Immunostimulatory Action Research Articles

RNA and a cell wall component of Enterococcus faecalis IC-1 are required for phagocytosis and interleukin 12 production by the mouse macrophage cell line J774.1

Enterococcus faecalis is a resident lactic acid bacterium in the human intestine. Its immunostimulatory action was reported to be enhanced by heat sterilization. To investigate its beneficial actions, we evaluated the ability of 10 E. faecalis strains to induce interleukin-12 (IL-12) production in a mouse macrophage cell line, J774.1 and found that the strain, E. faecalis IC-1, had a potent IL-12-inducing ability. Furthermore, we investigated the underlying mechanism by treating IC-1 cells with RNase or lysozyme. Its activity almost disappeared and an antagonist of Toll-like receptor (TLR) 7 inhibited this activity. Moreover, lysozyme-treated IC-1 bacteria were not phagocytized by J774.1 cells, and did not induce IL-12 production. Based on our results, we propose that macrophages recognize the cell wall components of IC-1, leading to phagocytosis. The IC-1 RNA is then recognized by TLR7, which induces the production of IL-12.

Recombinant Human IFNα-2b Response Promotes Vaginal Epithelial Cells Defense against Candida albicans.

Classical antifungal drugs have been subjected to restrictions due to drug toxicity, drug resistance, bioavailability, and detrimental drug interactions. Type I interferon (IFN) exerts direct distinct immunostimulatory or immunomodulatory actions; however, little is known regarding the anti-fungal reactions of vaginal epithelial cells (VECs) induced by the type I IFN response. Therefore, in the present study, we evaluated the cytotoxic activity, immunocompetent cytokine responses, and non-B IgG production of the VK2/E6E7 VEC line following recombinant human IFN α-2b (rhIFNα-2b) treatment in response to Candida albicans. When treated with rhIFNα-2b, the production of IL-2, IL-4, and IL-17 were significantly up-regulated compared to the infected control cells (P < 0.05). Our scanning electron microscopy results revealed that C. albicans can invade VECs by inducing both endocytosis and active penetration. RhIFNα-2b was able to transform the VECs into a thallus and stretched pattern, promoting the fusion of filopodia to form a lamellipodium and enhancing the mobility and the repair capacity of the VECs. In addition, rhIFNα-2b could effectively inhibit the adhesion, hyphal formation, and proliferation of C. albicans. Collectively, these responses restored the immune function of the infected VECs against C. albicans in vitro, providing a theoretical basis for this novel treatment strategy.

Imiquimod triggering varicella zoster reactivation in an immunocompetent patient

Imiquimod is widely used for the treatment of superficial basal cell carcinoma as well as several other dermatologic conditions. While its local immunostimulatory action is well proven, its potential for systemic immunomodulation has been less well described. We report the first case (to our knowledge) of varicella zoster reactivation caused by Imiquimod in an immunocompetent patient.

The identification of novel natural immunostimulatory compounds that limit bacterial infection.

Francisella tularensis is a highly infectious bacterium that causes the disease, tularemia. Tularemia can be transmitted to humans in many ways including: inhalation, biting insects, contact with infected animal tissue, and contaminated food and water. The Centers for Disease Control and Prevention classified F. tularensis as a category A bioterror agent due to its virulence and ease of aerosolization. The intentional release of a resistant strain of F. tularensis could be devastating. Consequently, there is a need for novel treatments effective against F. tularensis infections. We previously tested a cataloged natural compound library with THP-1 cells (a monocyte line) infected with an engineered, fluorescent F. tularensis (LVS/pTC3D) to look for inhibition of bacterial growth. The focus of current investigations is to determine whether the compounds that exhibited inhibition were acting as traditional antibiotics or as immunostimulants. Disk diffusion assays indicated which compounds inhibited growth of F. tularensis in the absence of immune cells meaning these extracts exhibited were acting as antibiotics. Compounds that did not show inhibition on the diffusion assays are being tested again in the THP-1 infection model to verify their immunostimulatory action. These compounds could potentially be used as novel antimicrobial treatments.

Spirulina elicits the activation of innate immunity and increases resistance against Vibrio alginolyticus in shrimp.

