This study aimed to investigate the therapeutic effect of Leonuri Herba aqueous decoction on primary dysmenorrhea(PD) and explore the underlying mechanism in conjunction with untargeted metabolomics. Forty adult female rats were randomly divi-ded into a normal group, a model control group, ibuprofen(0.12 g·kg~(-1)) group, and high-and low-dose Leonuri Herba aqueous decoction(5 and 2.5 g·kg~(-1)) groups, with eight rats in each group. The PD rat model was prepared using intramuscular injection of estradiol benzoate combined with intraperitoneal injection of pitocin. Drugs were administered by gavage from the 4th day of modeling for 7 d. After the last administration, pitocin was injected intraperitoneally, and the writhing latency and writhing times within 30 min were recorded. The uterine and ovarian coefficients were determined. Estradiol(E_2), progesterone(Prog), oxytocin(OT), cyclooxyge-nase 2(COX-2), prostaglandin E_2(PGE_2), prostaglandin F_(2α)(PGF_(2α)), and Ca~(2+) levels in uterine tissues were measured by ELISA and biochemical kits. Morphological changes in uterine and ovarian tissues were observed by hematoxylin-eosin(HE) staining. The protein expression of oxytocin receptor(OTR), prostaglandin E_2 receptor 3(EP3), and estrogen receptor alpha(ERα) in uterine tissues was detected by immunohistochemistry. The mRNA expression of OTR, PGE_2 receptors 1-4(EP1, EP2, EP3, and EP4), and PGF_(2α) receptor(FP) in uterine tissues was detected by quantitative real-time PCR. Untargeted metabolomics analysis was performed by ultra-high-performance liquid chromatography-quadrupole-time-of-flight mass spectrometry(LC-QTOF-MS) technology to screen potential biomarkers and enrich metabolic pathways. The results showed that Leonuri Herba was able to significantly reduce the writhing times in PD rats(P<0.05 or P<0.01), significantly reduce the uterine and ovarian coefficients(P<0.01), and improve their histomorphology. After treatment with Leonuri Herba, PGE_2 content was significantly increased(P<0.05), COX-2, PGF_(2α) and Ca~(2+) content, and PGF_(2α)/PGE_2 was significantly decreased(P<0.05 or P<0.01), and OT content was decreased, while E_2 and Prog content tended to further increase in uterine tissues of PD rats. Correspondingly, OTR and EP3 protein expression was significantly downregulated(P<0.05 or P<0.01) and ERα protein expression was upregulated(P<0.05) in uterine tissues. The mRNA expression of FP and EP4 in uterine tissues was significantly downregulated(P<0.01), and the mRNA expression of EP1, EP3, and OTR showed a decreasing trend. The untargeted metabolomics results showed that 10 differential metabolites were restored in the plasma of PD rats after Leonuri Herba treatment. The results indicate that Leonuri Herba is effective in the prevention and treatment of PD, and the underlying mechanism may be attributed to the regulation of PGs synthesis and corresponding receptor binding.
Humans ; Rats ; Female ; Animals ; Estrogen Receptor alpha ; Oxytocin ; Dysmenorrhea/metabolism* ; Cyclooxygenase 2 ; Dinoprostone ; RNA, Messenger/metabolism* ; Dinoprost

