MNK2 governs the macrophage antiinflammatory phenotype.

Tumor-associated macrophages (TAMs) continuously fine tune their immune modulatory properties, but how gene expression programs coordinate this immune cell plasticity is largely unknown. Selective mRNA translation, controlled by MNK1/MNK2 and mTOR pathways impinging on eIF4E, facilitates reshaping of proteomes without changes in abundance of corresponding mRNAs. Using polysome profiling developed for small samples we show that, during tumor growth, gene expression in TAMs is predominately modulated via mRNA-selective changes in translational efficiencies. These alterations in gene expression paralleled accumulation of antiinflammatory macrophages with augmented phosphorylation of eIF4E, a target of the MNK1 and MNK2 kinases, known to selectively modulate mRNA translation. Furthermore, suppression of the MNK2, but not the mTOR signaling pathway, reprogrammed antiinflammatory macrophages toward a proinflammatory phenotype with the ability to activate CD8+ T cells. Thus, selective changes of mRNA translation depending on MNK2 signaling represents a key node regulating macrophage antiinflammatory functions.

Bionic Bilayer Scaffold for Synchronous Hyperthermia Therapy of Orthotopic Osteosarcoma and Osteochondral Regeneration.

Large osseous void, postsurgical neoplastic recurrence, and slow bone-cartilage repair rate raise an imperative need to develop functional scaffold in clinical osteosarcoma treatment. Herein, a bionic bilayer scaffold constituting croconaine dye-polyethylene glycol@sodium alginate hydrogel and poly(l-lactide)/hydroxyapatite polymer matrix is fabricated to simultaneously achieve a highly efficient killing of osteosarcoma and an accelerated osteochondral regeneration. First, biomimetic osteochondral structure along with adequate interfacial interaction of the bilayer scaffold provide a structural reinforcement for transverse osseointegration and osteochondral regeneration, as evidenced by upregulated specific expressions of collagen type-I, osteopontin, and runt-related transcription factor 2. Meanwhile, thermal ablation of the synthesized nanoparticles and mitochondrial dysfunction caused by continuously released hydroxyapatite induce residual tumor necrosis synergistically. To validate the capabilities of inhibiting tumor growth and promoting osteochondral regeneration of our proposed scaffold, a novel orthotopic osteosarcoma model simulating clinical treatment scenarios of bone tumors is established on rats. Based on amounts of in vitro and in vivo results, an effective killing of osteosarcoma and a suitable osteal-microenvironment modulation of such bionic bilayer composite scaffold are achieved, which provides insightful implications for photonic hyperthermia therapy against osteosarcoma and following osseous tissue regeneration.

A subset of type-II collagen-binding antibodies prevents experimental arthritis by inhibiting FCGR3 signaling in neutrophils.

Rheumatoid arthritis (RA) involves several classes of pathogenic autoantibodies, some of which react with type-II collagen (COL2) in articular cartilage. We previously described a subset of COL2 antibodies targeting the F4 epitope (ERGLKGHRGFT) that could be regulatory. Here, using phage display, we developed recombinant antibodies against this epitope and examined the underlying mechanism of action. One of these antibodies, R69-4, protected against cartilage antibody- and collagen-induced arthritis in mice, but not autoimmune disease models independent of arthritogenic autoantibodies. R69-4 was further shown to cross-react with a large range of proteins within the inflamed synovial fluid, such as the complement protein C1q. Complexed R69-4 inhibited neutrophil FCGR3 signaling, thereby impairing downstream IL-1ß secretion and neutrophil self-orchestrated recruitment. Likewise, human isotypes of R69-4 protected against arthritis with comparable efficiency. We conclude that R69-4 abrogates autoantibody-mediated arthritis mainly by hindering FCGR3 signaling, highlighting its potential clinical utility in acute RA.

A subset of antibodies targeting citrullinated proteins confers protection from rheumatoid arthritis.

Although elevated levels of anti-citrullinated protein antibodies (ACPAs) are a hallmark of rheumatoid arthritis (RA), the in vivo functions of these antibodies remain unclear. Here, we have expressed monoclonal ACPAs derived from patients with RA, and analyzed their functions in mice, as well as their specificities. None of the ACPAs showed arthritogenicity nor induced pain-associated behavior in mice. However, one of the antibodies, clone E4, protected mice from antibody-induced arthritis. E4 showed a binding pattern restricted to skin, macrophages and dendritic cells in lymphoid tissue, and cartilage derived from mouse and human arthritic joints. Proteomic analysis confirmed that E4 strongly binds to macrophages and certain RA synovial fluid proteins such as α-enolase. The protective effect of E4 was epitope-specific and dependent on the interaction between E4-citrullinated α-enolase immune complexes with FCGR2B on macrophages, resulting in increased IL-10 secretion and reduced osteoclastogenesis. These findings suggest that a subset of ACPAs have therapeutic potential in RA.

