Exosomal miRNA-155-5p from M1-polarized macrophages suppresses angiogenesis by targeting GDF6 to interrupt diabetic wound healing.

Unprogrammed macrophage polarization, especially prolonged activation of proinflammatory macrophages, is associated with delayed wound healing in diabetic objectives. Macrophage-derived exosomes cargo a variety of microRNAs (miRNAs), participating in different stages in wound healing. Here, exosomes were isolated from naive bone marrow-derived macrophages (BMDMs) (M0-Exos), interferon-γ plus lipopolysaccharide-polarized BMDMs (M1-Exos), and interleukin-4-polarized BMDMs (M2-Exos). M1-Exos impaired migration and tube formation in human umbilical vein endothelial cells (HUVECs) compared to M0-Exos, whereas M2-Exos exhibited the opposite effects. High-throughput sequencing was performed to decipher the miRNA expression profiles in M0-Exos, M1-Exos, and M2-Exos. A total of 63 miRNAs were identified to be differentially expressed in exosomes derived from polarized BMDMs. Among them, miRNA-155-5p is highly expressed in M1-Exos, which interrupted angiogenesis in HUVECs. Furthermore, miRNA-155-5p directly binds to the 3' UTR of growth differentiation factor 6 (GDF6) mRNA to suppress its protein expression. Lastly, local administration of a temperature-sensitive hydrogel Pluronic F-127 loading miRNA-155-5p antagomiR promoted angiogenesis and accelerated wound healing in diabetic db/db mice via enhancing GDF6. In summary, this study deciphered the miRNA expression profiles in exosomes from polarized macrophages. M2-like macrophage-derived exosomes and miRNA-155-5p inhibitors could be promising therapeutics against diabetic foot ulcers.

Limonoids from the roots of Melia azedarach and their anti-inflammatory activity.

Twelve undescribed limonoids, meliazedarines J-U (1-12), along with a known one, were isolated from the roots of Melia azedarach. Their structures were elucidated by extensive spectroscopic investigations, X-ray diffraction analyses, and ECD calculations. Compounds 1-8 were identified as ring intact limonoids, while compounds 9-12 were established as ring C-seco ones. The anti-inflammatory potential of compounds 1-4, 6, 8, 9, and 11-13 was evaluated on macrophages. Compounds 1, 3, 4, 6, and 9 significantly suppressed nitric oxide production in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages, among them compound 3 showed the best inhibitory effect with an IC50 value of 7.07 ± 0.48 µΜ. Furthermore, compound 3 effectively reduced interleukin-1ß secretion in LPS plus nigericin-induced THP-1 macrophages by inhibiting NLRP3 inflammasome activation. The results strongly suggested that limonoids from the roots of M. azedarach might be candidates for treating inflammation-related diseases.

Bisabolane-Type Sesquiterpenoids with a Tetrahydrofuran or Tetrahydropyran Ring from Vernonia solanifolia.

Phytochemical investigation of the aerial parts of Vernonia solanifolia resulted in the isolation of 23 new highly oxidized bisabolane-type sesquiterpenoids (1-23). Structures were determined by interpretation of spectroscopic data, single-crystal X-ray diffraction analysis, and time-dependent density functional theory electronic circular dichroism calculations. Most compounds possess a rare tetrahydrofuran (1-17) or tetrahydropyran ring (18-21). Compounds 1/2 and 11/12 are pairs of epimers isomerized at C-10, while compounds 9/10 and 15/16 are isomerized at C-11 and C-2, respectively. The anti-inflammatory effect in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages was evaluated for pure compounds. Compound 9 inhibited LPS-stimulated NO production at the concentration of 80 µM. It showed an anti-inflammatory effect by suppressing the activation of the NF-κB signaling pathway.

Germacrane-type sesquiterpenoids from the flowers of Chrysanthemum indicum with hepatoprotective activity.

