PGF2 Research Articles

IMMU-10. IDENTIFICATION OF A NEW THERAPEUTIC ANTIGEN FOR GLIOBLASTOMA USING A MONOCLONAL ANTIBODY LIBRARY FROM PATIENT-DERIVED TUMOR SPHERES

Abstract Glioblastoma (GBM) has a poor prognosis despite intensive treatment by surgery, radiation, and chemotherapy, thus new treatment strategies are urgently needed. Various innovative cancer therapies targeting cancer-specific cell surface antigens, such as chimeric antigen receptor T cell therapy, are developed and more cell surface antigens that is highly specific for GBM cells are needed. Although extensive transcriptome analyses or proteomic analysis of GBM cells were performed, few transcripts specific for GBM cells have been identified. However, GBM cell-specific antigen epitopes formed by post-translational modifications of proteins may have been missed by transcriptome or proteomic analysis, and could still be discovered by thoroughly searching for cancer-specific monoclonal antibodies. In this study, we aimed to identify GBM-specific antigens using a monoclonal antibody library from patient-derived tumor spheres, create CAR-T cells against the antigen, and check antitumor efficiency of the CAR-T cells. Approximately 25,000 monoclonal antibody-producing hybridomas were establishment using BALB/c mice and patient derived tumor spheres. Among them, a antibodies that bind to plural glioblastomas and not to plural normal brain cells were selected and we identified the antigen as Prostaglandin F2 receptor negative regulator (PTGFRN) by the expression cloning method. PTGFRN witch is expressed in several cancers regulate migration, angiogenesis, and cell polarity and decreases the radiosensitivity of GBM cells. We created CAR-T cells against the antigen and co-cultured it with GBM cells, which significantly increased the production of IL-2 and INF-γ. Furthermore, the CAR-T cells injected intracranial in an orthotopic xenograft model with patient-derived GBM cells, significantly reduced tumor growth. PTGFRN CAR-T-cell therapy may be a potential novel therapeutic target for GBM.

Effect of Gonadotropin-Releasing Hormone Treatment on Superovulation Response and Embryo Recovery in Holstein Heifers: A Field Trial at Zagros Milk and Meat Company, Iran

Background: Superovulation is a crucial component of assisted reproductive technology. Inducing superovulation with Gonadotropin-releasing hormone (GnRH) can lead to ovarian hyperstimulation, potentially affecting reproductive outcomes. Objectives: This study aimed to examine the effects of GnRH treatment on superovulation response and embryo recovery in Holstein heifers. Methods: Twenty-one Holstein heifers (age: 135.15 months; weight: 361.05 kg) were selected based on their general health status and ovarian function. All heifers received two consecutive doses of prostaglandin F2α and underwent superovulation. The heifers were inseminated twice: Once at the onset of standing estrus and again 12 hours later. In the treatment group (13 heifers), a single dose of GnRH was administered simultaneously with the second insemination. The superovulation response was evaluated based on the number of corpus luteum (CL), unovulated follicles (UoF), total recovered embryos/ova, and the number of transferable embryos, using logistic regression with the GENMOD procedure in SAS. Results: The mean number of CL was not significantly greater in the control group (13.6) compared to the GnRH group (11.4). The average number of UoF was similar between the two groups (P = 0.1853). However, the control group had a significantly higher average total number of recovered embryos/ova (7.7) compared to the GnRH group (2.1). Additionally, the control group produced more transferable embryos, with an average of 2.5, while the GnRH group averaged 0.7 (P < 0.0062). Conclusions: Gonadotropin-releasing hormone likely deactivated the oviduct by disrupting the balance of estradiol and estrogen, leading to a reduction in both the total number of embryos and the number of transferable embryos in heifers.

Effects of thiacloprid, a neonicotinoid pesticide, on rat reproductive system: Pregnancy hormone disruption and abortion trends

Thiacloprid (TCL), commonly known as Biscaya, is among the most widely used pesticides in agriculture, designed to eliminate insects by targeting their nicotinic receptors. This study explores the effects of orally administering TCL (at a dose of 50 mg/kg) on the hormone secretion crucial for pregnancy and the factors influencing abortion throughout the early, middle, and late stages of pregnancy in female rats.Following TCL exposure, there were significant increases in levels of 17β-Estradiol, prostaglandins F2α and E2, and serum oxytocin hormone in different stages of pregnancy. In contrast, progesterone and endothelin-1 serum levels notably decreased during the initial and final stages of pregnancy. Additionally, TCL led to a substantial rise in lipid peroxidation levels and a decrease in total thiol molecules and total antioxidant capacity, especially in uterine tissue. Although TCL did not significantly affect the morphological characteristics of the delivered fetuses, it notably increased the number of abortions, especially during the second and third stages of pregnancy.In summary, our findings suggest that TCL elevates the risk of abortion in pregnant rats by disrupting the secretion of hormones crucial for fertility (such as 17β-Estradiol/progesterone) and by increasing the secretion of abortion-inducing hormones like prostaglandins and oxytocin. Furthermore, these effects may be associated with disruptions in the oxidant/antioxidant balance within the ovaries and uterus.

