accumulation is helpful in reducing the inflammation stage caused by S. aureus.

CAN ACUTE MUSCLE DAMAGE LEAD TO OSTEOARTHRITIS?

Abstract Skeletal muscles are crucial for maintaining musculoskeletal homeostasis, as they provide mechanical support, joint stability, and biochemical functions via myokine secretion. We investigated whether acute muscle damage or weakness could contribute to the development of osteoarthritis (OA). Using a chemical damage model, we induced muscle damage by injecting 1.2% barium chloride into the tibialis anterior muscle, resulting in an immune response characterized by a significant increase in the concentration of Prostaglandin E2 (PGE2) and other lipid mediators derived from the EPA and DHA signaling pathways, as quantified using our innovative lipidomics methods. The muscle damage was confirmed and visualized by muscle histomorphometry. The damage resulted in muscle weakness, detected by a significant decrease in grip strength at 4-days post-injury, but without changes in mice activity measured with force-plate actimeter. After 1-month post-injury, grip strength and muscle histology returned to normal, but histological changes were observed in the articulating cartilage, indicating surface erosion and cartilage matrix loss (OARSI grade 4). We used Raman Spectroscopy and micro-CT to confirm the joint alterations observed in the changes in subchondral bone and increased bone volume fraction in the cortical shell of the subchondral bone, respectively, features characteristic of OA in rodents and humans. These findings suggest that muscle weakness or imbalance due to injury can contribute to joint degeneration. This study highlights the significance of muscles in preventing OA and provides a novel approach for exploring the relationship between muscle damage and OA development. This new model could prove valuable for aging research.

Prostaglandin E2 promotes Th17 differentiation induces corneal epithelial cell apoptosis and participates in the progression of dry eye

This study is mainly based on T helper type 17 (Th17) cells analysis of the mechanism of prostaglandin E2 (PGE2) promoting the progression of dry eye (DE). Scopolamine and dry environment were used to induce mice DE model. Celecoxib was used to inhibit PGE2. Corneal epithelial cells and CD4+ T cells were used to construct a co-culture system. The osmotic pressure was increased by adding NaCl to simulate DE in vitro. AH6809 and E7046 were used to pre-culture to inhibit EP2/4 in T cells to verify the effect of exogenous PGE2 on Th17 cell differentiation and corneal epithelial cell apoptosis. The function of Th17 cells was analyzed by detecting RORγt and interleukin-17 (IL-17). PGE2 was instilled on the ocular surface to induce DE symptoms of mice. AH6809 and E7046 were used to inhibit EP2/4. The corneal epithelial cell apoptosis was observed by TUNEL. The proportion of Th17 cells in corneal tissue and draining lymph nodes (DLNs) was detected by flow cytometry. In DE mice, the concentration of PGE2 and IL-17 increased in tears, and the proportion of Th17 increased, while inhibition of PGE2 alleviated the symptoms of DE and inhibited Th17 differentiation. Hypertonic environment induces corneal epithelial cells to secrete PGE2. PGE2 promoted the expression of EP2/4 and the differentiation of Th17 cells in vitro. The hypertonic environment promoted PGE2 level and the apoptosis of corneal epithelial cells in the co-culture system. PGE2 alone did not cause corneal epithelial cell apoptosis, while PGE2 promoted apoptosis by promoting Th17. Blocking EP2/4 reduced the induction of Th17 differentiation by PGE2 and the promoted corneal epithelial cell apoptosis. Animal experiments showed that exogenous PGE2 induced DE symptoms. Blocking EP2/4 not only inhibited the proportion of Th17, but also alleviated the apoptosis of corneal epithelial cells caused by PGE2. PGE2 induces aggravation of inflammation by promoting the level of Th17 in the ocular surface, and causes corneal epithelial cell apoptosis, thereby participating in the progression of DE.