OBJECTIVE: To observe the effects of electroacupuncture (EA) on NLRP3 inflammasome and its downstream protein gastermin D (GSDMD) in rats with primary dysmenorrhea (PDM), and to explore the potential mechanism of EA on the treatment of PDM. METHODS: Forty healthy female SD rats without pregnancy were randomly divided into a control group, a model group, an EA group and an ibuprofen group, 10 rats in each group. PDM model was prepared by injection of estradiol benzoate and oxytocin. Except the control group, the rats in each group were subcutaneously injected with estradiol benzoate for 10 days, and oxytocin was injected on the 11th day. The rats in the EA group were intervened with EA (dense wave, frequency of 50 Hz) at "Guanyuan" (CV 4) and "Sanyinjiao" (SP 6) at the same time of modeling, once a day, 20 min each time, for 10 consecutive days. The rats in the ibuprofen group were treated with 0.8 mL of ibuprofen by gavage (concentration of ibuprofen solution was 1.25 mg/mL) for 10 consecutive days. After modeling, the writhing reaction was observed. After intervention, the HE staining method was used to observe the histological morphology of uterus and evaluate the pathological damage score of uterus; ELISA method was used to detect the serum levels of prostaglandin E2 (PGE2) and prostaglandin F2α (PGF2α); Western blot method was used to detect the protein expression of NLRP3, apoptosis related spot like protein (ASC), caspase-1, GSDMD, GSDMD-N and inflammatory factors (interleukin [IL]-1β, IL-18) in uterine tissue. RESULTS: In the model group, a large number of vacuolar degeneration and death of endometrial epithelial cells, spiral arterioles congestion in lamina propria and neutrophil infiltration were observed. In the EA group, there was a small amount of vacuolar degeneration and death of endometrial epithelial cells, a small amount of spiral arterioles congestion in the lamina propria, and a small amount of neutrophils infiltration. In the ibuprofen group, there was very small number of degeneration and death of endometrial epithelial cells, and no obvious arterial congestion was found in lamina propria, and neutrophil infiltration was occasionally seen. Compared with the control group, in the model group the number of writhing was increased (P<0.01), the writhing reaction score and serum level of PGF2α and PGF2α/PGE2 value were increased (P<0.01), the level of PGE2 was decreased (P<0.01). Compared with the model group, in the EA group and the ibuprofen group the number of writhing were decreased (P<0.05), the latency of writhing was prolonged (P<0.01), the writhing reaction scores and serum levels of PGF2α and PGF2α/PGE2 values were decreased (P<0.05, P<0.01), the levels of PGE2 were increased (P<0.01). Compared with the control group, the protein expression of NLRP3, ASC, caspase-1, GSDMD, GSDMD-N, IL-1β and IL-18 in the uterine tissues of rats was increased in the model group (P<0.01). Compared with the model group, the protein expression of NLRP3, ASC, caspase-1, GSDMD, GSDMD-N, IL-1β and IL-18 in the uterine tissues of rats was decreased in the EA group and the ibuprofen group (P<0.01, P<0.05). There was no significant difference between the EA group and the ibuprofen group in the above indexes (P>0.05). CONCLUSION EA could alleviate pain and uterine tissue injury in rats with PDM. The mechanism may be related to the inhibition of the activation of NLRP3 inflammasome in rat uterine tissues, thereby inhibiting pyroptosis and its inflammatory factors release.
Animals ; Female ; Pregnancy ; Rats ; Caspases ; Dinoprost ; Dinoprostone ; Dysmenorrhea ; Electroacupuncture ; Ibuprofen ; Inflammasomes ; Interleukin-18 ; NLR Family, Pyrin Domain-Containing 3 Protein ; Oxytocin ; Phosphate-Binding Proteins ; Pyroptosis ; Rats, Sprague-Dawley ; Uterus

To explore the anti-platelet aggregation and anti-thrombotic mechanisms of Trichosanthis Fructus combined with aspirin based on network pharmacology and the validation of arteriovenous by pass model in rats. The databases of Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),Drug Repositioning and Adverse Drug Reaction Chemical-Protein Interactome(DRAR-CPI),Universal Protein Resource(Uniprot) and the Database for Annotation,Visualization,and Integrated Discovery(DAVID) were used to predict protein targets and analyze biological pathway and signal pathway in the combination of Trichosanthis Fructus with aspirin. The effects of pretreatment with Trichosanthis Fructus pellets,aspirin pellets and their combination on thromboxane B2(TXB2),6-keto prostaglandin F1α(6-keto-PGF1α) and cyclic adenosine monophosphate(c AMP) in rat thrombotic model were studied. Through the study of network pharmacology,12 components of aspirin and Trichosanthis Fructus,including hydroxygenkwanin,quercetin and adenosine,were found to show the anti-platelet aggregation and anti-thrombosis mechanisms through9 common protein targets,such as SRC,RAC1,MAPK14,MAPK1,AKT1,and 14 common signaling pathways,such as VEGF signaling pathway. After the intervention with Trichosanthis Fructus pellets combined with aspirin pellets,the vascular endothslia growth factor(VEGF) signaling pathway can be activated to inhibit platelet aggregation and improve vascular endothelial function,and show the anti-platelet aggregation and anti-thrombosis mechanisms,which verify the results of the network pharmacology,and explain the anti-platelet aggregation and anti-thrombotic mechanisms of the combination of Trichosanthis Fructus pellets with aspirin pellets.
6-Ketoprostaglandin F1 alpha ; metabolism ; Animals ; Aspirin ; pharmacology ; Cyclic AMP ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Fruit ; chemistry ; Platelet Aggregation ; drug effects ; Platelet Aggregation Inhibitors ; pharmacology ; Rats ; Signal Transduction ; Thrombosis ; drug therapy ; Thromboxane B2 ; metabolism ; Trichosanthes ; chemistry

