Splenic Phagocytosis Research Articles

Chemotherapeutic Mechanism of Action of the Synthetic Resorcinolic Methyl 3,5-dimethoxy-2-octanoylbenzoate.

Resorcinolic lipids are described as potential examples of selective chemotherapeutic adjuvants that can enhance the effects of cyclophosphamide (CYC) while promoting cell death without causing DNA damage. Therefore, the current study attempted to describe how the resorcinolic lipid methyl 3,5-dimethoxy-2-octanoylbenzoate (AMS35BB) interacted with DNA (DNA docking) and how this compound affected genetic toxicology models and other biological characteristics when combined with CYC. We observed that AMS35BB, used alone (7.5 and 10 mg/kg), increases the frequency of genomic damage (comet assay) but not chromosomal damage (micronuclei assay), lowers phagocytosis, and promotes cell death in Swiss male mice. When used in association with CYC, AMS35BB can reduce the risk of genomic damage by up to 33.8% as well as chromosomal damage, splenic phagocytosis, cell death, and lymphocyte frequency. Molecular docking showed that AMS35BB had a higher affinity than the active metabolite of CYC for binding to the DNA double helix major groove. As a result, AMS35BB has the potential to be both an adjuvant when used in association with CYC and a therapeutic candidate for the development of a selective chemotherapeutic drug.

3-Heptylidene-4,6-Dimethoxy-3H-Isobenzofuran-1-One Is Genotoxic, Increases the Frequency of Cell Death, and Potentiates the Effects of Cyclophosphamide and Cisplatin.

3-heptylidene-4,6-dimethoxy-3H-isobenzofuran-1-one (Phthalide 1) is the precursor of three resorcinol lipids that have been described as potential chemotherapeutic agents and capable of potentiating the effects of cyclophosphamide. In this study, we evaluated the genotoxic potential, cell-killing potential, and interactions with cyclophosphamide and cisplatin of phthalide 1. Twelve groups were created from 120 mice: Negative Control, cyclophosphamide (100 mg/kg), cisplatin (6 mg/kg), Phthalide 1 (5, 10 and 20 mg/kg), and associations of 1 with cyclophosphamide and 1 with cisplatin. The results demonstrate that 1 increases (p < 0.05) the frequency of chromosomal damage, liver and kidney cell death, and splenic phagocytosis. The association of 1 with cyclophosphamide and cisplatin demonstrated a chemopreventive effect and, therefore, a reduction (p < 0.05) in the frequency of chromosomal damage. However, cell death and splenic phagocytosis did not suffer significant variations. As a result of the above, 1 has potential chemotherapeutic application and may be a candidate for developing a new generation of chemotherapeutics. In addition, it has characteristics to be used as a chemotherapy adjuvant in association with cyclophosphamide and cisplatin since it increases the frequency of cell death induced by chemotherapy. We also reported that the chemopreventive effect of 1, in association with cyclophosphamide and cisplatin, can prevent adverse effects (induction of DNA damage in non-tumor cells) without interfering with the mode of action of chemotherapy drugs and, therefore, without reducing the induction of cell death.

ZIM, a Norbornene Derived from 4-Aminoantipyrine, Induces DNA Damage and Cell Death but in Association Reduces the Effect of Commercial Chemotherapeutics

Cancer incidence is increasing, and the drugs are not very selective. These drugs cause adverse effects, and the cells become resistant. Therefore, new drugs are needed. Here, we evaluated the effects of ZIM, a candidate for chemotherapy, and 4-AA alone and in association with commercial chemotherapeutic agents. Subsequently, the results of ZIM and 4-AA were compared. Male Swiss mice were treated with doses of 12, 24, or 48 mg/kg ZIM or 4-AA alone or in association with cisplatin (6 mg/kg), doxorubicin (16 mg/kg), and cyclophosphamide (100 mg/kg). Biometric parameters, DNA damage (comet and micronuclei), cell death, and splenic phagocytosis were evaluated. DNA docking was also performed to confirm the possible interactions of ZIM and 4-AA with DNA. 4-AA has been shown to have low genotoxic potential, increase the frequency of cell death, and activate phagocytosis. ZIM causes genomic and chromosomal damage in addition to causing cell death and activating phagocytosis. In association with chemotherapeutical agents, both 4-AA and ZIM have a chemopreventive effect and, therefore, reduce the frequency of DNA damage, cell death, and splenic phagocytosis. The association of 4-AA and ZIM with commercial chemotherapeutic agents increased the frequency of lymphocytes compared to chemotherapeutic agents alone. Molecular docking demonstrated that ZIM has more affinity for DNA than 4-AA and its precursors (1 and 2). This was confirmed by the lower interaction energy of the complex (-119.83 kcal/mol). ZIM can break the DNA molecule and, therefore, its chemotherapeutic effect can be related to DNA damage. It is considered that ZIM has chemotherapeutic potential. However, it should not be used in combination with cisplatin, doxorubicin, and cyclophosphamide as it reduces the effects of these drugs.

