Maoto, a traditional Japanese medicine, controls acute systemic inflammation induced by polyI:C administration through noradrenergic function.

Maoto, a traditional Japanese medicine (Kampo), is widely used to treat upper respiratory tract infections, including influenza virus infection. Although maoto is known to inhibit pro-inflammatory responses in a rodent model of acute inflammation, its underlying mechanism remains to be determined. In this study, we investigated the involvement of immune responses and noradrenergic function in the inhibitory action of maoto. In a mouse model of polyI:C-induced acute inflammation, maoto was administered orally in conjunction with intraperitoneal injection of PolyI:C (6 mg/kg), and blood was collected after 2 h for measurement of plasma cytokines by ELISA. Maoto significantly decreased PolyI:C-induced TNF-α levels and increased IL-10 production. Neither pretreatment with IL-10 neutralizing antibodies nor T-cell deficiency using nude mice modified the inhibitory effect of maoto, indicating that the anti-inflammatory effects of maoto are independent of IL-10 and T cells. Furthermore, the inhibitory effects of maoto on PolyI:C-induced TNF-α production were not observed in ex vivo splenocytes, suggesting that maoto does not act directly on inflammatory cells. Lastly, pretreatment with a ß-adrenergic receptor antagonist partially cancelled the anti-inflammatory effects of maoto. Collectively, these results suggest that maoto mediates its anti-inflammatory effects via ß-adrenergic receptors in vivo.

Thalidomide alleviates neuropathic pain through microglial IL-10/ß-endorphin signaling pathway.

Thalidomide is an antiinflammatory, antiangiogenic and immunomodulatory agent which has been used for the treatment of erythema nodosum leprosum and multiple myeloma. It has also been employed in treating complex regional pain syndromes. The current study aimed to reveal the molecular mechanisms underlying thalidomide-induced pain antihypersensitive effects in neuropathic pain. Thalidomide gavage, but not its more potent analogs lenalidomide and pomalidomide, inhibited mechanical allodynia and thermal hyperalgesia in neuropathic pain rats induced by tight ligation of spinal nerves, with ED50 values of 44.9 and 23.5 mg/kg, and Emax values of 74% and 84% MPE respectively. Intrathecal injection of thalidomide also inhibited mechanical allodynia and thermal hyperalgesia in neuropathic pain. Treatment with thalidomide, lenalidomide and pomalidomide reduced peripheral nerve injury-induced proinflammatory cytokines (TNFα, IL-1ß and IL-6) in the ipsilateral spinal cords of neuropathic rats and LPS-treated primary microglial cells. In contrast, treatment with thalidomide, but not lenalidomide or pomalidomide, stimulated spinal expressions of IL-10 and ß-endorphin in neuropathic rats. Particularly, thalidomide specifically stimulated IL-10 and ß-endorphin expressions in microglia but not astrocytes or neurons. Furthermore, pretreatment with the IL-10 antibody blocked upregulation of ß-endorphin in neuropathic rats and cultured microglial cells, whereas it did not restore thalidomide-induced downregulation of proinflammatory cytokine expression. Importantly, pretreatment with intrathecal injection of the microglial metabolic inhibitor minocycline, IL-10 antibody, ß-endorphin antiserum, and preferred or selective µ-opioid receptor antagonist naloxone or CTAP entirely blocked thalidomide gavage-induced mechanical antiallodynia. Our results demonstrate that thalidomide, but not lenalidomide or pomalidomide, alleviates neuropathic pain, which is mediated by upregulation of spinal microglial IL-10/ß-endorphin expression, rather than downregulation of TNFα expression.

Cynandione A and PHA-543613 inhibit inflammation and stimulate macrophageal IL-10 expression following α7 nAChR activation.

