Macrophages exhibit phenotypic diversity permitting wide-ranging roles in maintaining physiologic homeostasis. Hyaluronic acid, a major glycosaminoglycan of the extracellular matrix, has been shown to have differential signaling based on its molecular weight. With this in mind, the main objective of this study was to elucidate the role of hyaluronic acid molecular weight on macrophage activation and reprogramming. Changes in macrophage activation were assessed by activation state selective marker measurement, specifically quantitative real time polymerase chain reaction, and cytokine enzyme-linked immunoassays, after macrophage treatment with differing molecular weights of hyaluronic acid under four conditions: the resting state, concurrent with classical activation, and following inflammation involving either classically or alternatively activated macrophages. Regardless of initial polarization state, low molecular weight hyaluronic acid induced a classically activated-like state, confirmed by up-regulation of pro-inflammatory genes, including nos2, tnf, il12b, and cd80, and enhanced secretion of nitric oxide and TNF-α. High molecular weight hyaluronic acid promoted an alternatively activated-like state, confirmed by up regulation of pro-resolving gene transcription, including arg1, il10, and mrc1, and enhanced arginase activity. Overall, our observations suggest that macrophages undergo phenotypic changes dependent on molecular weight of hyaluronan that correspond to either (1) pro-inflammatory response for low molecular weight HA or (2) pro-resolving response for high molecular weight HA. These observations bring significant further understanding of the influence of extracellular matrix polymers, hyaluronic acid in particular, on regulating the inflammatory response of macrophages. This knowledge can be used to guide the design of HA-containing biomaterials to better utilize the natural response to HAs.

Cellular α-glucosidases I and II are enzymes that sequentially trim the three terminal glucoses in the N-linked oligosaccharides of viral envelope glycoproteins. This process is essential for the proper folding of viral glycoproteins and subsequent assembly of many enveloped viruses, including dengue virus (DENV). Imino sugars are substrate mimics of α-glucosidases I and II. In this report, we show that two oxygenated alkyl imino sugar derivatives, CM-9-78 and CM-10-18, are potent inhibitors of both α-glucosidases I and II in vitro and in treated animals, and efficiently inhibit DENV infection of cultured human cells. Pharmacokinetic studies reveal that both compounds are well tolerated at doses up to 100mg/kg in rats and have favorable pharmacokinetic properties and bioavailability in mice. Moreover, we showed that oral administration of either CM-9-78 or CM-10-18 reduces the peak viremia of DENV in mice. Interestingly, while treatment of DENV infected mice with ribavirin alone did not reduce the viremia, combination therapy of ribavirin with sub-effective dose of CM-10-18 demonstrated a significantly enhanced antiviral activity, as indicated by a profound reduction of the viremia. Our findings thus suggest that combination therapy of two broad-spectrum antiviral agents may provide a practically useful approach for the treatment of DENV infection.

Chemokine receptor-ligand interactions are important to support functioning of both normal and pathologic cells. The expression and function of chemokine receptors in epithelial ovarian carcinoma (EOC) is largely unknown. Here, we report that the lymphotactin receptor (XCR1) was expressed in primary and metastatic human epithelial ovarian carcinoma (EOC) specimens and cell lines. In contrast, expression of XCR1 was not detected in the normal ovary or in human normal ovarian surface epithelial cells. Our data indicate that XCL1 and XCL2 are either present in the malignant ascites or expressed by the ovarian carcinoma cells. The addition of lymphotactin (XCL1 and XCL2) stimulated migration and proliferation of XCR1-positive cells. Reduction of XCR1 expression in ovarian carcinoma cell line SKOV-3 resulted in abrogated diaphragm and peritoneal wall tumor formation and in reduced frequency of colonic, splenetic, and liver nodules in an in vivo xenograft mouse model. Taken together, our data suggest that XCR1 is expressed early during the course of tumorigenic transformation and contributes towards increased cell migration and proliferation, which can facilitate the prometastatic behavior of EOC cells.