Toxicological analysis and efficacy of 2-phenylchromone on mycobacteria viability and inflammatory response induced by Mycobacterium bovis

BackgroundStudies have reported that compounds similar to flavone present antimicrobial and anti-inflammatory properties; however these effects are poorly described in the context of mycobacterium infection. Purpose: This study investigated the efficacy of 2-phenylchromone (Flavone) on mycobacteria viability and inflammatory response induced by Mycobacterium bovis, as well as its toxicological potential in in vitro and in vivo models. Methods: In vitro antimycobacterial activity was performed using the resazurin microtiter assay method. In order to investigate the anti-inflammatory action, C57Bl/6 mice were pretreated orally with 2-phenylchromone (1–100 mg/kg) 1 h before the pleurisy induction. The leukocyte migration and cytokine production were evaluated. Acute oral toxicity test, genotoxicity evaluations and splenic phagocytosis assay were also evaluated. ResultsThe minimum inhibitory concentration of 2-phenylchromone in the presence of the Mycobacterium tuberculosis strain was 28.90 µg/mL. In the BCG-induced pleurisy model, the oral treatment with 2-phenylchromone (1, 10, or 100 mg/kg) caused a significant reduction in the tumor necrosis factor (TNF) levels in the pleural exudate, as well as in the total and differential leukocyte counts. The oral treatment with 2-phenylchromone also did not present acute toxicity or genotoxicity in comet assay and micronucleus tests. However, 2-phenylchromone induced an increase of 1.48-times in splenic phagocytosis in the animals. ConclusionOur results showed that 2-phenylchromone did not induced toxicity signs and is effective against Mycobacterium viability and reduced the inflammatory response elicited by mycobacteria.

Antimicrobial activity of plant extracts against carbapenem-producing Klebsiella pneumoniae and in vivo toxicological assessment

ABSTRACT The global spread of multidrug-resistant strains has prompted the scientific community to explore novel sources of chemicals with antimicrobial activity. The aim of the study was to examine the antimicrobial activity in vitro of 28 extracts against carbapenem-producing Klebsiella pneumoniae, individually and in combination with antibiotics and in vivo toxicological assessment of the most active product. The multi-resistant K. pneumoniae strain was submitted for phenotypic and molecular characterization. The antibacterial activity of 28 plant extracts was evaluated alone and in combination with antibiotics against this strain through the agar disk diffusion. Of these, 16 extracts showed synergism against carbapenem-producing K. pneumoniae, being that B. crassifolia extract exhibited synergism with three antibiotics. Based on this assessment, B. crassifolia-extract-induced toxicity on Swiss male mice was evaluated by administering this extract and subsequently determining apoptosis and splenic phagocytosis using the comet and micronucleus assays. The results of this study showed that B. crassifolia extract had synergistic activity promising and groups treated with B. crassifolia exhibited no genotoxic or mutagenic activity, indicating that B. crassifolia extract exerted beneficial effects and appeared safe to use at the studied concentrations.

Assessment of the toxicogenic effects and cell death potential of the ester (Z)-methyl 4-((1,5-dimethyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl)amino)-4-oxobut-2-anoate in combination with cisplatin, cyclophosphamide and doxorubicin.

Despite rapid advances in both the early detection and treatment of cancer, the mortality from this disease remains high, which justifies the development of new products that are more selective and effective and have fewer side effects. Accordingly, a novel ester was synthesized that contains two pharmacophores with important biological activities: (I) 4-aminoantipyrine, which has anti-inflammatory and antioxidant effects, and (II) the pharmacophore 1,4-dioxo-butenyl, which has cytotoxic activity. When administered alone, this compound is non-genotoxic, and it does not cause an increasing in splenic phagocytosis. Nevertheless, it can induce cell death. When administered in combination with commercial chemotherapeutic agents, such as doxorubicin, cisplatin, and cyclophosphamide, the ester shows antigenotoxic activity and decreases phagocytosis and reduces the potential to cause cell death. These results indicate that the compound should not be used in combination with chemotherapeutic agents that exert their effect through DNA damage, an important feature of antitumor drugs.