Cynandione A, an acetophenone isolated from Cynanchum Wilfordii Radix, attenuates inflammation. The present study aimed to study the mechanisms underlying cynandione A-induced antiinflammation. Treatment with cynandione A and the specific α7 nicotinic acetylcholine receptor (α7 nAChR) agonist PHA-543613 remarkably reduced overexpression of proinflammatory cytokines, including tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1ß in lipopolysaccharide (LPS)-treated RAW264.7 cells and primary peritoneal macrophages, and endotoxemic mice. Both cynandione A and PHA-543613 also stimulated IL-10 expression in naïve and LPS-treated macrophages and endotoxemic mice. Cynandione A- and PHA-543613-inhibited proinflammatory cytokine expression was completely blocked by the α7 nAChR antagonist methyllycaconitine and the IL-10 antibody. The stimulatory effect of cynandione A and PHA-543613 on IL-10 expression were suppressed by methyllycaconitine and knockdown of α7 nAChRs using siRNA/α7 nAChR. Cynandione A significantly stimulated STAT3 phosphorylation, which was attenuated by methyllycaconitine and the IL-10 neutralizing antibody. The STAT3 activation inhibitor NSC74859 also blocked cynandione A-inhibited proinflammatory cytokine expression. Taken together, our results, for the first time, demonstrate that cynandione A and PHA-543613 inhibit inflammation through macrophageal α7 nAChR activation and subsequent IL-10 expression.

Anti-rheumatic activity of pseudoephedrine (a substituted phenethylamine) in complete Freund's adjuvant-induced arthritic rats by down regulating IL-1ß, IL-6 and TNF-α as well as upregulating IL-4 and IL-10.

Pseudoephedrine (substituted phenethylamine) is well known as psychotic and bronchodilator. Numerous studies on phenethylamine derivatives indicated that these agents have the potential to abolish inflammatory responses in the non-biological and biological systems. These facts provided the basis to conduct a study on pseudoephedrine to explore its therapeutics in Complete Freund's Adjuvant (CFA)-induced arthritis. Furthermore, existing treatment approaches for RA associated with limited effect on chronic immunological models. Real-time polymerase chain reaction (q-PCR) was performed to execute the expression of pro and anti-inflammatory cytokines in treated and non-treated arthritic rats. These findings were further co investigate by histological observations. The paw volume, paw diameter, weight variations and arthritic score were determined at specific days throughout the experiment of 28 days. Pseudoephedrine at all doses significantly (p < 0.001) suppressed the expression of PGE2, TNF-α, IL-1ß and IL-6. Moreover, pseudoephedrine (20 and 40 mg/kg) caused significant augmentation of IL-4 and IL-10. Similarly, the drug expressed a significant anti-arthritic effect by reducing the paw volume, paw diameter and arthritic score. Similarly, it also reverts the reduction in body weight of arthritic rats at all above-mentioned doses. These findings supported the anti-arthritic potential of pseudoephedrine and recommended it for clinical trials.

Structure-based decoupling of the pro- and anti-inflammatory functions of interleukin-10.

Interleukin-10 (IL-10) is an immunoregulatory cytokine with both anti-inflammatory and immunostimulatory properties and is frequently dysregulated in disease. We used a structure-based approach to deconvolute IL-10 pleiotropy by determining the structure of the IL-10 receptor (IL-10R) complex by cryo-electron microscopy at a resolution of 3.5 angstroms. The hexameric structure shows how IL-10 and IL-10Rα form a composite surface to engage the shared signaling receptor IL-10Rß, enabling the design of partial agonists. IL-10 variants with a range of IL-10Rß binding strengths uncovered substantial differences in response thresholds across immune cell populations, providing a means of manipulating IL-10 cell type selectivity. Some variants displayed myeloid-biased activity by suppressing macrophage activation without stimulating inflammatory CD8+ T cells, thereby uncoupling the major opposing functions of IL-10. These results provide a mechanistic blueprint for tuning the pleiotropic actions of IL-10.

Blocking IL-10 signaling with soluble IL-10 receptor restores in vitro specific lymphoproliferative response in dogs with leishmaniasis caused by Leishmania infantum.

rIL-10 plays a major role in restricting exaggerated inflammatory and immune responses, thus preventing tissue damage. However, the restriction of inflammatory and immune responses by IL-10 can also favor the development and/or persistence of chronic infections or neoplasms. Dogs that succumb to canine leishmaniasis (CanL) caused by L. infantum develop exhaustion of T lymphocytes and are unable to mount appropriate cellular immune responses to control the infection. These animals fail to mount specific lymphoproliferative responses and produce interferon gamma and TNF-alpha that would activate macrophages and promote destruction of intracellular parasites. Blocking IL-10 signaling may contribute to the treatment of CanL. In order to obtain a tool for this blockage, the present work endeavored to identify the canine casIL-10R1 amino acid sequence, generate a recombinant baculovirus chromosome encoding this molecule, which was expressed in insect cells and subsequently purified to obtain rcasIL-10R1. In addition, rcasIL-10R1 was able to bind to homologous IL-10 and block IL-10 signaling pathway, as well as to promote lymphoproliferation in dogs with leishmaniasis caused by L. infantum.