The effect of Spirulina dried powder (SDP) on the immune response of white shrimp Litopenaeus vannamei was studied invitro and invivo. Incubating shrimp haemocytes in 0.5mgml(-1) SDP caused the degranulation of haemocytes and a reduction in the percentage of large cells within 30min. Shrimp haemocytes incubated in 1mgml(-1) SDP significantly increased their phenoloxidase (PO) activity, serine proteinase activity, and respiratory burst activity (RB, release of superoxide anion). A recombinant protein of lipopolysaccharide and β-1,3-glucan binding protein (LGBP) of the white shrimp was produced, named rLvLGBP, and examined for its binding with SDP. An ELISA binding assay showed that rLvLGBP binds to SDP with a dissociation constant of 0.0507μM. In another experiment, shrimp fed diets containing SDP at 0 (control), 30, and 60gkg(-1) after four weeks were examined for LGBP transcript level and lysozyme activity, as well as phagocytic activity, clearance efficiency, and resistance to Vibrio alginolyticus. These parameters were significantly higher in shrimp receiving diets containing SDP at 60gkg(-1) or 30gkg(-1) than in controls. In conclusion, shrimp haemocytes receiving SDP provoked the activation of innate immunity as evidenced by the recognition and binding of LGBP, degranulation of haemocytes, reduction in the percentage of large cells, increases in PO activity, serine proteinase activity, superoxide anion levels, and up-regulated LGBP transcript levels. Shrimp receiving diets containing SDP had increased lysozyme activity and resistance against V. alginolyticus infection. This study showed the mechanism underlying the immunostimulatory action of Spirulina and its immune response in shrimp.

The TLR-specific adjuvants R-848 and CpG-B endorse the immunological reaction of neonatal antigen-presenting cells.

Preterm neonates display an impaired vaccine response. Neonatal antigen-presenting cells (APCs) are less effective to induce an adaptive immune response and to promote the development of immunological memory. Efficient adjuvantal toll-like receptor (TLR)-triggering may overcome the neonatal immunological impairment. Accordingly, the aim of this study was to investigate the immunostimulatory action of R-848 and CpG-B on neonatal APCs. Surface marker and cytokine secretion of APCs were evaluated after incubation of cord blood and peripheral blood mononuclear cells with the indicated adjuvants and were analyzed using flow cytometry. TLR-specific stimulation resulted in a significant induction of costimulatory molecules on neonatal APCs. Stimulation with R-848 resulted in significant higher secretion of TNFα, IL-6, IL-10, IL-12/IL-23p40, IL-12p70, and IFN-γ. Interestingly, CpG-B resulted in significant higher secretion of TNFα and IL-6. In summary, the incubation of TLR-agonists induced activation and maturation of neonatal APCs. These data show that modern TLR-specific adjuvants achieve a direct effect and potent upregulation of activation and maturation markers and cytokines in preterm neonates. We thus conclude that agents triggering TLRs might possibly overcome neonatal lack of vaccine responses.

Deletion of Nuclear Localizing Signal Attenuates Proinflammatory Activity of Prothymosin-Alpha and Enhances Its Neuroprotective Effect on Transient Ischemic Stroke.

Post-ischemic inflammation plays an important role in the progression of ischemia/reperfusion injuries. Prothymosin-α (ProT) can protect cells from necrotic death following ischemia; however, its immunostimulatory actions may counteract the neuroprotective effect. We proposed that ProTΔNLS, synthesized by deleting its nuclear localizing signal (NLS) at the C-terminal of ProT, can attenuate the immunostimulatory activity and has more salient neuroprotective effect. In this study, we examined the therapeutic effects of ProT and ProTΔNLS in a transient middle cerebral artery occlusion (tMCAO) model of rats. Rats that had sustained 90min of tMCAO were treated with GST-vehicle, ProT, or ProTΔNLS. Therapeutic outcomes were evaluated by infarction volume assay and behavioral assessment. Changes to inflammatory mediators, including tumor necrosis factor α (TNF-α), interleukin-10 (IL-10), and myeloperoxidase (MPO) were evaluated by enzyme-linked immunosorbent assay. Activated matrix metalloproteinases 2 (MMP-2) and 9 (MMP-9) levels were evaluated by gelatin zymography. Microglial activation was identified by double-immunostaining for Iba-1 and CD68. Our results showed that while both ProT and ProTΔNLS reduce infarction volume and improve functional outcome, ProTΔNLS provides the best therapeutic outcome. ProT increases TNF-α but decreases IL-10 secretion after ischemic injury, reflecting its pro-inflammatory activity. ProTΔNLS suppresses expression of TNF-α, MPO, and activity of MMPs in ischemic brain tissue. It also suppresses activation of microglia in penumbral cortex. These data demonstrate the immunesuppressive activities of ProTΔNLS. In conclusion, ProT has pro-inflammatory effect that may counteract its neuroprotective effect. Deletion of NLS from ProT may attenuate post-ischemic inflammation and enhance the neuroprotective effects of ProT.

