Paeonol alleviates postmenopause-induced neuropsychiatric symptoms through the modulation of GPR30 in ovariectomized mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The Moutan cortex (MC), the root bark of Paeonia suffruticosa Anderws (Paeoniaceae), has been historically employed in traditional herbal medicine for addressing women's ailments by replenishing kidney Yin. AIM OF THE STUDY: We aimed to explore if paeonol, an active constituent of MC, could ameliorate neuropsychiatric symptoms, such as anxiety, depression, and cognitive impairments, associated with post-menopausal syndrome (PMS) in an ovariectomized (OVX) mouse model. MATERIALS AND METHODS: The experimental design comprised 6 groups, including a sham group, OVX group, paeonol administration groups (3, 10 or 30 mg/kg, p.o.), and an estradiol (E2)-treated positive control group. Behavioral tests including the open field, novel object recognition, Y-maze, elevated plus-maze, splash, and forced swimming tests were conducted. In addition, we investigated the effets of paeonol on the phosphorylated levels of phosphatidylinositol 3-kinase (PI3K), Akt, and mammalian target of rapamycin (mTOR), as well as on the expression levels of G protein-coupled receptor (GPR30) and brain-derived neurotrophic factor (BDNF) in the prefrontal cortex and hippocampus. RESULTS: Paeonol treatment (10 and 30 mg/kg, p.o.) effectively reversed the cognitive decline in OVX mice, measured by the novel object recognition and Y-maze tests, similar to that in the positive control group. Additionally, it alleviated anxiety- and depressive-like behaviors, as evaluated by the elevated plus-maze test, splash test, and forced swimming test. Paeonol restored GPR30 expression levels in the prefrontal cortex and hippocampus, mirroring the effects of E2 administration. Furthermore, it reversed the reduced expression levels of the PI3K-Akt-mTOR signaling pathway in the prefrontal cortex and hippocampus and increased BDNF expression in the hippocampus of OVX mice. CONCLUSION: This research suggests that paeonol would be beneficial for alleviating PMS-associated cognitive impairment, anxiety and depression.

MicroRNA let-7g links foam cell formation and adipogenic differentiation: A key regulator of Paeonol treating atherosclerosis-osteoporosis.

BACKGROUD: High comorbidity rates have been reported in patients with atherosclerosis and osteoporosis, posing a serious risk to the health and well-being of elderly patients. To improve and update clinical practice regarding the joint treatment of these two diseases, the common mechanisms of atherosclerosis and osteoporosis need to be clarified. MicroRNAs (miRNAs), are importance molecules in the pathogenesis of human diseases, including in cardiovascular and orthopedic fields. They have garnered interest as potential targets for novel therapeutic strategies. However, the key miRNAs involved in atherosclerosis and osteoporosis and their precise regulation mechanisms remain unknown. Paeonol (Pae), an active ingredient in Cortex Moutan, has shown promising results in improving both lipid and bone metabolic abnormalities. However, it is uncertain whether this agent can exert a cotherapeutic effect on atherosclerosis and osteoporosis. OBJECTIVE: This study aimed to screen important shared miRNAs in atherosclerotic and osteoporotic complications, and explore the mechanism of the protective effects of Pae against atherosclerosis and osteoporosis in high-fat diet (HFD)-fed ApoE-/- mice. METHODS: An experimental atherosclerosis and osteoporosis model was established in 40-week-old HFD ApoE-/- mice. Various techniques such as Oil Red O staining, HE staining and micro-CT were used to confirm the co-occurrence of these two diseases and efficacy of Pae in addition to the associated biochemical changes. Bioinformatics was used to screen key miRNAs in the atherosclerosis and osteoporosis model, and gene involvement was assessed through serum analyses, qRT-PCR, and western blot. To investigate the effect of Pae on the modulation of the miR let-7g/HMGA2/CEBPß pathway, Raw 264.7 cells were cocultured with bone marrow mesenchymal stem cells (BMSCs) and treated with an miR let-7g mimic/inhibitor. RESULTS: miR let-7g identified using bioinformatics was assessed to evaluate its participation in atherosclerosis-osteoporosis. Experimental analysis showed reduced miR let-7g levels in the atherosclerosis-osteoporosis mice model. Moreover, miR let-7g was required for BMSC - Raw 264.7 cell crosstalk, thereby promoting foam cell formation and adipocyte differentiation. Treatment with Pae significantly reduced plaque accumulation and foam cell number in the aorta while increasing bone density and improving trabecular bone microarchitecture in HFD ApoE-/- mice. Pae also increased the level of miR let-7g in the bloodstream of model mice. In vitro studies, Pae enhanced miR let-7g expression in BMSCs, thereby suppressing the HMGA2/CEBPß pathway to prevent the formation of foam cells and differentiation of adipocytes induced by oxidized low-density lipoprotein (ox-LDL). CONCLUSION: The study results suggested that miR let-7g participates in atherosclerosis -osteoporosis regulation and that Pae acts as a potential therapeutic agent for preventing atherosclerosis-osteoporosis through regulatory effects on the miR let-7g/HMGA2/CEBPß pathway to hinder foam cell formation and adipocyte differentiation.

