3, 3'- (3, 5-DCPBC) Down-Regulates Multiple Phosphokinase Dependent Signal Transduction Pathways in Malignant Melanoma Cells through Specific Diminution of EGFRY1086 Phosphorylation.

Melanoma is the most dangerous skin malignancy due to its strong metastatic potential with high mortality. Activation of crucial signaling pathways enforcing melanoma progression depends on phosphorylation of distinct tyrosine kinases and oxidative stress. We here investigated the effect of a bis-coumarin derivative [3, 3'- ((3″, 5'-Dichlorophenyl) methylene) bis (4-hydroxy-2H-chromen-2-one)] [3, 3'- (3, 5-DCPBC)] on human melanoma cell survival, growth, proliferation, migration, intracellular redox state, and deciphered associated signaling pathways. This derivative is toxic for melanoma cells and non-toxic for melanocytes, their benign counterpart, and fibroblasts. 3, 3'- (3, 5-DCPBC) inhibits cell survival, migration, and proliferation of different metastatic and non-metastatic melanoma cell lines through profound suppression of the phosphorylation of Epidermal Growth Factor receptor (EGFR) and proto-oncogene cellular sarcoma (c-SRC) related downstream pathways. Thus, 3, 3'- (3, 5-DCPBC) endowed with the unique property to simultaneously suppress phosphorylation of multiple downstream kinases, such as EGFR/JAK/STAT and EGFR/SRC and their corresponding transcription factors.

Platinum complexes as inhibitors of DNA repair protein Ku70 and topoisomerase IIα in cancer cells.

Ku70 protein and topoisomerase IIα (Topo IIα) are promising targets of anticancer drugs, which play critical roles in DNA repair and replication processes. Three platinum(II) complexes, [PtCl(NH3)2(9-(pyridin-2-ylmethyl)-9H-carbazole)]NO3 (OPPC), [PtCl(NH3)2(9-(pyridin-3-ylmethyl)-9H-carbazole)]NO3 (MPPC), and [PtCl(NH3)2(9-(pyridin-4-ylmethyl)-9H-carbazole)]NO3 (PPPC), were designed as inhibitors of Ku70 and Topo IIα. Their antitumor activity and inhibitory efficacy on Ku70 and Topo IIα were investigated on cellular and molecular levels. OPPC exhibited high antiproliferative activity against various cancer cell lines, with acute toxicity to mice being lower than that of cisplatin. Moreover, OPPC could enter cancer cells effectively and cause DNA damage, which was evidenced by the enhanced expression of γ-H2AX, Chk1/2 phosphorylation, p53 and cell cycle arrest. OPPC also downregulated the DNA damage repair protein Ku70 and inhibited the formation of Ku70 foci-the central points or loci of Ku70, which would suppress DNA repair and induce a nonhomologous end joining response in cancer cells. More importantly, these complexes showed inhibition towards Topo IIα; in particular, OPPC was more effective than MPPC and PPPC. In the Topo IIα knockdown cells, Ku70 and Topo IIα were directly associated with the DNA damage and apoptotic response. The molecular docking provided detailed structural insights into the interactions of the complexes with Topo IIα. This study demonstrates that the cytotoxicity of these complexes is associated with the DNA damage and repair pathways mediated by Ku70 and Topo IIα; OPPC is an effective inhibitor of Ku70 and Topo IIα and restrains cancer cells via a mechanism utterly distinct from that of cisplatin.

Insecticide resistance in Australian Spodoptera frugiperda (J.E. Smith) and development of testing procedures for resistance surveillance.