Rotavirus Recombinant VP6 Nanotubes Act as an Immunomodulator and Delivery Vehicle for Norovirus Virus-Like Particles.

We have recently shown that tubular form of rotavirus (RV) recombinant VP6 protein has an in vivo adjuvant effect on the immunogenicity of norovirus (NoV) virus-like particle (VLP) vaccine candidate. In here, we investigated in vitro effect of VP6 on antigen presenting cell (APC) activation and maturation and whether VP6 facilitates NoV VLP uptake by these APCs. Mouse macrophage cell line RAW 264.7 and dendritic cell line JAWSII were used as model APCs. Internalization of VP6, cell surface expression of CD40, CD80, CD86, and major histocompatibility class II molecules, and cytokine and chemokine production were analyzed. VP6 nanotubes were efficiently internalized by APCs. VP6 upregulated the expression of cell surface activation and maturation molecules and induced secretion of several proinflammatory cytokines and chemokines. The mechanism of VP6 action was shown to be partially dependent on lipid raft-mediated endocytic pathway as shown by methyl-β-cyclodextrin inhibition on tumor necrosis factor α secretion. These findings add to the understanding of mechanism by which VP6 exerts its immunostimulatory and immunomodulatory actions and further support its use as a part of nonlive RV-NoV combination vaccine.

Immunostimulatory activity of glycopeptides from Paecilomyces sinensis under normal and cyclophosphamide induced immunosuppressive conditions in mice models.

The present study was designed to evaluate immune-modulating effects of the glycopeptide from Paecilomyces sinensis (CPS-II) by using mouse peritoneal macrophage and cytoxan (CTX) induced immunosuppression models. Our results from phagocytotic and mononuclear phagocytic system function assays showed that CPS-II stimulated phagocytosis of the phagocytes. A splenocyte proliferation assay showed that CPS-II acted to combine Concanavalin A (ConA) or lipopolysaccharides (LPS) in splenocyte proliferation. The results demonstrated that CPS-II increased the indices of the thymus and spleen. Hematological and histopathological analysis revealed the protective effect of CPS-II against CTX induced immunosuppression. CPS-II also significantly increased the expression of CD4(+) and CD8(+) splenic T lymphocytes, which were suppressed by CTX in peripheral blood. The expressions of serum cytokines related to immune function, including TNF-α, IL-6, and IFN-γ, were up-regulated in a dose-dependent manner. The expression of the transcription factor NF-κB in the spleen was enhanced after CPS-II-treatment. In conclusion, our results indicated that CPS-II was involved in immunostimulatory actions leading to its modulatory effects on immunosuppression, and one possible mechanism of action was to activate NF-κB.

Therapeutic Potential of Interferon-γ and Its Antagonists in Autoinflammation: Lessons from Murine Models of Systemic Juvenile Idiopathic Arthritis and Macrophage Activation Syndrome.

Interferon-γ (IFN-γ) affects immune responses in a complex fashion. Its immunostimulatory actions, such as macrophage activation and induction of T helper 1-type responsiveness, are widely acknowledged, however, as documented by a large body of literature, IFN-γ has also the potential to temper inflammatory processes via other pathways. In autoimmune and autoinflammatory disorders, IFN-γ can either play a disease-enforcing role or act as protective agent, depending on the nature of the disease. In animal models of any particular autoimmune disease, certain changes in the induction procedure can reverse the net outcome of introduction or ablation of IFN-γ. Here, we review the role of endogenous IFN-γ in inflammatory disorders and related murine models, with a focus on systemic juvenile idiopathic arthritis (sJIA) and macrophage activation syndrome (MAS). In particular, we discuss our recent findings in a mouse model of sJIA, in which endogenous IFN-γ acts as a regulatory agent, and compare with results from mouse models of MAS. Also, we elaborate on the complexity in the activity of IFN-γ and the resulting difficulty of predicting its value or that of its antagonists as treatment option.