Ultrasound-Assisted Extraction of Paeonol from Moutan Cortex: Purification and Component Identification of Extract.

Moutan Cortex (MC) is a traditional Chinese medicine that contains abundant medicinal components, such as paeonol, paeoniflorin, etc. Paeonol is the main active component of MC. In this study, paeonol was extracted from MC through an ultrasound-assisted extraction process, which is based on single-factor experiments and response surface methodology (RSM). Subsequently, eight macroporous resins of different properties were used to purify paeonol from MC. The main components of the purified extract were identified by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS/MS). The results indicate the optimal parameters are as follows: liquid-to-material ratio 21:1 mL/g, ethanol concentration 62%, ultrasonic time 31 min, ultrasonic temperature 36 °C, ultrasonic power 420 W. Under these extraction conditions, the actual yield of paeonol was 14.01 mg/g. Among the eight tested macroporous resins, HPD-300 macroporous resin was verified to possess the highest adsorption and desorption qualities. The content of paeonol increased from 6.93% (crude extract) to 41.40% (purified extract) after the HPD-300 macroporous resin treatment. A total of five major phenolic compounds and two principal monoterpene glycosides were characterized by comparison with reference compounds. These findings will make a contribution to the isolation and utilization of the active components from MC.

Protective mechanism of Paeonol on central nervous system.

Cerebrovascular diseases involve neuronal damage, resulting in degenerative neuropathy and posing a serious threat to human health. The discovery of effective drug components from natural plants and the study of their mechanism are a research idea different from chemical synthetic medicines. Paeonol is the main active component of traditional Chinese medicine Paeonia lactiflora Pall. It widely exists in many medicinal plants and has pharmacological effects such as anti-atherosclerosis, antiplatelet aggregation, anti-oxidation, and anti-inflammatory, which keeps generally used in the treatment of cardiovascular and cerebrovascular diseases. Based on the therapeutic effects of Paeonol for cardiovascular and cerebrovascular diseases, this article reviewed the pharmacological effects of Paeonol in Alzheimer's disease, Parkinson's disease, stroke, epilepsy, diabetes encephalopathy, and other neurological diseases, providing a reference for the research of the mechanism of Paeonol in central nervous system diseases.

[Study on biomarkers of acteoside in treating puromycin aminonucleoside nephropathy in young rats based on non-targeted urine metabolomics technology].

This study aims to reveal the endogenous metabolic characteristics of acteoside in the young rat model of purinomycin aminonucleoside nephropathy(PAN) by non-targeted urine metabolomics and decipher the potential mechanism of action. Biochemical indicators in the urine of rats from each group were determined by an automatic biochemical analyzer. The potential biomarkers and related core metabolic pathways were identified by ultra-high performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry(UHPLC-LTQ-Orbitrap MS) combined with principal component analysis(PCA) and orthogonal partial least squares-discriminant analysis(OPLS-DA). MetaboAnalyst 5.0 was used to establish the receiver operating characteristic(ROC) curve for evaluating the clinical diagnostic performance of core metabolites. The results showed that acteoside significantly decreased urinary protein-to-creatinine ratio in PAN young rats. A total of 17 differential metabolites were screened out by non-targeted urine metabolomics in PAN young rats and they were involved in phenylalanine metabolism and phenylalanine, tyrosine and tryptophan biosynthesis. Thirtten differential metabolites were screened by acteoside intervention in PAN young rats, and they were involved in phenylalanine metabolism and arginine and proline metabolism. Among them, leucylproline and acetophenone were the differential metabolites that were significantly recovered after acteoside treatment. These pathways suggest that acteoside treats PAN in young rats by regulating amino acid metabolism. The area under the curve of two core biomarkers, leucylproline and acetophenone, were both greater than 0.9. In summary, acteoside may restore amino acid metabolism by regulating endogenous differential metabolites in PAN young rats, which will help to clarify the mechanism of acteoside in treating chronic glomerulonephritis in children. The characteristic biomarkers screened out have a high diagnostic value for evaluating the treatment of chronic glomerulonephritis in children with acteoside.

