Allergoid-mannan conjugates reprogram monocytes into tolerogenic dendritic cells via epigenetic and metabolic rewiring.

BACKGROUND: Allergoid-mannan conjugates are novel vaccines for allergen-specific immunotherapy being currently assayed in phase 2 clinical trials. Allergoid-mannan conjugates target dendritic cells (DCs) and generate functional forkhead box P3 (FOXP3)-positive Treg cells, but their capacity to reprogram monocyte differentiation remains unknown. OBJECTIVE: We studied whether allergoid-mannan conjugates could reprogram monocyte differentiation into tolerogenic DCs and the underlying molecular mechanisms. METHODS: Monocytes from nonatopic and allergic subjects were differentiated into DCs under conventional protocols in the absence or presence of allergoid-mannan conjugates. ELISA, real-time quantitative PCR, coculture, flow cytometry, and suppression assay were performed. Metabolic and epigenetic techniques were also used. RESULTS: Monocyte differentiation from nonatopic and allergic subjects into DCs in the presence of allergoid-mannan conjugates yields stable tolerogenic DCs. Lipopolysaccharide-stimulated mannan-tolDCs show a significantly lower cytokine production, lower TNF-α/IL-10 ratio, and higher expression of the tolerogenic molecules PDL1, IDO, SOCS1, SOCS3, and IL10; and they induce higher numbers of functional FOXP3+ Treg cells than conventional DC counterparts. Mannan-tolDCs shift glucose metabolism from Warburg effect and lactate production to mitochondrial oxidative phosphorylation. They also display epigenetic reprogramming involving specific histone marks within tolerogenic loci and lower expression levels of histone deacetylase genes. Mannan-tolDCs significantly increase the expression of the anti-inflammatory miRNA-146a/b and decrease proinflammatory miRNA-155. CONCLUSIONS: Allergoid-mannan conjugates reprogram monocyte differentiation into stable tolerogenic DCs via epigenetic and metabolic reprogramming. Our findings shed light on the novel mechanisms by which allergoid-mannan conjugates might contribute to allergen tolerance induction during allergen-specific immunotherapy.

Inhibition of SYK and cSrc kinases can protect bone and cartilage in preclinical models of osteoarthritis and rheumatoid arthritis.

The pathophysiology of osteoarthritis (OA) includes the destruction of subchondral bone tissue and inflammation of the synovium. Thus, an effective disease-modifying treatment should act on both of these pathogenetic components. It is known that cSrc kinase is involved in bone and cartilage remodeling, and SYK kinase is associated with the inflammatory component. Thus the aim of this study was to characterize the mechanism of action and efficacy of a small molecule multikinase inhibitor MT-SYK-03 targeting SYK and cSrc kinases among others in different in vitro and in vivo arthritis models. The selectivity of MT-SYK-03 kinase inhibition was assayed on a panel of 341 kinases. The compound was evaluated in a set of in vitro models of OA and in vivo OA and RA models: surgically-induced arthritis (SIA), monosodium iodoacetate-induced arthritis (MIA), collagen-induced arthritis (CIA), adjuvant-induced arthritis (AIA). MT-SYK-03 inhibited cSrc and SYK with IC50 of 14.2 and 23 nM respectively. Only five kinases were inhibited > 90% at 500 nM of MT-SYK-03. In in vitro OA models MT-SYK-03 reduced hypertrophic changes of chondrocytes, bone resorption, and inhibited SYK-mediated inflammatory signaling. MT-SYK-03 showed preferential distribution to joint and bone tissue (in rats) and revealed disease-modifying activity in vivo by halving the depth of cartilage erosion in rat SIA model, and increasing the pain threshold in rat MIA model. Chondroprotective and antiresorptive effects were shown in a monotherapy regime and in combination with methotrexate (MTX) in murine and rat CIA models; an immune-mediated inflammation in rat AIA model was decreased. The obtained preclinical data support inhibition of cSrc and SYK as a viable strategy for disease-modifying treatment of OA. A Phase 2 clinical study of MT-SYK-03 is to be started.