The Mechanism Involved in the Inhibition of Resveratrol and Genistein on the Contractility of Isolated Rat Uterus Smooth Muscle

Purpose: This study aimed to compare the effects of the phytoestrogens resveratrol (RES) and genistein (GEN) on the contractility of isolated uterine smooth muscle from rats, focusing on both spontaneous and stimulated contractions, and to investigate the underlying mechanisms. Methods: Uterine strips were suspended vertically in perfusion chambers containing Kreb’s solution, various concentrations of RES and GEN were added to the ex vivo uterine strips, and contractions were measured before and after incubation with RES or GEN. Results: (1) Both RES and GEN inhibited K+-induced contractions in a dose-dependent manner; the β/β2-adrenoceptor antagonist propranolol (PRO), ICI118551, the ATP-dependent K+ channel blocker glibenclamide (HB-419) and the NO synthase inhibitor N-nitro-L-arginine (L-NNA) diminished the inhibitory effects of RES and GEN on K+-induced contractions. (2) RES and GEN also dose-dependently inhibited PGF2α-induced uterine contractions. (3) The inhibitory effects of RES and GEN were observed in spontaneous contractile activities as well; PRO, ICI118551, HB-419 and L-NNA attenuated the inhibitory effects of RES and GEN on the spontaneous contractions of isolated uterine muscle strips. (4) RES and GEN significantly decreased the cumulative concentration response of Ca2+ and shifted the Ca2+ cumulative concentration–response curves to the right in high-K+ Ca2+-free Kreb’s solution. (5) RES and GEN markedly reduced the first phasic contraction induced by oxytocin, acetylcholine, and prostaglandin F2α but did not alter the second phasic contraction caused by CaCl2 in Ca2+-free Kreb’s solution. Conclusions: RES and GEN can directly inhibit both spontaneous and activated contractions of isolated uterine smooth muscle. The mechanisms underlying the inhibitory effects of RES and GEN likely involve β adrenergic receptor activation, reduced Ca2+ influx and release, the activation of ATP-dependent K+ channels and increased NO production.

7624 m6A mRNA Methylation in Brown Adipose Tissue Regulates Systemic Insulin Sensitivity via an Inter-Organ Prostaglandin Signaling Axis

Abstract Disclosure: L. Xiao: None. D. F De Jesus: None. C. Ju: None. J. Wei: None. J. Hu: None. A. DiStefano-Forti: None. T. Tsuji: None. C. Cero: None. V. Männistö: None. S.M. Manninen: None. S. Wei: None. O. Ijaduola: None. M. Blüher: Consulting Fee; Self; Amgen Inc, AstraZeneca, Bayer, Inc., Boehringer Ingelheim, Lilly USA, LLC, Novo Nordisk, Pfizer, Inc., Sanofi. A.M. Cypess: None. J. Pihlajamäki: None. Y. Tseng: None. C. He: Advisory Board Member; Self; Accent Therapeutics. R.N. Kulkarni: Advisory Board Member; Self; Novo Nordisk, Biomea, Inversago, REDD. Brown adipose tissue (BAT) has the capacity to regulate systemic metabolism through the secretion of “Batokines” including signaling lipids. N6-methyladenosine (m6A) is the most prevalent and abundant post-transcriptional mRNA modification and has been reported to regulate BAT adipogenesis and energy expenditure. Here, we demonstrate that the absence of the m6A writer, methyltransferase-like 14 (METTL14), modifies the BAT secretome to initiate inter-organ communication to improve systemic insulin sensitivity. Importantly, these phenotypes are independent of UCP1-mediated energy expenditure and thermogenesis. Using lipidomics, we identified prostaglandin E2 (PGE2) and prostaglandin F2a (PGF2a) as M14KO-BAT-secreted insulin sensitizers. Notably, circulatory PGE2 and PGF2a levels are inversely correlated with insulin sensitivity in humans. Furthermore, in vivo administration of PGE2 and PGF2a in high-fat diet-induced insulin-resistant obese mice recapitulates the phenotypes of Mettl14-deficient animals. PGE2 or PGF2a improves insulin signaling by suppressing the expression of specific AKT phosphatases. Mechanistically, METTL14-mediated m6A installation promotes decay of transcripts encoding prostaglandin synthases and their regulators in human and mouse brown adipocytes in a YTHDF2/3-dependent manner. Collectively, these findings reveal a novel biological mechanism in the m6A-dependent regulation of the BAT secretome in modulating systemic insulin sensitivity in both mice and humans. Presentation: 6/1/2024