Paeonol ameliorates hippocampal neuronal damage by inhibiting GRM5/GABBR2/ß-arrestin2 and activating the cAMP-PKA signaling pathway in premenstrual irritability rats.

Premenstrual dysphoric disorder (PMDD) is a periodic psychiatric disorder with high prevalence in women of childbearing age, seriously affecting patients' work and life. Currently, the international first-line drugs for PMDD have low efficiency and increased side effects. Paeonol, a major component of the traditional Chinese medicine Cortex Moutan, has been applied in treating PMDD in China with satisfactory results, but the therapeutic mechanism is not fully understood. This study aims to evaluate the therapeutic effects and pharmacological mechanisms of paeonol on the main psychiatric symptoms and hippocampal damage in PMDD. We established a premenstrual irritability rat model by the resident-intruder paradigm and performed elevated plus maze and social interactions. And we employed the HE and Nissl staining techniques to observe the therapeutic effect of paeonol on hippocampal damage in PMDD rats. Subsequently, Elisa, qRT-PCR Array, Western Blotting, and cell models were utilized to elucidate the underlying molecular mechanisms through which paeonol intervenes in treating PMDD. In this study, we demonstrated the therapeutic effects of paeonol on irritability, anxiety, and social withdrawal behaviors in rats. In addition, we found that paeonol significantly reduced the serum corticosterone (CORT) level, improved hippocampal morphological structure and neuron number, and reduced hippocampal neuron apoptosis in PMDD rats. Paeonol reduced GRM5, GABBR2, ß-arrestin2, and GRK3 expression levels in hippocampal brain regions of PMDD rats and activated the cAMP/PKA signaling pathway. Inhibitor cell experiments showed that paeonol specifically ameliorated hippocampal injury by modulating the ß-arrestin2/PDE4-cAMP/PKA signaling pathway. The present study demonstrates, for the first time, that paeonol exerts a therapeutic effect on periodic psychotic symptoms and hippocampal injury in PMDD through inhibiting GRM5/GABBR2/ß-arrestin2 and activating cAMP-PKA signaling pathway. These findings enhance our understanding of the pharmacological mechanism underlying paeonol and provide a solid scientific foundation for its future clinical application.

Review of the Protective Mechanism of Paeonol on Cardiovascular Disease.

Cardiovascular disease (CVD) is one of the leading causes of death in the world. Paeonol(Pae) is a phenolic component extracted from peony bark, peony root and Xu Changqing. Studies have shown that Pae can protect cardiomyocytes by inhibiting oxidative stress, promoting mitochondrial fusion, regulating mitochondrial autophagy and inhibiting inflammation. In addition, Pae improves ventricular remodeling by inhibiting myocardial apoptosis, hypertrophy and fibrosis. Pae also has a good protective effect on blood vessels by inhibiting vascular inflammation, reducing the expression of adhesion molecules, inhibiting vascular proliferation, and inhibiting oxidative stress and endoplasmic reticulum stress(ERS). Pae also has the effect of anti-endothelial cell senescence, promoting thrombus recanalization and vasodilating. In conclusion, the molecular targets of Pae are very complex, and the relationship between different targets and signaling pathways cannot be clearly explained, which requires us to use systems biology methods to further study specific molecular targets of Pae. It has to be mentioned that the bioavailability of Pae is poor, and some nanotechnology-assisted drug delivery systems improve the therapeutic effect of Pae. We reviewed the protective mechanism of paeonol on the cardiovascular system, hoping to provide help for drug development in the treatment of CVD.

Paeonol repurposing for cancer therapy: From mechanism to clinical translation.