Two new germacrane-type sesquiterpenoids, chrysanthemolides A (1) and B (2), and four known germacrane-type sesquiterpenoids, hanphyllin (3), 3ß-hydroxy-11α,13-dihydro-costunolide (4), costunolide (5), and 6,7-dimethylmethylene-4-aldehyde-1ß-hydroxy-10(15)-ene-(4Z)-dicyclodecylene (6), were isolated and identified from the flowers of Chrysanthemum indicum. The structures of the new compounds were elucidated via high resolution electrospray ionization mass spectrometry (HR-ESI-MS), 1D and 2D nuclear magnetic resonance (NMR) spectra and electronic circular dichroism (ECD). Meanwhile, all the isolates were tested for their hepatoprotective activity in tert-butyl hydroperoxide (t-BHP) injured AML12 cells. Compounds 1, 2, and 4 showed significant protective effects at 40 µM, comparable with the positive control resveratrol at 10 µM. As the most potent one, compound 1 was chosen for further studies. Compound 1 dose-dependently increased the viability of t-BHP-injured AML12 cells. Furthermore, compound 1 decreased reactive oxygen species accumulation, while increased glutathione level, heme oxygenase-1 level and superoxide dismutase activity, through anchoring in the binding site of Kelch domain of the Kelch-like ECH-associated protein 1 (Keap1) to promote the dissociation of nuclear factor erythroid 2-related factor 2 from Keap1 and translocation to nuclei. In summary, germacrane-type sesquiterpenoids from C. indicum might be further developed to protect liver against oxidative damage.

Fighting cancer by triggering non-canonical mitochondrial permeability transition-driven necrosis through reactive oxygen species induction.

Non-apoptotic necrosis shows therapeutic potential for the treatment of various diseases, especially cancer. Mitochondrial permeability transition (MPT)-driven necrosis is a form of non-apoptotic cell death triggered by oxidative stress and cytosolic Ca2+ overload, and relies on cyclophilin D (CypD). Previous reports demonstrated that isobavachalcone (IBC), a natural chalcone, has anticancer effect by apoptosis induction. Here, we found that IBC induced regulated necrosis in cancer cells. IBC triggered non-apoptotic cell death in lung and breast cancer cells mediated by reactive oxygen species (ROS). IBC caused mitochondrial injury and dysfunction as evidenced by mitochondrial Ca2+ overload, the opening of MPT pore, mitochondrial membrane potential collapse, and structural damages. IBC-triggered cell death could be remarkably reversed by the ROS scavengers, cyclosporin A (CsA) and hemin, whereas CypD silence and heme oxygenase-1 overexpression failed to do so. Protein kinase B, dihydroorotate dehydrogenase, and mitogen-activated protein kinases were not involved in IBC-induced necrosis as well. In addition, IBC showed an anticancer effect in a 4T1 breast cancer cell-derived allograft mouse model, and this effect was considerably reversed by CsA. Collectively, our results showed that IBC triggered non-canonical MPT-driven necrosis mediated by ROS in cancer cells, which might provide a novel strategy for fighting against cancer.

Lipophagy: Molecular Mechanisms and Implications in Hepatic Lipid Metabolism.

The liver is the most significant metabolic organ in the body and plays an important role in lipid metabolism. Liver lipid metabolism disorders cause hepatic diseases such as hepatitis, hepatic cirrhosis, and hepatoma. Autophagy is a process of generating energy and building blocks by degrading redundant or damaged proteins and organelles. Thus, it helps in the maintenance of cellular homeostasis. Recent discoveries revealed that lipophagy plays a vital role in hepatic cellular homeostasis and lipid metabolism. Its imbalance is always associated with the perturbation of lipid metabolism in the liver. This article reviewed the molecular mechanisms involved in lipophagy and the interaction between lipophagy and hepatic lipid metabolism. Increasing evidence suggests that lipophagy is an effective method to resolve liver diseases.

Affinity-based protein profiling-driven discovery of myricanol as a Nampt activator.