6828 Exploring Metabolomic Clues in Diabetic Retinopathy

Abstract Disclosure: M.W. Simonson: None. Y. Li: None. J.J. McAnany: None. B. Prasad: None. E.C. Hanlon: None. S. Pannain: None. B.T. Layden: None. E. Van Cauter: None. J.C. Park: None. S.J. Crowley: None. F.Y. Chau: None. K.K. Danielson: None. H. Chen: None. G.E. Chlipala: None. C. Martinez: None. S. Reutrakul: Speaker; Self; Eli Lilly & Company. Background: Diabetic retinopathy (DR) is a common microvascular complication of diabetes. Metabolomics offers the possibility for novel biomarker discovery. The purpose of this study was to determine metabolomic differences that characterized patients with versus without DR. Methods: A total of 62 serum samples were collected from 36 type 2 diabetes (T2D) patients with DR and 26 T2D patients without DR, and analyzed via UPLC-MS. The metabolite data were normalized using median normalization. The differential expression of metabolites was compared among the groups using the limma package in R. The covariates (age, sex, BMI and sleep apnea severity) were accounted for by using a generalized linear model fitted to the data. Significantly different metabolites were filtered by an absolute log2 Fold Change (FC) of 1.5 or greater and Q values < 0.05. Results: Mean (SD) age for DR and no-DR group was 55 (±5.9) versus 54 (±6.7) years, while BMI was 33 (±5.8) versus 33 (±6.3) kg/m2 respectively. Both DR and no-DR groups were 58% female. A1C levels in DR group was marginally higher than in no-DR group (p=0.09). There were significant differences in metabolomic changes between DR versus no-DR, with 166 total differential metabolites after adjusting for covariates and filtering by FC. Notable metabolites included those involved in fatty acid metabolism, acyl carnitines, prostaglandins, and phospholipids. There was an overall upregulation of serum acyl carnitines in DR as compared to no-DR, including 2,6 dimethylheptanoyl carnitine (FC = 3.0) and cis-5-Tetradecenoylcarnitine (FC = 2.0). Additionally, there was a decrease in long chain fatty acids in DR as compared to no-DR, which included 6-hydroxypentadecanedioic acid (FC = -6.6), 10,16-dihydroxy-palmitic acid (FC = -6.0), and tetracosatetraenoyl CoA (FC = -6.1). The 11-beta prostaglandin F2 (11-PGF2) was increased in DR (FC = 1.9). Circulating phospholipids including lysophosphatidylcholine (22:2) (lysoPC) (FC = 1.8) and phosphatidylcholine (24:1/24:1) (PC) (FC = 5.9) were also significantly dysregulated. Conclusions: We identified a subset of metabolites that discriminate T2D patients with or without DR. The changes in acyl carnitines and long chain fatty acids may reflect mitochondrial dysfunction in DR. The presence of certain prostaglandins highlights the inflammatory environment which may facilitate DR progression. Increases in phospholipids such as lysoPC suggest an atherogenic state and higher LDL oxidation, while increases in PC may indicate altered lipogenesis in DR. These findings may serve as new potential serum biomarkers to promote efficient preventive care and provide mechanistic insight into understanding DR pathophysiology. Presentation: 6/2/2024

Effect of the Application of PGF2α Associated With Ovulation Induction in a Fixed-Time Superovulation Programme for Precocious Nellore Heifers.

This study evaluated the effect of prostaglandin F2α (PGF2α) associated with gonadotropin-releasing hormone (GnRH) for ovulation induction in precocious indicus heifers submitted to a fixed-time superovulation (SOV) programme. Precocious Nellore heifers (n = 35), aged 13 months, were subjected to the SOV protocol. On day 0 (D0), all animals received intravaginal insertion of a progesterone (P4) device along with intramuscular administration of 2 mg of oestradiol benzoate, plus 200 IU of follicle-stimulating hormone in decreasing doses, with 12-h intervals between D4 and D7, in addition to 150 μg of D-cloprostenol on D6 and device removal on D7. On D8, the donors received 10.5 μg of buserelin acetate and the treatment group received 300 μg of D-cloprostenol/PGF2α. Artificial insemination was performed 12 h and 24 h after GnRH administration using frozen semen. On D15 of the protocol (i.e., D7 after insemination), the embryos were collected and evaluated. All animals passed through the control and treatment groups. Results were evaluated by analysis of variance using an adjusted mixed-effects model (p < 0.05). There was no difference in the total number of embryos between the control and treatment groups (10.40 ± 1.52 vs. 9.60 ± 1.36; p = 0.63) or viable embryos (6.30 ± 1.22 vs. 4.30 ± 0.71). For precocious indicus heifers, treatment with PGF2α in association with GnRH did not affect embryo production in the fixed-time SOV protocol.