Combined immune score predicts the prognosis of newly diagnosed multiple myeloma patients in the bortezomib-based therapy era.

ABSTRACT: To investigate the effect of a combined immune score including the lymphocyte-to-monocyte ratio (LMR) and uninvolved immunoglobulin (u-Ig) levels on the prognosis of newly diagnosed multiple myeloma (NDMM) patients treated with bortezomib.Clinical data of 201 NDMM patients were retrospectively analyzed. Patients with LMR ≥ 3.6 and LMR < 3.6 were scored 0 and 1, respectively. Patients with preserved u-Ig levels, suppression of 1 u-Ig, and suppression of at least 2 u-Igs were scored 0, 1, and 2, respectively. The immune score, established from these individual scores, was used to separate patients into good (0-1 points), intermediate (2 points), and poor (3 points) risk groups. The baseline data, objective remission rate (ORR), whether receive maintenance treatment regularly and overall survival of patients before treatment were analyzed.The ORR of the good-risk group was significantly higher than that of the intermediate-risk group (75.6% vs 57.7%, P = .044) and the poor-risk group (75.6% vs 48.2%, P = .007). The multivariate analysis results showed that age ≥ 65 years, International Staging System stage III, platelet count ≤ 100 × 109/L, lactate dehydrogenase (LDH) > 250 U/L, serum calcium > 2.75 mmol/L, no receipt of regular maintenance treatment, LMR < 3.6, suppressed u-Igs = 1, suppressed u-Igs ≥ 2, intermediate-risk group and poor-risk group were independent predictors of poor overall survival.In the bortezomib era, the LMR, u-Ig levels, and the immune score play an important role in the prognosis of NDMM patients. Among them, the immune score showed the strongest prognostic value, and it could be a beneficial supplement for the early identification of high-risk patients.

α-Ketoglutarate Inhibits Thrombosis and Inflammation by Prolyl Hydroxylase-2 Mediated Inactivation of Phospho-Akt.

BACKGROUND: Phospho-Akt1 (pAkt1) undergoes prolyl hydroxylation at Pro125 and Pro313 by the prolyl hydroxylase-2 (PHD2) in a reaction decarboxylating α-ketoglutarate (αKG). We investigated whether the αKG supplementation could inhibit Akt-mediated activation of platelets and monocytes, in vitro as well as in vivo, by augmenting PHD2 activity. METHODS: We treated platelets or monocytes isolated from healthy individuals with αKG in presence of agonists in vitro and assessed the signalling molecules including pAkt1. We supplemented mice with dietary αKG and estimated the functional responses of platelets and monocytes ex vivo. Further, we investigated the impact of dietary αKG on inflammation and thrombosis in lungs of mice either treated with thrombosis-inducing agent carrageenan or infected with SARS-CoV-2. FINDINGS: Octyl αKG supplementation to platelets promoted PHD2 activity through elevated intracellular αKG to succinate ratio, and reduced aggregation in vitro by suppressing pAkt1(Thr308). Augmented PHD2 activity was confirmed by increased hydroxylated-proline and enhanced binding of PHD2 to pAkt in αKG-treated platelets. Contrastingly, inhibitors of PHD2 significantly increased pAkt1 in platelets. Octyl-αKG followed similar mechanism in monocytes to inhibit cytokine secretion in vitro. Our data also describe a suppressed pAkt1 and reduced activation of platelets and leukocytes ex vivo from mice supplemented with dietary αKG, unaccompanied by alteration in their number. Dietary αKG significantly reduced clot formation and leukocyte accumulation in various organs including lungs of mice treated with thrombosis-inducing agent carrageenan. Importantly, in SARS-CoV-2 infected hamsters, we observed a significant rescue effect of dietary αKG on inflamed lungs with significantly reduced leukocyte accumulation, clot formation and viral load alongside down-modulation of pAkt in the lung of the infected animals. INTERPRETATION: Our study suggests that dietary αKG supplementation prevents Akt-driven maladies such as thrombosis and inflammation and rescues pathology of COVID19-infected lungs. FUNDING: Study was funded by the Department of Biotechnology (DBT), Govt. of India (grants: BT/PR22881 and BT/PR22985); and the Science and Engineering Research Board, Govt. of India (CRG/000092).

