Salidroside protects against ventilation-induced lung injury by inhibiting the expression of matrix metalloproteinase-9

ContextSalidroside, a compound extracted from Rhodiola rosea L. (Crassulaceae), possesses many beneficial pathological effects.ObjectiveTo explore the effect of salidroside on ventilator-induced lung endothelial dysfunction in vivo and in vitro.Materials and methodsIn vivo, male ICR mice were divided into sham, ventilation, salidroside, and ventilation plus salidroside groups. The mice were ventilated for 4 h, salidroside (50 mg/kg) was administrated intraperitoneally before ventilation, dexamethasone (Dex) (5 mg/kg) was used as a positive control. In vitro, mouse lung vascular endothelial cells (MLVECs) were treated with salidroside, MMP-9 siRNA, and BAY11-7082 (10 μM), and then exposed to cyclic stretch for 4 h. Afterward, lung tissues and MLVECs were collected for further analysis.ResultsSalidroside pre-treatment significantly reversed the expression of vascular endothelial cadherin (VE-cadherin) and zonula occluden-1 (ZO-1) proteins in cyclic stretch-treated MLVECs (0.46 ± 0.09 vs. 0.80 ± 0.14, 0.49 ± 0.05 vs. 0.88 ± 0.08) and ventilated lung tissues (0.56 ± 0.06 vs. 0.83 ± 0.46, 0.49 ± 0.08 vs. 0.80 ± 0.12). The results further indicated that salidroside inhibited the expression of matrix metalloproteinase-9 (MMP-9), whereas knockdown of its expression restored the expression levels of VE-cadherin (0.37 ± 0.08 vs. 0.85 ± 0.74) and ZO-1 (0.48 ± 0.08 vs. 0.81 ± 0.11) in stretched MLVECs. Meanwhile, salidroside inhibited the NF-κB signalling pathway and alleviated lung injury.ConclusionsSalidroside protected against stretch-induced endothelial barrier function, improving lung injury after ventilation. Thus, salidroside may be a promising therapeutic agent for patients with MV-induced lung injury.     

Rosmarinic acid ameliorates acetaminophen-induced acute liver injury in mice via RACK1/TNF-α mediated antioxidant effect

ContextRosmarinic acid (RA) dose-dependently ameliorates acetaminophen (APAP) induced hepatotoxicity in rats. However, whether RA hepatoprotective effect is by regulating RACK1 and its downstream signals is still unclear.ObjectiveThis study explores the RA protective effect on APAP-induced ALI and its mechanism.Materials and methodsSixty Kunming mice 6–8 weeks old were randomly separated into six groups (n = 10) and pre-treated with normal saline, ammonium glycyrrhetate (AG) or RA (10, 20 or 40 mg/kg i.p./day) for two consecutive weeks. Then, APAP (300 mg/kg, i.g.) was administrated to induce ALI, except for the control. Serum alanine/aspartate aminotransferases (ALT and AST), malondialdehyde (MDA), superoxide dismutase (SOD) and histopathology were used to authenticate RA effect. The liver RACK1 and TNF-α were measured by western blot.ResultsCompared with the APAP group, different dosages RA significantly decreased ALT (52.09 ± 7.98, 55.13 ± 10.19, 65.08 ± 27.61 U/L, p < 0.05), AST (114.78 ± 19.87, 115.29 ± 31.91, 101.78 ± 21.85 U/L, p < 0.05), MDA (2.37 ± 0.87, 2.13 ± 0.87, 1.86 ± 0.39 nmol/mg, p < 0.01) and increased SOD (306.178 ± 90.80, 459.21 ± 58.54, 444.01 ± 78.09 U/mg, p < 0.05). With increasing doses of RA, RACK1 and TNF-α expression decreased. Moreover, the RACK1 and TNF-α levels were positively correlated with MDA (r = 0.8453 and r = 0.9391, p < 0.01).Discussion and conclusionsOur findings support RA as a hepatoprotective agent to improve APAP-induced ALI and the antioxidant effect mediated through RACK1/TNF-α pathway.     