Osteoimmunity-Regulating Biomimetically Hierarchical Scaffold for Augmented Bone Regeneration.

Engineering a proper immune response following biomaterial implantation is essential to bone tissue regeneration. Herein, a biomimetically hierarchical scaffold composed of deferoxamine@poly(ε-caprolactone) nanoparticles (DFO@PCL NPs), manganese carbonyl (MnCO) nanosheets, gelatin methacryloyl hydrogel, and a polylactide/hydroxyapatite (HA) matrix is fabricated to augment bone repair by facilitating the balance of the immune system and bone metabolism. First, a 3D printed stiff scaffold with a well-organized gradient structure mimics the cortical and cancellous bone tissues; meanwhile, an inside infusion of a soft hydrogel further endows the scaffold with characteristics of the extracellular matrix. A Fenton-like reaction between MnCO and endogenous hydrogen peroxide generated at the implant-tissue site triggers continuous release of carbon monoxide and Mn2+ , thus significantly lessening inflammatory response by upregulating the M2 phenotype of macrophages, which also secretes vascular endothelial growth factor to induce vascular formation. Through activating the hypoxia-inducible factor-1α pathway, Mn2+ and DFO@PCL NP further promote angiogenesis. Moreover, DFO inhibits osteoclast differentiation and synergistically collaborates with the osteoinductive activity of HA. Based on amounts of data in vitro and in vivo, strong immunomodulatory, intensive angiogenic, weak osteoclastogenic, and superior osteogenic abilities of such an osteoimmunity-regulating scaffold present a profound effect on improving bone regeneration, which puts forward a worthy base and positive enlightenment for large-scale bone defect repair.

Antibodies to Cartilage Oligomeric Matrix Protein Are Pathogenic in Mice and May Be Clinically Relevant in Rheumatoid Arthritis.

OBJECTIVE: Cartilage oligomeric matrix protein (COMP) is an autoantigen in rheumatoid arthritis (RA) and experimental models of arthritis. This study was undertaken to investigate the structure, function, and relevance of anti-COMP antibodies. METHODS: We investigated the pathogenicity of monoclonal anti-COMP antibodies in mice using passive transfer experiments, and we explored the interaction of anti-COMP antibodies with cartilage using immunohistochemical staining. The interaction of the monoclonal antibody 15A11 in complex with its specific COMP epitope P6 was determined by x-ray crystallography. An enzyme-linked immunosorbent assay and a surface plasma resonance technique were used to study the modulation of calcium ion binding to 15A11. The clinical relevance and value of serum IgG specific to the COMP P6 epitope and its citrullinated variants were evaluated in a large Swedish cohort of RA patients. RESULTS: The murine monoclonal anti-COMP antibody 15A11 induced arthritis in naive mice. The crystal structure of the 15A11-P6 complex explained how the antibody could bind to COMP, which can be modulated by calcium ions. Moreover, serum IgG specific to the COMP P6 peptide and its citrullinated variants was detectable at significantly higher levels in RA patients compared to healthy controls and correlated with a higher disease activity score. CONCLUSION: Our findings provide the structural basis for binding a pathogenic anti-COMP antibody to cartilage. The recognized epitope can be citrullinated, and levels of antibodies to this epitope are elevated in RA patients and correlate with higher disease activity, implicating a pathogenic role of anti-COMP antibodies in a subset of RA patients.

Tumor MHC class I expression alters cancer-associated myelopoiesis driven by host NK cells.

Downregulation of MHC class I (MHCI) molecules on tumor cells is recognized as a resistance mechanism of cancer immunotherapy. Given that MHCI molecules are potent regulators of immune responses, we postulated that the expression of MHCI by tumor cells influences systemic immune responses. Accordingly, mice-bearing MHCI-deficient tumor cells showed reduced tumor-associated extramedullary myelopoiesis in the spleen. Depletion of natural killer (NK) cells abrogated these differences, suggesting an integral role of immune-regulatory NK cells during tumor progression. Cytokine-profiling revealed an upregulation of TNF-α by NK cells in tumors and spleen in mice-bearing MHCI expressing tumors, and inhibition of TNF-α enhanced host myelopoiesis in mice receiving adoptive transfer of tumor-experienced NK cells. Our study highlights a critical role of NK cells beyond its identity as a killer lymphocyte and more importantly, the potential host responses to a localized tumor as determined by its MHCI expression.