Evaluation of pain intensity with prostaglandin E2 biomarker and visual analog scale during initiation of orthodontic treatment: A prospective study

The aim of this study was to assess subjectively and objectively, at which timepoint the patient undergoing fixed orthodontic treatment will have maximum pain during the initiation of orthodontic treatment. A total of 22 patients (21 ± 2.7 years, [15 female and 7 male]) requiring extraction of maxillary first premolar participated in the study. Following bonding, an initial archwire of 0.016" NiTi was placed with a distal force of 100 g applied to the canines. The concentration of prostaglandin E2 (PGE2) in the gingival crevicular fluid was assessed at four time points: 1 hour before (T0); 24 hours (T1); 48 hours (T2); 72 hours (T3) after initial archwire placement. The pain intensity was recorded with visual analog scale. The concentration of PGE2 increased from T0 to T1. The highest concentration was observed at T2 following which there was a mild decline at T3. Statistically significant difference in the PGE2 concentration was observed between T0 to T1 and T1 to T2. The visual analogue scale (VAS) scores increased from T0 to T1. The highest score was observed at T2 followed by a decline at T3. A statistically significant difference in the VAS score was noted between T0 to T1 and T2 to T3. On correlating PGE2 and VAS score, a weak negative correlation was noted between T0 to T1 and no correlation was noted between T1 to T2 and T2 to T3. The patients experience maximum pain between 24 hours (T1) and 48 hours (T2) following initial archwire placement. The highest concentration of PGE2 and VAS score was observed at 48 hours (T2).

PSIII-8 Extra-Label Bisphosphonate Effects on Intra-Articular Inflammation in Juvenile Horses Challenged with Intra-Articular Lipopolysaccharide

Abstract Anecdotal assertions that bisphosphonates, such as clodronate disodium (CD), exhibit anti-inflammatory properties has led to extra-label use, despite a lack of scientific evidence of its efficacy. The objective of this study was to determine the effects of CD on intra-articular inflammation, hypothesizing that intra-muscular administration of CD would reduce intra-articular inflammation following an acute inflammatory challenge by lipopolysaccharide (LPS). To test this, 32 yearling Quarter horses were stratified by age (500 ± 13 d), BW (336 ± 26 kg), sex (n = 16 female; n = 16 male) and initial bone optical density into 4 treatment groups. The 140-d study consisted of two phases: Phase 1 (d 0 to 83) and Phase 2 (d 84 to 140) emulated sale preparation and early performance training, respectively. Horses were housed individually (3.6 m × 7.3 m) and fed to meet requirements of growing horses undergoing moderate training. Treatments consisted of control (CON; n = 8; no CD), single-dose (1X; n = 8; d 84), two-doses (2X; n = 8; d 0, 84), and four-doses (4X; n = 8; d 0, 42, 84, 126) of CD. All horses received iso-volumetric intramuscular injections of 1.8mg/kg BW clodronate disodium (OSPHOS) or saline (placebo) on d 0, 42, 84, and 126. Following treatment administration on d 126, radial carpal joints of each horse were injected with 0.8 mL of either 0.5 ng sterile LPS derived from Escherichia coli O55:B5 or sterile lactated Ringer’s solution as a contralateral control. Synovial fluid was collected before LPS injection (h 0) and at 6, 12, 24, and 336 h post injection. Synovial fluid was analyzed using an ELISA for prostaglandin E2 (PGE2), an indicator of joint inflammation. Data were log transformed and analyzed using the MIXED procedure of SAS. There was an h × LPS interaction (P &amp;lt; 0.01) confirming an acute, transient inflammatory response with increased PGE2 at 6, 12, and 24 h compared with the contralateral control joint, and resolved (P = 0.62) by h 336 post-injection. There was a treatment × h × LPS interaction (P = 0.02) in which 1X, 2X and 4X CD groups had greater synovial PGE2 concentrations at 6 h post-injection in the LPS joint compared with CON. In summary, intra-articular LPS induced localized inflammation, and the PGE2 response was greater in all horses treated with CD. The results of this study reject the hypothesis, suggesting clodronate disodium administration does not reduce intra-articular inflammation following acute induced synovitis.

Histological, Physiological and Biomechanical Effects of Low-Level Laser Therapy on Tendon Healing in Animals and Humans: A Systematic Review.