Paeonol (PAE) is a natural phenolic monomer isolated from the root bark of Paeonia suffruticosa that has been widely used in the clinical treatment of some inflammatory-related diseases and cardiovascular diseases. Much preclinical evidence has demonstrated that PAE not only exhibits a broad spectrum of anticancer effects by inhibiting cell proliferation, invasion and migration and inducing cell apoptosis and cycle arrest through multiple molecular pathways, but also shows excellent performance in improving cancer drug sensitivity, reversing chemoresistance and reducing the toxic side effects of anticancer drugs. However, studies indicate that PAE has the characteristics of poor stability, low bioavailability and short half-life, which makes the effective dose of PAE in many cancers usually high and greatly limits its clinical translation. Fortunately, nanomaterials and derivatives are being developed to ameliorate PAE's shortcomings. This review aims to systematically cover the anticancer advances of PAE in pharmacology, pharmacokinetics, nano delivery systems and derivatives, to provide researchers with the latest and comprehensive information, and to point out the limitations of current studies and areas that need to be strengthened in future studies. We believe this work will be beneficial for further exploration and repurposing of this natural compound as a new clinical anticancer drug.

UPLC-Q-TOF/MS Based Plasma Metabolomics for Identification of Paeonol's Metabolic Target in Endometriosis.

Endometriosis is a common gynecological illness in women of reproductive age that significantly decreases life quality and fertility. Paeonol has been shown to play an important part in endometriosis treatments. Understanding the mechanism is critical for treating endometriosis. In this study, autologous transplantation combined with a 28 day ice water bath was used to create a rat model of endometriosis with cold clotting and blood stagnation. The levels of estradiol and progesterone in plasma were detected by ELISA, and the pathological changes of ectopic endometrial tissue were examined by H&E staining, which proved the efficacy of paeonol. For metabolomic analysis of plasma samples, UPLC-Q/TOF-MS was combined with multivariate statistical analysis to identify the influence of paeonol on small molecule metabolites relevant to endometriosis. Finally, the key targets were screened using a combination of network pharmacology and molecular docking approaches. The results showed that the pathological indexes of rats were improved and returned to normal levels after treatment with paeonol, which was the basis for confirming the efficacy of paeonol. Metabolomics results identified 13 potential biomarkers, and paeonol callbacks 7 of them, involving six metabolic pathways. Finally, four key genes were found for paeonol therapy of endometriosis, and the results of molecular docking revealed a significant interaction between paeonol and the four key genes. This study was successful in establishing a rat model of endometriosis with cold coagulation and blood stagnation. GCH1, RPL8, PKLR, and MAOA were the key targets of paeonol in the treatment of endometriosis. It is also demonstrated that metabolomic techniques give the potential and environment for comprehensively understanding drug onset processes.

A hydroalcoholic extract of Senecio nutans SCh. Bip (Asteraceae); its effects on cardiac function and chemical characterization.

ETHNOPHARMACOLOGY RELEVANCE: The plant Senecio nutans SCh. Bip. is used by Andean communities to treat altitude sickness. Recent evidence suggests it may produce vasodilation and negative cardiac inotropy, though the cellular mechanisms have not been elucidated. PURPOSE: To determinate the mechanisms action of S. nutans on cardiovascular function in normotensive animals. METHODS: The effect of the extract on rat blood pressure was measured with a transducer in the carotid artery and intraventricular pressure by a Langendorff system. The effects on sheep ventricular intracellular calcium handling and contractility were evaluated using photometry. Ultra-high-performance liquid-chromatography with diode array detection coupled with heated electrospray-ionization quadrupole-orbitrap mass spectrometric detection (UHPLC-DAD-ESI-Q-OT-MSn) was used for extract chemical characterization. RESULTS: In normotensive rats, S. nutans (10 mg/kg) reduced mean arterial pressure (MAP) by 40% (p < 0.05), causing a dose-dependent coronary artery dilation and decreased left ventricular pressure. In isolated cells, S. nutans extract (1 µg/ml) rapidly reduced the [Ca2+]i transient amplitude and sarcomere shorting by 40 and 49% (p < 0.001), respectively. The amplitude of the caffeine evoked [Ca2+]i transient was reduced by 24% (p < 0.001), indicating reduced sarcoplasmic reticulum (SR) Ca2+ content. Sodium-calcium exchanger (NCX) activity increased by 17% (p < 0.05), while sarcoendoplasmic reticulum Ca2+-ATPase (SERCA) activity was decreased by 21% (p < 0.05). LC-MS results showed the presence of vitamin C, malic acid, and several antioxidant phenolic acids reported for the first time. Dihydroeuparin and 4-hydroxy-3-(3-methylbut-2-enyl) acetophenone were abundant in the extract. CONCLUSION: In normotensive animals, S. nutans partially reduces MAP by decreasing heart rate and cardiac contractility. This negative inotropy is accounted for by decreased SERCA activity and increased NCX activity which reduces SR Ca2+ content. These results highlight the plant's potential as a source of novel cardio-active phytopharmaceuticals or nutraceuticals.