A mussel-inspired supramolecular hydrogel with robust tissue anchor for rapid hemostasis of arterial and visceral bleedings.

In recent years, the developed hemostatic technologies are still difficult to be applied to the hemostasis of massive arterial and visceral hemorrhage, owing to their weak hemostatic function, inferior wet tissue adhesion, and low mechanical properties. Herein, a mussel-inspired supramolecular interaction-cross-linked hydrogel with robust mechanical property (308.47 ± 29.20 kPa) and excellent hemostatic efficiency (96.5% ± 2.1%) was constructed as a hemostatic sealant. Typically, we combined chitosan (CS) with silk fibroin (SF) by cross-linking them through tannic acid (TA) to maintain the structural stability of the hydrogel, especially for wet tissue adhesion ability (shear adhesive strength = 29.66 ± 0.36 kPa). Compared with other materials reported previously, the obtained CS/TA/SF hydrogel yielded a lower amount of blood loss and shorter time to hemostasis in various arterial and visceral bleeding models, which could be ascribed to the synergistic effect of wound closure under wet state as well as intrinsic hemostatic activity of CS. As a superior hemostatic sealant, the unique hydrogel proposed in this work can be exploited to offer significant advantages in the acute wound and massive hemorrhage with the restrictive access of therapeutic moieties.

[Investigation on occupational exposure to 5-fluorouracil in pharmacy intravenous admixture service of a hospital].

OBJECTIVE: To investigate the level of occupational exposure to 5-fluorouracil (5-Fu) in the pharmacy intravenous admixture service (PIVAS) of a hospital, and identify the sources of 5-Fu contamination. METHODS: The 5-Fu concentrations in air, on the surface of different areas in PIVAS and personal protective equipments were detected using UV-vis spectrophotometry. RESULTS: The 5-Fu in air could not be detected. The 5-Fu concentrations on five different surfaces of biological safety cabinets were (22.00 +/- 6.35), (13.99 +/- 2.46), (14.13 +/- 0.72), (7.25 +/- 1.19) and (9.87 +/- 1.23) ng/cm2, respectively, which were significantly higher than those [(3.14 +/- 0.04), (5.43 +/- 0.65), (2.26 +/- 0.17), (2.26 +/- 0.17) and (3.63 +/- 0.46) ng/cm2] of corresponding controls (P < 0.05 or P < 0.01). The 5-Fu concentrations of the floor under cabinets [(18.19 +/- 5.22) ng/cm2], the floor in front of cabinets [(10.25 +/- 2.57)ng/cm2], the office floor [(11.64 +/- 2.53) ng/cm2], the terrace floor [(99.89 +/- 14.06 ) ng/cm2], the floor beside trash can in dressing room [(24.54 +/- 0.23) ng/cm2] were significantly higher than those of control [(3.36 +/- 0.11 ) ng/cm2] (P < 0.05 or P < 0.01). The 5-Fu concentrations of the tables in preparation room [(7.22 +/- l.04) ng/cm2] and the tables in office [(11.81 +/- 1.18) ng/cm2] were significantly higher than those of control [(5.56 +/- 0.14) ng/cm2] (P < 0.05 or P < 0.01). The 5-Fu concentrations of the indoor handle in preparation room were significantly higher than those of controls (P < 0.05 or P < 0.01). 5-Fu concentrations on the surfaces of outdoor handle and floor beside door in preparation room were not significantly increased compared with controls (P > 0.05). The 5-Fu concentrations on the surfaces of infusion bags, transfer box, transfer trays were significantly higher than those of controls (P < 0.05). The differences of 5-Fu concentrations between outer and inner masks and controls were not significant (P > 0.05). The 5-Fu concentrations of gloves of preparing and checking staffs were significantly higher than those of controls (P < 0.05 or P < 0.01). CONCLUSION: The preparing and checking process of 5-Fu and the treatment of medical wastes are major sources of 5-Fu contamination.

Cartilage-binding antibodies initiate joint inflammation and promote chronic erosive arthritis.