Combined Administration of TLR4 (LPS) and TLR3 (Poly I:C) Ligands to CBA Mice Elevates the Content of Osteogenic MSC by 1.6 Times and Increases Content of Bone Marrow MSC to Intermediate Level between Values Attained by Their Individual Administration.

In 1 and 24 h after combined administration of TLR4 (LPS) and TLR3 (Poly I:C) ligands to CBA mice, the content of MSC in bone marrow increased to intermediate value between the levels attained by their individual injections. The content of osteogenic MSC assessed in 24 h postinjection corresponded to the control level in Poly I:C group, decreased in LPS group by 2.5 times relatively to the control, and increased by 1.6 times (relatively to control) after combined administration of the ligands. In 3 h after combined addition of LPS and Poly I:C in vitro to 12-day-old primary culture of mouse bone marrow stromal cells, the concentration of TNFα in culture medium was intermediate between the levels attained by their individual application. The data revealed dependence of activation of stromal tissue on intensity of innate immunity reactions; they also attested to marked elevation of osteogenicity of MSC pool after costimulation with Poly I:C and LPS, which can underlie augmented calcification of the tissues during combined viral and bacterial infections.

The role of B regulatory (B10) cells in inflammatory disorders and their potential as therapeutic targets.

Over the past decade, studies have identified subset of B cells, which play suppressive functions in additions to the conventional functions of B cells: antigen processing and presentation, activation of T cells and antibody productions. Because of their regulatory function, they were named as B regulatory cells (Bregs). Bregs restrict the severity of autoimmune disorders in animal disease models such as experimental autoimmune myocarditis (EAM), experimental autoimmune encephalitis (EAE), and collagen-induced arthritis (CIA) but can contribute to the development of infection and cancer. In humans, the roles of B regulatory cells in autoimmune diseases have not been clearly established because of the inconsistent findings from many researchers. This is believed to arise from the speculated fact that Bregs lack specific marker, which can be used to identify and characterize them in human diseases. The CD19+CD24hiCD38hiCD1dhiB cells have been associated with the regulatory function. Available evidences highlight the relevance of increasing IL-10-producing B cells in autoimmune diseases and the possibility of serving as new therapeutic targets in inflammatory disorders. This review empanels the functions of Bregs in autoimmune diseases in both human and animal models, and further evaluates the possibility of Bregs as therapeutic targets in inflammatory disorders. Consequently, this might help identify possible research gaps, which need to be clarified as researchers speculate the possibility of targeting some subsets of Bregs in the treatment of inflammatory disorders.

Immune regulation and cytotoxic T cell activation of IL-10 agonists - Preclinical and clinical experience.

The expansion and activation of tumor antigen reactive CD8+ T cells are primary goals of immunotherapies for cancer. IL-10 is an anti-inflammatory cytokine with an essential role in the development and proliferation of regulatory T cells, restricting myeloid and chronic inflammatory T cell responses. However, IL-10 is also essential for the expansion of antigen activated, tumor specific CD8+ T cells, leading to spontaneous tumor development in IL-10 deficient patients and mice. IL-10 induces IFNγ and cytotoxic mediators in antigen activated T cells. In clinical trials, monotherapy with recombinant, pegylated IL-10 (Pegilodecakin) induced objective responses in cancer patients. Patients receiving pegilodecakin had a systemic increase of IFNγ and granzymes, proliferation and expansion of immune checkpoint positive CD8+ T cells. Combination of pegilodecakin with anti-PD-1 appeared to improve on the efficacy of the single agents.

Real-time monitoring of IL-6 and IL-10 reporter expression for anti-inflammation activity in live RAW 264.7 cells.