Herein, we synthesized an affinity-based probe of myricanol (pMY) with a photo-affinity cross-linker to initiate a bioconjugation reaction, which was applied for target identification in live C2C12 myotubes. Pull-down of biotinylated pMY coupled with mass spectroscopy and Western blotting revealed that pMY can bind with nicotinamide phosphoribosyltransferase (Nampt), a rate-limiting enzyme in the nicotinamide adenine dinucleotide salvage pathway. Cellular thermal shift assay, drug affinity responsive target stability assay and recombinant protein labeling further validated the direct interaction between myricanol and Nampt. Myricanol did not affect the protein expression of Nampt, but enhanced its activity. Knock-down of Nampt totally abolished the promoting effect of myricanol on insulin-stimulated glucose uptake in C2C12 myotubes. Taken together, myricanol sensitizes insulin action in myotubes through binding with and activating Nampt.

Comparison of Ophiopogon japonicus and Liriope spicata var. prolifera from Different Origins Based on Multi-Component Quantification and Anticancer Activity.

The tuberous root of Ophiopogon japonicus (Thunb.) Ker-Gawl. is a well-known Chinese medicine also called Maidong (MD) in Chinese. It could be divided into "Chuanmaidong" (CMD) and "Zhemaidong" (ZMD), according to the geographic origins. Meanwhile, the root of Liriope spicata (Thunb.) Lour. var. prolifera Y. T. Ma (SMD) is occasionally used as a substitute for MD in the market. In this study, a reliable pressurized liquid extraction and HPLC-DAD-ELSD method was developed for the simultaneous determination of nine chemical components, including four steroidal saponins (ophiopojaponin C, ophiopogonin D, liriopesides B and ophiopogonin D'), four homoisoflavonoids (methylophiopogonone A, methylophiopogonone B, methylophiopogonanone A and methylophiopogonanone B) and one sapogenin (ruscogenin) in CMD, ZMD and SMD. The method was validated in terms of linearity, sensitivity, precision, repeatability and accuracy, and then applied to the real samples from different origins. The results indicated that there were significant differences in the contents of the investigated compounds in CMD, ZMD and SMD. Ruscogenin was not detected in all the samples, and liriopesides B was only found in SMD samples. CMD contained higher ophiopogonin D and ophiopogonin D', while the other compounds were more abundant in ZMD. Moreover, the anticancer effects of the herbal extracts and selected components against A2780 human ovarian cancer cells were also compared. CMD and ZMD showed similar cytotoxic effects, which were stronger than those of SMD. The effects of MD may be due to the significant anticancer potential of ophiopognin D' and homoisoflavonoids. These results suggested that there were great differences in the chemical composition and pharmacological activity among CMD, ZMD and SMD; thus, their origins should be carefully considered in clinical application.

14-Deoxygarcinol improves insulin sensitivity in high-fat diet-induced obese mice via mitigating NF-κB/Sirtuin 2-NLRP3-mediated adipose tissue remodeling.

Interleukin (IL)-1ß is a culprit of adipose tissue inflammation, which in turn causes systematic inflammation and insulin resistance in obese individuals. IL-1ß is mainly produced in monocytes and macrophages and marginally in adipocytes, through cleavage of the inactive pro-IL-1ß precursor by caspase-1, which is activated via the NLRP3 inflammasome complex. The nuclear factor-κB (NF-κB) transcription factor is the master regulator of inflammatory responses. Brindle berry (Garcinia cambogia) has been widely used as health products for treating obesity and related metabolic disorders, but its active principles remain unclear. We previously found a series of polyisoprenylated benzophenones from brindle berry with anti-inflammatory activities. In this study we investigated whether 14-deoxygarcinol (DOG), a major polyisoprenylated benzophenone from brindle berry, alleviated adipose tissue inflammation and insulin sensitivity in high-fat diet fed mice. The mice were administered DOG (2.5, 5 mg · kg-1 · d-1, i.p.) for 4 weeks. We showed that DOG injection dose-dependently improved insulin resistance and hyperlipidemia, but not adiposity in high-fat diet-fed mice. We found that DOG injection significantly alleviated adipose tissue inflammation via preventing macrophage infiltration and pro-inflammatory polarization of macrophages, and adipose tissue fibrosis via reducing the abnormal deposition of extracellular matrix. In LPS plus nigericin-stimulated THP-1 macrophages, DOG (1.25, 2.5, 5 µM) dose-dependently suppressed the activation of NLRP3 inflammasome and NF-κB signaling pathway. We demonstrated that DOG bound to and activated the deacetylase Sirtuin 2, which in turn deacetylated and inactivated NLRP3 inflammasome to reduce IL-1ß secretion. Moreover, DOG (1.25, 2.5, 5 µM) dose-dependently mitigated inflammatory responses in macrophage conditioned media-treated adipocytes and suppressed macrophage migration toward adipocytes. Taken together, DOG might be a drug candidate to treat metabolic disorders through modulation of adipose tissue remodeling.