Resorcinolic lipid 3-heptyl-3,4,6-trimethoxy-3H-isobenzofuran-1-one is a strategy for melanoma treatment

AimsPrevious studies performed by our research group indicated that cytosporone analogues are capable of prevent or repair DNA damages. This work presents the evaluation of the activity of AMS35AA for metastatic murine melanoma cells (B16F10) in experimental model in vitro and, in pre-clinic assay of metastatic melanoma in vivo, using mice lineage C57BL/6. Main methodsIn vitro assays were performed: MTT and comet assay, flow cytometry evaluation, gene expression assay by RT-PCR, qualitative evaluation of cell death using B16F10 cells. In vivo assays: micronucleus and comet assay, splenic phagocytosis, melanoma murine model and histopathological analysis, using mice lineage C57BL/6 (n = 20). Key findingsIn vitro results performed by MTT assay showed that AMS35AA is cytotoxic for B16F10 cells (p < 0.05). Based on comet assay the genotoxicity of the IC50 was determined (95.83 μg/mL) (p < 0.05). These data were corroborated by flow cytometry analysis after the treatment with AMS35AA, which indicates the cellular death by apoptosis (p < 0.05) and increasing of ATR, p53, p21 and GADD45 gene expressions verified using RT-PCR. With respect to in vivo results, it was observed that AMS35AA did not show genotoxic activity. Data of tumor volume ex vivo indicate reduction of tumor for the treated animals with AMS35AA up to 15.84×, which is superior to Dacarbazina (50 mg/Kg, p.c.; i.p.). SignificanceIn summary, the study showed that AMS35AA reveals relevant results regarding to cytotoxicity of B16F10 murine melanoma cells, inducing death by apoptosis via mitochondrial and/or mediated by DNA damages.

Assessment of genetic integrity, splenic phagocytosis and cell death potential of (Z)-4-((1,5-dimethyl-3-oxo-2-phenyl-2,3dihydro-1H-pyrazol-4-yl) amino)-4-oxobut-2-enoic acid and its effect when combined with commercial chemotherapeutics.

The increased incidence of cancer and its high treatment costs have encouraged the search for new compounds to be used in adjuvant therapies for this disease. This study discloses the synthesis of (Z)-4-((1,5-dimethyl-3-oxo-2-phenyl-2,3dihydro-1H-pyrazol-4-yl) amino)-4-oxobut-2-enoic acid (IR-01) and evaluates not only the action of this compound on genetic integrity, increase in splenic phagocytosis and induction of cell death but also its effects in combination with the commercial chemotherapeutic agents doxorubicin, cisplatin and cyclophosphamide. IR-01 was designed and synthesized based on two multifunctionalyzed structural fragments: 4-aminoantipyrine, an active dipyrone metabolite, described as an antioxidant and anti-inflammatory agent; and the pharmacophore fragment 1,4-dioxo-2-butenyl, a cytotoxic agent. The results indicated that IR-01 is an effective chemoprotector because it can prevent clastogenic and/or aneugenic damage, has good potential to prevent genomic damage, can increase splenic phagocytosis and lymphocyte frequency and induces cell death. However, its use as an adjuvant in combination with chemotherapy is discouraged since IR-01 interferes in the effectiveness of the tested chemotherapeutic agents. This is a pioneer study as it demonstrates the chemopreventive effects of IR-01, which may be associated with the higher antioxidant activity of the precursor structure of 4-aminoantipyrine over the effects of the 1,4-dioxo-2-butenyl fragment.