PSXII-11 Impact of pre-breeding and breeding supplementation in May-born range heifers on growth rates and reproductive performance

Abstract Our objectives were to determine the impact rumen undegradable protein (RUP) supplementation prior to and during the breeding season had on growth rates and reproductive performance in May-born yearling range heifers. This study was conducted over a 3-yr period at the University of Nebraska, Gudmundsen Sandhills Laboratory (GSL) near Whitman, NE and utilized May-born Red Angus x Simmental crossbred heifers [n = 192; weaning body weight (BW) = 200 kg ± 28 kg]. Heifers were randomly assigned by pre-breeding BW to one of two treatments: 1) received 0.908 kg of a dried distillers grain supplement (DDGS; Supp, n = 66) pre-breeding through the breeding season or 2) received no supplement (NoSupp, n = 138). Each year, treatments were initiated in July and ended in August (55 ± 3 d). Using a C-Lock Smart Feeder (Rapid City, SD) heifers had individual access to their allotted DDGS supplementation. On average, NoSupp heifers consumed 0.09 kg/d (yr 1) and 0.02 kg/d (yr 2 and 3); whereas, Supp heifers averaged 0.59, 0.74, and 0.76 kg/d in yr 1, 2, and 3, respectively. Heifer BW was collected pre-trial (May), pre-breeding (July), breeding (August), pregnancy diagnosis (October), and pre-calving (April). Body condition score (BCS) was collected at pregnancy diagnosis (October) and pre-calving (April; Yr 1 and 2). Before each breeding season, two blood samples were collected 10 d apart to determine pubertal status. Heifers were synchronized with a single 5 mL intramuscular injection of prostaglandin F2 alpha (Lutalyse, Zoetis, Parsippany, NJ) 5 d after bull turnout during a 30 d natural service breeding season. Pregnancy diagnosis was determined via rectal ultrasound in October. No differences (P &amp;gt; 0.12) in BW were noted among treatments for all measurement timepoints. Moreover, SUPP heifers had greater (P &amp;lt; 0.02) average daily gain (ADG) from pre- trial to pregnancy diagnosis compared with NoSUPP heifers. However, there was no differences (P = 0.11) in ADG from pregnancy diagnosis to pre- calving. At pregnancy diagnosis (October), a treatment x year interaction (P = 0.02) for BCS was present where Supp heifers had a greater BCS than NoSupp heifers in yr 3. Heifer BCS pre-calving was not affected (P = 0.12) by treatment. There was a greater percentage (P &amp;lt; 0.01) of SUPP heifers cycling compared with NoSUPP heifers. However, there were no differences (P &amp;gt; 0.24) in pregnancy rates, the percentage that calved in the first 21 d, calf birth BW, or subsequent calving date. Ultimately, this study implies that providing a RUP supplement prior to and during the breeding season will increase the percentage of heifers that are cycling before breeding and have a greater ADG through pregnancy diagnosis; however, supplementation will not impact other reproductive parameters or BW gains.

PSIII-6 Multi-sire artificial insemination of beef heifers in the Nebraska Sandhills effect on pregnancy rate compared with single-sire insemination

Abstract The objective of this study was to compare pregnancy outcomes resulting from artificial insemination (AI) with multi-sire semen (MS) compared with single-sire semen (SS) in beef heifers. Another study observed pregnancy and paternity outcomes to sexed MS without a control treatment. In the first study, unrelated ABS Global Angus bulls (n = 3) were selected for AI to 441 (Yr 1) and 391 (Yr 2) Angus crossbred heifers. Heifers were synchronized with the melengestrol acetate – prostaglandin F2α timed-AI protocol. Conventional semen, from each bull, were labeled A, B, or C. Remaining semen was pooled in equal amounts to form conventional MS straws. Heifers were AI using alternating treatment of MS or SS: A, B, or C with one-half of inseminated heifers receiving either MS or SS. The GLIMMIX procedure in SAS 9.4 analyzed responses to treatments using the BIN distribution option and reported as LSMEANS. Contrasts and estimates analyzed responses between treatments. Ultrasound determined SS pregnancy rate averaged 60% (SS-A = 64%, SS-B = 58%, SS-C = 58%) and MS average 62% (P = 0.63). Paternity was determined in 57 MS calves and confirmed in 49 SS calves. In Yr 1, Bull A sired 7%, bull B sired 56%, and bull C sired 37%. Although bull A was much less successful than bull B or C, the values are not significantly different due to the small sample size (P = 0.11 and P = 0.24, respectively). Few conclusions can be made from the ongoing study, but it is hypothesized MS may improve consistency of pregnancy rate by equilibrating semen quality in herds subjected to timed AI. In the observational study, multiple 5-way cross bulls born from the ranch herd were selected for AI of 951 ± 41 heifers for 3 yr. Heifers were synchronized using the previously mentioned protocol with split-time AI. Unequal amounts of semen were collected, pooled, and sexed to favor heifer progeny at an expected ratio of 9:1. Semen from 2 to 4 random bulls were pooled making 3 to 4 MS groups each year. Heifers exhibiting estrus were administered sexed MS, but there was no control treatment. The observed ultrasound determined average pregnancy rate of heifers administered sexed MS was 65% (Yr 1), 75% (Yr 2), and 72% (Yr 3). Previous literature suggests average pregnancy rate to AI with sexed SS is 53%. Paternity testing in Yr 2 found semen contribution mirrored paternity ratios within some groups but the sire group with 2 bulls found one sired 15% and the other sired 85%. This study exemplified a viable method for AI that requires further research. Further research into semen quality of a single sire, and pooled semen could lead to optimization of fertilization among individuals within a herd.