Simple Nano-Luciferase-Based Assay for the Rapid and High-Throughput Detection of SARS-CoV-2 3C-Like Protease.

In this study, we described an easy-to-perform nano-luciferase (nLuc) sensor for the rapid detection of 3-chymotrypsin-like protease (3CLpro) encoded by SARS-CoV-2. The technology is based on the cleavage reaction of recombinant-nLuc via 3CLpro. The nLuc-based assay is a general, one-step method and is naturally specific in detection. The stability, sensitivity, detection range, and response time are fully characterized. The application of 3CLpro detection in artificial and human saliva as well as antiviral drug screening demonstrates that the method can quantify 3CLpro with high sensitivity in one step. With its unique features, the nLuc-based assay may find broad applications in the auxiliary diagnosis of SARS-CoV-2, as well as other types of coronavirus infection.

Bioactivity-based molecular networking-guided identification of guttiferone J from Garcinia cambogia as an anti-obesity candidate.

BACKGROUND AND PURPOSE: Pharmacological intervention to induce browning of white adipose tissue provides a promising anti-obesity therapy. The fruits of Garcinia cambogia (Clusiaceae) have been widely applied to manage body weight; however, the chemical principles remain unclear. The current study aims to discover browning inducers from the fruits of G. cambogia and investigate the underlying mechanisms. EXPERIMENTAL APPROACH: The bioactivity-based molecular networking and Oil Red O staining on 3T3-L1 and C3H10T1/2 adipocytes were applied for guided isolation. High-fat diet-induced obese mice were recruited to evaluate the anti-obesity activity. KEY RESULTS: The bioactivity-based molecular networking-guided isolation yielded several polycyclic polyprenylated acylphloroglucinols from the fruits of G. cambogia with lipid-lowering effect in adipocytes, including guttiferone J (GOJ), garcinol and 14-deoxygarcinol. As the most potent one, GOJ (10 µM) reduced lipid accumulation by 70% and 76% in 3T3-L1 and C3H10T1/2 adipocytes, respectively. Furthermore, GOJ (2.5-10 µM) increased the expression of the deacetylase sirtuin 3 (SIRT3) and activated it, which, in turn, reduced the acetylation level of PPARγ coactivator-1α to boost mitochondrial biogenesis and promoted uncoupling protein 1 expression to enhance thermogenesis, resulting in browning of adipocytes. In high-fat diet-induced-obese mice, GOJ (10 and 20 mg·kg-1 ·day-1 for 12 weeks) protected against adiposity, hyperlipidaemia, insulin resistance and liver lipotoxicity, through boosting SIRT3-mediated browning of inguinal adipose tissue. CONCLUSION AND IMPLICATIONS: GOJ represents a new scaffold of thermogenic inducer, which is responsible for the anti-obesity property of G. cambogia and can be further developed as a candidate for treating obesity and its related disorders.

Sirtuins: Key players in obesity-associated adipose tissue remodeling.