OBJECTIVETo investigate the effect of total flavone of haw leaves (TFHL) on the expression of nuclear factor erythroid-2 related factor (Nrf2) and other related factors in nonalcoholic steatohepatitis (NASH) rats induced by high-fat diet and then to further discuss the mechanism of TFHL's prevention against NASH.

METHODSHigh-fat diet was fed to 40 rats to establish the NASH model. Then model rats were intragastrically administrated with 40, 80, 160 mg/(kg•day) TFHL, respectively. The pathological changes of liver tissues in NASH rats were detected by oil red O and hematoxylin-eosin (HE) stainings. The expression of Nrf2 in rat liver was examined through immunohistochemistry. The level of 8-iso-prostaglandin F2&alpha; in serum was detected through enzyme linked immunosorbent assay (ELISA). The mRNA and protein levels of Nrf2 and other related factors in liver tissue were measured by real-time reverse transcriptionpolymerase chain reaction and western blot.

RESULTSLipid deposition, hepatic steatosis, focal necrosis in lobular inflammation and ballooning degeneration were emerged in livers of NASH rats. The 8-iso-prostaglandin F2&alpha; in the serum of NASH rats increased significantly compared with the control group (P<0.05). The mRNA of Nrf2, hemeoxyenase1 (HO-1) and the mRNA and protein levels of quinine oxidoreductase (NQO1) in NASH rats liver tissue showed a striking increase, while the mRNA levels of Keap1, r-glutamylcysteine synthethase (rGCS) and glutathione S-transferase (GST) were significantly decreased compared with the control group (P<0.05). After TFHL treatment, 8-iso-prostaglandin F2&alpha; level in serum significantly decreased, and Nrf2 mRNA and protein levels in hepatocytes nucleus enhanced compared with the model group (P<0.05 or 0.01). Meanwhile the Keap1 mRNA, the mRNA and protein levels of HO-1, NQO1 antibody, rGCS antibody, GST increased after TFHL treatment (P<0.05 or 0.01).

CONCLUSIONSNrf2 and other related factors were involved in development of NASH, and they also served as an important part in its occurrence. By regulating expression of Nrf2 and other related factors, TFHL may play a role in antioxidative stress and prevention of NASH.

Animals ; Cell Nucleus ; drug effects ; metabolism ; Crataegus ; chemistry ; Dinoprost ; metabolism ; Flavones ; pharmacology ; therapeutic use ; Lipids ; chemistry ; Liver ; drug effects ; metabolism ; pathology ; NF-E2-Related Factor 2 ; genetics ; metabolism ; Non-alcoholic Fatty Liver Disease ; drug therapy ; genetics ; pathology ; Phytotherapy ; Plant Leaves ; chemistry ; RNA, Messenger ; genetics ; metabolism ; Rats, Sprague-Dawley

OBJECTIVETo observe the effects of Danhong Injection (丹红注射液) and its main components, including daiclzein and hydroxysafflor yellow A (HSYA), on the anticoagulation, fibrinolysis, anti-apoptosis in hypoxia model of vein endothelial cells (VECs).