Extraction process optimization of sulfated galactan-rich fractions from Hypnea musciformis in order to obtain antioxidant, anticoagulant, or immunomodulatory polysaccharides

The seaweed Hypnea musciformis synthesizes sulfated galactans and these galactans have been reported to have antioxidant, anticoagulant, and immunostimulatory properties. However, efficient extraction of these polysaccharides depends on the particular extraction method used, some of these polysaccharides are not extracted. Previously, we obtained galactans with different properties by using a simple extraction method. Here, we performed slight modifications to the extraction method, which led to variation in the type of polysaccharides obtained and their biological activity. We obtained four sulfated galactan-rich fractions named as FT4v, FT6v, FT8v, and FT10v. The chemical composition data showed that the fractions had high sugar and sulfate contents. Fraction FT4v exhibited the highest sulfate/sugar ratio (0.47), while FT6v (0.37), FT8v (0.39), and FT10v (0.39) presented lower sulfate/sugar ratios. Fractions FT4v and FT6v showed the highest antioxidant activity. Fraction FT4v also exhibited the greatest anticoagulant potential; however, other fractions presented marked immunomodulatory action. FT10v stimulated the highest levels of NO production whereas FT6v stimulated the highest expression of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) in RAW cells. Fraction FT10v exhibited the greatest cytotoxic action of the obtained fractions against HeLa and 786 cell lines. In vitro tests showed that the sulfated galactan-rich fractions isolated from H. musciformis with low sulfate/sugar ratios, i.e., the fractions containing fewer sulfated polysaccharides (FT6v, FT8v, and FT10v), showed marked immunostimulatory action while the fraction with the high sulfate/sugar ratio (FT4v) exhibited improved anticoagulant activity. Therefore, our results suggest that different extraction conditions promote the acquisition of sulfated galactan-rich fractions from H. musciformis with different biological activities.

Structural Basis for Polymer Packing and Solvation Properties of the Organogermanium Crystalline Polymer Propagermanium and Its Derivatives

Of organogermanium compounds known to have an immunostimulatory action, propagermanium [PGe; 3-oxygermylpropionic acid polymer, (C3 H5 GeO3.5 )n] is the only one used as a pharmaceutical agent, to treat the hepatitis B virus in Japan. However, because of lack of information about its structure, PGe has been confused with a polymeric solid, repagermanium (RGe, Ge-132, poly-trans-[(2-carboxyethyl) germasesquioxane], (C18 H30 Ge6 O21 )n), which has the same essential formula as PGe. To clarify this issue, the structure of PGe was analyzed using X-ray diffraction (XRD). PGe has a polymeric ladder-shaped structure of a concatenated eight-membered ring composed of Ge-O bonds, which is clearly distinguished from the infinite sheet structure in RGe. Moreover, we observed temperature or moisture-dependent transformations among these compounds using powder XRD. For instance, PGe was easily dissolved in water, and transformed to RGe by exposure to water vapor, but transformed into another straight-chain structure when exposed to aqueous solution. As a result of these findings, PGe was indicated to have labile polymer packing against RGe. These characteristics of PGe may affect pharmaceutical properties such as respective stability and solubility, which indicate its unique impact on physiological activity.