[Chemical and nutrient differences between medicinal material, residues, and residue compost of Moutan Cortex].

Moutan Cortex(MC) residues produced after the extraction of MC can be re-extracted for active components and used to produce organic fertilizer and animal feed. However, they are currently disposed as domestic waste, which pollutes the environment. This study analyzed the chemical composition of the medicinal material, residues, and residue compost of MC by UPLC-UV-Q-TOF-MS. Furthermore, the nutrient composition of MC residues and the residue compost was analyzed. The results showed that:(1)MC residues had lower content of chemicals than the medicinal material, and content of paeonol, gallic acid, and galloylglucose in MC residues were about 1/3 of that in the medicinal material. The content of chemicals were further reduced after residue composting, and the quantitative compounds were all below the limits of detection.(2)Compared with MC residues, the residue compost showed the total nitrogen, total phosphorus, total potassium, and organic matter content increasing by 122.67%, 31.32%, 120.39%, and 32.06%, respectively. Therefore, we concluded that the MC residues can be used to re-extract active compounds such as paeonol, gallic acid, and galloylglucose. The MC residue compost is a high-quality organic fertilizer containing minimal content of chemicals and can be widely used in the cultivation of Chinese medicinal herbs.

Acronyrones A-C, unusual prenylated acetophenones from Acronychia pedunculata.

Two novel prenylated acetophenones with new carbon skeletons, acronyrones A and B (1 and 2), and a new analogue, acronyrone C (3), together with two known compounds (4 and 5) were isolated from the leaves of Acronychia pedunculata. Their structures with absolute configurations were identified by interpretation of spectroscopic data, single crystal X-ray diffraction, and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 represent the first example of prenylated acetophenones possessed a C7 (1) and a C6 (2) side chain, forming a 4-isobutylchroman-2-one unit and a 3-(2-methylpropylidene)benzofuran-2(3H)-one moiety with the acetophenone core, respectively. In addition, compound 4 exhibited significant dose-dependent transcriptional activation effect against retinoid X receptor-α (RXRα), and could be regarded as a new type of non-classical RXR ligand.

Diterpene glycosides, acetophenone glycosides and tannins from polar extracts of the root of Euphorbia fischeriana with cytotoxicity and antibacterial activities.

The phytochemical study on Euphorbia fischeriana, a folk medicinal plant in China, led to the isolation of eight undescribed glycosides, including two diterpene glycosides, three acetophenone glycosides and three tannins together with eight known ones. Their planar structures were elucidated by extensive analyses of 1D, 2D NMR experiments and HRESIMS. The absolute configurations were determined by NOESY experiments, ECD calculations. All undescribed compounds were evaluated for their cytotoxicity and antibacterial activities in vitro. Two diterpene glycosides (1-2) showed cytotoxic activity with IC50 values ranging from 5.4 to 16.2 µM toward Hep-G2, Hep-3B, A549, NCI-H460 and AGS cells. Tannins (6-8) showed the significant antibacterial activity with MIC values in the range of 1.56-6.25 µg/mL.

Antifungal and Antibiofilm Efficacy of Paeonol Treatment Against Biofilms Comprising Candida albicans and/or Cryptococcus neoformans.