Genetic toxicological assessment and anti-arthritic effects of an ethanolic extract obtained from Salvia lachnostachys Benth leaves in mice

In the present study, mice were subjected to prolonged treatment with ethanolic extract of Salvia lachnostachys Benth leaves (SLEE), and the inflammatory and arthritic parameters were evaluated using the Complete Freund's Adjuvant (CFA) model. The genotoxicity of SLEE were also assayed using genetic toxicological tests. For the CFA model, 28 male C57BL/6 mice were distributed randomly into four groups (control, 50 mg/kg of SLEE, 100 mg/kg of SLEE and dexamethasone) for the evaluation of hyperalgesia and paw edema for 21 days after injection of CFA into the paw. To conduct the toxicogenetic assessments (comet assay and micronuclei assay), apoptosis and splenic phagocytosis were evaluated in male Swiss mice after the administration of saline (control group), cyclophosphamide (positive control group) and SLEE (10, 100 and 1000 mg/kg). SLEE significantly reduced the mechanical hyperalgesia and edema caused by CFA injection. The results of the toxicogenetic assessment revealed no toxicogenetic potential in the mice, and the evaluation of apoptosis showed an increase in apoptotic cells in the spleen after 72 h of treatment with SLEE (1000 mg/kg). SLEE exhibited anti-arthritic activity with no toxicogenetic damage. These toxicogenic results support the safety of SLEE.

In vivo chemotherapeutic insight of a novel isocoumarin (3-hexyl-5,7-dimethoxy-isochromen-1-one): Genotoxicity, cell death induction, leukometry and phagocytic evaluation.

Chemotherapy is one of the major approaches for the treatment of cancer. Therefore, the development of new chemotherapy drugs is an important aspect of medicinal chemistry. Chemotherapeutic agents include isocoumarins, which are privileged structures with potential antitumoral activity. Herein, a new 3-substituted isocoumarin was synthesized from 2-iodo-3,5-dimethoxy-benzoic acid and oct-1-yne in a cross-coupling Sonogashira reaction followed by a copper iodide-catalyzed intramolecular cyclization as key step using MeOH/Et3N as the solvent system. The present study also evaluated the leukometry, phagocytic activity, genotoxic potential and cell death induction of three different doses (5 mg/kg, 10 mg/kg and 20 mg/kg) of this newly synthesized isocoumarin, alone and in combination with the commercial chemotherapeutic agents cyclophosphamide (100 mg/kg) and cisplatin (6 mg/kg) in male Swiss mice. The results suggest that the isocoumarin has genotoxicity and causes cell death. Noteworthy, this new compound can increase splenic phagocytosis and lymphocyte frequency, which are related to immunomodulatory activity. When combined with either cyclophosphamide or cisplatin, chemopreventive activity led to a reduction in the effects of both chemotherapeutic drugs. Thus, the new isocoumarin is not a candidate for chemotherapeutic adjuvant in treatments using cyclophosphamide or cisplatin. Nevertheless, the compound itself is an important prototype for the development of new antitumor drugs.

4-Aminoantipyrine reduces toxic and genotoxic effects of doxorubicin, cisplatin, and cyclophosphamide in male mice

The analgesic drug dipyrone is used to treat side effects (including pain and fever) of cancer chemotherapeutic agents. Dipyrone is metabolized to 4-aminoantipyrine (4-AA), a PGE2-dependent blocker and inhibitor of cyclooxygenase (COX). We evaluated the genotoxic, mutagenic, apoptotic, and immunomodulatory activities of 4-AA in vivo and the effects of its combination with the antineoplastic drugs doxorubicin, cisplatin, and cyclophosphamide. 4-AA did not cause genotoxic/mutagenic damage, splenic phagocytosis, or leukocyte alterations. However, when combined with the antineoplastic agents, 4-AA decreased their genotoxic, mutagenic, apoptotic, and phagocytic effects. These results suggest that 4-AA might interfere with DNA damage-mediated chemotherapy.

Campomanesia adamantium extract induces DNA damage, apoptosis, and affects cyclophosphamide metabolism.

Campomanesia adamantium (Cambess.) O. Berg. is originally from Brazil. Its leaves and fruits have medicinal properties such as anti-inflammatory, antidiarrheal and antiseptic properties. However, the mutagenic potential of this species has been reported in few studies. This study describes the mutagenic/antimutagenic, splenic phagocytic, and apoptotic activities of C. adamantium hydroethanolic extract with or without cyclophosphamide in Swiss mice. The animals orally received the hydroethanolic extract at doses of 30, 100, or 300 mg/kg with or without 100 mg/kg cyclophosphamide. Mutagenesis was evaluated by performing the micronucleus assay after treatment for 24, 48, and 72 h, while splenic phagocytic and apoptotic effects were investigated after 72 h. Short-term exposure of 30 and 100 mg/kg extract induced mild clastogenic/aneugenic effects and increased splenic phagocytosis and apoptosis in the liver, spleen, and kidneys. When the extract was administered in combination with cyclophosphamide, micronucleus frequency and apoptosis reduced. Extract components might affect cyclophosphamide metabolism, which possibly leads to increased clearance of this chemotherapeutic agent. C. adamantium showed mutagenic activity and it may decrease the effectiveness of drugs with metabolic pathways similar to those associated with cyclophosphamide. Thus, caution should be exercised while consuming these extracts, especially when received in combination with other drugs.