TNF-α inhibition, antioxidant effects and chemical analysis of extracts and fraction from Brazilian guaraná seed powder.

Paullinia cupana Kunth., commonly named Guaraná, is a plant from Brazil used as stimulant. The aim of this study was to evaluate the potential of extracts and tannins-rich and methylxanthines-free fraction from guaraná in the anti-inflammatory and antioxidant effect in vitro. Extract 1 obtained good yields of tannins and methylxanthines and was used to identify a type-A procyanidin trimer by LC-ESI-MS. Fraction 4 was rich in tannins and absent of methylxanthines. The extracts and fraction exhibited strong capacity for scavenging DPPH radical with IC50 between 5.88 and 42.75-µg/mL and inhibited TNF-α release by LPS-activated THP-1 cells when compared with control cells and did not present toxicity to THP-1 cells. The fraction 4, rich in tannins, was highly active, with IC50 5.88 µg/mL by DPPH method and inhibited TNF-α release in 83.50% at 90 µg/mL. These results reinforced potential anti-inflammatory of guaraná and data for new therapeutic approaches.

Berberine Directly Targets the NEK7 Protein to Block the NEK7-NLRP3 Interaction and Exert Anti-inflammatory Activity.

Berberine (BBR), a traditional Chinese medicine, has therapeutic effects on a variety of inflammation-related diseases, but its direct proteomic targets remain unknown. Using activity-based protein profiling, we first demonstrated that BBR directly targets the NEK7 protein via the hydrogen bond between the 2,3-methylenedioxy and 121-arginine (R121) residues. The fact that R121 is located precisely within the key domain involved in the NEK7-NLRP3 interaction allows BBR to specifically block the NEK7-NLRP3 interaction and successively inhibit IL-1ß release, independent of the NF-κB and TLR4 signaling pathways. Moreover, BBR displays in vivo anti-inflammatory efficacy in a NEK7-dependent manner. Therefore, we consider NEK7 to be a key target of BBR in the treatment of NLRP3-related inflammatory diseases, and the development of novel NEK7-NLRP3 interaction inhibitors might be easily achieved using NEK7 as a target.

Protective effects of citrulline supplementation in ulcerative colitis rats.

It has been reported that supplementing certain amino acids has therapeutic effects on ulcerative colitis (UC). We intend to explore whether citrulline (Cit) supplementation has protective effects on UC. Fifteen male Wistar rats were divided into normal control group (NC group), UC group and UC+Cit group, with five rats in each group. The UC model was established by TNBS/ethanol method. Rats in UC+Cit group were intragastrically administered with Cit for 7 consecutive days after modeling. All rats were sacrificed after 7 days. Blood samples were collected to detect the number of monocytes. Colon tissues were taken for HE staining. Immunohistochemistry staining for CD68 and p-STAT3 were performed to detect the infiltration of monocytes and the phosphorylation of STAT3 in colon tissues. The concentrations of MCP-1, IL-6 and IL-17A and the protein expression of p-STAT3 in colon tissues were measured by ELISA and western blot methods, respectively. The body weight of UC group rats decreased significantly after 7 days (p<0.05). However, the weight loss of UC+Cit group rats was not statistically significant (p>0.05). The number of peripheral blood monocytes in UC+Cit group was significantly lower than that in UC group (p<0.05), and the infiltration of CD68-positive monocytes in the colon tissue of UC+Cit group was significantly reduced than that in UC group. The concentrations of MCP-1, IL-6 and IL-17A and the expression of p-STAT3 in colon tissues of UC+Cit group rats were significantly lower than those in UC group (both p<0.05). Our study suggests that Cit supplementation may be a potential therapy for UC.

In Vitro and In Vivo Screening of Wild Bitter Melon Leaf for Anti-Inflammatory Activity against Cutibacterium acnes.