Spodoptera frugiperda (J.E. Smith) is a highly invasive noctuid pest first reported in northern Australia during early 2020. To document current status of resistance in S. frugiperda in Australia, insecticide toxicity was tested in field populations collected during the first year of establishment, between March 2020 and March 2021. Dose-response was measured by larval bioassay in 11 populations of S. frugiperda and a susceptible laboratory strain of Helicoverpa armigera. Emamectin benzoate was the most efficacious insecticide (LC50 0.023µg/ml) followed by chlorantraniliprole (LC50 0.055µg/ml), spinetoram (LC50 0.098µg/ml), spinosad (LC50 0.526µg/ml), and methoxyfenozide (1.413µg/ml). Indoxacarb was the least toxic selective insecticide on S. frugiperda (LC50 3.789µg/ml). Emamectin benzoate, chlorantraniliprole and methoxyfenozide were 2- to 7-fold less toxic on S. frugiperda compared with H. armigera while spinosyns were equally toxic on both species. Indoxacarb was 28-fold less toxic on S. frugiperda compared with H. armigera. There was decreased sensitivity to Group 1 insecticides and synthetic pyrethroids in S. frugiperda compared with H. armigera: toxicity was reduced up to 11-fold for methomyl, 56 to 199-fold for cyhalothrin, and 44 to 132-fold for alpha cypermethrin. Synergism bioassays with metabolic inhibitors suggest involvement of mixed function oxidase in pyrethroid resistance. Recommended diagnostic doses for emamectin benzoate, chlorantraniliprole, spinetoram, spinosad, methoxyfenozide and indoxacarb are 0.19, 1.0, 0.75, 6, 12 and 48µg/µl, respectively.

Decarboxylases as hypothetical targets for actions of organophosphates: Molecular modeling for prediction of hidden and unexpected health threats.

The rise of various neurodegenerative disorders are somewhat correlating with the worldwide application of multiple anthropogenic toxicants. Though different possible targets were revealed to date, for example, for organophosphorus compounds (OPs), plenty of questions remain. Several decarboxylases (aromatic amino acid decarboxylase, AADC; histidine decarboxylase, HDC; glutamate decarboxylase, GAD) catalyze the biosynthesis of neurotransmitters and neuromodulators and contain pyridoxal phosphate (PLP) as a cofactor. In the current work, 18 OPs which have different neurotoxicity (chemical warfare agents and pesticides) and can penetrate through the blood-brain barrier, were selected. Then, their possible interaction with these decarboxylases in both apo- and holoforms was revealed using computer modeling methods (molecular docking and dynamics). The main amino acid residues of the enzymes responsible for binding OPs have been identified. Individual substances that are most dangerous from the point of view of a possible negative effect on the activity of several decarboxylases were revealed among studied OPs. Glyphosate should be of special interest, since it is not highly toxic towards serine hydrolases, but may prove to be a strong inhibitor for decarboxylases. Holo-AADC could be the most inhibition-prone enzyme among all those investigated.

Oviposition behaviour and biochemical response of an insect pest, Zeugodacus cucurbitae (Coquillett) (Diptera: Tephritidae) to plant phenolic compound phloroglucinol.

Phenolic compounds are the secondary metabolites (SMs) present in plants carrying different bioactivities. In the present study, we explored the influence of a phenolic compound namely phloroglucinol on oviposition behaviour and different biochemical entities of an insect pest Zeugodacus cucurbitae (Coquillett) (Diptera: Tephritidae) using artificial diet. Phloroglucinol (IUPAC name: benzene-1,3,5-triol) affected the activity of antioxidant and detoxifying enzymes viz. superoxide dismutases (SOD), catalase (CAT), ascorbate peroxidases (APOX). dehydroascorbate reductase (DHAR), peroxidases (POX), phenol oxidase (PO), glutathione peroxidase (GPOX), glutathione S-transferase peroxidase (GSTpox), glutathione reductase (GR), glutathione S-transferase (GST) and esterases (EST) as well as the biological antioxidants viz. ascorbate content and glutathione. The lipid peroxide content (LP) and hydrogen peroxide content (H2O2) were significantly enhanced in the treated larvae indicating oxidative stress in the insect. Significant inhibition in oviposition was observed and effective repellency percentage increased with phloroglucinol treatment as compared to control. The oviposition deterrent activity and toxic effects of phloroglucinol on various biochemical parameters of Z. cucurbitae larvae revealed in the present study clearly confirms its suitability for use in pest management.