METHODSVECs were prepared and were put in a hypoxia environment, which consisted of mixed gas of 95% N and 5% CO mixed gas, when reached confluent culture. Five groups used different treatments, including normal control group, hypoxia group, daiclzein group, HSYA group and Danhong Injection group. The VECs were identified by fluorescence double labeling methods. The morphology was observed by a phase contrast microscopy. The effects of Danhong Injection, daiclzein and HSYA on 6 keto prostaglandin F1&alpha; (6-keto-PGF1&alpha;) level was measured by the method of radioimmunoassay (RIA). Superoxide dismutase (SOD) activity was tested by water soluble tetrazolium salt. The content of malondialdehyde (MDA) was measured by thiobarbituric acid. The activities of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI) were measured by the method of chromogenic substrate. The contents of endothelin (ET) and nitric oxide (NO) were detected by non-equilibrium RIA and enzymelinked immunosorbent assay. Cells apoptosis rate was determined by flow cytometry.

RESULTSCompared with the normal control group, the floating cells number, PAI activity, ET and MDA contents, and cells apoptosis rate in the culture solution of hypoxia group were all significantly increased, whereas the 6-keto-PGF1&alpha; and NO contents, and t-PA and SOD activities were decreased significantly (P<0.01). Compared with the hypoxia group, Danhong Injection markedly increased the 6-keto-PGF1&alpha; content and SOD activity, regulated PAI and t-PA activities, ET and NO contents, and decreased MDA content and cells apoptosis rate (P<0.05 or P<0.01).

CONCLUSIONSDanhong Injection and its main components played an important role in protecting primary VECs from hypoxic damage by regulating the secretion and vasomotor function of VECs. The function of Danhong Injection was most remarkable.

6-Ketoprostaglandin F1 alpha ; metabolism ; Animals ; Apoptosis ; drug effects ; Blood Coagulation ; drug effects ; Cell Count ; Cells, Cultured ; Drugs, Chinese Herbal ; pharmacology ; Endothelial Cells ; drug effects ; metabolism ; Endothelins ; metabolism ; Factor VIII ; metabolism ; Fibrinolysis ; drug effects ; Fluorescent Antibody Technique ; Humans ; Infant, Newborn ; Injections ; Malondialdehyde ; metabolism ; Nitric Oxide ; metabolism ; Plasminogen Inactivators ; metabolism ; Rabbits ; Superoxide Dismutase ; metabolism ; Tissue Plasminogen Activator ; metabolism ; Umbilical Veins ; cytology

Cold Temperature ; Dinoprost ; analogs & derivatives ; urine ; Dust ; Humans ; Interleukin-6 ; urine ; Thromboxane B2 ; analogs & derivatives ; urine ; Weather

Bovine embryos (day 5) were cultured to day 10 with or without 100 ng/mL PGF2α in medium supplemented with control; 100 nM Dex; 1,000 U/mL recombinant human leukemia inhibitory factor (rhLIF); or Dex+rhLIF. Although the rates to development to the blastocyst were not significantly different among groups, the hatching rate after additional culture with Dex +/or rhLIF was significantly higher in all supplemented groups than the control (p < 0.05). In the presence of PGF2α, the hatching rate was significantly restored in all supplemented groups relative to the group treated with only PGF2α and the control (p < 0.05). Embryo transfer (ET) was performed with blastocysts (day 7). PGF2α levels of control recipient cows were significantly higher in the circulatory blood samples collected 60 min after ET than in samples collected 60 min before ET (p < 0.005), and were decreased in cows injected with loading medium supplemented with Dex+rhLIF (p < 0.005). Pregnancy rate was significantly higher in the ET group that received supplemented embryo-loading medium than in the non-supplemented control (p < 0.05). The intrauterine administration of Dex and rhLIF at ET prevented increased PGF2α in circulatory blood and resulted in enhanced pregnancy rate.
Animals ; Blastocyst ; Cattle* ; Dexamethasone* ; Embryo Transfer* ; Embryonic Structures* ; Fertilization in Vitro ; Humans* ; Leukemia Inhibitory Factor* ; Leukemia* ; Pregnancy Rate* ; Pregnancy* ; Prostaglandins F