Methyl-β-cyclodextrin suppresses the monocyte-endothelial adhesion triggered by lipopolysaccharide (LPS) or oxidized low-density lipoprotein (oxLDL)

ContextRecent studies demonstrated the anti-atherosclerotic efficacy of cyclodextrin. However, it remains unclear whether cyclodextrin exerts the anti-atherosclerotic effect via regulating monocyte-endothelial adhesion.ObjectiveTo answer that question by recruiting methyl-β-cyclodextrin (MβCD) as a cyclodextrin representative.Materials and methodsHuman umbilical vein endothelial cells (HUVECs) were not treated, or treated with 1 µg/mL liposaccharide (LPS) or 50 µg/mL oxidized low-density lipoprotein (oxLDL) for 12 h, 5 mM MβCD for 1 h, and LPS/oxLDL (1 and 50 µg/mL, respectively for 12 h) plus MβCD (5 mM for 1 h), respectively. The effects of MβCD on LPS/oxLDL-triggered monocyte-endothelial adhesion and related molecules in signalling pathways were evaluated via confocal microscopy, flow cytometry, RT-PCR, western blotting, and cell adhesion assay.ResultsMβCD with an IC₅₀ of 27.66 mM (1 h treatment) exerted no significant cytotoxicity at ≤5 mM for ≤2 h. Compared with the control, both LPS and oxLDL induced an ∼2–3-fold increase in adhesion molecule expression (ICAM-1 and VCAM-1 at protein and mRNA levels) and NF-κB phosphorylation (p-NF-κB/pP65), an increase in IκB kinase (IKK), and a decrease in phosphorylated protein kinase B (p-Akt), respectively. Moreover, more monocytes (2-fold higher for LPS and 15% higher for oxLDL) were attached on LPS/oxLDL-stimulated HUVECs. 5 mM MβCD reversed the LPS/oxLDL-induced changes back to the control levels.ConclusionsMβCD significantly suppresses the LPS/oxLDL-triggered monocyte-endothelial adhesion by downregulating adhesion molecule expression probably via LPS-IKK-NF-κB or oxLDL-Akt-NF-κB pathway. This study demonstrates a potential mechanism of the anti-atherosclerotic efficacy of cyclodextrin from the angle of monocyte-endothelial adhesion.     

Edaravone prevents high glucose-induced injury in retinal Müller cells through thioredoxin1 and the PGC-1α/NRF1/TFAM pathway

ContextOxidative injury in a high-glucose (HG) environment may be a mechanism of diabetic retinopathy (DR) and edaravone can protect retinal ganglion cells by scavenging ROS.ObjectiveTo explore the effect of edaravone on HG-induced injury.Materials and methodsFirst, Müller cells were cultured by different concentrations of glucose for different durations to obtain a suitable culture concentrations and duration. Müller cells were then divided into Control, HG + Vehicle, HG + Eda-5 μM, HG + Eda-10 μM, HG + Eda-20 μM, and HG + Eda-40 μM groups. Cells were cultured by 20 mM glucose and different concentrations of edaravone for 72 h.ResultsThe IC₅₀ of glucose at 12–72 h is 489.3, 103.5, 27.92 and 20.71 mM, respectively. When Müller cells were cultured in 20 mM glucose for 72 h, the cell viability was 52.3%. Edaravone significantly increased cell viability compared to Vehicle (68.4% vs 53.3%; 78.6% vs 53.3%). The EC₅₀ of edaravone is 34.38 μM. HG induced high apoptosis rate (25.5%), while edaravone (20 and 40 μM) reduced it to 12.5% and 6.89%. HG increased the DCF fluorescence signal (189% of Control) and decreased the mitochondrial membrane potential by 57%. Edaravone significantly decreased the DCF fluorescence signal (144% and 132% of Control) and recovered the mitochondrial membrane potential to 68% and 89% of Control. Furthermore, HG decreased the expression of TRX1, PGC-1α, NRF1 and TFAM, which were restored by edaravone.Discussion and conclusionThese findings provide a new potential approach for the treatment of DR and indicated new molecular targets in the prevention of DR.     

α-Amylase and dipeptidyl peptidase-4 (DPP-4) inhibitory effects of Melicope latifolia bark extracts and identification of bioactive constituents using in vitro and in silico approaches