Cutibacterium acnes (formerly Propionibacterium acnes) is a key pathogen involved in the development and progression of acne inflammation. The numerous bioactive properties of wild bitter melon (WBM) leaf extract and their medicinal applications have been recognized for many years. In this study, we examined the suppressive effect of a methanolic extract (ME) of WBM leaf and fractionated components thereof on live C. acnes-induced in vitro and in vivo inflammation. Following methanol extraction of WBM leaves, we confirmed anti-inflammatory properties of ME in C. acnes-treated human THP-1 monocyte and mouse ear edema models. Using a bioassay-monitored isolation approach and a combination of liquid-liquid extraction and column chromatography, the ME was then separated into n-hexane, ethyl acetate, n-butanol and water-soluble fractions. The hexane fraction exerted the most potent anti-inflammatory effect, suppressing C. acnes-induced interleukin-8 (IL-8) production by 36%. The ethanol-soluble fraction (ESF), which was separated from the n-hexane fraction, significantly inhibited C. acnes-induced activation of mitogen-activated protein kinase (MAPK)-mediated cellular IL-8 production. Similarly, the ESF protected against C. acnes-stimulated mouse ear swelling, as measured by ear thickness (20%) and biopsy weight (23%). Twenty-four compounds in the ESF were identified using gas chromatograph-mass spectrum (GC/MS) analysis. Using co-cultures of C. acnes and THP-1 cells, ß-ionone, a compound of the ESF, reduced the production of IL-1ß and IL-8 up to 40% and 18%, respectively. ß-ionone also reduced epidermal microabscess, neutrophilic infiltration and IL-1ß expression in mouse ear. We also found evidence of the presence of anti-inflammatory substances in an unfractionated phenolic extract of WBM leaf, and demonstrated that the ESF is a potential anti-inflammatory agent for modulating in vitro and in vivo C. acnes-induced inflammatory responses.

The coadministration of trehalose dibehenate and monosodium urate crystals promotes an antitumor phenotype in human-derived myeloid cells.

Trehalose dibehenate (TDB), a ligand for the macrophage-inducible C-type lectin, has shown promise as an adjuvant for preventative vaccines and also as an anticancer agent in murine assays. The potential for TDB to affect the antitumor immune response of human myeloid cells, however, has not been studied. We investigated the effect of the adjuvants TDB and monosodium urate (MSU) crystals on the protumor or antitumor immune phenotype of human monocytes, macrophages and monocyte-derived dendritic cells (Mo-DCs). TDB treatment alone led to an inflammatory response in all three cell types, which was most pronounced when using human monocytes, with MSU augmenting this response. TDB also decreased cell surface markers associated with a protumorigenic phenotype, with MSU showing some ability to augment this response. Notably, a significant reduction in CD115 was observed for all antigen-presenting cells upon TDB or MSU + TDB treatment. The potential to increase the antigen-presenting capabilities of the myeloid cells was also observed upon treatment with TDB and MSU + TDB, as indicated by the upregulation of cell surface markers such as CD86 for all three cell types and a favorable ratio of interleukin (IL)-12p40 to IL-10 for monocytes stimulated with MSU + TDB. There was no significant production of IL-12p40 by Mo-DC; however, in a mixed lymphocyte assay, MSU + TDB costimulation of Mo-DC led to a significant increase in CD4+ T-cell numbers and in the IL-12p40-to-IL-10 ratio. Taken together, these findings show for the first time the potential of MSU + TDB costimulation to favor a tumor-suppressive phenotype in human-derived myeloid cells.

Autophagy in endothelial cells regulates their haematopoiesis-supporting ability.