PSLBI-3 The use of meloxicam oral suspension (15 mg/mL) to control post-partum pain and inflammation in dairy cattle following dystocia

Abstract Dystocia is painful, given the increased time, tissue damage, and stress associated with handling and forced extraction. Around parturition, dry matter intake is reduced, and cows experience increased inflammatory mediators. This could lead to subsequent health issues, including an increased risk of metabolic disease, reduced reproductive performance, and decreased milk production. Controlling pain and disease are important for animal welfare so it is critical to alleviate dystocia pain and inflammation. The administration of a non-steroidal anti-inflammatory drug (NSAID) could aid in alleviating the negative sequelae of dystotic parturition. Meloxicam (MOS) is an NSAID with greater oral bioavailability and inhibits cyclooxygenase, the enzyme responsible for converting arachidonic acid to prostaglandin. The reduction in prostaglandin, specifically prostaglandin F2α, resulting from meloxicam administration following a dystotic calving could minimize pain and control systemic inflammation. Objectives were: 1) to demonstrate post-partum administration of MOS in dystotic lactating dairy cows reduces pain (for up to 3 d) as measured by a significant reduction in plasma substance P expression levels (P &amp;lt; 0.05); 2) to demonstrate post-partum administration of MOS in dystotic lactating dairy cattle reduces inflammation (for up to 14 d) as measured by a significant reduction in plasma haptoglobin expression levels (P &amp;lt; 0.05); and 3) to evaluate the safety of administering MOS in post-partum dairy cattle, especially as it pertains to reproductive health. The study included dairy farms in three geographical areas in Canada, with nine farms and 145 cows. Primiparous and multiparous cows identified as having dystotic calvings were enrolled based on observation by the producer or study technician. Cows were randomly assigned to a treatment group of MOS or the placebo within each farm site. Animals received test or control treatment 0 to 12 h post-dystotic calving. The calving and treatment times were recorded. No analgesics was given before calving. Data collected included: 1) pain-related behaviors; 2) physiological markers of pain; 3) physiological markers of inflammation measured as acute phase proteins; 4) vulvovaginal inflammation; 5) physical evaluation including daily observations, body weight, rumination, blood and urinalysis, reproductive evaluation, time to first insemination and daily milk weights. The data are being evaluated and initial results show favorable responses to the treatment.

420 Automated identification and behavioral estrus detection in freestall-housed dairy cows using computer vision and machine learning: A two-layered algorithm approach

Abstract A critical aspect of dairy management involves maintaining accurate records of individual animals and promptly detecting estrus behavior, which is vital for successful breeding and reproductive outcomes. Recent advancements in precision livestock management, utilizing computer vision and machine learning technologies, have paved the way for automated and precise solutions in these areas. The objective of this study is to investigate the application of computer vision and machine learning for animal identification and behavioral estrus detection in dairy cows housed in freestall settings, employing a two-layered algorithmic approach. The research was conducted at the Joe Bearden Dairy Research Center, where a total of 10 lactating Holstein-Friesian cows were assessed across a 7-d trial period. For synchronization efforts, cows were provided a single injection of Prostaglandin F2α at trial initiation with expected estrus behaviors to be exhibited within the following 56 h. Continuous video was captured in 4k resolution using a five-camera closed caption media recording system strategically placed in the pen corners and over the feed bunk alleyway to capture multiple perspectives. Video data extrapolated into frames and manual annotation was utilized to generate the machine learning algorithm’s training data. Frames were randomly selected from a scattering of 56 h of video across all five camera viewpoints and down sampled to 384 x 640 p for both layers. A two-layer approach was employed utilizing an algorithm for each aspect of the study with the first model detecting the behavior, and the second model using the detected region of interest to identify specific cows. The first layer was trained on 682 frames and validated against 123 manually annotated frames. The second layer was trained on 102 frames and validated against 16 frames manually annotated for individual animals. Results demonstrated that the model used for mounting behavior identification has high precision (0.819) and recall (0.89), achieving a 0.926 mAP at an IoU of 0.5. This model achieved an F1 score of 0.86 at a confidence threshold of 0.268. The results demonstrated that the model used for individual animal identification across different classes achieved high precision (0.861) and recall (0.842), with a mAP of 0.897 at an IoU of 0.5. The F1 score for all classes was 0.89 at a confidence threshold of 0.788. In conclusion, these promising findings underscore the potential of computer vision and machine learning technologies to revolutionize dairy cow management practices. This innovative approach not only has potential to enhance operational efficiency, but also contributes significantly to improved reproductive management, thereby advancing precision and productivity in dairy operations.