ContextMelicope latifolia (DC.) T. G. Hartley (Rutaceae) was reported to contain various phytochemicals including coumarins, flavonoids, and acetophenones.ObjectiveThis study investigates the antidiabetic and antioxidant effects of M. latifolia bark extracts, fractions, and isolated constituents.Materials and methodsMelicope latifolia extracts (hexane, chloroform, and methanol), fractions, and isolated constituents with varying concentrations (0.078–10 mg/mL) were subjected to in vitro α-amylase and dipeptidyl peptidase-4 (DPP-4) inhibitory assay. Molecular docking was performed to study the binding mechanism of active compounds towards α-amylase and DPP-4 enzymes. The antioxidant activity of M. latifolia fractions and compounds were determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging and β-carotene bleaching assays.ResultsMelicope latifolia chloroform extract showed the highest antidiabetic activity (α-amylase IC₅₀: 1464.32 μg/mL; DPP-4 IC₅₀: 221.58 μg/mL). Fractionation of chloroform extract yielded four major fractions (CF₁–CF₄) whereby CF₃ showed the highest antidiabetic activity (α-amylase IC₅₀: 397.68 μg/mL; DPP-4 IC₅₀: 37.16 μg/mL) and resulted in β-sitosterol (1), halfordin (2), methyl p-coumarate (3), and protocatechuic acid (4). Isolation of compounds 2–4 from the species and their DPP-4 inhibitory were reported for the first time. Compound 2 showed the highest α-amylase (IC₅₀: 197.53 μM) and β-carotene (88.48%) inhibition, and formed the highest number of molecular interactions with critical amino acid residues of α-amylase. The highest DPP-4 inhibition was exhibited by compound 3 (IC₅₀: 911.44 μM).Discussion and conclusionsThe in vitro and in silico analyses indicated the potential of M. latifolia as an alternative source of α-amylase and DPP-4 inhibitors. Further pharmacological studies on the compounds are recommended.     

Rhodiola rosea polysaccharides promote the proliferation of bone marrow haematopoietic progenitor cells and stromal cells in mice with aplastic anaemia

ContextThe effects of Rhodiola rosea L. (Crassulaceae) polysaccharides (RRPs) on haematopoiesis are poorly understood.ObjectiveTo determine the effects of RRPs on haematopoiesis in mice with aplastic anaemia.Materials and methodsAplastic anaemia was induced in Kunming mice by ⁶⁰Coγ (2.0 Gy) irradiation and cyclophosphamide administration (50 mg/kg/day for 3 consecutive days; intraperitoneal injection). The in vivo effects of RRPs (10, 20, and 40 mg/kg; intraperitoneal injection) on haematopoiesis were analyzed using peripheral blood tests, histopathological examination of haematopoietic tissues, culture of haematopoietic progenitors and bone marrow stromal cells (BMSCs), and Western blotting of Fas and Fas ligand (FasL). The in vitro effects of RRPs on bone-marrow haematopoietic progenitors and BMSCs were also evaluated.ResultsCompared to anaemic controls, high-dose RRPs (40 mg/kg) significantly increased red blood cells (8.21 ± 0.57835 versus 6.13 ± 1.34623 × 10¹²/L), white blood cells (5.11 ± 1.6141 versus l.54 ± 1.1539 × 10⁹/L), and BMSCs (10.33 ± 1.5542 versus 5.87 ± 3.1567 × 10¹²/L) in mice with aplastic anaemia (all p < 0.01). High-dose RRPs significantly increased the formation of colony-forming unit-granulocyte macrophage (CFU-GM), burst-forming unit-erythroid (BFU-E), and colony-forming unit-erythroid (CFU-E; p < 0.01). Fas and FasL protein expression in BMSCs decreased after RRPs administration. Especially at the high dose, RRPs (150 μg/mL) significantly promoted in vitro CFUs-E, BFUs-E, and CFUs-GM formation. RRPs (150–300 μg/mL) also promoted BMSC proliferation.Discussion and conclusionsRRPs helped to promote haematopoietic recovery in mice with aplastic anaemia, facilitating haematopoietic tissue recovery. This study indicated some mechanisms of the haematopoietic regulatory effects of RRPs. Our findings provide a laboratory basis for clinical research on RRPs.     

Effects of ginkgo leaf tablet on the pharmacokinetics of rosiglitazone in rats and its potential mechanism

ContextGinkgo leaf tablet (GLT), a traditional Chinese herbal formula, is often combined with rosiglitazone (ROS) for type 2 diabetes mellitus treatment. However, the drug-drug interaction between GLT and ROS remains unknown.ObjectiveTo investigate the effects of GLT on the pharmacokinetics of ROS and its potential mechanism.Materials and methodsThe pharmacokinetics of 10 mg/kg ROS with 100/200 mg/kg GLT as single-dose and 10-day multiple-dose administration were investigated in Sprague-Dawley rats. In vitro, the effects of GLT on the activity of CYP2C8 and CYP2C9 were determined in recombinant human yeast microsomes and rat liver microsomes with probe substrates.ResultsThe t_1/2 of ROS increased from 2.14 ± 0.38 (control) to 2.79 ± 0.37 (100 mg/kg) and 3.26 ± 1.08 h (200 mg/kg) in the single-dose GLT administration. The AUC_0-t (139.69 ± 45.46 vs. 84.58 ± 39.87 vs. 66.60 ± 15.90 h·μg/mL) and t_1/2 (2.75 ± 0.70 vs. 1.99 ± 0.44 vs. 1.68 ± 0.35 h) decreased significantly after multiple-dose GLT treatment. The IC₅₀ values of quercetin, kaempferol, and isorhamnetin, GLT main constituents, were 9.32, 7.67, and 11.90 μmol/L for CYP2C8, and 27.31, 7.57, and 4.59 μmol/L for CYP2C9. The multiple-dose GLT increased rat CYP2C8 activity by 44% and 88%, respectively.Discussion and conclusionsThe metabolism of ROS is attenuated in the single dose of GLT by inhibiting CYP2C8 and CYP2C9 activity, and accelerated after the multiple-dose GLT treatment via inducing CYP2C8 activity in rats, indicating that the clinical dose of ROS should be adjusted when co-administrated with GLT.     