BACKGROUND: Endothelial cells (ECs) function as an instructive platform to support haematopoietic stem cell (HSC) homeostasis. Our recent studies found that impaired bone marrow (BM) ECs are responsible for the defective haematopoiesis in patients with poor graft function (PGF), which is characterised by pancytopenia post-allotransplant. Although activated autophagy was reported to benefit ECs, whether EC autophagy plays a critical role in supporting HSCs and its effect on PGF patients post-allotransplant remain unclear. METHODS: To evaluate whether the autophagy status of ECs modulates their ability to support haematopoiesis, human umbilical vein endothelial cells (HUVECs) and primary BM ECs derived from healthy donors were subjected to knockdown or overexpression of Beclin-1 (an autophagy-related protein). Moreover, BM ECs derived from PGF patients were studied. FINDINGS: Beclin-1 knockdown significantly reduced the haematopoiesis-supporting ability of ECs by suppressing autophagy, which could be restored by activating autophagy via Beclin-1 upregulation. Moreover, autophagy positively regulated haematopoiesis-related genes in HUVECs. Subsequently, a prospective case-control study demonstrated that defective autophagy reduced Beclin-1 expression and the colony-forming unit (CFU) plating efficiency in BM ECs from PGF patients compared to matched patients with good graft function. Rapamycin, an autophagy activator, quantitatively and functionally improved BM ECs from PGF patients in vitro and enhanced their ability to support HSCs by activating the Beclin-1 pathway. INTERPRETATION: Our results suggest that the autophagy status of ECs modulates their ability to support haematopoiesis by regulating the Beclin-1 pathway. Defective autophagy in BM ECs may be involved in the pathogenesis of PGF post-allotransplant. Rapamycin provides a promising therapeutic approach for PGF patients. FUNDING: Please see funding sources.

T cell-derived soluble glycoprotein GPIbα mediates PGE2 production in human monocytes activated with the vaccine adjuvant MDP.

Vaccine adjuvants containing analogs of microbial products activate pattern recognition receptors (PRRs) on antigen-presenting cells, including monocytes and macrophages, which can cause prostaglandin E2 (PGE2) release and consequently undesired inflammatory responses and fever in vaccine recipients. Here, we studied the mechanism of PGE2 production by human monocytes activated with muramyl dipeptide (MDP) adjuvant, which activates cytosolic nucleotide-binding oligomerization domain 2 (NOD2). In rabbits, administration of MDP elicited an early increase in PGE2 followed by fever. In human monocytes, MDP alone did not induce PGE2 production. However, high amounts of PGE2 and the proinflammatory cytokines IL-1ß and IL-6 were secreted by monocytes activated with MDP in the presence of conditioned medium obtained from CD3 bead-isolated T cells (Tc CM) but not from those isolated without CD3 beads. Mass spectrometry and immunoblotting revealed that the costimulatory factor in Tc CM was glycoprotein Ib α (GPIbα). Antibody-mediated blockade of GPIbα or of its receptor, Mac-1 integrin, inhibited the secretion of PGE2, IL-1ß, and IL-6 in MDP + Tc CM-activated monocytes, whereas recombinant GPIbα protein increased PGE2 production by MDP-treated monocytes. In vivo, COX2 mRNA abundance was reduced in the liver and spleen of Mac-1 KO mice after administration of MDP compared with that of treated wild-type mice. Our findings suggest that the production of PGE2 and proinflammatory cytokines by MDP-activated monocytes is mediated by cooperation between two signaling pathways: one delivered by MDP through NOD2 and a second through activation of Mac-1 by T cell-derived GPIbα.

Re-analysis of herbal extracts data reveals that inflammatory processes are mediated by hormetic mechanisms.

Using data from Schink et al. (2018), a large number of herbal extracts were assessed for their capacity to induce pro- and anti-inflammatory effects based on TLR4 expression normalized for cell viability in two immune cell models (i.e., HeLa-TLR4 transfected reporter cell line, and THP-1 monocytes) applying seven concentrations (0.01-3.0%). The analysis revealed that 70-80% of the extracts satisfying the a priori entry criteria also satisfied a priori evaluative criteria for hormetic concentration responses. These findings demonstrate that a large proportion of herbal extracts display hormetic dose responses in immune cells, indicating that hormetic mechanisms mediate pro- and anti-inflammatory processes and may provide a means to guide optimal dosing strategies. The identification of doses eliciting only anti-inflammatory therapeutic activity as well as the use of dose-variable herbal extracts in the treatment of inflammatory diseases will be challenging.

Assessment of cytotoxicity and immune compatibility of phytochemicals-mediated biosynthesised silver nanoparticles using Cynara scolymus.