In vivo administration of the frog skin peptide frenatin 2.1S induces immunostimulatory phenotypes of mouse mononuclear cells

Host-defense peptides secreted by epithelial cells exhibit cytotoxic and immunoregulatory effects in order to protect the organism against invading microorganisms. Antimicrobial peptides derived from frog skin display both immunostimulatory and immunosuppressive actions as demonstrated by in vitro cytokine production by macrophages. Frenatin 2.1S, first isolated from skin secretions of the frog, Sphaenorhynchus lacteus (Hylidae), enhances the in vitro production of pro-inflammatory IL-1β, TNF-α and IL-23 by mouse peritoneal cells. In order to test whether the immunostimulatory action of frenatin 2.1S may be reproduced in vivo, effects of intraperitoneal injections of this peptide on mononuclear cells in the peritoneum and spleen were determined 24h after administration. The data indicate that frenatin 2.1S enhances the activation state and homing capacity of Th1 type lymphocytes and NKT cells in the mouse peritoneal cavity, as evaluated by increased expression of early activation marker CD69 among T and NKT cells and chemokine receptor CXCR3 among T cells. Frenatin 2.1S significantly increases the percentage of (F4/80+CD11c+CD206+) pro-inflammatory M1 macrophages and enhances the expression of MHC class II molecules on F4/80+CD11c+ macrophages in the mouse peritoneal cavity. Additionally, injection of frenatin 2.1S, in the presence or absence of lipopolysaccharide, increases the percentage of peritoneal B cells of the (CD19+CD11b+CD5+) B1a phenotype thus contributing to an inflammatory milieu. We suggest that the immunostimulatory effect of frenatin 2.1S may have therapeutic relevance in disease states, such as certain types of cancer, in which an enhanced inflammatory response may be beneficial.

Parenteral immunization of PLA/PLGA nanoparticle encapsulating outer membrane protein (Omp) from Aeromonas hydrophila: Evaluation of immunostimulatory action in Labeo rohita (rohu)

Advanced vaccine research approaches needs to explore on biodegradable nanoparticles (NPs) based vaccine carrier that can serve as antigen delivery systems as well as immuno-stimulatory action to induce both innate and adaptive immune response in fish. Immunogenicity of PLA and PLGA NPs encapsulating outer membrane protein (Omp) antigen of Aeromonas hydrophila were evaluated through intra-peritoneal injection in fish, Labeo rohita. Antigen loaded PLA-Omp (223.5 ± 13.19 nm) and PLGA-Omp (166.4 ± 21.23 nm) NPs were prepared using double emulsion method by efficiently encapsulating the antigen reaching the encapsulation efficiency 44 ± 4.58% and 59.33 ± 5.13% respectively. Our formulated PLA Omp and PLGA-Omp NPs were in nanometer range (<500 nm) and could be successfully endocyted in the body. Despite low antigen loading in PLA-Omp, it showed considerably slower antigen release in vitro than PLGA-Omp NPs. Other physical properties like zetapotential values and poly dispersity index (PDI) confirmed the stability as well as monodisperse nature of the formulated nanoparticles. The spherical and isolated nature of PLA-Omp and PLGA-Omp NPs were revealed by SEM analysis. Upon immunization of all antigenic formulations (PLA-Omp NP, PLGA-Omp NP, FIA-Omp, PLA NP, PLGA NP, PBS as control), significant higher bacterial agglutination titre and haemolytic activity were observed in case of PLA-Omp and PLGA-Omp immunized groups than rest groups at both 21 days and 42 days. The specific antibody response was significantly increased and persisted up to 42 days of post immunization by PLA-Omp, PLGA-Omp, FIA-Omp. PLA-Omp NPs showed better immune response (higher bacterial agglutination titre, haemolytic activity, specific antibody titre, higher percent survival upon A. hydrophila challenge) than PLGA-Omp in L. rohita confirming its better efficacy. Comparable antibody response of PLA-Omp and PLGA-Omp with FIA-Omp treated groups suggested that PLA and PLGA could be replacement for Freund's adjuvant (for stimulating antibody response) to overcome many side effects offering long lasting immunity. Our encouraging results suggest that PLA/PLGA nanoparticles based delivery system could be a novel antigen carrier for parenteral immunization in fish.