Low-level Laser Therapy (LLLT) was widely used in clinical practice for tendon disorders. However, the underlying mechanisms and effectiveness of LLLT in treating tendon injury remain unclear. Therefore, the present study was conducted aiming to summarize the evidence regarding the histological, physiological, and biomechanical effects of LLLT on tendon healing in animal and human models. Four databases were searched for relevant literature. Four independent reviewers screened abstracts and full-text articles, extracted relevant data, evaluated the risk of bias, and quantified the quality of evidence. Database searches yielded 1400 non-duplicated citations. Fifty-five studies were included (50 animal and five human studies). Animal studies revealed that LT had stimulating effects on collagen organization, collagen I and collagen II formation, matrix metalloproteinase (MMP)-8, transforming growth factor β1, vascular endothelial growth factor, hydroxyproline, maximum load, maximum elongation before breaking, and tendon stiffness. However, LLLT had inhibitory effects on the number of inflammatory cells, histological scores, relative amount of collagen III, cyclooxygenase-2, prostaglandin E2 (PGE2), interleukin-6, tumor necrosis factor-α, MMP-1, and MMP-3. Although one human study found that LLLT reduced the concentration of PGE2 in peritendinous tissue of the Achilles tendon, other human studies revealed that the effects of LLLT on the physiology and biomechanics of human tendons remained uncertain. LLLT facilitates tendon healing through various histological, physiological, and biomechanical effects in animal models. Only post-LLLT anti-inflammatory effects were found in human studies.

Botulinum neurotoxin type A does not exert concentration-dependent effects on equine articular cartilage in vitro.

To determine whether Botulinum neurotoxin type A (BoNT-A) ameliorates the effects of interleukin 1 (IL-1) on equine articular cartilage, or exerts negative effects on normal equine articular cartilage homeostasis in vitro. Articular cartilage explants from 6 healthy femoropatellar joints of 3 adult horses. Explants were allocated to the IL-1 challenged or unchallenged group, then exposed to 1 of 6 concentrations of BoNT-A (0, 1, 10, 50, 100, or 500 pg/mL) for 96 hours. To assess BoNT-A's effects on inflammation, prostaglandin E2 (PGE2) was measured in media via ELISA. Matrix degradation was determined as the percentage of sulfated glycosaminoglycans (sGAG) released from explants via dimethylmethylene blue assay. Aggrecan synthesis was estimated using CS846 ELISA and collagen type II degradation was estimated using C2C ELISA on media. Chondrocyte apoptosis was assessed via in-situ TUNEL assay. Generalized linear mixed models were fitted to determine treatment effects using α = 0.05. The challenge with IL-1 resulted in increased concentrations of PGE2 and CS846 in media and increased release of sGAG from explants. BoNT-A did not significantly impact PGE2 or CS846 concentration in media, percentage of sGAG released, or chondrocyte apoptosis in IL-1 challenged or unchallenged cartilage explants. The concentration of C2C in media was below the quantifiable limit of the ELISA in all samples. BoNT-A did not show chondroprotective effects or have negative effects on cartilage homeostasis in vitro at the concentrations tested. While chondroprotective effects were not observed, BoNT-A may be safe for intraarticular use. In vivo testing is warranted before clinical use.

EP4 Receptor Conformation Sensor Suited for Ligand Screening and Imaging of Extracellular Prostaglandins.

Prostaglandins are important lipid mediators with a wide range of functions in the human body. They act mainly via plasma membrane localized prostaglandin receptors, which belong to the G-protein coupled receptor class. Due to their localized formation and short lifetime, it is important to be able to measure the distribution and abundance of prostaglandins in time and/or space. In this study, we present a Foerster resonance energy transfer (FRET)-based conformation sensor of the human prostaglandin E receptor subtype 4 (EP4 receptor), which was capable of detecting prostaglandin E2 (PGE2)-induced receptor activation in the low nanomolar range with a good signal-to-noise ratio. The sensor retained the typical selectivity for PGE2 among arachidonic acid products. Human embryonic kidney cells stably expressing the sensor did not produce detectable amounts of prostaglandins making them suitable for a coculture approach allowing us, over time, to detect prostaglandin formation in Madin-Darby canine kidney cells and primary mouse macrophages. Furthermore, the EP4 receptor sensor proved to be suited to detect experimentally generated PGE2 gradients by means of FRET-microscopy, indicating the potential to measure gradients of PGE2 within tissues. In addition to FRET-based imaging of prostanoid release, the sensor allowed not only for determination of PGE2 concentrations, but also proved to be capable of measuring ligand binding kinetics. The good signal-to-noise ratio at a commercial plate reader and the ability to directly determine ligand efficacy shows the obvious potential of this sensor interest for screening and characterization of novel ligands of the pharmacologically important human EP4 receptor. SIGNIFICANCE STATEMENT: The authors present a biosensor based on the prostaglandin E receptor subtype 4, which is well suited to measure extracellular prostaglandin E2 (PGE2) concentration with high temporal and spatial resolution. It can be used for the imaging of PGE2 levels and gradients by means of Foerster resonance energy transfer microscopy, and for determining PGE2 release of primary cells as well as for screening purposes in a plate reader setting.