BACKGROUND: Antibodies binding to cartilage proteins are present in the blood and synovial fluid of early rheumatoid arthritis patients. In order to develop animal models mimicking the human disease, we have characterized the arthritogenic capacity of monoclonal antibodies directed towards different joint proteins in the cartilage. METHODS: Purified antibodies specific to unmodified or citrullinated collagen type II (CII), collagen type XI (CXI), and cartilage oligomeric matrix protein (COMP) were produced as culture supernatant, affinity purified, pooled as antibody cocktails (Cab3 and Cab4), and injected intravenously into mice to induce arthritis. An adjuvant (lipopolysaccharide or mannan) was subsequently injected intraperitoneally on either day 5 or day 60 to enhance arthritis. Antibody binding and complement activation on the cartilage surface were analyzed by immunohistochemical methods. Bone erosions and joint deformations were analyzed by histological assessments, enzyme-linked immunosorbent assays, and micro-CT. Luminex was used to detect CII-triple helical epitope-specific antibody responses. RESULTS: The new cartilage antibody cocktails induced an earlier and more severe disease than anti-CII antibody cocktail. Many of the mouse strains used developed severe arthritis with 3 antibodies, binding to collagen II, collagen XI, and cartilage oligomeric matrix protein (the Cab3 cocktail). Two new models of arthritis including Cab3-induced LPS-enhanced arthritis (lpsCAIA) and Cab3-induced mannan-enhanced arthritis (mCAIA) were established, causing severe bone erosions and bone loss, as well as epitope spreading of the B cell response. Cab4, with addition of an antibody to citrullinated collagen II, induced arthritis more efficiently in moderately susceptible C57BL/6 J mice. CONCLUSIONS: The new mouse model for RA induced with cartilage antibodies allows studies of chronic development of arthritis and epitope spreading of the autoimmune response and bone erosion.

Selective Conversion of Hemicellulose in Macroalgae Enteromorpha prolifera to Rhamnose.

Direct hydrothermal conversion (HC) of macroalgae Enteromorpha prolifera was conducted over the temperature range of 140-240 °C. At 160 °C, monosaccharides and small molecular acids began to generate. A high yield (18.8%) of monosaccharides was obtained at 180 °C, whereas 29.6% of small molecular organic acids was attained at 200 °C. Formic acid (FA) was then employed as a catalyst, which could selectively catalyze the conversion of hemicellulose at low temperature (94.1%, 140 °C). Rhamnose (45.2%) based on the mass of carbohydrates in E. prolifera was produced by the catalysis of 0.7 mL of FA (160 °C, 60 min, 1 g of biomass loading). A low ratio of biomass amount to water was beneficial to the solution of water-soluble components of hemicellulose in E. prolifera to get high yields to monosaccharides. HC showed promise to be an applicable and efficient method in the treatment of E. prolifera with high conversion of carbohydrates.

A Shared Epitope of Collagen Type XI and Type II Is Recognized by Pathogenic Antibodies in Mice and Humans with Arthritis.

Background: Collagen XI (CXI) is a heterotrimeric molecule with triple helical structure in which the α3(XI) chain is identical to the α1(II) chain of collagen II (CII), but with extensive posttranslational modifications. CXI molecules are intermingled in the cartilage collagen fibers, which are mainly composed of CII. One of the alpha chains in CXI is shared with CII and contains the immunodominant T cell epitope, but it is unclear whether there are shared B cell epitopes as the antibodies tend to recognize the triple helical structures. Methods: Mice expressing the susceptible immune response gene Aq were immunized with CII or CXI. Serum antibody responses were measured, monoclonal antibodies were isolated and analyzed for specificity to CII, CXI, and triple helical collagen peptides using bead-based multiplex immunoassays, enzyme-linked immunosorbent assays, and Western blots. Arthritogenicity of the antibodies was investigated by passive transfer experiments. Results: Immunization with CII or CXI leads to a strong T and B cell response, including a cross-reactive response to both collagen types. Immunization with CII leads to severe arthritis in mice, with a response toward CXI at the chronic stage, whereas CXI immunization induces very mild arthritis only. A series of monoclonal antibodies to CXI were isolated and of these, the L10D9 antibody bound to both CXI and CII equally strong, with a specific binding for the D3 epitope region of α3(XI) or α1(II) chain. The L10D9 antibody binds cartilage in vivo and induced severe arthritis. In contrast, the L5F3 antibody only showed weak binding and L7D8 antibody has no binding to cartilage and did not induce arthritis. The arthritogenic L10D9 antibody bound to an epitope shared with CII, the triple helical D3 epitope. Antibody levels to the shared D3 epitope were elevated in the sera from mice with arthritis as well as in rheumatoid arthritis. Conclusion: CXI is immunologically not exposed in healthy cartilage but contains T and B cell epitopes cross-reactive with CII, which could be activated in both mouse and human arthritis and could evoke an arthritogenic response.

Anti-citrullinated protein antibodies cause arthritis by cross-reactivity to joint cartilage.