Anti-Inflammatory and Immunostimulatory Effects of Extract from Culture of Effective Microorganisms (ECEM) Revealed by Functional Genomics and Metabolome Analyses

The extract from culture of effective microorganisms (ECEM) is a secondary metabolite produced in symbiotic culture of a mixture of yeasts and lactic bacteria followed by photosynthetic bacteria. Metabolomics studies have demonstrated that ECEM contained several active ingredients associated with anti-inflammation through interactions with nuclear factor-κB. Functional genomics using DNA microarrays revealed that ECEM had distinctive anti-inflammatory and immunostimulatory actions in RAW264.7 cells. ECEM inhibited the LPS-upregulated expression of one group of pro-inflammatory genes (iNos, Il6 and Cox2) but not another group genes (Tnf, Ccl7 and Dusp2); it also upregulated the expression of Cd1d1, which participates in antigen presentation on lipids, resulting in activation of natural killer T (NKT) cells related to infection defense and cancer. Bacterial glycosylated lipid and other related complex lipids in ECEM are thought to trigger this upregulation of Cd1d1 expression. These results suggest that ECEM functions in the immune response in both inflammation and antigen presentation and that it contains components important for these functions. These novel effects of ECEM could be invaluable for developing functional drinks or health beverages.

Immunomodulatory effects of Stachytarpheta cayennensis leaf extract and its synergistic effect with artesunate.

BackgroundThe leaves of Stachytarpheta cayennensis C. Rich. (Verbenaceae) have been reported to possess potent anti-inflammatory and anti-malarial activities. Due to close association between inflammatory and immune responses, we evaluated the immunomodulatory activity of leaves extract of S. cayennensis. The combined effects of the leaves extract and artesunate, a standard antimalarial agent with immunomodulatory effects, were also evaluated.MethodsThe immunomodulatory activity of the methanol extract of the leaves of S. cayennensis (MESC) was evaluated in mice using the Delayed-Type hypersensitivity response (DTHR), primary and secondary humoral immune responses and the in vivo leucocyte mobilization tests. The immunomodulatory effect of artesunate and the combined effects of MESC and artesunate were evaluated using the phagocytic activity of polymorphonuclear neutrophils. Acute toxicity and lethality test in addition to the preliminary phytochemical studies of MESC were also performed.ResultsThe MESC exhibited 64.21% inhibition of DTHR at 500 mg/kg dose and evoked 139.64% of phagocytic stimulation at 100 μg/ml dose. Also MESC significantly (p < 0.05) showed dose related stimulation of humoral immunity and a highest percentage leucocyte mobilization of 10.15% at 250 mg/kg dose. Artesunate offered a non-significant (p > 0.05) percentage phagocytic stimulation (PPS) while the combined effect of artesunate and MESC exhibited a significant (p < 0.05) dose dependent PPS with highest PPS of 393.77% at 100 μg/ml. The LD50 of the MESC was estimated to be greater than 5000 mg/kg since there were no lethality and signs of acute intoxication after 48 h observation. Preliminary phytochemical analysis revealed the presence of carbohydrates, glycosides, flavonoids, saponins, alkaloids, terpenoids and steroids.ConclusionsThe results of this study indicated that MESC possesses immunostimulatory action with significant synergistic effects with artesunate, and can therefore, offer immune boosting activities in disorders of immune suppression.

Melatonin and atopy: role in atopic dermatitis and asthma.

Melatonin may have important immunostimulatory actions in allergic diseases, in addition to its well-known antioxidant and cytoprotective effects in several inflammatory conditions. The activation of the immune system leads to free radical production associated with decreased melatonin levels and depressed antioxidant enzyme activities in several inflammatory diseases. Many skin disorders, including atopic dermatitis, are accompanied by infiltration and activation of mast cells, which release vasoactive and proinflammatory mediators. Experimental data suggest that melatonin inhibits development of atopic eczema and reduces serum total IgE and IL-4. Allergic asthma is a condition characterized by bronchial hyperresponsiveness and the presence of IgE antibodies in response to inhaled allergens; often there is also enhanced total serum IgE levels. Melatonin regulates smooth muscle tone and influences the immune response. Melatonin may, however, act as a pro-inflammatory agent in asthma leading to bronchial constriction. The safety of melatonin as a sleep-inducing agent has been confirmed in asthmatic subjects, but its routine use is not recommended in bronchial asthma. This review summarizes what is known about the role of melatonin as an immunomodulatory agent in asthma and atopic eczema.