Ibuprofen, Flurbiprofen, Etoricoxib or Paracetamol Do Not Influence ACE2 Expression and Activity In Vitro or in Mice and Do Not Exacerbate In-Vitro SARS-CoV-2 Infection.

SARS-CoV-2 uses the human cell surface protein angiotensin converting enzyme 2 (ACE2) as the receptor by which it gains access into lung and other tissue. Early in the pandemic, there was speculation that a number of commonly used medications-including ibuprofen and other non-steroidal anti-inflammatory drugs (NSAIDs)-have the potential to upregulate ACE2, thereby possibly facilitating viral entry and increasing the severity of COVID-19. We investigated the influence of the NSAIDS with a range of cyclooxygenase (COX)1 and COX2 selectivity (ibuprofen, flurbiprofen, etoricoxib) and paracetamol on the level of ACE2 mRNA/protein expression and activity as well as their influence on SARS-CoV-2 infection levels in a Caco-2 cell model. We also analysed the ACE2 mRNA/protein levels and activity in lung, heart and aorta in ibuprofen treated mice. The drugs had no effect on ACE2 mRNA/protein expression and activity in the Caco-2 cell model. There was no up-regulation of ACE2 mRNA/protein expression and activity in lung, heart and aorta tissue in ibuprofen-treated mice in comparison to untreated mice. Viral load was significantly reduced by both flurbiprofen and ibuprofen at high concentrations. Ibuprofen, flurbiprofen, etoricoxib and paracetamol demonstrated no effects on ACE2 expression or activity in vitro or in vivo. Higher concentrations of ibuprofen and flurbiprofen reduced SARS-CoV-2 replication in vitro.

Functional Roles of JNK and p38 MAPK Signaling in Nasopharyngeal Carcinoma.

c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) family members integrate signals that affect proliferation, differentiation, survival, and migration in a cell context- and cell type-specific way. JNK and p38 MAPK activities are found upregulated in nasopharyngeal carcinoma (NPC). Studies have shown that activation of JNK and p38 MAPK signaling can promote NPC oncogenesis by mechanisms within the cancer cells and interactions with the tumor microenvironment. They regulate multiple transcription activities and contribute to tumor-promoting processes, ranging from cell proliferation to apoptosis, inflammation, metastasis, and angiogenesis. Current literature suggests that JNK and p38 MAPK activation may exert pro-tumorigenic functions in NPC, though the underlying mechanisms are not well documented and have yet to be fully explored. Here, we aim to provide a narrative review of JNK and p38 MAPK pathways in human cancers with a primary focus on NPC. We also discuss the potential therapeutic agents that could be used to target JNK and p38 MAPK signaling in NPC, along with perspectives for future works. We aim to inspire future studies further delineating JNK and p38 MAPK signaling in NPC oncogenesis which might offer important insights for better strategies in diagnosis, prognosis, and treatment decision-making in NPC patients.

Kinome-Wide Profiling Identifies Human WNK3 as a Target of Cajanin Stilbene Acid from Cajanus cajan (L.) Millsp.

Pigeon Pea (Cajanus cajan (L.) Millsp.) is a common food crop used in many parts of the world for nutritional purposes. One of its chemical constituents is cajanin stilbene acid (CSA), which exerts anticancer activity in vitro and in vivo. In an effort to identify molecular targets of CSA, we performed a kinome-wide approach based on the measurement of the enzymatic activities of 252 human kinases. The serine-threonine kinase WNK3 (also known as protein kinase lysine-deficient 3) was identified as the most promising target of CSA with the strongest enzymatic activity inhibition in vitro and the highest binding affinity in molecular docking in silico. The lowest binding affinity and the predicted binding constant pKi of CSA (-9.65 kcal/mol and 0.084 µM) were comparable or even better than those of the known WNK3 inhibitor PP-121 (-9.42 kcal/mol and 0.123 µM). The statistically significant association between WNK3 mRNA expression and cellular responsiveness to several clinically established anticancer drugs in a panel of 60 tumor cell lines and the prognostic value of WNK3 mRNA expression in sarcoma biopsies for the survival time of 230 patients can be taken as clues that CSA-based inhibition of WNK3 may improve treatment outcomes of cancer patients and that CSA may serve as a valuable supplement to the currently used combination therapy protocols in oncology.