Obesity, a complex disease involving an excessive amount of body fat and a major threat to public health all over the world, is the determining factor of the onset and development of metabolic disorders, including type 2 diabetes, cardiovascular diseases, and non-alcoholic fatty liver disease. Long-term overnutrition results in excessive expansion and dysfunction of adipose tissue, inflammatory responses and over-accumulation of extracellular matrix in adipose tissue, and ectopic lipid deposit in other organs, termed adipose tissue remodeling. The mammalian Sirtuins (SIRT1-7) are a family of conserved NAD+-dependent protein deacetylases. Mounting evidence has disclosed that Sirtuins and their prominent substrates participate in a variety of physiological and pathological processes, including cell cycle regulation, mitochondrial biogenesis and function, glucose and lipid metabolism, insulin action, inflammatory responses, and energy homeostasis. In this review, we provided up-to-date and comprehensive knowledge about the roles of Sirtuins in adipose tissue remodeling, focusing on the fate of adipocytes, lipid mobilization, adipose tissue inflammation and fibrosis, and browning of adipose tissue, and we summarized the clinical trials of Sirtuin activators and inhibitors in treating metabolic diseases, which might shed light on new therapeutic strategies for obesity and its associated metabolic diseases.

Chantriolides F-P, Highly Oxidized Withanolides with Hepatoprotective Activity from Tacca chantrieri.

Eleven highly oxidized withanolides, chantriolides F-P (1-11), together with six known analogues (12-17), were isolated from the rhizomes of Tacca chantrieri. Their structures were established on the basis of comprehensive spectroscopic data analysis and comparison with published NMR data, and their absolute configurations were further confirmed by experimental ECD data and single crystal X-ray diffraction analysis. The structures of compounds 5-8 contained a chlorine atom substituted at C-3. Compounds 1 and 12 are a pair of epimers isomerized at C-24 and C-25, while compounds 9 and 16 are isomerized at C-1, C-7, C-24, and C-25. Next, the hepatoprotective effect of all the isolates was evaluated on tert-butyl hydroperoxide (t-BHP)-injured AML12 hepatocytes. Compounds 5-11 and 16 significantly enhanced cell viability. Compound 8 decreased reactive oxygen species accumulation and increased glutathione level in t-BHP injured AML12 hepatocytes through promoting nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2).

Mechanistic insights into the clinical Y96D mutation with acquired resistance to AMG510 in the KRASG12C.

Special oncogenic mutations in the RAS proteins lead to the aberrant activation of RAS and its downstream signaling pathways. AMG510, the first approval drug for KRAS, covalently binds to the mutated cysteine 12 of KRASG12C protein and has shown promising antitumor activity in clinical trials. Recent studies have reported that the clinically acquired Y96D mutation could severely affect the effectiveness of AMG510. However, the underlying mechanism of the drug-resistance remains unclear. To address this, we performed multiple microsecond molecular dynamics simulations on the KRASG12C-AMG510 and KRASG12C/Y96D-AMG510 complexes at the atomic level. The direct interaction between the residue 96 and AMG510 was impaired owing to the Y96D mutation. Moreover, the mutation yielded higher flexibility and more coupled motion of the switch II and α3-helix, which led to the departing motion of the switch II and α3-helix. The resulting departing motion impaired the interaction between the switch II and α3-helix and subsequently induced the opening and loosening of the AMG510 binding pocket, which further disrupted the interaction between the key residues in the pocket and AMG510 and induced an increased solvent exposure of AMG510. These findings reveal the resistance mechanism of AMG510 to KRASG12C/Y96D, which will help to offer guidance for the development of KRAS targeted drugs to overcome acquired resistance.

Ablation of Ghrelin Receptor Mitigates the Metabolic Decline of Aging Skeletal Muscle.