Fungal populations are commonly found in natural environments and present enormous health care challenges, due to increased resistance to antifungal agents. Paeonol exhibits antifungal activities; nevertheless, the antifungal and antibiofilm activities of paeonol against Candida albicans and Cryptococcus neoformans remain largely unexplored. Here, we aimed to evaluate the antifungal and antibiofilm activities of paeonol against C. albicans and/or C. neoformans (i.e., against mono- or dual-species). The minimum inhibitory concentrations (MICs) of paeonol for mono-species comprising C. albicans or C. neoformans were 250 µg ml-1, whereas the MIC values of paeonol for dual-species were 500 µg ml-1. Paeonol disrupted cell membrane integrity and increased the influx of gatifloxacin into cells of mono- and dual-species cells, indicating an antifungal mode of action. Moreover, paeonol at 8 times the MIC damaged mono- and dual-species cells within C. albicans and C. neoformans biofilms, as it did planktonic cells. In particular, at 4 and 8 mg ml-1, paeonol efficiently dispersed preformed 48-h biofilms formed by mono- and dual-species cells, respectively. Paeonol inhibited effectively the yeast-to-hyphal-form transition of C. albicans and impaired capsule and melanin production of C. neoformans. The addition of 10 MIC paeonol to the medium did not shorten the lifespan of C. elegans, and 2 MIC paeonol could effectively protect the growth of C. albicans and C. neoformans-infected C. elegans. Furthermore, RNA sequencing was employed to examine the transcript profiling of C. albicans and C. neoformans biofilm cells in response to 1/2 MIC paeonol. RNA sequencing data revealed that paeonol treatment impaired biofilm formation of C. albicans by presumably downregulating the expression level of initial filamentation, adhesion, and growth-related genes, as well as biofilm biosynthesis genes, whereas paeonol inhibited biofilm formation of C. neoformans by presumably upregulating the expression level of ergosterol biosynthesis-related genes. Together, the findings of this study indicate that paeonol can be explored as a candidate antifungal agent for combating serious single and mixed infections caused by C. albicans and C. neoformans.

New paeonol derivative C302 reduces hypertension in spontaneously hypertensive rats through endothelium-dependent and endothelium-independent vasodilation.

Hypertension is a major risk factor for cardiovascular disease and Chinese herb monomers could provide new structural skeletons for anti-hypertension new drug development. Paeonol is a Chinese herbal monomer extracted from Cortex moutan, exhibited some anti-hypertensive activity. The study focused on the structural optimization of paeonol to provide promising lead compounds for anti-hypertension new drug development. Herein, twelve new paeonol derivatives (PD) were designed and synthesized and their vasodilation activity was evaluated by in vitro vasodilation drug screening platform based on Myograph. Its anti-hypertension activity, PD-C302 (2-hydroxy-4-methoxyvalerophenone) as a representative with the optimal vasodilation activity, was determined by its response to blood pressure in spontaneously hypertensive rats (SHR) in vivo. Moreover, its molecular mechanism was probed by the vasodilation activity of rat superior mesenteric artery rings with or without endothelium pre-contracted by potassium chloride (KCl) or phenylephrine hydrochloride (PE). It was indicated that PD-C302 significantly reduced the blood pressure in SHR, which would involve in PD-C302-induced vasodilation. Furthermore, endothelium-dependent pathways and endothelium-independent pathways both contributed importantly to PD-C302-induced vasodilation at low concentration of PD-C302. Endothelium-independent pathways (vascular smooth muscle cell-mediated vasodilation), were mainly responsible for the PD-C302-induced vasodilation at high concentration of PD-C302, which involved in opening multiple K+ channels to restrain Ca2+ channels, and then triggered vasodilation to reduce blood pressure. PD-C302 has a simple structure and favorable anti-hypertensive activity in vivo, which could be a promising lead compound for anti-hypertension new drug development.

Supplementation of Mito TEMPO and acetovanillone in semen extender improves freezability of buffalo spermatozoa.