Involvement of Mitochondrial Mechanisms and Cyclooxygenase-2 Activation in the Effect of Desethylamiodarone on 4T1 Triple-Negative Breast Cancer Line.

Novel compounds significantly interfering with the mitochondrial energy production may have therapeutic value in triple-negative breast cancer (TNBC). This criterion is clearly fulfilled by desethylamiodarone (DEA), which is a major metabolite of amiodarone, a widely used antiarrhythmic drug, since the DEA previously demonstrated anti-neoplastic, anti-metastasizing, and direct mitochondrial effects in B16F10 melanoma cells. Additionally, the more than fifty years of clinical experience with amiodarone should answer most of the safety concerns about DEA. Accordingly, in the present study, we investigated DEA's potential in TNBC by using a TN and a hormone receptor positive (HR+) BC cell line. DEA reduced the viability, colony formation, and invasive growth of the 4T1 cell line and led to a higher extent of the MCF-7 cell line. It lowered mitochondrial transmembrane potential and induced mitochondrial fragmentation. On the other hand, DEA failed to significantly affect various parameters of the cellular energy metabolism as determined by a Seahorse live cell respirometer. Cyclooxygenase 2 (COX-2), which was upregulated by DEA in the TNBC cell line only, accounted for most of 4T1's DEA resistance, which was counteracted by the selective COX-2 inhibitor celecoxib. All these data indicate that DEA may have potentiality in the therapy of TNBC.

Drug Repurposing of the Unithiol: Inhibition of Metallo-ß-Lactamases for the Treatment of Carbapenem-Resistant Gram-Negative Bacterial Infections.

The increasing antibiotic resistance is a clinical problem worldwide. Numerous Gram-negative bacteria have already become resistant to the most widely used class of antibacterial drugs, ß-lactams. One of the main mechanisms is inactivation of ß-lactam antibiotics by bacterial ß-lactamases. Appearance and spread of these enzymes represent a continuous challenge for the clinical treatment of infections and for the design of new antibiotics and inhibitors. Drug repurposing is a prospective approach for finding new targets for drugs already approved for use. We describe here the inhibitory potency of known detoxifying antidote 2,3-dimercaptopropane-1-sulfonate (unithiol) against metallo-ß-lactamases. Unithiol acts as a competitive inhibitor of meropenem hydrolysis by recombinant metallo-ß-lactamase NDM-1 with the KI of 16.7 µM. It is an order of magnitude lower than the KI for l-captopril, the inhibitor of angiotensin-converting enzyme approved as a drug for the treatment of hypertension. Phenotypic methods demonstrate that the unithiol inhibits natural metallo-ß-lactamases NDM-1 and VIM-2 produced by carbapenem-resistant K. pneumoniae and P. aeruginosa bacterial strains. The 3D full atom structures of unithiol complexes with NDM-1 and VIM-2 are obtained using QM/MM modeling. The thiol group is located between zinc cations of the active site occupying the same place as the catalytic hydroxide anion in the enzyme-substrate complex. The sulfate group forms both a coordination bond with a zinc cation and hydrogen bonds with the positively charged residue, lysine or arginine, responsible for proper orientation of antibiotics upon binding to the active site prior to hydrolysis. Thus, we demonstrate both experimentally and theoretically that the unithiol is a prospective competitive inhibitor of metallo-ß-lactamases and it can be utilized in complex therapy together with the known ß-lactam antibiotics.

Comprehensive Detoxification Mechanism Assessment of Red Imported Fire Ant (Solenopsis invicta) against Indoxacarb.