Evodiamine decreased the systemic exposure of pravastatin in non-alcoholic steatohepatitis rats due to the up-regulation of hepatic OATPs

ContextPatients with non-alcoholic steatohepatitis (NASH) may have a simultaneous intake of pravastatin and evodiamine-containing herbs.ObjectiveThe effect of evodiamine on the pharmacokinetics of pravastatin and its potential mechanisms were investigated in NASH rats.Materials and methodsThe NASH model was conducted with feeding a methionine choline-deficient (MCD) diet for 8 weeks. Sprague-Dawley rats were randomised equally (n = 6) into NASH group, evodiamine group (10 mg/kg), pravastatin group (10 mg/kg), and evodiamine (10 mg/kg) + pravastatin (10 mg/kg) group. Normal control rats were fed a standard diet. Effects of evodiamine on the pharmacokinetics, distribution, and uptake of pravastatin were investigated.ResultsEvodiamine decreased C_max (159.43 ± 26.63 vs. 125.61 ± 22.17 μg/L), AUC_0-t (18.17 ± 2.52 vs. 14.91 ± 2.03 mg/min/L) and AUC_0-∞ (22.99 ± 2.62 vs. 19.50 ± 2.31 mg/min/L) of orally administered pravastatin in NASH rats, but had no significant effect in normal rats. Evodiamine enhanced the uptake (from 154.85 ± 23.17 to 198.48 ± 26.31 pmol/mg protein) and distribution (from 736.61 ± 108.07 to 911.89 ± 124.64 ng/g tissue) of pravastatin in NASH rat liver. The expression of Oatp1a1, Oatp1a4, and Oatp1b2 was up-regulated 1.48-, 1.38-, and 1.51-fold by evodiamine. Evodiamine decreased the levels of IL-1β, IL-6, and TNF-α by 27.82%, 24.76%, and 29.72% in NASH rats, respectively.Discussion and conclusionsEvodiamine decreased the systemic exposure of pravastatin by up-regulating the expression of OATPs. These results provide a reference for further validation of this interaction in humans.     

Huoxue Qianyang Qutan recipe attenuates cardiac fibrosis by inhibiting the NLRP3 inflammasome signalling pathway in obese hypertensive rats

ContextHuoXue QianYang QuTan Recipe (HQQR) is used to manage hypertension and cardiac remodelling, but the mechanism is elusive.ObjectiveTo determine the mechanism of HQQR on obesity hypertension (OBH)-related myocardial fibrosis.Materials and methodsOBH models were prepared using spontaneously hypertensive rats (SHRs) and divided (n = 6) into saline, low-dose (19.35 g/kg/d) HQQR, high-dose (38.7 g/kg/d) HQQR, and valsartan (30 mg/kg/d) groups for 10 weeks. Systolic blood pressure (SBP), and Lee’s index were measured. Heart tissues were examined by histology. HQQR’s effects were examined on cardiac fibroblasts (CFs) stimulated with angiotensin II and treated with HQQR, a caspase-1 inhibitor, siNLRP3, and oeNLRP3.ResultsHQQR(H) reduced SBP (201.67 ± 21.00 vs. 169.00 ± 10.00), Lee’s index (321.50 ± 3.87 vs. 314.58 ± 3.88), and left ventricle mass index (3.26 ± 0.27 vs. 2.71 ± 0.12) in vivo. HQQR reduced percentage of fibrosis area (18.99 ± 3.90 vs. 13.37 ± 3.39), IL-1β (10.07 ± 1.16 vs. 5.35 ± 1.29), and inhibited activation of NLRP3/caspase-1/IL-1β pathway. HQQR also inhibiting the proliferation (1.09 ± 0.02 vs. 0.84 ± 0.01), fibroblast to myofibroblast transition (14.74 ± 3.39 vs. 3.97 ± 0.53), and collagen deposition (Col I; 0.50 ± 0.02 vs. 0.27 ± 0.05 and Col III; 0.48 ± 0.21 vs. 0.26 ± 0.11) with different concentrations selected based on IC₅₀ in vitro (all ps < 0.05). NLRP3 interference further confirmed HQQR inhibiting NLRP3 inflammasome signalling.ConclusionHQQR blunted cardiac fibrosis development in OBH and suppressed CFs proliferation by directly interfering with the NLRP3/caspase-1/IL-1β pathway.     