Seasonal expressions of COX-1, COX-2 and EP4 in the scent glands of muskrats (Ondatra zibethicus).

Prostaglandins (PGs) serve as signaling molecules that regulate various physiological processes, including inflammation, immune response, blood clotting and reproduction. The aim of this study was to investigate the immunolocalizations and expression patterns of prostaglandin-E2 (PGE2), cyclooxygenase (COX)-1 and COX-2, as well as its receptor subtypes 4 (EP4) in the scent glands of muskrats (Ondatra zibethicus) during the breeding and non-breeding periods. There were significant seasonal differences in the scent glandular mass, with higher values in the breeding season and relatively low in the non-breeding season. PGE2, EP4, COX-1 and COX-2 have been immunolocalized in the scent glandular and epithelial cells in both breeding and non-breeding seasons, whereas no immunostaining was observed in the interstitial cells. The protein and mRNA expression levels of EP4, COX-1 and COX-2 were higher in the scent glands of the breeding season than those of the non-breeding season. The mean mRNA levels of EP4, COX-1 and COX-2 were positively correlated with the scent glandular weights. The circulating follicle stimulating hormone (FSH), luteinizing hormone (LH), testosterone (T) and PGE2, as well as scent glandular PGE2 and dihydrotestosterone (DHT) concentrations were also significantly higher in the breeding season. Additionally, the transcriptomic study in the scent glands identified that differentially expressed genes might be related to fatty carboxylic monocarboxylic acid, steroidogenic-related pathway and prostanoid metabolic process. These findings suggested that prostaglandin-E2 might play an essential autocrine or paracrine role in regulating seasonal changes in the scent glandular functions of the muskrats.

Measurement of Cyclooxygenase Products in Plasma as Markers for Inhibition of Cyclooxygenase Isoforms by Oral Meloxicam in New Zealand White Rabbits (Oryctolagus cuniculus ).

Pain management in rabbits is a challenging task that is complicated by the rabbit's ability to hide signs of distress and the limited pharmacologic data available for this species. Pharmacokinetic data has shown that in rabbits, meloxicam, a nonsteroidal anti-inflammatory NSAID, reaches plasma concentrations that are known to provide analgesia in dogs and cats; these concentrations could theoretically alleviate pain in rabbits. However, the inhibitory effects of meloxicam on cyclooxygenase (COX) isoforms have not been studied in rabbits. In this study, we measured the products of COX-1 and COX-2 after the oral administration of a single 1 mg/kg dose of meloxicam to New Zealand White rabbits (n = 6). Blood samples were collected before drug administration (T0) and then at predetermined time points over 48 h. Plasma prostaglandin E₂ (PGE₂ ) and thromboxane (TxB₂) concentrations were measured as surrogate markers for COX-1 and COX-2, respectively, by using commercial ELISA kits. After meloxicam administration, both TxB₂ and PGE₂ plasma concentrations fell significantly below baseline, with maximal mean reductions to 80% and 60% of baseline at 8 h, respectively. The reduction in PGE₂ concentrations was followed by a significant increase that moved its mean plasma concentrations toward baseline between 8 and 24 h. Adverse effects such as lethargy, inappetence, or changes in fecal production were not observed in any rabbits. In conclusion, meloxicam appeared to significantly inhibit both COX-1 and COX-2 with a time course similar to previously reported meloxicam plasma concentration-time profiles in rabbits. Our data suggest that a dosage of 1 mg/kg given orally could provide analgesia to rabbits, but a more frequent dosing interval than the currently recommended daily dosing may be required to maintain clinical efficacy.