In previous study, we suggested that the interleukin (IL)-6 and IL-10 could serve as a good biomarker for anti-inflammation that related to chronic inflammatory disease. Recently, we are finding new anti-inflammation compounds from natural products by screening of IL-6 and IL-10 levels. Although, we could measure IL-6 and IL-10 levels by several methods. However, all methods could not measure continuous kinetic of IL-6 and IL-10 levels. Most methods have multiple steps and take a long time. Therefore, there is no a suitable method for screening. To this end, we established IL-6 and IL-10 promoter assay which can monitor with reference gene as Glyceraldehyde 3-phosphate dehydrogenase (gapdh) promoter in living single cell. It could determine IL-6 and IL-10 levels continuously in real-time within two steps. We evaluated IL-6 and IL-10 reporter expression in LPS-induced RAW 264.7 cells with well-known anti-inflammatory compounds such as quercetin, xanthones, ß-D-glucan and dexamethasone. As the results, the expression of IL-6 and IL-10 reporters were strongly induced by LPS. The expression of IL-6 reporter was inhibited by all anti-inflammation compounds in LPS-induced RAW 264.7 cells. The expression of IL-10 reporter was inhibited by quercetin, xanthones and dexamethasone in LPS-induced RAW 264.7 cells. While, expression of IL-10 reporter was induced by ß-D-glucan. These results indicated that this assay could use for determination of IL-6 and IL-10 reporter expression in LPS-induced RAW 264.7 cells for anti-inflammation activity. Moreover, the results showed that natural compounds have an effect on the time course of IL-6 and IL-10 expressions. Therefore, real-time monitoring has a merit for natural compounds screening. We suggested that this assay could serve as a compound screening assay for anti-inflammation activity.

[A COMPARATIVE EVALUATION OF THE EFFECT OF DENTURES FROM VARIOUS MATERIALS ON THE ORAL CAVITY'S IMMUNOLOGICAL AND REDOX-DEPENDENT HOMEOSTASIS].

The purpose of our study was a comparative analysis of the effect of dentures from various materials on the immunological and redox-dependent homeostasis of the oral cavity. We studied 60 patients with removable dentures made based on plastics Prothyl Hot, Vertex BasiQ 20 (differing by polymerization regime) and elastic thermoplastic polymer Perflex Flexi Nylon. The control group consisted of 15 volunteers with a practically healthy oral cavity, who did not use dentures. Saliva collected on an empty stomach in a glass tube without the use of a stimulator before the establishment of a denture and 3 days and 1 month after. The content of the protein P-53 in saliva determined by immunoenzymatic assay with use of "Cusabio" reagent. The cytokines (IL1ß, IL10) content in saliva was determined immunoenzymatic assay. To determine the redox balance in the saliva of patients, the lipoperoxydradicals content (LOO.) content (by EPR method, using the spin-labeled α-phenyl-tertbutylnitron (PBN) (SIGMA)) and the activity of antioxidant enzymes (catalase and SOD) (by spectrophotometry) studied. Statistical processing of the results was carried out using the software package SPSS (version 10.0). Results of analysis show that defects associated with a lack of teeth do not affect the immune and oxidative balance of the oral cavity, but contribute to the development of destructive changes in the oral cavity's soft tissues, which manifested by an increase in the content of the proapoptotic protein P-53 in the saliva. After establishment of a denture, the intensity of apoptosis in the oral cavity tissues reduced. Establishment of a denture induced development of an inflammatory reaction during the first days, the intensity of which gradually decreased and completely disappeared at the end of the first month of the observation (manifested by the normalization of the parameters of the immune balance and antioxidant system). Minimal traumatic effects observed during establishment of a denture made based on Perflex Flexi Nylon.

Anti-inflammatory Effect of Somatostatin Analogue Octreotide on Rheumatoid Arthritis Synoviocytes.

Somatostatin and its analogues are known to have modulatory effects on immune response and their anti-proliferative, anti-angiogenic, and analgesic properties make them attractive candidates for a therapeutic use in immune-mediated diseases, such as rheumatoid arthritis. Here, we demonstrate the ability of the somatostatin analogue octreotide to inhibit interleukin-15 and to increase interleukin-10 production by rheumatoid arthritis fibroblast-like synovial cells maintained in a chronic inflammatory state. We also prove that the inhibitory effect of octreotide on interleukin-15 and tumor necrosis factor-α production depended on the increase in interleukin-10, since neutralizing anti-interleukin-10 antibody was able to partially reverse this inhibition. In addition, our observations suggest an octreotide control on purinergic signaling, with an inhibitory effect on purinergic P2X and P2Y receptors activation. This would have great implications, considering the roles of P2 receptors in the onset of inflammation. Data here reported extend knowledge on the biological action of octreotide and underline its multiple effects on immune response, which could make octreotide an attractive and valid support for the therapy of diseases where several inflammatory mediators are involved, such as rheumatoid arthritis, and in which the simultaneous action on different aspects can be a successful strategy.