The orexigenic hormone ghrelin has multifaceted roles in health and disease. We have reported that ablation of the ghrelin receptor, growth hormone secretagogue receptor (GHS-R), protects against metabolic dysfunction of adipose tissues in aging. Our further observation interestingly revealed that GHS-R deficiency phenocopies the effects of myokine irisin. In this study, we aim to determine whether GHS-R affects the metabolic functions of aging skeletal muscle and whether GHS-R regulates the muscular functions via irisin. We first studied the expression of metabolic signature genes in gastrocnemius muscle of young, middle-aged and old mice. Then, old GHS-R knockout (Ghsr-/-) mice and their wild type counterparts were used to assess the impact of GHS-R ablation on the metabolic characteristics of gastrocnemius and soleus muscle. There was an increase of GHS-R expression in skeletal muscle during aging, inversely correlated with the decline of metabolic functions. Remarkedly the muscle of old GHS-R knockout (Ghsr-/-) mice exhibited a youthful metabolic profile and better maintenance of oxidative type 2 muscle fibers. Furthermore, old Ghsr-/- mice showed improved treadmill performance, supporting better functionality. Also intriguing to note was the fact that old GHS-R-ablated mice showed increased expression of the irisin precursor FNDC5 in the muscle and elevated plasma irisin levels in circulation, which supports a potential interrelationship between GHS-R and irisin. Overall, our work suggests that GHS-R has deleterious effects on the metabolism of aging muscle, which may be at least partially mediated by myokine irisin.

Homocysteine-targeting compounds as a new treatment strategy for diabetic wounds via inhibition of the histone methyltransferase SET7/9.

In hypoxia and hyperglycemia, SET7/9 plays an important role in controlling HIF-1α methylation and regulating the transcription of HIF-1α target genes, which are responsible for angiogenesis and wound healing. Here, we report the Ir(III) complex Set7_1a bearing acetonitrile (ACN) ligands as a SET7/9 methyltransferase inhibitor and HIF-1α stabilizer. Interestingly, Set7_1a could engage SET7/9 and strongly inhibit SET7/9 activity, especially after preincubation with homocysteine (Hcy), which is elevated in diabetes. We hypothesize that Set7_1a exchanges ACN subunits for Hcy to disrupt the interaction between SET7/9 and SAM/SAH, which are structurally related to Hcy. Inhibition of SET7/9 methyltransferase activity by Set7_1a led to reduced HIF-1α methylation at the lysine 32 residue, causing increased HIF-1α level and recruitment of HIF-1α target genes that promote angiogenesis, such as VEGF, GLUT1, and EPO, in hypoxia and hyperglycemia. Significantly, Set7_1a improved wound healing in a type 2 diabetic mouse model by activating HIF-1α signaling and downstream proangiogenic factors. To our knowledge, this is the first Hcy-targeting iridium compound shown to be a SET7/9 antagonist that can accelerate diabetic wound healing. More importantly, this study opens a therapeutic avenue for the treatment of diabetic wounds by the inhibition of SET7/9 lysine methyltransferase activity.

Exosome-cargoed microRNAs: Potential therapeutic molecules for diabetic wound healing.

Diabetic foot ulcers are one of the most common complications of diabetes, requiring repeated surgical intervention and leading to amputation. Owing to the lack of effective drugs, novel therapeutics need to be explored. Decreased angiogenic factors, endothelial cell dysfunction and vascular lumen stenosis impair angiogenesis in diabetic wounds. Exosome-cargoed microRNAs are emerging as pivotal regulators of angiogenesis during wound closure. Herein, we summarize the up-to-date knowledge of exosomal microRNAs in modulating angiogenesis and accelerating diabetic wound healing, as well as their targets and underlying mechanisms. Exosomal microRNAs could be therapeutics with negligible rejection complications and good compatibility to treat diabetic foot ulcers.

Triterpenoids from the fruits of Melia azedarach L. and their cytotoxic activities.