The study was focused on the phytochemicals-mediated biosynthesis of silver nanoparticles using leaf extracts and infusions from Cynara scolymus. To identify the antioxidant activity and total phenolic content, the 1,1-diphenyl-1-picrylhydrazyl and Folin-Ciocalteau methods were applied, respectively. The formation and stability of the reduced silver ions were monitored by UV-vis spectrophotometer. The particle sizes of the silver nanoparticles were characterised using the dynamic light scattering technique and scanning electron microscope. The phase composition of the obtained silver nanoparticles was characterised by X-ray diffraction. The silver nanoparticles suspension, artichoke infusion, and silver ions were separately tested towards potential cytotoxicity and pro-inflammatory effect using mouse fibroblasts and human monocytes cell line, respectively. The total phenolic content and antioxidant activity of ethanol extract and infusion were found significantly higher as compared to aqueous extract and infusion. The UV-visible spectrophotometric analysis revealed the presence of the characteristic absorption band of the Ag nanoparticles. Moreover, it was found that with the increasing volume of plant extract, the average size of particles was increased. Biocompatibility results evidently showed that silver nanoparticles do not induce monocyte activation, however in order to avoid their cytotoxicity suspension at a concentration <2 ppm should be applied.

Useful prognostic tools based on complete blood cell counts in diffuse large B-cell lymphoma.

INTRODUCTION: Diffuse large B-cell lymphoma (DLBCL) is the most frequent lymphoma. Three prognostic factors are widely used in DLBCL: International Prognostic Index (IPI), Revised-IPI (R-IPI), and National Comprehensive Cancer Network-IPI (NCCN-IPI). METHOD: We established a prognostic model using peripheral blood absolute lymphocyte/absolute monocyte counts ratio (LMR), hemoglobin, and platelet counts obtained from complete blood cell counts (CBC) data at diagnosis based on 214 cases of DLBCL who received more than one course of R-CHOP therapy at a single institution. RESULTS: The cutoff values for using the receiver operating characteristics (ROC) curve for LMR, hemoglobin, and platelet counts were 1.6, 100 g/L, and 150 × 109 /L, respectively. Stratification was performed using the three factors (LMR < 1.6, hemoglobin < 100 g/L, and platelet counts < 150 × 109 /L). CBC Group 1 (none of the 3 factors) included 92 cases, CBC Group 2 (1 or 2 of these factors) included 108 cases, and CBC Group 3 (all 3 factors) included 11 cases. The 5-year OS rates were 78.2%, 60.9%, and 10.1%, respectively. In multivariate analysis, CBC Group 3 (hazard ratio, 2.9760; 95% confidence interval, 1.2670-6.991; P = .01) were prognostic factors for OS. CBC Group 3 had factors based on which the further stratification of the poor prognosis group into IPI high-risk and R-IPI poor-risk groups (P = .01, <.0001, respectively) was possible. CONCLUSIONS: In DLBCL, combination of three CBC parameters has the potential to be a useful prognostic tool.

Pectin-rich extracts from olives inhibit proliferation of Caco-2 and THP-1 cells.

Three olive modified pectin extracts have been produced by heat and acid treatment of the major by-product of olive oil production. Their effect on proliferation of the colon carcinoma Caco-2 and the leukemia monocytic THP-1 cell lines has been studied in order to determine possible anti-tumor properties. All extracts inhibited proliferation at some of the concentrations ranging from 1 to 10 mg ml-1. Interestingly none of the extracts inhibited the growth of confluent Caco-2 cells, showing the specificity of the antiproliferative effect for the transformed Caco-2 phenotype. All the extracts inhibited agglutination of red blood cells by galectin-3, a lectin involved in tumor growth, metastasis, and immune cell regulation that has been proposed as a mediator of the anti-tumor effects of modified pectins. In addition, activation of caspase-3 in THP-1 cells indicates that treatment with the pectin-rich extracts triggers apoptosis. These results point to a possible use as health-promoting food ingredients or supplements.

Microwave diathermy induces mitogen-activated protein kinases and tumor necrosis factor-α in cultured human monocytes.