Lactobacillus acidophilus Improves Intestinal Inflammation in an Acute Colitis Mouse Model by Regulation of Th17 and Treg Cell Balance and Fibrosis Development.

Disruption of the balance among the microbiota, epithelial cells, and resident immune cells in the intestine is involved in the pathogenesis of inflammatory bowel disease (IBD). Probiotics exert protective effects against IBD, and probiotic commensal Lactobacillus species are common inhabitants of the natural microbiota, especially in the gut. To investigate the effects of Lactobacillus acidophilus on the development of IBD, L. acidophilus was administered orally in mice with dextran sodium sulfate (DSS)-induced colitis. DSS-induced damage and the therapeutic effect of L. acidophilus were investigated. Treatment with L. acidophilus attenuated the severity of DSS-induced colitis. Specifically, it suppressed proinflammatory cytokines such as interleukin (IL)-6, tumor necrosis factor-α, IL-1ß, and IL-17 in the colon tissues, which are produced by T helper (Th) 17 cells. Moreover, in vitro L. acidophilus treatment directly induced T regulatory (Treg) cells and the production of IL-10, whereas the production of IL-17 was suppressed in splenocytes. In addition, we found that L. acidophilus treatment decreased the levels of α-smooth muscle actin, a marker of activated myofibroblasts, and type I collagen compared with control mice. These results suggest that L. acidophilus may be a novel treatment for IBD by modulating the balance between Th17 and Treg cells, as well as fibrosis development.

Current concepts on immunopathogenesis of hepatitis B virus infection.

Hepatitis B virus (HBV) infection is a leading cause of liver damage and hepatic inflammation. Upon infection, effective antiviral responses by CD8+ T cells, CD4+ T cells, Natural killer (NK) cells, and monocytes can lead to partial or complete eradication of the viral infection. To date, many studies have shown that the production of inhibitory cytokines such as Interleukin 10 (IL-10), Transforming growth factor beta (TGF-ß), along with dysfunction of the dendritic cells (DCs), and the absence of efficient innate immune responses could lead to T cell exhaustion, development of persistent infection, and inability to eradicate the viral infection from liver. Understanding the immunopathogenesis of the virus could be useful in providing further insights toward novel strategies in the eradication of HBV infection.

Pretreatment with Antiviral Preparation Kagocel Normalizes the Content of Bone Marrow Multipotent Stromal Cells and TNFα in Blood Serum of CBA Mice Disturbed by Administration of S. typhimurium Antigen Complex In Vivo and Maintains High Concentration of IL-10 and Th1 Cytokines.

Pretreatment with the active substance of antiviral preparation Kagocel, inductor of type I endogenous IFN, in a daily therapeutic dose (30 µg/mouse) 3 h prior to administration of S. typhimurium antigens to CBA mice reduced the number of bone marrow multipotent stromal cell (significantly increased by 3.2 times on the next day after antigen injection) to the initial level. Thus, activation of the stromal tissue induced by administration of the bacterial antigen was blocked. In addition, preliminary administration of Kagocel modulated the cytokine profile of the blood serum affected by S. typhimurium antigens: reduced 1.6-fold elevated concentration a proinflammatory cytokine TNFα to the control level (in 4 h after antigen injection) and maintained this level in 20 h after antigen administration. Kagocel also maintained the concentration of anti-inflammatory cytokine IL-10 at the level surpassing the normal by 1.6 times and high concentrations of Th1 cytokines (IL-2, IFNγ, and IL-12). These results suggest that Kagocel can reduce the immune response to bacterial antigens (similar to type I IFN [7]), which can contribute to its therapeutic and preventive effects in addition to its well documented antiviral activity and then this preparation can be used for the therapy of diseases accompanied by excessive or chronic inflammation.

Electro-Acupuncture at Zusanli Acupoint (ST36) Suppresses Inflammation in Allergic Contact Dermatitis Via Triggering Local IL-10 Production and Inhibiting p38 MAPK Activation.