Objective Few strategies to improve pain outcome in knee OA exist, in part because how best to evaluate pain over the long-term is unclear. Our objectives were: determine frequency of a good pain experience outcome based on previously formulated OA pain stages; and test the hypothesis that less depression and pain catastrophizing and greater self-efficacy and social support are each associated with greater likelihood of a good outcome. Methods Study participants all with knee OA reported pain stage at baseline and 2 years. Baseline assessments utilized the Geriatric Depression Scale, Pain Catastrophizing Scale, Arthritis Self-Efficacy Scale, and MOS Social Support Survey. Using pain experience stages, good outcome was defined b persistence in or movement to no pain or stage 1 (predictable pain, known trigger) at 2 years. A multivariable logistic regression model was developed to identify independent predictors of a good outcome. Results Of 212, 136 (64%) had a good pain outcome and 76 (36%) a poor outcome. In multivariable analysis, higher self-efficacy was associated with a significantly higher likelihood of good outcome (adjusted OR 1.14, 95% CI: 1.04–1.24); higher pain catastrophizing was associated with a significantly lower likelihood of good outcome (adjusted OR 0.88, 95% CI: 0.83–0.94). Conclusion This stage-based measure provides a meaningful and interpretable means to assess pain outcome in knee OA. The odds of a good 2-year outcome in knee OA were lower in persons with greater pain catastrophizing and higher in persons with greater self-efficacy. Targeting these factors may help to improve pain outcome in knee OA.

Macrophages are the initial biologic responders to biomaterials. These highly plastic immune sentinels control and modulate responses to materials, foreign or natural. The responses may vary from immune stimulatory to immune suppressive. Several parameters have been identified that influence macrophage response to biomaterials, specifically size, geometry, surface topography, hydrophobicity, surface chemistry, material mechanics, and protein adsorption. In this review, the influence of these parameters is supported with examples of both synthetic and naturally derived materials and illustrates that a combination of these parameters ultimately influences macrophage responses to the biomaterial. Having an understanding of these properties may lead to highly efficient design of biomaterials with desirable biologic response properties.

Glioblastoma multiforme (GBM) and other central nervous system (CNS) cancers have poor long-term prognosis, and there is a significant need for improved treatments. GBM initiation and progression are mediated, in part, by microRNA (miRNA), which are endogenous posttranscriptional gene regulators. Misregulation of miRNAs is a potential target for therapeutic intervention in GBM. In this work, a micelle-like nanoparticle delivery system based upon the block copolymer poly(ethylene glycol-b-lactide-b-arginine) was designed with and without a reducible linkage between the lactide and RNA-binding peptide, R15, to assess the ability of the micelle-like particles to disassemble. Using confocal live cell imaging, intracellular dissociation was pronounced for the reducible micelleplexes. This dissociation was also supported by higher efficiency in a dual luciferase assay specific for the miRNA of interest, miR-21. Notably, micelleplexes were found to have significantly better stability and higher anti-miRNA activity in cerebrospinal fluid than in human plasma, suggesting an advantage for applying micelleplexes to CNS diseases and in vivo CNS therapeutics. The reducible delivery system was determined to be a promising delivery platform for the treatment of CNS diseases with miRNA therapy.

Cell lytic peptides are a class of drugs that can be used to selectively kill invading organisms or diseased cells. Several of these peptides have been identified as potential therapeutics. Herein, we report a novel process for purifying recombinant melittin, a cell lytic peptide that inserts into the membranes of cells causing cell lysis, from Escherichia coli. The process involves surfactant and low pH to solubilize melittin fusion proteins from the insoluble fraction of bacterial lysates. We are able to significantly improve purity of the final product and confirm the activity of the peptide. The process yields recombinant melittin that is effective when used to treat U-87 MG glioma cells and inhibits growth of the Gram-positive pathogenic bacterium Streptococcus pyogenes. We demonstrate a method of repeated extraction of the insoluble protein fraction with mild detergent at a low pH that is able to generate a yield of pure, soluble melittin of approximately 0.5 to 1 mg/L of E. coli culture.

Hydrogels are finding increased clinical utility as advances continue to exploit their favorable material properties. Hydrogels can be adapted for many applications, including surface coatings and drug delivery. Anti-infectious surfaces and delivery systems that actively destroy invading organisms are alternative ways to exploit the favorable material properties offered by hydrogels. Sterilization techniques are commonly employed to ensure the materials are non-infectious upon placement, but sterilization is not absolute and infections are still expected. Natural, anti-bacterial proteins have been discovered which have the potential to act as anti-infectious agents; however, the proteins are toxic and need localized release to have therapeutic efficacy without toxicity. In these studies, we explore the use of the glutathione s-transferase (GST) to anchor the bactericidal peptide, melittin, to the surface of poly(ethylene glycol) diacrylate (PEGDA) hydrogel microspheres. We show that therapeutic levels of protein can be anchored to the surface of the microspheres using the GST anchor. We compared the therapeutic efficacy of recombinant melittin released from PEGDA microspheres to melittin. We found that, when released by an activating enzyme, thrombin, recombinant melittin efficiently inhibits growth of the pathogenic bacterium Streptococcus pyogenes as effectively as melittin created by solid phase peptide synthesis. We conclude that a GST protein anchor can be used to immobilize functional protein to PEGDA microspheres and the protein will remain immobilized under physiological conditions until the protein is enzymatically released.