The red imported fire ant (Solenopsis invicta) is one of the deadliest invasive ant species that threatens the world by disrupting biodiversity, important functions within a natural ecosystem, and community structure. They are responsible for huge economic losses in the infested countries every year. Synthetic insecticides, especially indoxacarb, have been broadly used to control S. invicta for many years. However, the biochemical response of S. invicta to indoxacarb remains largely undiscovered. Here, we used the sublethal doses of indoxacarb on the S. invicta collected from the eight different cities of Southern China. The alteration in the transcriptome profile of S. invicta following sublethal dosages of indoxacarb was characterized using high-throughput RNA-seq technology. We created 2 libraries, with 50.93 million and 47.44 million clean reads for indoxacarb treatment and control, respectively. A total of 2018 unigenes were regulated after insecticide treatment. Results indicated that a total of 158 differentially expressed genes (DEGs) were identified in the indoxacarb-treated group, of which 100 were significantly upregulated and 58 were downregulated, mostly belonging to the detoxification enzymes, such as AChE, CarE, and GSTs. Furthermore, results showed that most of these DEGs were found in several KEGG pathways, including steroid biosynthesis, other drug metabolizing enzymes, glycerolipid metabolism, chemical carcinogenesis, drug-metabolizing cytochrome P450, glutathione metabolism, glycerophospholipid metabolism, glycolysis/gluconeogenesis, and metabolism of xenobiotics. Together, these findings indicated that indoxacarb causes significant alteration in the transcriptome profile and signaling pathways of S. invicta, providing a foundation for further molecular inquiry.

Phosphodiesterase 4D promotes angiotensin II-induced hypertension in mice via smooth muscle cell contraction.

Hypertension is a common chronic disease, which leads to cardio-cerebrovascular diseases, and its prevalence is increasing. The cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) pathway participates in multiple cardiovascular diseases. Phosphodiesterase (PDE) 4 has been shown to regulate PKA activity via cAMP specific hydrolysis. However, whether PDE4-cAMP-PKA pathway influences hypertension remains unknown. Herein, we reveal that PDE4D (one of PDE4 isoforms) expression is upregulated in the aortas of experimental hypertension induced by angiotensin II (Ang II). Furthermore, knockout of Pde4d in mouse smooth muscle cells (SMCs) attenuates Ang II-induced hypertension, arterial wall media thickening, vascular fibrosis and vasocontraction. Additionally, we find that PDE4D deficiency activates PKA-AMP-activated protein kinase (AMPK) signaling pathway to inhibit myosin phosphatase targeting subunit 1 (MYPT1)-myosin light chain (MLC) phosphorylation, relieving Ang II-induced SMC contraction in vitro and in vivo. Our results also indicate that rolipram, a PDE4 inhibitor, may be a potential drug for hypertension therapy.

Anti-Phototoxicity Effect of Phenolic Compounds from Acetone Extract of Entada phaseoloides Leaves via Activation of COX-2 and iNOS in Human Epidermal Keratinocytes.

The extract from Entada phaseoloides was employed as active ingredients of natural origin into cosmetic products, while the components analysis was barely reported. Using LC-DAD-MS/qTOF analysis, eleven compounds (1-11) were proposed or identified from acetone extract of E. phaseoloides leaves (AE). Among them, six phenolic compounds, protocatechuic acid (2), 4-hydroxybenzoic acid (3), luteolin-7-O-ß-d-glucoside (5), cirsimaritin (6), dihydrokaempferol (9), and apigenin (10), were isolated by various chromatographic techniques. Protocatechuic acid (2), epicatechin (4), and kaempferol (11) at a concentration 100 µM increased the HaCaT cells viability of the UVB-irradiated cell without any cytotoxicity effect and reduced the expression of COX-2 and iNOS inflammation gene. Moreover, compounds 2 and 4 could have potent effects on cell migration during wound closure. These results suggest that compounds 2, 4, and 11 from AE have anti-photoaging properties and could be employed in pharmaceutical and cosmeceutical products.