This study was to investigate whether working in conditions of elevated concentrations of mine gases (CO2, CO, CH4, DMS) and dust may result in oxidative stress. Coal miners (n=94) from the Velenje Coal mine who were arranged into control group and three groups according to a number of consecutive working days. 8-isoprostane as a biological marker of oxidative stress was measured in exhaled breath condensate (EBC). Miners who worked for three consecutive days had higher 8-isoprostane values in EBC compared to the control group. Gas/dust concentrations and exposure time of a single/two day shift seem too low to trigger immediate oxidative stress.
Adult ; Biomarkers ; analysis ; Breath Tests ; Coal ; adverse effects ; Coal Mining ; manpower ; Dinoprost ; analogs & derivatives ; analysis ; Dust ; analysis ; Humans ; Male ; Middle Aged ; Miners ; statistics & numerical data ; Occupational Exposure ; analysis ; Oxidative Stress

The objective of this study was to investigate factors that influence the success of resynchronization protocols for bovines with and without progesterone supplementation. Cow synchronized and not found pregnant were randomly assigned to two resynchronization protocols: ovsynch without progesterone (P4) supplementation (n = 66) or with exogenous P4 administered from Days 0 to 7 (n = 67). Progesterone levels were measured on Days 0 and 7 of these protocols as well as 4 and 5 days post-insemination. Progesterone supplementation raised the P4 levels on Day 7 (p < 0.05), but had no overall effect on resynchronization rates (RRs) or pregnancy per artificial insemination (P/AI). However, cows with Body Condition Score (BCS) > 3.5 had increased P/AI values while cows with BCS < 2.75 had decreased P/AI rates after P4 supplementation. Primiparous cows had higher P4 values on Day 7 than pluriparous animals (p = 0.04) and tended to have higher RRs (p = 0.06). Results of this study indicate that progesterone supplementation in resynchronization protocols has minimal effects on outcomes. Parity had an effect on the levels of circulating progesterone at initiation of the protocol, which in turn influenced the RR.
Animals ; Cattle/*physiology ; Dinoprost/administration & dosage/*pharmacology ; Estrus Synchronization/*drug effects/methods ; Female ; Fertility Agents/administration & dosage/pharmacology ; Gonadotropin-Releasing Hormone/administration & dosage/*pharmacology ; Insemination, Artificial/veterinary ; Ovulation/drug effects ; Pregnancy ; Progesterone/administration & dosage/*pharmacology ; Tromethamine/administration & dosage/*pharmacology

OBJECTIVETo investigate the effect of chloroquine on airway hyperresponsiveness in asthmatic mice and explore the possible mechanism.

METHODSBalb/c mouse models of asthma established using OVA received intraperitoneal injections of chloroquine, dexamethasone, or both prior to OVA challenge. Within 24 h after the final challenge, airway hyper- responsiveness (AHR) of the mice was assessed, and the total cell count and the counts of different cell populations in the bronchoalveolar lavage fluid (BALF) were determined under light microscopy. The severity of lung inflammation was evaluated using HE staining, and the concentrations of IL-6 and PGF2&alpha; in the BALF were detected by enzyme-linked immunosorbent assay (ELISA).

RESULTSChloroquine pretreatment significantly decreased AHR (P<0.001) in the asthmatic mice and reduced the total cell count (P<0.01), eosinophils (P<0.001), neutrophils (P<0.01), and PGF2&alpha; levels in the BALF. Chloroquine combined with low-dose dexamethasone significantly lessened inflammations around the bronchioles (P<0.05) and blood vessels (P<0.01) in the lung tissue, and obviously lowered IL-6 (P<0.05) and PGF2&alpha; (P<0.001) in the BALF in the asthmatic mice.

CONCLUSIONChloroquine can inhibit AHR in asthmatic mice and produce better anti-inflammatory effect when combined with dexamethasone for treatment of neutrophilic asthma.

Animals ; Asthma ; chemically induced ; drug therapy ; Bronchoalveolar Lavage Fluid ; cytology ; Chloroquine ; pharmacology ; Dexamethasone ; pharmacology ; Dinoprost ; metabolism ; Disease Models, Animal ; Enzyme-Linked Immunosorbent Assay ; Eosinophils ; cytology ; Inflammation ; pathology ; Interleukin-6 ; metabolism ; Leukocyte Count ; Lung ; pathology ; Mice ; Mice, Inbred BALB C ; Neutrophils ; cytology