182 Converting spring-born heifers into a summer-calving herd increases subsequent cow longevity and productivity

Abstract Developing replacement heifers is one of the most expensive and complex management decisions for a cow-calf producer, which also has long-term implications for profitability. The objective of this study was to determine the impact of converting spring-born heifers into a summer-calving herd on growth, reproductive performance, longevity, and productivity compared with early or late-born May heifers. Over a 3 yr period, Red Angus/Simmental crossbred heifers (n = 273) were utilized to determine the impact of converting March-born heifers (Convert; n = 90) to a May-calving herd compared with May-born heifers on reproductive performance, body weight (BW), and calf performance from a yearling to 5 yr of age. May-born heifers were retrospectively grouped into 2 different groups: heifers born in the first 21 d of calving as a heifer calf (Early; n = 123) or heifers born after the 21 d of calving as a heifer calf (Late; n = 60). Heifers were exposed to bulls for a 45-d breeding season with a bull-to-heifer ratio of 1:20. Heifers were synchronized with a single injection of prostaglandin F2α (5-mL i.m.; Lutalyse, Zoetis, Parsippany, NJ) 5 d after bulls were introduced in the pasture for breeding. Data were analyzed as a randomized design using the MIXED procedure (SAS Inst. Inc., Cary, NC, USA). All binomial data were analyzed using PROC GLIMMIX. Heifer served as the experimental unit with treatment and year developed set as fixed effects. Heifer BW at December, May, July, and pregnancy check (Oct) were greater (P &amp;lt; 0.01) for Convert heifers with no difference between Early and Late born heifers. Heifer BCS at pregnancy check was not different (P = 0.13) among heifer treatment groups. However, pregnancy rate tended (P = 0.08) to increase for Convert heifers with no difference between Early and Late born May heifers. After 5 yr of age, retention rate was increased (P = 0.04) in Convert cows compared with Early and Late May-born cows with no difference (P = 0.84) between the two May-born heifer groups. Total kilograms of calf weaned was greater (P = 0.04) in Convert cows than their counterparts with no differences (P = 0.94) between Early and Late May-born cows. This study implies that selecting replacement heifers from an earlier calving season for replacements in a later calving season increases the longevity and long-term productivity of the cowherd than selecting heifers within the same calving season. This may be even more important in resource-poor breeding environments that create reproductive challenges for heifers and rebreeding young cows.

M6A mRNA methylation in brown fat regulates systemic insulin sensitivity via an inter-organ prostaglandin signaling axis independent of UCP1

Brown adipose tissue (BAT) regulates systemic metabolism by releasing signaling lipids. N6-methyladenosine (m6A) is the most prevalent and abundant post-transcriptional mRNA modification and has been reported to regulate BAT adipogenesis and energy expenditure. Here, we demonstrate that the absence of m6A methyltransferase-like 14 (METTL14) modifies the BAT secretome to improve systemic insulin sensitivity independent of UCP1. Using lipidomics, we identify prostaglandin E2 (PGE2) and prostaglandin F2a (PGF2a) as BAT-secreted insulin sensitizers. PGE2 and PGF2a inversely correlate with insulin sensitivity in humans and protect mice from high-fat-diet-induced insulin resistance by suppressing specific AKT phosphatases. Mechanistically, METTL14-mediated m6A promotes the decay of PTGES2 and CBR1, the genes encoding PGE2 and PGF2a biosynthesis enzymes, in brown adipocytes via YTHDF2/3. Consistently, BAT-specific knockdown of Ptges2 or Cbr1 reverses the insulin-sensitizing effects in M14KO mice. Overall, these findings reveal a novel biological mechanism through which m6A-dependent regulation of the BAT secretome regulates systemic insulin sensitivity.

Integrated metabolomics and lipidomics investigation of the mechanism of Danggui Sini Decoction on improving lipid homeostasis in primary dysmenorrhea

BackgroundDanggui Sini Decoction (DGSND) is a classic prescription for treating primary dysmenorrhea (PD), while, the ameliorating effects of DGSND on PD and its mechanisms are not yet fully understood. PurposeThe present study is devoted to investigate the protective effect of DGSND against PD and the possible mechanism from the perspective of metabolomics as well as lipidomics. MethodsDGSND was characterized by UPLC-Q-TOF/MS. The PD rat model was induced by estradiol benzoate and oxytocin, and traditional pharmacology, including writhing times, latency time, biochemical index, organ index, and histopathology were performed to evaluated the efficacy of DGSND on PD. Urine metabolomics strategy combined with functional analysis was adopted to delineate the therapeutic effect of DGSND on PD rats and anchor the crucial pathway, and lipidomics analysis was further performed with the uterine tissue as the research object to elucidate the protective mechanism of DGSND from the perspective of lipid homeostasis. Finally, western blot analysis was used to validate the expression of key metabolic enzymes in lipid metabolism. ResultsDGSND was effective in ameliorating writhing times, latency time, the value of prostaglandin F2α (PGF2α)/PGE2, uterus index, and morphological changes of PD rats. Metabolic signature of PD rats was primarily characterized by the disturbance of steroid hormone metabolism, amino acid metabolism, and lipid metabolism. Functional analysis revealed the urine biomarkers of PD were most related with lipid abnormality. Further lipidomics analysis indicated DGSND exerted anti-PD effects by remodeling lipid homeostasis, which might be due to the significant correlations between different kinds of lipids, especially the extremely high correlation of phosphatidylethanolamine, phosphatidylcholine, and fatty acids. Moreover, the key metabolic enzymes expression of CK, PLA2, LPCAT3, COX-2, and 5-LOX can be greatly downregulated by DGSND. ConclusionOur findings demonstrated a novel protective mechanism of DGSND against PD by regulating lipid homeostasis.