BACKGROUND: Oxidative stress is one of the leading factors responsible for poor post-thaw semen quality because of overproduction of reactive oxygen species (ROS) over neutralizing antioxidants present in semen. Mainly two ROS generation sites are present in spermatozoa, that is, mitochondria and plasma membrane. Therefore, the idea of targeting these specific sites for minimization of ROS production with the compounds having known mechanism of actions was built up as a core for this research. OBJECTIVE: Present study was done to investigate the effects of Mito TEMPO and acetovanillone individually and in combination on freezability of buffalo spermatozoa. MATERIALS AND METHODS: For the experiment, semen extender was supplemented with Mito TEMPO (50 µM), acetovanillone (50 µM), and a combination of Mito TEMPO + acetovanillone (50 µM+ 50 µM), designated as Group II, Group III, and Group IV, respectively. Control group without any supplementation was designated as Group I. A total of 24 ejaculates with individual progressive motility (IPM) of ≥70% were selected for the study. After final dilution, filling-sealing of straws, equilibration, and freezing were done as per the standard procedure. Semen samples were evaluated for IPM, plasma membrane integrity, lipid peroxidation, total antioxidant capacity (TAC), and cholesterol to phospholipids (C/P) ratio at both fresh and post-thaw stages. Evaluation of ROS, mitochondrial membrane potential (MMP), capacitation status (CTC assay), and in vitro fertility potential were conducted only on frozen-thawed samples. RESULTS: The addition of Mito TEMPO (50 µM) and acetovanillone (50 µM) individually and in combination significantly (p < 0.05) improved post-thaw semen quality in terms of IPM, plasma membrane integrity, TAC, cholesterol content, C/P ratio, MMP, Chlortetracycline (CTC)-Full (F) pattern, and zona binding ability of buffalo spermatozoa, while significantly (p < 0.05) reduced ROS production, lipid peroxidation, and capacitation like changes as compared to the control group. DISCUSSION: As Mito TEMPO acts as an SOD mimetic and also detoxifies ferrous iron at the mitochondria level, it aids in neutralization of excessive ROS production and minimizes oxidative stress-related damages that enhances the antioxidant potential of sperm mitochondria. Earlier studies also indicated improved post-thaw semen quality in 50 µM supplemented group. The improvement observed in acetovanillone (50 µM) group might be because of inhibition of Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase as this enzyme activation by various physical/chemical inducers during cryopreservation process leads to activation of CatSper channel resulting in calcium influx, premature capacitation, and acrosomal reaction like changes through activation of adenylate cyclase and cAMP/PKA-mediated tyrosine phosphorylation of sperm proteins. Acetovanillone also prevents NADPH oxidase-mediated inhibition of glutathione reductase activity, which has a vital role in protecting the structural and functional integrity of sperm plasma membrane. CONCLUSION: Results indicated beneficial effects of supplementation of Mito TEMPO and acetovanillone on sperm freezability and individual supplementation was as efficient as the combination group for sustaining post-thaw semen quality.

Paeonol enhances treatment of fluconazole and amphotericin B against oropharyngeal candidiasis through HIF-1α related IL-17 signaling.

Oropharyngeal candidiasis (OPC) is an oral infection mainly caused by Candida albicans, a dimorphic human opportunistic pathogen that can proliferate and invade the superficial oral epithelium using its hyphae. The filamentation of C. albicans is a hallmark of biofilm formation, accompanied by the occurrence of a hypoxic microenvironment. Paeonol (PAE) is a traditional medicine with multiple properties. In a previous study, we demonstrated the synergism of PAE plus Fluconazole (FLU) or Amphotericin B (AmB) against C. albicans in vitro and in vivo. This study aimed to explore the therapeutic mechanisms of drug combinations on OPC. In an established OPC mouse model, the culture of hypoxia was observed by calcofluor white and hypoxyprobe staining. The expression and levels of IL-17 signaling-associated genes and proteins (IL-17A and IL-23) were evaluated in tissue homogenates and EC109 cells. The results show that compared with the single therapy, PAE plus FLU or AmB can decrease fungal burden, restore mucosal integrity, and reduce the hypoxic microenvironment and inflammation in the OPC mice. Relative to infected mice, the drug combinations can also rectify the abnormal expression of hypoxia inducible factor (hif)-1α, il-17a, and il-23 mRNA. Meanwhile, compared with the infected EC109 cells treated with a single drug, PAE plus FLU or AmB significantly inhibited the mRNA and protein expression of HIF-1α, IL-17A, and IL-23. Taken together, the possible mechanism of PAE plus FLU or AmB can be attributed to the regulation of hypoxia-associated IL-17 signaling in OPC treatment.

Paeonol inhibits the malignancy of Apatinib-resistant gastric cancer cells via LINC00665/miR-665/MAPK1 axis.