Today, it is known that autoimmune diseases start a long time before clinical symptoms appear. Anti-citrullinated protein antibodies (ACPAs) appear many years before the clinical onset of rheumatoid arthritis (RA). However, it is still unclear if and how ACPAs are arthritogenic. To better understand the molecular basis of pathogenicity of ACPAs, we investigated autoantibodies reactive against the C1 epitope of collagen type II (CII) and its citrullinated variants. We found that these antibodies are commonly occurring in RA. A mAb (ACC1) against citrullinated C1 was found to cross-react with several noncitrullinated epitopes on native CII, causing proteoglycan depletion of cartilage and severe arthritis in mice. Structural studies by X-ray crystallography showed that such recognition is governed by a shared structural motif "RG-TG" within all the epitopes, including electrostatic potential-controlled citrulline specificity. Overall, we have demonstrated a molecular mechanism that explains how ACPAs trigger arthritis.

Microwave-enhanced pyrolysis of natural algae from water blooms.

Microwave-enhanced pyrolysis (MEP) of natural algae under different reaction conditions was carried out. The optimal conditions for bio-oil production were the following: algae particle size of 20-5 mesh, microwave power of 600W, and 10% of activated carbon as microwave absorber and catalyst. The maximum liquid yield obtained under N2, 10% H2/Ar, and CO2 atmosphere was 49.1%, 51.7%, and 54.3% respectively. The energy yield of bio-products was 216.7%, 236.9% and 208.7% respectively. More long chain fatty acids were converted into hydrocarbons by hydrodeoxygenation under 10% H2/Ar atmosphere assisted by microwave over activated carbon containing small amounts of metals. Under CO2 atmosphere, carboxylic acids (66.6%) were the main products in bio-oil because the existence of CO2 vastly inhibited the decarboxylation. The MEP of algae was quick and efficient for bio-oil production, which provided a way to not only ameliorate the environment but also obtain fuel or chemicals at the same time.

One-pot synthesis of 5-hydroxymethylfurfural directly from starch over SO(4)(2-)/ZrO2-Al2O3 solid catalyst.

The synthesis of 5-hydroxymethylfurfural (HMF) directly from starch was studied in dimethyl sulfoxide-water. The effects of catalyst variation, reaction time, water content, catalyst loading and temperature on the reaction were investigated. The SO(4)(2-)/ZrO(2)-Al(2)O(3) catalyst was found to act as a bifunctional catalyst with high activity for both hydrolysis and dehydration of starch. HMF yield of 55% was obtained after 6h at 423K for the reaction of starch (the molar ratio of water to glucose in starch is 44/1) over the SO(4)(2-)/ZrO(2)-Al(2)O(3) catalyst, which bears high acidity and moderate basicity with Zr/Al molar ratio of 1:1.

Two-step hydrothermal conversion of Pubescens to obtain furans and phenol compounds separately.

The stepwise hydrothermal conversion of Pubescens selectively to furans and phenol compounds was investigated in a sealed autoclave reactor. The influence of reaction temperature and reaction time on the product distribution was studied. The results were compared to those from direct pyrolysis process. The residues from Pubescens via hydrothermal route and pyrolytic route were characterized by SEM and component analysis. A new method for obtaining furans and phenols separately through a two-step process was proposed. The first step was suggested performing at a moderate temperature for a shorter time to obtain liquid product with high furan content, while the second step was designed proceeding at a higher temperature for a comparatively longer time to produce more phenol compounds.

The direct pyrolysis and catalytic pyrolysis of Nannochloropsis sp. residue for renewable bio-oils.

Nannochloropsis sp. (a kind of green microalga) residue was pyrolyzed without catalyst or with different amount of HZSM-5 catalyst in a fixed bed reactor in nitrogen flow. The effects of pyrolysis parameters such as temperature and catalyst-to-material ratio on product yields were studied. The bio-oils obtained were analyzed by elemental, GC-MS and FTIR analysis. The results indicated that the bio-oils from catalytic pyrolysis of Nannochloropsis sp. residue (BOCP) had lower oxygen content (19.5 wt.%) and higher heating-value (32.7 MJ kg(-1)) than those obtained from direct pyrolysis (BODP) which had an oxygen content of 30.1 wt.% and heating-value of 24.6 MJ kg(-1). The BODP mainly consisted of long carbon chain compounds with various terminal groups (LCTG), while the BOCP mainly consisted of aromatic hydrocarbons. These properties of bio-oils demonstrated that the Nannochloropsis sp. residue can be used as a renewable energy resource and chemical feedstock.