Hormonal profiles and biomarkers leading to parturition in cattle.

This study aimed to understand the physiological mechanisms regulating parturition and to identify potential biomarkers to predict onset of birth. Additionally, we compared hormone profiles between cows with shorter and longer gestation lengths. Twenty-eight days before due date until 3d postpartum, cows (n = 18) were blood sampled daily. Circulating concentrations were measured for progesterone (P4) and estradiol (E2) by RIA, testosterone, prostaglandin F2α metabolite (PGFM), cortisol, pregnancy-specific protein B (PSPB) by ELISA and lactate concentrations by colorimetric assay. At end of gestation, P4 decreased from d-14 to d-4 (from 3.6 to 1.4ng/mL), most likely from rapid loss of placental P4 production (64% of decline in 24h). A second rapid decrease in P4 to undetectable concentrations was observed from d-2 to parturition (from 1.4 to 0.1ng/ml; most likely luteal origin) corresponding to increase in PGFM from d-2 to parturition (249.7 to 2868.4pg/mL). Estradiol and PSPB increased ~8-fold from ~13d before parturition with acute rise in E2 but not PSPB (45% vs 13% in first 24h). Testosterone decreased slightly during the same period. Cortisol and lactate increased only at calving. Comparison of cows with shorter vs longer gestation, when data were normalized to parturition day, a difference was detected in circulating E2 and PGFM patterns, but not P4 and PSPB. Thus, the first significant hormonal changes associated with parturition begin at d-14 with E2 and PSPB as two clear biomarkers of impending parturition. Cows with shorter and longer gestation had hormonal differences indicative of identifiable earlier placental maturation.

Comparative study between intralesional injection of prostaglandin F2α alone or combined with corticosteroids in the treatment of alopecia areata

Background Alopecia areata (AA) is an autoimmune disorder causing hair loss, yet its treatment remains challenging due to limited Food and Drug Administration approved therapies. Prostaglandin F2α (PGF2α) has emerged as a potential therapy. Objective To assess the efficacy of PGF2α intralesional injection in AA treatment. Patients and methods This case comparative work was performed on 75 localized AA patients who were split into three groups receiving different intralesional injections: group A received triamcinolone acetonide (TRA), group B received a combination of TRA and latanoprost (a PGF2α analog), and group C received latanoprost alone. Treatment response was assessed using the Severity of Alopecia Tool score, Severity of Alopecia Tool II, and Alopecia Areata Severity Index. Results High treatment response was observed in 76% of group B, compared to 64% in group A and 40% in group C. Group B showed the most significant reduction in severity score (median=0), followed by group A (median=1), and group C (median=3). Multivariate analysis identified group B as a significant predictor of a high treatment response (odds ratio=10.292, 95% confidence interval=1.904–55.632, P=0.007). Conclusion Intralesional injection of TRA and latanoprost (PGF2α analog) demonstrated superior efficacy in treating localized AA compared to other treatment modalities.

Effect of PG-600 dose in cows with persistent corpus luteum treated with PGF2α and hCG

The purpose of this study was to determine the effect of administering a dose of gonadotropin (PG-600) in cows with persistent corpus luteum (PCL) which were treated with prostaglandin F2 α (PGF2α) and human chorionic gonadotropin (hCG). This study used 20, three-year-old cows diagnosed with PCL, had recently given birth, and were not pregnant. Cows were randomly allocated into groups T0, T1, T2 and T3. Cows in all groups were injected with PGF2α (25 mg/cow). Simultaneously, cows in groups T1, T2 and T3 were also injected with PG-600 at doses of 100, 150 and 200 IU/cow respectively. PGF2α and PG-600 were injected when PCL was identified. After that, cows showing signs of estrus were injected with hCG (100 IU/cow) and artificially inseminated 12-18 hours after the appearance of signs of estrus. AI was conducted using frozen thawed semen. Sixty days following AI, a pregnancy examination was conducted using USG. Estrus was detected in all treated cows, while the pregnancy rates varied. Compared with other groups, the groups given 100 and 150 IU PG-600 produced higher pregnancy rates. It could be concluded that the combination of 25 mg PGF2α, 150 IU PG-600, and 100 IU hCG was the most effective treatment to produce high pregnancy rates in cows with persistent corpus luteum.