Isolation of Mare's Milk Oligosaccharide Fraction of Colostrum, Transitional, and Mature Phases Promotes In Vitro Oxidative Burst in Murine Macrophages

Present preliminary findings demonstrate in vitro stimulation of splenic lymphocytes of BALB/c mice after exposure to oligosaccharide fractions isolated from mare's milk during colostrum, transitional, and mature lactation periods, which shows immunostimulatory action of oligosaccharide content present in mares' milk. Oligosaccharide fractions caused more pronounced oxidative burst at a much lower concentration (0.1 μg/mL) compared with that shown by levamisole (standard immunostimulant) at a much higher concentration of 10 μg/mL. In this study, it was observed that oligosaccharides fraction of colostrum phase attaining high potency of immunostimulatory property.

Rapid systemic and local treatments with the antibacterial peptide dimer A3-APO and its monomeric metabolite eliminate bacteria and reduce inflammation in intradermal lesions infected with Propionibacterium acnes and meticillin-resistant Staphylococcus aureus

When administered intramuscularly, the designer antibacterial peptide dimer A3-APO is highly efficacious in mouse models of Acinetobacter baumannii and Staphylococcus aureus burn infections. Here we compared the efficacy of A3-APO and its monomeric metabolite in mouse models of S. aureus and Propionibacterium acnes intradermal infections following administration as intramuscular (i.m.) or topical treatments. In the animal models, either (i) the ears of CD-1 mice were infected with P. acnes or (ii) S. aureus was injected into burn wounds inflicted to the back. A3-APO or the monomer were injected intramuscularly at 5mg/kg one to three times or were applied three times as 1% local treatment in phosphate-buffered saline or Vaseline®. Despite being inactive against the strains in vitro, in vivo the skin conditions of the mice were dramatically improved upon peptide treatment regardless of dosing frequency, administration mode or drug valency. In the P. acnes study, A3-APO statistically significantly reduced ear thickness and ear bacterial counts. The amount of ear connective tissue and epithelial macrophages correlated with therapeutic success. Bacterial load in the lesions was more representative of physical improvement than ear dimensions. In the S. aureus model, both peptides eliminated wound bacteria from >107 CFU/mg to almost background levels, with monomer treatment being somewhat more successful. In conclusion, A3-APO and its monomeric metabolite very efficiently ameliorate resistant aerobic and anaerobic intradermal infections, but the protection is apparently not due to direct bacterial killing. Immunostimulatory and anti-inflammatory actions are likely involved. Nevertheless, topical and i.m. administrations are equally effective.

Antioxidant potential of all-trans retinoic acid (ATRA) and enhanced activity of liposome encapsulated ATRA against inflammation and tumor-directed angiogenesis

The purpose of this study was to investigate whether all-trans retinoic acid (ATRA) has antioxidant property. The study was also focused on its inhibitory effect on the acute and chronic inflammation and tumor-associated capillary formation in terms of angiogenesis in C57BL/6 mice after incorporated in liposome composed of distearoylphosphatidylcholine (DSPC/cholesterol). ATRA possesses a number of important biologic activities including oncostatic, antioxidant and immunostimulatory actions. Our study was designed to evaluate the antioxidant activity of free ATRA by nitric oxide scavenging, superoxide radical scavenging, hydroxyl radical scavenging and lipid peroxide scavenging assays. The ATRA showed significant scavenging activities in all these antioxidant assays comparable to the standard antioxidant. We have also evaluated the activity of encapsulated ATRA against anti-inflammatory activity in C57BL/6 mice. The paw oedema inhibition was found in carrageenan model as 55.56% and 66.67% for free ATRA and encapsulated ATRA treatment respectively and for formaldehyde model it was found to be 60.87% and 69.57% respectively compared with saline treated control mice. Encapsulated ATRA inhibited the tumor-associated capillary formation in mice induced by highly metastatic B16F10 melanoma cells significantly than the free ATRA did. In this study the inhibition of tumor-directed capillary formation was found to be 56.25% and 62.50% for free ATRA and encapsulated ATRA treatment respectively. In conclusion, ATRA showed a significant antioxidant property in vitro. Free ATRA has anti-inflammatory activity as proved by us in animal model of acute and chronic inflammation and antiangiogenesis activity. Furthermore, its activity was boosted by encapsulation in liposome.