Acupuncture has shown beneficial effect in the treatment of multiple dermatologic conditions including dermatitis, pruritus, urticaria, and hyperhidrosis; however, the detailed mechanisms are still kept unclear. This study aimed to investigate if electro-acupuncture (EA) treatment prevents 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD) in rats and explore its underlying mechanisms. ACD was induced by sensitizing and challenging with DNFB topically. Rats were treated daily following bilateral subcutaneous stimulation of EA at Zusanli acupoint (ST36) for 1 week. Ear swelling and serum IgE levels were measured. The ear biopsies were obtained for histology. Inflammatory cytokines on the dermatological ear and local acupoint tissue were assayed. Spleen lymphocytes and the homogenized supernatant of local acupuncture area were used to co-culture for flow cytology and immune analysis, respectively. EA treatment at ST36 notably inhibited ear swelling and inflammatory cell infiltration on DNFB-induced ACD. EA also decreased serum IgE concentrations and alleviated the production of inflammatory cytokines in dermatological ear. Additionally, EA treatment attenuated the percentage of CD4+IFN-γ+ and CD4+IL-4+ T cells associated with ACD. Interestingly, secretion of interleukin (IL)-10 in the local acupoint tissue following EA stimulation was increased and showed suppressive function when co-cultured with the spleen lymphocytes from DNFB group. Lastly, EA treatment demonstrably suppressed p38 MAPK activation in DNFB-treated rats. Our findings suggest that EA treatment at ST36 may ameliorate inflammation associated with DNFB-induced ACD via triggering local IL-10 production and inhibiting p38 MAPK activation, which provide an alternative and promising therapy for ACD.

Modulation of functional characteristics of resident and thioglycollate-elicited peritoneal murine macrophages by a recombinant banana lectin.

We demonstrated that a recombinant banana lectin (rBanLec), which structural characteristics and physiological impacts highly resemble those reported for its natural counterparts, binds murine peritoneal macrophages and specifically modulates their functional characteristics. By using rBanLec in concentrations ranging from 1 µg to 10 µg to stimulate resident (RMs) and thioglycollate-elicited (TGMs) peritoneal macrophages from BALB/c and C57BL/6 mice, we have shown that effects of rBanLec stimulation depend on its concentration but also on the functional status of macrophages and their genetic background. rBanLec, in a positive dose-dependent manner, promotes the proliferation of TGMs from both BALB/c and C57BL/6 mice, while its mitogenic influence on RMs is significantly lower (BALB/c mice) or not detectable (C57BL/6 mice). In all peritoneal macrophages, irrespective of their type and genetic background, rBanLec, in a positive dose dependent manner, enhances the secretion of IL-10. rBanLec stimulation of RMs from both BALB/c and C57BL/6 resulted in a positive dose-dependent promotion of proinflammatory phenotype (enhancement of NO production and IL-12 and TNFα secretion, reduction of arginase activity). Positive dose-dependent skewing toward proinflammatory phenotype was also observed in TGMs from C57BL/6 mice. However, the enhancement of rBanLec stimulation promotes skewing of TGMs from BALB/c mice towards anti-inflammatory profile (reduction of NO production and IL-12 secretion, enhancement of arginase activity and TGFß and IL-4 secretion). Moreover, we established that rBanLec binds oligosaccharide structures of TLR2 and CD14 and that blocking of signaling via these receptors significantly impairs the production of TNFα and NO in BALB/c macrophages. Since the outcome of rBanLec stimulation depends on rBanLec concentration as well as on the functional characteristics of its target cells and their genetic background, further studies are needed to investigate its effects under physiological and specific pathological conditions.

Nanoliposomal Buparvaquone Immunomodulates Leishmania infantum-Infected Macrophages and Is Highly Effective in a Murine Model.

Visceral leishmaniasis is a fatal parasitic neglected disease affecting 1.5 million people worldwide. Based on a drug repositioning approach, the aim of this work was to investigate the in vitro immunomodulatory potential of buparvaquone (BPQ) and to establish a safe regimen to evaluate the in vivo efficacy of BPQ entrapped by negatively charged nanoliposomes (BPQ-LP) in Leishmania infantum-infected hamsters. Small-angle X-ray scattering, dynamic light scattering, and the ζ-potential were applied in order to study the influence of BPQ on the liposome structure. Our data revealed that BPQ was located in the polar-apolar interface, snorkeling the polar region, and protected against aggregation inside the lipophilic region. The presence of BPQ also decreased the Z-average hydrodynamic diameter and increased the surface charge. Compared to intravenous and intramuscular administration, a subcutaneous route was a more effective route for BPQ-LP; at 0.4 mg/kg, BPQ-LP reduced infection in the spleen and liver by 98 and 96%, respectively. Treatment for 5 days resulted in limited efficacy, but 10 days of treatment resulted in an efficacy similar to that of a 15-day regimen. The nanoliposomal drug was highly effective, with a mean 50% effective dose of 0.25 mg/kg, reducing the parasite load in bone marrow by 80%, as detected using quantitative PCR analysis. In addition, flow cytometry studies showed that BPQ upregulated cytokines as tumor necrosis factor, monocyte chemoattractant protein 1, interleukin-10 (IL-10), and IL-6 in Leishmania-infected macrophages, eliminating the parasites via a nitric oxide-independent mechanism. This new formulation proved to be a safe and effective treatment for murine leishmaniasis that could be a useful candidate against visceral leishmaniasis.