Combining the Vaginal Microbiome and Serum Metabolome to Screen for Potential Biomarkers of Early Pregnancy in Cows.

Early pregnancy diagnostic techniques are of significant importance in livestock farming, particularly in dairy farming. This study aimed to screen artificially inseminated cows for potential biomarkers at day 21 of pregnancy using microbiota-metabolomics analysis. The microbiome analysis revealed significant changes (p < 0.05) in the composition and abundance of the vaginal microbiota in cows after pregnancy. Notably, there was an increase in the abundance of [Eubacterium]_hallii_group (p < 0.05) associated with the production of short-chain fatty acids in the pregnant group compared with the non-pregnant group. Furthermore, significant alterations were observed in the serum metabolome, with notable changes in the concentrations of prolyl-hydroxyproline (Pro-Hyp) (p < 0.01) and bonactin (p < 0.01). The majority of differential metabolites clustered within the pathways of amino acid metabolism and lipid metabolism, with lipid metabolism exhibiting a higher proportion and playing a pivotal role in early pregnancy. An enzyme-linked immunosorbent assay was employed to quantify three key metabolites of the arachidonic acid pathway. The results demonstrated significant decreases in serum concentrations of leukotriene B4 (LTB4) (p < 0.05) and prostaglandin F2α (PGF2α) (p < 0.01) and no significant changes in arachidonic acid (AA) (NS) concentrations after 21 days of gestation in cows. Spearman's correlation analysis was utilized to investigate the interrelationship between the vaginal microbiota and serum metabolites. In conclusion, the present study demonstrated that biomaterials such as bonactin, Pro-hyp, LTB4, PGF2α in serum metabolites and [Eubacterium]_hallii_group in the vaginal flora of cows could be utilized as potential biomarkers for 21 days of gestation in cows.

Restoration of cervical lymphatic vessel function in aging rescues cerebrospinal fluid drainage.

Cervical lymphatic vessels (cLVs) have been shown to drain solutes and cerebrospinal fluid (CSF) from the brain. However, their hydrodynamical properties have never been evaluated in vivo. Here, we developed two-photon optical imaging with particle tracking in vivo of CSF tracers (2P-OPTIC) in superficial and deep cLVs of mice, characterizing their flow and showing that the major driver is intrinsic pumping by contraction of the lymphatic vessel wall. Moreover, contraction frequency and flow velocity were reduced in aged mice, which coincided with a reduction in smooth muscle actin expression. Slowed flow in aged mice was rescued using topical application of prostaglandin F2α, a prostanoid that increases smooth muscle contractility, which restored lymphatic function in aged mice and enhanced central nervous system clearance. We show that cLVs are important regulators of CSF drainage and that restoring their function is an effective therapy for improving clearance in aging.

Fu's subcutaneous needling combined with monkshood cake-separated moxibustion for primary dysmenorrhea with cold congealing and blood stasis: a randomized controlled trial

To observe the clinical efficacy of Fu's subcutaneous needling combined with monkshood cake-separated moxibustion for primary dysmenorrhea with cold congealing and blood stasis. Sixty patients with primary dysmenorrhea of cold congealing and blood stasis were randomly divided into an observation group (30 cases, 1 case dropped out) and a control group (30 cases, 2 cases dropped out). The control group received monkshood cake-separated moxibustion at Shenque (CV 8) and bilateral Zigong (EX-CA 1), while the observation group received Fu's subcutaneous needling based on the control group. The muscles were palpated and the affected muscles were determined. Needles were inserted 5-10 cm away from the affected muscles and reperfusion activity was performed simultaneously. All the treatment started on the first day of menstrual cycle pain, once a day, for 3 days, totaling for 3 menstrual cycles. The visual analogue scale (VAS) score, Cox menstrual symptom scale (CMSS) score, and traditional Chinese medicine (TCM) syndrome score in the two groups were observed before treatment, after 2 treatment courses and after 3 treatment courses. The serum prostaglandin F2α(PGF2α) levels before and after 3 treatment courses were measured, and the clinical efficacy of the two groups was evaluated. After 2 and 3 treatment courses, the VAS scores, CMSS scores, and TCM syndrome scores in the two groups were lower than those before treatment (P<0.05), and the scores in the observation group were lower than those in the control group (P<0.05). After 3 treatment courses, the PGF2α level in the observation group was decreased (P<0.05), and were lower than that in the control group (P<0.05). The total effective rate was 96.6% (28/29) in the observation group, which was higher than 64.3% (18/28) in the control group (P<0.05). Fu's subcutaneous needling combined with monkshood cake-separated moxibustion could effectively reduce the pain intensity, improve clinical symptoms of dysmenorrhea, and lower PGF2α level in patients with primary dysmenorrhea of cold congealing and blood stasis.