Diaryl sulfide analogs of combretastatin A-4: Toxicogenetic, immunomodulatory and apoptotic evaluations and prospects for use as a new chemotherapeutic drug.

Combretastatin A-4 exhibits efficient anti-cancer potential in human tumors, including multidrug-resistant tumors. We evaluated the mutagenic, apoptotic and immunomodulatory potential of two diaryl sulfide analogs of combretastatin A-4, 1,2,3-trimethoxy-5-([4-methoxy-3-nitrophenyl]thio)benzene (analog 1) and 1,2,3-trimethoxy-5-([3-amino-4-methoxyphenyl]thio)benzene (analog 2), as well as their association with the anti-tumor agent cyclophosphamide, in Swiss mice. Such evaluation was achieved using the comet assay, peripheral blood micronucleus test, splenic phagocytosis assay, and apoptosis assay. Both analogs were found to be genotoxic, mutagenic and to induce apoptosis. They also increased splenic phagocytosis, although this increase was more pronounced for analog 2. When combined with cyclophosphamide, analog 1 enhanced the mutagenic and apoptotic effects of this anti-tumor agent. In contrast, analog 2 did not enhance the effects of cyclophosphamide and prevented apoptosis at lower doses. These data suggest that analog 1 could be an adjuvant chemotherapeutic agent and possibly improve the anti-neoplastic effect of cyclophosphamide. Additionally, this compound could be a candidate chemotherapeutic agent and/or an adjuvant for use in combined anti-cancer therapy.

A novel cytosporone 3-Heptyl-4,6-dihydroxy-3H-isobenzofuran-1-one: synthesis; toxicological, apoptotic and immunomodulatory properties; and potentiation of mutagenic damage.

BackgroundA large number of studies are attempting to identify alternative products from natural sources or synthesized compounds that effectively interact with cancer cells without causing adverse effects on healthy cells. Resorcinolic lipids are a class of bioactive compounds that possess anticancer activity and are able to interact with the lipid bilayer. Therefore, the objective of this study was to synthesize a novel resorcinolic lipid and test its biological proprieties.MethodsWe aimed to synthesize a novel resorcinolic lipid belonging to the class of cytosporones, AMS049 (3-Heptyl-4,6-dihydroxy-3H-isobenzofuran-1-one) and to evaluate the toxicity of two concentrations of this lipid (7.5 and 10 mg/kg) by determining its genotoxic, mutagenic, immunomodulatory, and apoptotic effects, as well as any biochemical and histopathological alterations in mice treated with cyclophosphamide. The results were analyzed by ANOVA followed by the Tukey test A . level of significance of p < 0.05 was adopted.ResultsThe new cytosporone AMS049 was synthesized in only three steps and in satisfactory yields. The results indicate that the compound is neither genotoxic nor mutagenic and does not alter biochemical parameters. The histological alterations observed in the liver and kidneys did not compromise the function of these organs. Histology of the spleen suggested immunomodulation, although no changes were observed in splenic phagocytosis or differential blood cell count. The results also show that AMS049 potentiates the mutagenic effect of the chemotherapy drug cyclophosphamide and that the combination induces apoptosis.ConclusionThese facts indicate a potential therapeutic application of this novel cytosporone as an important chemotherapeutic adjuvant.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1532-2) contains supplementary material, which is available to authorized users.