ApoA-I/SR-BI modulates S1P/S1PR2-mediated inflammation through the PI3K/Akt signaling pathway in HUVECs.

Endothelial dysfunction plays a vital role during the initial stage of atherosclerosis. Oxidized low-density lipoprotein (ox-LDL) induces vascular endothelial injury and vessel wall inflammation. Sphingosine-1-phosphate (S1P) exerts numerous vasoprotective effects by binding to diverse S1P receptors (S1PRs; S1PR1-5). A number of studies have shown that in endothelial cells (ECs), S1PR2 acts as a pro-atherosclerotic mediator by stimulating vessel wall inflammation through the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. Scavenger receptor class B member I (SR-BI), a high-affinity receptor for apolipoprotein A-I (apoA-I)/high-density lipoprotein (HDL), inhibits nuclear factor-κB (NF-κB) translocation and decreases the plasma levels of inflammatory mediators via the PI3K/Akt pathway. We hypothesized that the inflammatory effects of S1P/S1PR2 on ECs may be regulated by apoA-I/SR-BI. The results showed that ox-LDL, a pro-inflammatory factor, augmented the S1PR2 level in human umbilical vein endothelial cells (HUVECs) in a dose- and time-dependent manner. In addition, S1P/S1PR2 signaling influenced the levels of inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-10, aggravating inflammation in HUVECs. Moreover, the pro-inflammatory effects induced by S1P/S1PR2 were attenuated by SR-BI overexpression and enhanced by an SR-BI inhibitor, BLT-1. Further experiments showed that the PI3K/Akt signaling pathway was involved in this process. Taken together, these results demonstrate that apoA-I/SR-BI negatively regulates S1P/S1PR2-mediated inflammation in HUVECs by activating the PI3K/Akt signaling pathway.

Phenotypic and Genetic Evaluation of the Influence of Pseudomonas aeruginosa Culture Fractions on the Human Mesenchymal Stem Cells Viability, Apoptotic Pathways and Cytokine Profile.

The objective of this study was to investigate the effects of P. aeruginosa PAO1 cellular and soluble culture fractions on human mesenchymal stem cells (MSCs) death signaling pathways and cytokine profile. The bone marrow isolated MSCs, incubated for different periods of time with one of the three P. aeruginosa PAO1 culture fractions, i.e. low density whole cultures, heat inactivated bacterial cultures sediments and sterile supernatants, were submitted to the following assays: i) fluorescence microscopy evaluation of cellular morphology and viability; ii) bax, caspase 9, relA and bcl-2 genes expression analysis by qRT-PCR; and iii) quantification of the level of IL-1ß, IL-6, IL-8 and IL-10 cytokines released in the MSCs supernatants determined by ELISA. Results were statistically analyzed using the GraphPad In Stat software. The PAO1 whole cultures exhibited the most relevant influences, impacting on MSCs morphology and viability, interfering with apoptotic pathways and significantly stimulating the production of IL-1ß and IL-10, while decreasing the production of IL-6 and IL-8. The culture supernatants increased the production of IL-1ß and reduced the secretion of all other tested cytokines, while heat-inactivated bacterial cells significantly stimulated both IL-1ß and IL-10 production. These data could suggest that in vivo, the fate of P. aeruginosa infection depends on the proportion between different bacterial culture fractions (i.e. the number of viable bacterial cells, the number of dead cells and the amount of bacterial soluble products accumulated locally) that could be influenced by the initial infective dose, by the host defense mechanisms, and also by the administered antimicrobial treatment that may thus interfere with the evolution and magnitude of the induced lesions.