Control of Expression of Key Cell Cycle Enzymes Drives Cell Line-Specific Functions of CDK7 in Human PDAC Cells.

Inhibition of the dual function cell cycle and transcription kinase CDK7 is known to affect the viability of cancer cells, but the mechanisms underlying cell line-specific growth control remain poorly understood. Here, we employed a previously developed, highly specific small molecule inhibitor that non-covalently blocks ATP binding to CDK7 (LDC4297) to study the mechanisms underlying cell line-specific growth using a panel of genetically heterogeneous human pancreatic tumor lines as model system. Although LDC4297 diminished both transcription rates and CDK T-loop phosphorylation in a comparable manner, some PDAC lines displayed significantly higher sensitivity than others. We focused our analyses on two well-responsive lines (Mia-Paca2 and Panc89) that, however, showed significant differences in their viability upon extended exposure to limiting LDC4297 concentrations. Biochemical and RNAseq analysis revealed striking differences in gene expression and cell cycle control. Especially the downregulation of a group of cell cycle control genes, among them CDK1/2 and CDC25A/C, correlated well to the observed viability differences in Panc89 versus Mia-Paca2 cells. A parallel downregulation of regulatory pathways supported the hypothesis of a feedforward programmatic effect of CDK7 inhibitors, eventually causing hypersensitivity of PDAC lines.

Expression of pyrethroid metabolizing P450 enzymes characterizes highly resistant Anopheles vector species targeted by successful deployment of PBO-treated bednets in Tanzania.

Long lasting insecticidal nets (LLINs) are a proven tool to reduce malaria transmission, but in Africa efficacy is being reduced by pyrethroid resistance in the major vectors. A previous study that was conducted in Muleba district, Tanzania indicated possible involvement of cytochrome P450 monooxygenases in a pyrethroid resistance in An. gambiae population where pre-exposure to piperonyl butoxide (PBO) followed by permethrin exposure in CDC bottle bioassays led to partial restoration of susceptibility. PBO is a synergist that can block pyrethroid-metabolizing enzymes in a mosquito. Insecticide resistance profiles and underlying mechanisms were investigated in Anopheles gambiae and An. funestus from Muleba during a cluster randomized trial. Diagnostic dose bioassays using permethrin, together with intensity assays, suggest pyrethroid resistance that is both strong and very common, but not extreme. Transcriptomic analysis found multiple P450 genes over expressed including CYP6M2, CYP6Z3, CYP6P3, CYP6P4, CYP6AA1 and CYP9K1 in An. gambiae and CYP6N1, CYP6M7, CYP6M1 and CYP6Z1 in An. funestus. Indeed, very similar suites of P450 enzymes commonly associated with resistant populations elsewhere in Africa were detected as over expressed suggesting a convergence of mechanisms across Sub-Saharan African malaria vectors. The findings give insight into factors that may correlate with pyrethroid PBO LLIN success, broadly supporting model predictions, but revision to guidelines previously issued by the World Health Organization is warranted.

LC-MS-based assay of granisetron 7-hydroxylation activity for the evaluation of CYP1A1 induction from diesel particulate matter-exposed hepatic and respiratory cell lines.

Particulate matter (PM) generally consists of aggregated particles containing trace metals and polycyclic aromatic hydrocarbons (PAHs). Cytochrome P450 (CYP) 1A1, one of the extensively investigated biomarkers, is highly inducible when PAHs activate the aryl hydrocarbon receptor (AhR). The present study focused on developing a LC-MS/MS-based assay to evaluate CYP1A1 induction potential following PM exposure. This assay adapted a CYP1A1 selective reaction of granisetron 7-hydroxylation in response to an AhR inducer, 6-formylindolo[3,2-b]carbazole (FICZ), in HepaRG and A549 cell lines. Exposure to FICZ (10 nM) increased the levels of granisetron 7-hydroxylation significantly, whereas no elevation of ethoxyresorufin-O-deethylation (EROD) activity was found in HepaRG cells. In A549 cells, granisetron 7-hydroxylation showed a better dose-response from 0 to 10000 nM FICZ treatment than EROD. EROD Additionally, the application of the assay with diesel PM exposure showed a concentration-dependent induction of CYP1A1 in HepaRG, A549, and human nasal epithelial cells. The granisetron assay has better selectivity for CYP1A1 than the conventional EROD assay, which is overlapped reaction with CYP1A2 and CYP1B1, with high correlations between AhR activation and CYP1A1 mRNA levels. Accompanying the great application potential to different organs and cell culture systems, future studies will implement the granisetron assay for the respiratory toxicity evaluation.