Cytotoxic furanosesquiterpenoids and steroids from Ircinia mutans sponges

ContextIrcinia mutans Wilson (Irciniidae) is a sponge with antimicrobial and cytotoxic constituents.ObjectiveOur objective was to characterise the cytotoxic constituents of two seasonal collections of I. mutans.Materials and methodsThe sponges were extracted in methanol-dichloromethane and their constituents were purified and characterised using column chromatography, GC-MS, 1 D and 2 D NMR. Anti-proliferative activities of the compounds, were evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay (0.25–100 μg/mL, 72 h) against leukaemia (MOLT-4), breast (MCF-7) and colon cancer (HT-29) human cells.ResultsThree furanosesquiterpoids; furodysin (1), ent-furodysinin (2) and furoircin (3) and ten sterols were characterised in I. mutans, for the first time. Cholesterol (4), cholesta-5, 7-dien-3β-ol (5) and ergosterol (6) were determined in the sponge from the winter collections, while cholesta-5, 22-dien-3β-ol (7), 24-methyldesmosterol (8), campesterol (9), stigmasterol (10), γ-ergostenol (11), chondrillasterol (12) and γ-sitosterol (13) were detected in the summer samples. The steroids from the winter collection exhibited cytotoxic activity with IC₅₀ values of 13.0 ± 0.9, 11.1 ± 1.7 and 1.1 ± 0.4 µg/mL, against the mentioned cancer cell lines, respectively, while those from the summer sample, showed greater activity, IC₅₀ = 1.1 ± 0.2 μg/mL against MOLT-4. The purified steroids showed potent MOLT-4 cytotoxic activity, IC₅₀ values = 2.3–7.8 µg/mL.Discussion and conclusionThe present study suggests that I. mutans is a rich source of cytotoxic steroids, and introduces 3 as new natural product. Considering the high cytotoxic activity of the steroids, these structures could be candidates for anticancer drug development in future research.     

Pharmacokinetic study on the interaction between succinic acid and irbesartan in rats and its potential mechanism

ContextSuccinic acid and irbesartan are commonly used drugs in cardiovascular disease treatment. The interaction might occur during their co-administration, which was still unclear.ObjectiveTo reveal the effect of succinic acid on the metabolism of irbesartan and its potential mechanism.Materials and methodsThe Sprague-Dawley rats (n = 6) were treated with a single dose of 30 mg/kg irbesartan (control) or the co-administration with the pre-treatment of 200 mg/kg succinic acid for 7 d. The effect of succinic acid on the metabolic stability and the activity of CYP2C9 was evaluated in rat liver microsomes.ResultsSuccinic acid increased the AUC (5328.71 ± 959.31 μg/L × h vs. 3340.23 ± 737.75 μg/L × h) and prolonged the half-life of irbesartan (from 12.79 ± 0.73 h to 20.59 ± 6.35 h). The T_max (2.83 ± 0.75 h vs. 3.83 ± 1.10 h) and clearance rate (3.46 ± 1.13 L/h/kg vs. 6.91 ± 1.65 L/h/kg) of irbesartan was reduced by succinic acid. Consistently, succinic acid improved the metabolic stability (half-life from 23.32 ± 3.46 to 27.35 ± 2.15 min, intrinsic clearance rate from 59.43 ± 6.12 to 50.68 ± 5.64 μL/min/mg protein). Succinic acid was also found to inhibit the activity of CYP2C9 with the IC₅₀ value of 13.87 μM.Discussion and conclusionsSuccinic acid increased the system exposure of irbesartan via inhibiting CYP2C9. The experiment design of this study also provides a reference for the further validation of this interaction in humans.     