Although rehabilitation practice for most patients consists of a combined use of thermotherapy that is produced from diathermy devices resulting faster and deeper heating to the patient, major concerns about occupational exposure to electromagnetic radiation for the operators must be considered. In most occasions, physiotherapists have involved multi-hour treatment sessions to different patients, resulting overuse of the diathermy device. Recently, our team along with other groups have raised serious concerns about the occupational safety aspects related to microwave diathermy (MWD) use. Driven by these recent reports, in this work, we tried to investigate the in vitro effects of a physiotherapist routine MWD device regarding its potential inflammatory biological effects that could be evoked in human cultured monocytes. Our results show that MWD does not alter the integrity of the cell membrane and, consequently, the viability of monocytes as assessed by Trypan blue and MTT measurements. Then again, members of the MAPK family (p38 and ERK1/2) were activated upon MWD exposure at 5-30 min, eventually leading to a time-dependent considerable increase in TNF-α production, a key pro-inflammatory mediator. Our results are indicative of a stress-activated phenomenon of monocytes upon MWD radiation, which could trigger potential hazardous cellular outcomes due to thermal and/or non-thermal bystander effects. Our results deserve further investigation, planned by our team in due course, to delineate the clinical correlations of these findings.

Engineered Cell-Assisted Photoactive Nanoparticle Delivery for Image-Guided Synergistic Photodynamic/Photothermal Therapy of Cancer.

Photoactivated therapy, including photodynamic therapy (PDT) and photothermal therapy (PTT), is a spatiotemporally precise, controllable, and noninvasive method for tumor therapy and has therefore attracted increasing attention in recent years. However, it is still a challenge to obtain highly efficient therapeutic photoactive agents (PAAs) and deliver them into tumor, especially the core of solid tumors. Here, we have developed a newly engineered monocyte (MNC)-based PAA system that realizes precise and highly efficient tumor diagnosis and therapy. First, a near-infrared emissive PAA molecule with both strong singlet oxygen (1O2) production and high photothermal conversion efficiency was precisely designed for realizing simultaneous PDT and PTT of tumor and was further fabricated to form PAA nanoparticles (NPs). After loading the PAA NPs into MNCs, the MNCs were then decorated with cyclic Arg-Gly-Asp (cRGD) groups through a metabolic labeling method to further improve their ability of targeting and homing into the deep regions of tumors. Using this strategy, we have achieved highly efficient solid tumor ablation results both in vitro and in vivo, indicating that our strategy has a promising prospect for solid tumor therapy.

Polyalthia Clerodane Diterpene Potentiates Hypoglycemia via Inhibition of Dipeptidyl Peptidase 4.

Serine protease dipeptidyl peptidase 4 (DPP-4) is involved in self/non-self-recognition and insulin sensitivity. DPP-4 inhibitors are conventional choices for diabetic treatment; however, side effects such as headache, bronchus infection, and nasopharyngitis might affect the daily lives of diabetic patients. Notably, natural compounds are believed to have a similar efficacy with lower adverse effects. This study aimed to validate the DPP-4 inhibitory activity of clerodane diterpene 16-hydroxycleroda-3,13-dien-15,16-olide (HCD) from Polyalthia longifolia, rutin, quercetin, and berberine, previously selected through molecular docking. The inhibitory potency of natural DPP-4 candidates was further determined by enzymatic, in vitro Caco-2, and ERK/PKA activation in myocyte and pancreatic cells. The hypoglycemic efficacy of the natural compounds was consecutively analyzed by single-dose and multiple-dose administration in diet-induced obese diabetic mice. All the natural-compounds could directly inhibit DPP-4 activity in enzymatic assay and Caco-2 inhibition assay, and HCD showed the highest inhibition of the compounds. HCD down-regulated LPS-induced ERK phosphorylation in myocyte but blocked GLP-1 induced PKA expression. For in vivo tests, HCD showed hypoglycemic efficacy only in single-dose administration. After 28-days administration, HCD exhibited hypolipidemic and hepatoprotective efficacy. These results revealed that HCD performed potential antidiabetic activity via inhibition of single-dose and long-term administrations, and could be a new prospective anti-diabetic drug candidate.