Quantitative analysis of resveratrol derivatives in the seed coats of tree peonies and their hypoglycemic activities in vitro/vivo.

Tree peonies are well-known horticultural and medicinal plants. The tree peony seeds, as emerging woody oil crops, recently have attracted great attention for their metabolites and bioactivities. In this study, the phytochemicals isolated from tree peony seed coats were systematically investigated. Seven polyphenolics were separated and prepared, mainly belonging to resveratrol derivatives. There was a great variation in the seed coat polyphenolic content among eight Paeonia species, and the contents of the resveratrol trimers and dimers were significantly higher in the seed coats of Paeonia ostii than other species. Based on the HPLC fingerprint characteristics and chemometric analysis, a clear discrimination among Paeonia plants was found, including the composition patterns and contents of the constituents. Moreover, the characteristic phytochemicals (vateriferol and trans-ε-viniferin) could significantly reduce the starch-mediated levels of postprandial blood glucose in diabetic/normal mice. In addition, in vitro enzyme tests showed that the two compounds could effectively and competitively inhibit α-glucosidase, with the IC50 values of 3.01 and 7.75 µM, respectively, indicating that vateriferol and trans-ε-viniferin could be therapeutic potential agents for hyperglycemia and diabetes mellitus.

Paeoniflorin alleviates NG-nitro-L-arginine methyl ester (L-NAME)-induced gestational hypertension and upregulates silent information regulator 2 related enzyme 1 (SIRT1) to reduce H2O2-induced endothelial cell damage.

Pregnancy-induced hypertension (PIH) is a leading cause of maternal mortality. Paeoniflorin has been reported to alleviate hypertension, thus relieving the injury of target organ. This study aimed to investigate the role of paeoniflorin in PIH development by regulating SIRT1 in rats. The mean arterial pressure (MAP), urine protein and histopathological damage of placenta in gestational hypertension rats were, respectively, detected by noninvasive tail-artery pressure measuring instrument, BCA method and H&E staining. The viability of human umbilical vein endothelial cells (HUVECs) treated with paeoniflorin or/and H2O2 was observed by CCK-8 assay. SIRT1 protein expression in HUVECs treated with paeoniflorin or/and H2O2 was analyzed by Western blot. Tunel assay, wound healing assay and tube formation assay were used to detect the apoptosis, migration and tube formation of HUVECs administrated with paeoniflorin or/and H2O2 or/and EX527 (SIRT1 inhibitor). As a result, MAP, urine protein and histopathological damage of placenta were enhanced in PIH rats, which were then alleviated by paeoniflorin. Paeoniflorin decreased the levels of sFlt-1, PlGF and VEGF in serum and placental tissues of gestational hypertension rats as well as the inflammatory response and oxidative stress. In addition, paeoniflorin promoted the expressions of SIRT1 and NO/eNOS and inhibited the production of iNOS in gestational hypertension rats to improve vascular endothelial cell injury. However, SIRT1 inhibition could suppress the protective effects of paeoniflorin on endothelial dysfunction of H2O2-induced HUVECs. In conclusion, paeoniflorin could improve gestational hypertension development by upregulating SIRT1.

The hsa_circRNA_102049 mediates the sorafenib sensitivity of hepatocellular carcinoma cells by regulating Reelin gene expression.