Baicalein inhibits the pharmacokinetics of simvastatin in rats via regulating the activity of CYP3A4

ContextBaicalein and simvastatin possess similar pharmacological activities and indications. The risk of their co-administration was unclear.ObjectiveThe interaction between baicalein and simvastatin was investigated to provide reference and guidance for the clinical application of the combination of these two drugs.Materials and methodsThe pharmacokinetics of simvastatin was investigated in Sprague–Dawley rats (n = 6). The rats were pre-treated with 20 mg/kg baicalein for 10 days and then administrated with 40 mg/kg simvastatin. The single administration of simvastatin was set as the control group. The rat liver microsomes were employed to assess the metabolic stability and the effect of baicalein on the activity of CYP3A4.ResultsBaicalein significantly increased the AUC_(0–t) (2018.58 ± 483.11 vs. 653.05 ± 160.10 μg/L × h) and C_max (173.69 ± 35.49 vs. 85.63 ± 13.28 μg/L) of simvastatin. The t_1/2 of simvastatin was prolonged by baicalein in vivo and in vitro. The metabolic stability of simvastatin was also improved by the co-administration of baicalein. Baicalein showed an inhibitory effect on the activity of CYP3A4 with the IC₅₀ value of 12.03 μM, which is responsible for the metabolism of simvastatin.Discussion and conclusionThe co-administration of baicalein and simvastatin may induce drug-drug interaction through inhibiting CYP3A4. The dose of baicalein and simvastatin should be adjusted when they are co-administrated.     

Esculin protects against methionine choline-deficient diet-induced non-alcoholic steatohepatitis by regulating the Sirt1/NF-[... formula ...]B p65 pathway

ContextEsculin, an active coumarin compound, has been demonstrated to exert anti-inflammatory effects. However, its potential role in non-alcoholic steatohepatitis (NASH) remains unclear.ObjectiveThis study explored the hepatoprotective effect and the molecular mechanism of esculin in methionine choline-deficient (MCD) diet-induced NASH.Materials and methodsFifty C57BL/6J mice were divided into five groups: control, model, low dosage esculin (oral, 20 mg/kg), high dosage esculin (oral, 40 mg/kg), and silybin (oral, 105 mg/kg). All animals were fed a MCD diet, except those in the control group (control diet), for 6 weeks.ResultsEsculin (20 and 40 mg/kg) inhibited MCD diet-induced hepatic lipid content (triglyceride: 16.95 ± 0.67 and 14.85 ± 0.78 vs. 21.21 ± 1.13 mg/g; total cholesterol: 5.10 ± 0.34 and 4.08 ± 0.47 vs. 7.31 ± 0.58 mg/g), fibrosis, and inflammation (ALT: 379.61 ± 40.30 and 312.72 ± 21.45 vs. 559.51 ± 37.01 U/L; AST: 428.22 ± 34.29 and 328.23 ± 23.21 vs. 579.36 ± 31.93 U/L). In vitro, esculin reduced tumour necrosis factor-α, interleukin-6, fibronectin, and collagen 4A1 levels, but had no effect on lipid levels in HepG2 cells induced by free fatty acid. Esculin increased Sirt1 expression levels and decreased NF-κB acetylation levels in vivo and in vitro. Interfering with Sirt1 expression attenuated the beneficial effect of esculin on inflammatory and fibrotic factor production in HepG2 cells.ConclusionsThese findings demonstrate that esculin ameliorates MCD diet-induced NASH by regulating the Sirt1/ac-NF-κB signalling pathway. Esculin could thus be employed as a therapy for NASH.     

The antithrombosis effect of dehydroandrographolide succinate: in vitro and in vivo studies