BACKGROUND: Paeonol is the extractive of Paeonia suffruticosa Andr and is reported to reverse the chemotherapy resistance of cancer cells. The present study explores the role of paeonol in inhibiting the malignant biological behaviors of Apatinib-resistant gastric cancer (GC) cells. METHODS: The Traditional Chinese Medicine Systems Pharmacology (TCMSP) database was adopted to screen the target genes of paeonol, and the STRING database was employed to construct a protein-protein interaction (PPI) network. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the target genes was performed employing DAVID online database. The expressions of these target genes in GC tissues and para-cancerous tissues were analyzed with GEPIA database, and GEO datasets (GSE109476 and GSE93415) were utilized to analyze differentially expressed lncRNAs and miRNAs in GC tissues and para-cancerous tissues. The expressions of LINC00665, miR-665 and MAPK1 mRNA in Apatinib-resistant GC cells were detected through quantitative real-time polymerase chain reaction (qRT-PCR). Cell counting kit-8 (CCK-8) assay was conducted to detect cell proliferation; Transwell assays were employed to detect cell migration and invasion, and TdT-mediated dUTP nick end labeling (TUNEL) assay was utilized to detect cell apoptosis. Dual-luciferase reporter gene assay was performed to detect the binding relationships between miR-665 and LINC00665, as well as between miR-665 and MAPK1 mRNA. The expressions of MAPK1 protein and glycolysis-associated proteins (GLUT1, LDHB and HK2) were detected by Western blot. Additionally, a tumor xenograft mice model was constructed to evaluate the effects of paeonol on lung metastasis. RESULTS: Paeonol could inhibit the proliferation, migration, invasion and glycolysis, and promote the apoptosis of Apatinib-resistant GC cells. TCMSP database suggested that Paeonol had 17 target genes, and 17 target genes were mainly enriched in signaling pathways related to apoptosis, glucose and lipid metabolism, etc.; GEPIA database suggests that MAPK1, among the 17 target genes, was markedly elevated in GC tissues. Paeonol could decrease LINC00665 and MAPK1 expressions in GC cells but increase the expression of miR-665. LINC00665 overexpression, MAPK1 overexpression or inhibition of miR-665 could abolish the inhibitive effects of paeonol on the malignant phenotypes of Apatinib-resistant GC cells. miR-665 is verified as an upstream regulator of MAPK1 and a target of LINC00665. Additionally, paeonol could significantly inhibit the lung metastasis in the tumor xenograft mice model. CONCLUSIONS: Paeonol can inhibit the malignancy of Apatinib-resistant GC cells through LINC00665/miR-665/MAPK1 axis. For the first time, our study imply that paeonol may be a potential drug to reverse Apatinib-resistant of GC cells.

Paeonol Ameliorates Cuprizone-Induced Hippocampal Demyelination and Cognitive Deficits through Inhibition of Oxidative and Inflammatory Events.

Multiple sclerosis (MS) is a chronic and inflammatory disorder of the central nervous system with autoimmune nature that is typified by varying degrees of demyelination and axonal damage. Paeonol is an active ingredient in some medicinal plants with anti-inflammatory and neuroprotective property. This study was conducted to reveal whether paeonol can alleviate hippocampal demyelination and cognitive deficits in cuprizone-induced murine model of demyelination as a model of MS. C57BL/6 mice received oral cuprizone (400 mg/kg) for 6 weeks, and paeonol was administered p.o. at two doses of 25 or 100 mg/kg, starting from the second week post-cuprizone for 5 weeks. After assessment of learning and memory in different tasks, oxidative stress and inflammation were evaluated besides immunohistochemical assessment of hippocampal myelin basic protein (MBP). Paeonol (100 mg/kg) properly ameliorated cognitive deficits in Y maze, novel object discrimination (NOD) test, and Barnes maze with no significant improvement of performance in passive avoidance task. In addition, paeonol treatment at the higher dose was also associated with partial restoration of hippocampal level of oxidative stress and inflammatory markers including MDA, ROS, GSH, SOD, catalase, NF-kB, and TNF. Besides, paeonol improved MMP as an index of mitochondrial integrity and health and reduced MPO as a factor of neutrophil infiltration. Furthermore, paeonol treatment prevented hippocampal MBP immunoreactivity, indicating its prevention of demyelination. In conclusion, the current study showed the preventive effect of paeonol against cuprizone-induced demyelination and cognitive deficits through reversing most oxidative stress- and inflammation-related parameters in addition to its improvement of mitochondrial health.

Exploring targets and signaling pathways of paeonol involved in relieving inflammation based on modern technology.

Paeonol, derived from natural plants (Moutan Cortex), has a wide range of biological effects, including anti-inflammatory and antitumor effects as well as favorable effects against cardiovascular and neurodegenerative diseases. The anti-inflammatory action is the main pharmacological activity of paeonol and has the greatest clinical relevance. However, the anti-inflammatory mechanism of paeonol has not been reported in sufficient detail. We systematically analyzed the anti-inflammatory mechanism of paeonol using network pharmacological databases and platforms, including TCMSP, Swiss TargetPrediction, OMIM, DrugBank, TTD, Jevnn, STRING11.0, and Metascape. Furthermore, we used high-throughput molecular docking method to prove the results of the above analyses, providing a reference for exploring the mechanism of paeonol and developing targeted drugs.