A growing body of research has illuminated that non-coding RNAs (ncRNAs) plays an important role in the development of drug resistance in hepatocellular carcinoma (HCC) cells. The expression profiles of differential expressed genes (DEGs) and ncRNAs related to the sorafenib resistance in HCC cells were analyzed according to the Gene Expression Omnibus (GEO) dataSets and The Cancer Genome Atlas (TCGA) datasets. Bioinformatics technology was used to construct the interaction network of DEGs and ncRNAs. Cell transfection, dual-luciferase reporter assay, Western blot, cell counting kit-8 (CCK-8), flow cytometry and quantitative real-time polymerase chain reaction(qRT-PCR) were used to study the mechanism of sorafenib resistance in HepG2 cells and Huh-7 cells. The expression of reelin (RELN) and secretagogin (SCGN) were the only down-regulated in sorafenib-resistant HCC cells. The results showed that RELN gene demethylation reversed the cytotoxic of sorafenib on HepG2 cells and Huh-7 cells. Hsa_circRNA_102049 over-expression promoted the sensitivity of HepG2 cells and Huh-7 cells to sorafenib, hsa_circRNA_102049 up-regulated the expression of RELN gene by sponging hsa-miR-214-3p. The resistance to sorafenib in RELN knockout HepG2 cells and Huh-7 cells could be reverted by has-circRNA_102049. These findings support targeting of hsa_circRNA_102049 and RELN in sorafenib-treated HCC cells as a novel intervention, which is expected to overcome sorafenib resistance of HCC cells.

Beneficial effects of metformin supplementation in hypothalamic paraventricular nucleus and arcuate nucleus of type 2 diabetic rats.

Background Oxidative stress and inflammation play important roles in the development of diabetes. Metformin (MET) is considered as the first-line therapy for patients with type 2 diabetes (T2D). Hypothalamic paraventricular nucleus (PVN) and hypothalamic arcuate nucleus (ARC) are vital in obesity and diabetes. However, there have been few studies on the effects of MET on inflammatory reaction and oxidative stress in the PVN and ARC of T2D diabetic rats. Methods Male Sprague-Dawley (SD) rats were fed with high-fat diet (HFD), and intraperitoneally injected with low-dose streptozotocin (STZ, 30 mg/kg) at 6th week to induce T2D diabetes. After injection of STZ, they were fed with HFD continually. Starting from the 8th week of HFD feeding, T2D rats received intragastrical administration of MET (150 mg/kg/day) in addition to the HFD for another 8 weeks. At the end of the 15th week, the rats were anaesthetized to record the sympathetic nerve activity and collect blood and tissue samples. Results In comparison with control rats, T2D diabetic rats had higher levels of pro-inflammatory cytokines (PICs) and excessive oxidative stress in the PVN and ARC, accompanied with more activated astrocytes. The renal sympathetic nerve activity (RSNA) and the plasma norepinephrine (NE) increased in T2D diabetic rats. The expression of tyrosine hydroxylase (TH) increased and the expression of 67-kDa isoform of glutamate decarboxylase (GAD67) decreased in T2D diabetic rats. Supplementation of MET decreased blood glucose, suppressed RSNA, decreased PICs (TNF-α, IL-1ß and IL-6) in PVN and ARC, attenuated oxidative stress and activation of astrocytes in ARC and PVN of T2D diabetic rats, as well as restored the balance of neurotransmitter synthetase. The number of Fra-LI (chronic neuronal excitation marker) positive neurons in the ARC and PVN of T2D diabetic rats increased. Chronic supplementation of MET also decreased the number of Fra-LI positive neurons in the ARC and PVN of T2D diabetic rats. Conclusion These findings suggest that the PVN and ARC participate in the beneficial effects of MET in T2D diabetic rats, which is possibly mediated via down-regulating of inflammatory molecules, attenuating oxidative stress and restoring the balance of neurotransmitter synthetase by MET in the PVN and ARC.