ContextDehydroandrographolide succinate (DAS) is mainly used in the clinical treatment of various infectious diseases. Its potential effects on platelet aggregation and blood coagulation systems have not been reported systematically.ObjectiveTo explore whether DAS exerts an antithrombotic effect and its internal mechanism.Materials and methodsHuman blood samples and Sprague-Dawley (SD) rats divided into control, aspirin (30 mg/kg), and DAS groups (200, 400 and 600 mg/kg) were used to measure the platelet aggregation rate, coagulation function, coagulation factor activity, and contents of thromboxane B₂ (TXB₂) and 6-keto-prostaglandin F_1α (6-keto-PGF_1α). The histopathology of the SD rat gastric mucosa was also observed. All rats were administered intragastric or intraperitoneal injections once a day for 3 consecutive days.ResultsCompared to control group, DAS significantly inhibited the platelet aggregation rate (ED₅₀ = 386.9 mg/kg) by decreasing TXB₂ levels (1531.95 ± 649.90 pg/mL to 511.08 ± 411.82 pg/mL) and activating antithrombin III (AT-III) (103.22 ± 16.22% to 146.46 ± 8.96%) (p < 0.05). In addition, DAS significantly enhanced the coagulation factors FV (304.12 ± 79.65% to 443.44 ± 75.04%), FVII (324.19 ± 48.03% to 790.66 ± 225.56%), FVIII (524.79 ± 115.47% to 679.92 ± 143.34%), FX (34.90 ± 7.40% to 102.76 ± 29.41%) and FXI (38.12 ± 10.33% to 65.47 ± 34.08%), increased the content of Fg (2.18 ± 0.39 to 3.61 ± 0.37 g/L), shorten the PT (10.42 ± 0.44 to 9.22 ± 0.21 s), APTT (16.43 ± 1.4 to 14.07 ± 0.75 s) and TT time (37.04 ± 2.13 to 32.68 ± 1.29 s) (p < 0.05), while the aspirin group showed no such effect on these items but showed reduced activity of FII (89.21 ± 21.72% to 61.83 ± 8.95%) and FVIII (524.79 ± 115.47% to 306.60 ± 29.96%) (p < 0.05). Histopathological changes showed aspirin-induced gastric mucosa haemorrhage and the protective effect of DAS in the gastric mucosa.ConclusionsDAS is more suitable than aspirin in thromboprophylaxis treatment, which provides a reliable theoretical and experimental basis for its clinical application.     

A novel nasal co-loaded loratadine and sulpiride nanoemulsion with improved downregulation of TNF-α, TGF-β and IL-1 in rabbit models of ovalbumin-induced allergic rhinitis

PurposeThe work aimed to develop a co-loaded loratadine and sulpiride nasal nanoemulsion for allergic rhinitis management.MethodsCompatibility studies were conducted adopting differential scanning calorimetry and Fourier transform infrared spectroscopy. Nanoemulsion formulations were prepared using soybean lecithin, olive oil and tween 80. Sodium cholate and glycerol were employed as co-surfactants. Nanoemulsions were assessed for viscosity, pH, droplet size, polydispersity index, zeta potential, electrical conductivity, entrapment, In vitro drug release and corresponding kinetics. Stability of the selected formulation was investigated. The biological effectiveness was evaluated in rabbit models of ovalbumin-induced allergic rhinitis by measuring TNF-α, TGF-β and IL-1.ResultsCompatibility studies revealed absence of drug/drug interactions. Nanoemulsions exhibited > 90% entrapment efficiency. The selected nanoemulsion demonstrated small droplet size (85.2 ± 0.2 nm), low PDI (0.35 ± 0.0) and appropriate Zeta Potential (−23.3 ± 0.2) and stability. It also displayed enhanced in vitro drug release following the Higuashi Diffusion and Baker–Lonsdale models. The mean relative mRNA expression of TNF-α, IL-1 and TGF-β significantly decreased from 9.59 ± 1.06, 4.15 ± 0.02 and 4.15 ± 0.02 to 1.28 ± 0.02, 1.93 ± 0.06 and 1.56 ± 0.02 respectively after treatment with the selected nanoemulsion formulation.ConclusionThe results reflected a promising potent effect of the combined loratadine and sulpiride nasal nanoemulsion in managing the symptoms of allergic rhinitis.     

The pharmacokinetic study on the interaction between nobiletin and anemarsaponin BII in vivo and in vitro

ContextThe interaction between nobiletin and anemarsaponin BII could affect the pharmacological activity of these two drugs during their combination.ObjectiveThe co-administration of nobiletin and anemarsaponin BII was investigated to explore the interaction and the potential mechanism.Materials and methodsMale Sprague-Dawley rats were only orally administrated with 50 mg/kg nobiletin as the control and another six rats were pre-treated with 100 mg/kg anemarsaponin BII for 7 d followed by the administration of nobiletin. The transport and metabolic stability of nobiletin were evaluated in vitro, and the effect of anemarsaponin BII on the activity of CYP3A4 was also assessed to explore the potential mechanism underlying the interaction.ResultsThe increasing C_max (2309.67 ± 68.06 μg/L vs. 1767.67 ± 68.86 μg/L), AUC (28.84 ± 1.34 mg/L × h vs. 19.57 ± 2.76 mg/L × h), prolonged t_1/2 (9.80 ± 2.33 h vs. 6.24 ± 1.53 h), and decreased clearance rate (1.46 ± 0.26 vs. 2.42 ± 0.40) of nobilein was observed in rats. Anemarsaponin BII significantly enhanced the metabolic stability of nobiletin in rat liver microsomes (half-life increased from 31.56 min to 39.44 min) and suppressed the transport of nobiletin in Caco-2 cells (efflux rate decreased from 1.57 ± 0.04 to 1.30 ± 0.03). The inhibitory effect of anemarsaponin BII on CYP3A4 was also found with an IC₅₀ value of 10.23 μM.Discussion and conclusionsThe interaction between anemarsaponin BII and nobiletin was induced by the inhibition of CYP3A4, which should draw special attention in their clinical co-administration.     