Glucose-6-phosphate dehydrogenase deficiency increases cell adhesion molecules and activates human monocyte-endothelial cell adhesion: Protective role of l-cysteine.

Glucose-6-phosphate dehydrogenase is a major enzyme that supplies the reducing agent nicotinamide adenine dinucleotide phosphate hydrogen (NADPH), which is required to recycle oxidized/glutathione disulfide (GSSH) to reduced glutathione (GSH). G6PD-deficient cells are susceptible to oxidative stress and a deficiency of GSH. Endothelial dysfunction is characterized by the loss of nitric oxide (NO) bioavailability, which regulates leukocyte adhesion to endothelium. G6PD-deficient endothelial cells (EC) demonstrate reduced expression of endothelial nitric oxide synthase (eNOS) and NO levels along with reduced GSH. Whether G6PD deficiency plays any role in EC dysfunction is unknown. The chronic inflammation commonly seen in those with metabolic syndrome, characterized by elevated levels of tumor necrosis factor (TNF) and monocyte chemoattractant protein 1 (MCP-1), provided an incentive for investigation of these cytokines as well. A GSH/G6PD-deficient model was created using human umbilical vein endothelial cells (HUVEC) treated with either buthionine sulfoximine (BSO), a pharmacological inhibitor of the rate-limiting enzyme of GSH biosynthesis (γ-glutamylcysteine synthetase), or with 6-aminonicotinamide (6-AN), an inhibitor of G6PD or G6PD siRNA. Normal and G6PD-deficient cells were also treated with pro-atherosclerotic stimuli such as high glucose, TNF, and MCP-1. After inhibiting or knocking down G6PD/GSH, the capacity of endothelial cells for monocyte recruitment was assessed by determining the expression of the adhesion molecules intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1), which was upregulated by G6PD deficiency and accompanied by the presence of the oxidative stress markers NADPH oxidase 4 (NOX4), inducible nitric oxide synthase (iNOS), and reactive oxygen species (ROS). Treatment with the inhibitors BSO and 6-AN caused increased levels of adhesion molecule mRNA and monocyte-EC adhesion. Following treatment with high glucose, G6PD-deficient cells showed an increase in levels of ICAM-1 and VCAM-1 mRNA, as well as monocyte-EC adherence, compared with results seen in control cells. Treatment with l-cysteine (a precursor of GSH) protected endothelial cells by increasing GSH and attenuating ROS, ICAM-1, VCAM-1, and monocyte-EC adhesion. These results suggest that G6PD/GSH deficiency plays a role in endothelial dysfunction and that supplementation with l-cysteine can restore GSH levels and reduce the EC activation markers in G6PD-deficient conditions.

Flavonoid composition, cellular antioxidant activity and (myelo)peroxidase inhibition of a Bryonia alba L. (Cucurbitaceae) leaves extract.

OBJECTIVES: The aim of the present study consisted in the isolation of flavonoids from the leaves of Bryonia alba L. and evaluation of their antioxidant activity and inhibition on peroxidase-catalysed reactions. METHODS: Flavonoids were isolated by preparative HPLC-DAD and their structures were elucidated by MS and NMR. Inhibitory effect was tested by the horseradish peroxidase and the myeloperoxidase assays. Cellular antioxidant assays consisted in testing the inhibitory activity on the reactive oxygen species released upon activation of neutrophils freshly isolated ex vivo from equine blood and of human monocytes-derived macrophages in vitro. Whole organism toxicity was assessed on zebrafish larvae. KEY FINDINGS: Four flavonoids (lutonarin, saponarin, isoorientin and isovitexin) were isolated. The performed assays showed significant antioxidant activity and inhibition for the peroxidase-catalysed reactions. Absence of cellular and zebrafish toxicity was confirmed. CONCLUSIONS: Bryonia alba L. leaves are particularly interesting for their flavonoids content and showed significant inhibitory effect on peroxidase-catalysed oxidation of substrates (Amplex Red and L012), as well as antioxidant/antiradical activity, proving that this species has a medicinal potential. Moreover, the present study highlights the absence of the toxicity of these leaves and offers though a novel perspective on the species, previously known as being toxic.