Eleven undescribed tetracyclic triterpenoids, meliazedarachins A-K, along with twenty-six known compounds were isolated from the fruits of Melia azedarach L.. Their structures were determined by HRESIMS, UV, IR, NMR, X-ray diffraction, electronic circular dichroism (ECD) spectra, and the modified Mosher's method. The cytotoxic activities of all the isolates were measured. Meliazedarachin K and mesendanin N showed cytotoxicity against five human cancer cell lines with IC50 values ranging from 9.02 to 31.31 µM. Meliazedarachin K showed significant cytotoxicity against HCT116 cell line with IC50 value of 9.02 ± 0.84 µM. 21α-methylmelianodiol showed significant cytotoxicity against HCT116 and RKO cell lines with IC50 values of 10.16 ± 1.22 and 8.57 ± 0.80 µM, respectively.

Canthin-6-one (CO) from Picrasma quassioides (D.Don) Benn. ameliorates lipopolysaccharide (LPS)-induced astrocyte activation and associated brain endothelial disruption.

BACKGROUND: Canthin-6-one (CO) is an active ingredient found in Picrasma quassioides (D.Don) Benn. (PQ) that displays various biological activities including anti-inflammatory properties. Several studies reported PQ displayed neuroprotective activities, but its effects on astrocytes have not yet been investigated. Astrocytes are crucial regulators of neuroinflammatory responses under pathological conditions in the central nervous system (CNS). Proinflammatory astrocytes can induce the blood-brain barrier (BBB) breakdown, which plays a key role in the progression of neurodegenerative disorder (ND). PURPOSE: This study aims to investigate the anti-neuroinflammatory effects of CO in LPS-induced astrocyte activation and its underlying mechanisms in protecting the blood-brain barrier (BBB) in vitro. METHODS: Mouse astrocytes (C8-D1A) were activated with lipopolysaccharide (LPS) with or without CO pretreatment. Effects of CO on astrocyte cell viability, secretions of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1ß (IL-1ß) and nitric oxide (NO) were determined. Intracellular transcriptions and translations of proinflammatory mediators, molecular signaling, [Ca2+] and the levels of reactive oxygen species (ROS) were evaluated by RT-PCR, western blotting, and flow cytometry, respectively. Astrocyte-conditioned medium (ACM) was further prepared for incubating endothelial monolayer (bEnd.3) grown on transwell. Endothelial disruptions were evaluated by transendothelial electrical resistance (TEER), FITC-dextran permeability and monocyte adhesion assays. Endothelial tight junctions (TJs) and molecular signaling pathways were evaluated by immunofluorescence staining and western blotting. RESULTS: CO attenuated LPS-induced expression of astrocytic proinflammatory mediators (TNF-α, IL-1ß, IL-6, NO) and inhibited deleterious molecular activities including inducible nitric oxide synthase (iNOS), p-NFκB and p-STAT3 in astrocytes. Incubation of ACM collected from CO-treated astrocytes significantly ameliorated endothelial disruptions, reduced expressions of endothelial cytokine receptors (IL-6R, gp130 (IL-6RB), TNFR and IL-1R), suppressed proinflammatory pathways, MAPKs (p-AKT, p-MEK, p-ERK, p-p38, p-JNK) and p-STAT3, restored endothelial stabilizing pathways (p-Rac 1) and upregulated beneficial endothelial nitric oxide synthase (eNOS). CONCLUSION: Our study demonstrates for the first time CO exhibited potent protective effects against astrocyte-mediated proinflammatory responses and associated endothelial barrier disruptions.

The role of irisin in metabolic flexibility: Beyond adipose tissue browning.

Metabolic flexibility is the ability to adapt to physiological and environmental changes in metabolic demand. Irisin was originally discovered as an exercise-induced myokine involved in fat browning. In this review, we summarize emerging evidence for the roles of irisin in regulating glucose metabolism and insulin sensitivity in skeletal muscle, neuroplasticity and satiety in central nervous system, ß cell function and insulin secretion in the pancreas, bone remodeling, and adipose tissue function, which together orchestrate whole-body metabolic flexibility. Irisin is a key communicating mediator between skeletal muscle and other organs, and its manipulation could be a promising therapeutic strategy for treating obesity and related metabolic disorders.