Cyanidin attenuates the apoptosis of rat nucleus pulposus cells and the degeneration of intervertebral disc via the JAK2/STAT3 signal pathway in vitro and in vivo

ContextCyanidin has been shown to have therapeutic potential in osteoarthritis. However, it is unclear whether cyanidin prevents the progression of intervertebral disc degeneration (IVDD).ObjectiveThis study evaluates the effects of cyanidin on IVDD in vitro and in vivo.Materials and methodsNucleus pulposus cells (NPCs) isolated from lumbar IVD of 4-week-old male Sprague-Dawley (SD) rats were exposed to 20 ng/mL IL-1β, and then treated with different doses (0-120 µM) of cyanidin for 24 h. SD rats were classified into three groups (n = 8) and treated as follows: control (normal saline), IVDD (vehicle), IVDD + cyanidin (50 mg/kg). Cyanidin was administered intraperitoneally for 8 weeks.ResultsThe IC₅₀ of cyanidin for NPCs was 94.78 µM, and cyanidin had no toxicity at concentrations up to 500 mg/kg in SD rats. Cyanidin inhibited the apoptosis of NPCs induced by IL-1β (12.73 ± 0.61% vs. 18.54 ± 0.60%), promoted collagen II (0.82-fold) and aggrecan (0.81-fold) expression, while reducing MMP-13 (1.02-fold) and ADAMTS-5 (1.40-fold) expression. Cyanidin increased the formation of autophagosomes in IL-1β-induced NPCs, and promoted LC3II/LC3I (0.83-fold) and beclin-1 (0.85-fold) expression, which could be reversed by chloroquine. Cyanidin inhibited the phosphorylation of JAK2 (0.47-fold) and STAT3 (0.53-fold) in IL-1β-induced NPCs. The effects of cyanidin could be enhanced by AG490. Furthermore, cyanidin mitigated disc degeneration in IVDD rats in vivo.Discussion and conclusionsCyanidin improved the function of NPCs in IVDD by regulating the JAK2/STAT3 pathway, which may provide a novel alternative strategy for IVDD. The mechanism of cyanidin improving IVDD still needs further work for in-depth investigation.     

Antiproliferative activity,cell-cycle arrest,apoptotic induction and LC-HRMS/MS analyses of extracts from two Linum species

ContextLinum is the largest genus of the Linaceae family; the species of this genus are known to have anticancer activity.ObjectiveIn this study, ethyl acetate extracts of L. numidicum Murb. (EAELN) and L. trigynum L. (EAELT) were examined, for the first time, for their anticancer capacity. The secondary metabolites compositions were analysed by LC-HRMS/MS.Materials and methodsThe antiproliferative effect of EAELN and EAELT (0–10.000 μg/mL) against PC3 and MDA-MB-231 cell lines were  evaluated by the MTT assay after 72 h of treatment. Flow cytometer analysis of apoptosis (Annexin V-FITC/PI) and cell cycle (PI/RNase) was also performed after treatment with EAELN and EAELT at 250, 500, and 1000 μg/mL, for 24 h.ResultsEAELN had the highest antiproliferative activity against PC3 (IC₅₀ 133.2 ± 5.73 μg/mL) and MDA-MB-231 (IC₅₀ 156.9 ± 2.83 μg/mL) lines, EAELN had also shown better apoptotic activity with 19 ± 2.47% (250 μg/mL), 87.5 ± 0.21% (500 μg/mL), and 92 ± 0.07% (1000 μg/mL), respectively, causing cell cycle arrest of PC3 cells in G2/M phase, whereas arrest in G0/G1 and G2/M phases was observed after treatment with EAELT. LC-HRMS/MS profiling of the extracts revealed the presence of known compounds that might be responsible for the observed anticancer activity such as chicoric acid, vicenin-2, vitexin and podophyllotoxin-β-d-glucoside.Discussion and conclusionsWe have shown, for the first time, that EAELN and EAELT exert anticancer activity through cell cycle arrest and induction of apoptosis. EAELN can be considered as a source to treat cancer. Further studies will be required to evaluate the effect of the active compounds, once identified, on other cancer cell lines.