Evaluation of anti-inflammatory, immunomodulatory, chemopreventive and wound healing potentials from Schinus terebinthifolius methanolic extract

Inflammatory and genetic alterations are related to the development of chronic diseases such as cancer. Schinus terebinthifolius Raddi, Anacardiaceae, is used in folk medicine to treat in- flammation, wounds and tumors. This study evaluated the anti-inflammatory, immunomod- ulatory, chemopreventive, and wound healing potentials of the methanolic extract from the leaves of Schinus terebinthifolius. The chemical composition of the extract was characterized using preliminary analytical LC methods. The results showed that the anti-inflammatory ac- tivity of the methanolic extract was similar to that of dexamethasone for edema reduction. Also, it inhibited the leukocyte migration into the air pouch and decreased plasma extrava- sation. In addition, the methanolic extract showed a healing action similar to that observed with collagenase. The methanolic extract is not genotoxic nor mutagenic, and in contrast it has chemopreventive activity, which elicits a high percentage of damage reduction by comet and micronucleus assay, preferably by bioantimutagenic action. The methanolic extract in- duced apoptosis and enhanced splenic phagocytosis in animals treated with cyclophospha- mide. The methanolic extract contents, resolved by LC, include phenolic acid and flavonoids. Our results suggest a therapeutic potential for the methanolic extract.

A new synthetic resorcinolic lipid 3-Heptyl-3,4,6-trimethoxy-3H-isobenzofuran-1-one: Evaluation of toxicology and ability to potentiate the mutagenic and apoptotic effects of cyclophosphamide

Resorcinolic lipids have important biological actions, including anti-carcinogenic activity. Therefore, we evaluated the mutagenic, genotoxic, immunomodulatory and apoptotic potential and the biochemical and histopathological changes caused by the synthetic resorcinolic lipid 3-Heptyl-3,4,6-trimethoxy-3H-isobenzofuran-1-one, (AMS35AA; 10, 20 and 40 mg/kg) alone or in combination with cyclophosphamide (100 mg/kg) in Swiss mice. The results indicated that AMS35AA is not genotoxic or mutagenic and does not alter liver or kidney histology. However, the compound does cause an increase (p < 0.05) in the levels of glutamic-oxaloacetic transaminase and creatinine and in splenic phagocytosis and liver and kidney apoptosis. When combined with cyclophosphamide, AMS35AA caused increased (p < 0.05) mutagenic damage (although the compound had anti-genotoxic activity), splenic phagocytosis, neutropenia and glutamic-oxaloacetic transaminase and creatinine levels (even in the absence of histological damage) and induced liver and kidney apoptosis. We conclude that this resorcinolic lipid may be an important chemotherapy adjuvant that can potentiate mutagenic damage and increase apoptosis caused by cyclophosphamide without causing adverse effects. In addition, the immunomodulatory activity of the compound should be noted, which counters reductions in lymphocyte number, a primary side effect of cyclophosphamide in cancer therapy.

Abstract 19046: RBC-Macrophage Inflammatory Axis: Potential Role in Atherosclerosis

Background: Red blood cells (RBC) modulate vasoreactivity, serve as a reservoir for chemokines via binding to Duffy antigen receptor for chemokines (DARC), and contribute to the pathology of atherosclerotic plaque rupture, suggesting a broad role in vascular pathophysiology. High fat diet (HFD) is known to negatively impact vascular function and modify RBC membrane composition, but how this affects RBC chemokine binding, and interaction with macrophages and blood vessels, is unknown. Methods and Results: RBC from C57BL/6J mice fed chow diet (chow RBC) or 60% HFD (HFD RBC) for 16 weeks were collected and studied in vitro and in vivo. Reactive oxygen species (DCFH fluorescence) and phosphatidylserine externalization (annexin V binding), a potential signal for RBC phagocytosis, were increased ~1.5 fold (p&lt;0.05) in HFD RBC compared to chow RBC. HFD RBC released ~1.5 fold more monocyte chemoattractant protein-1 (MCP-1) (p&lt;0.05) compared to chow RBC upon addition of heparin to disrupt DARC binding. In a transwell system, HFD RBC, as compared with chow RBC, produced a significant increase in the migration of thioglycollate-elicited macrophages through an endothelial monolayer (p&lt;0.05). Incubation of HFD RBC with murine macrophages demonstrated a ~1.5 fold (p&lt;0.05) increase in phagocytosis compared to chow RBC; upon their internalization by macrophages, HFD RBC more potently upregulated transcription of major pro-inflammatory chemokines Il-1b, Ccl3, and Cxcl2. As compared with chow RBC, exposure of murine aortic segments to HFD RBC under orbital shear conditions led to enhanced endothelial activation as evidenced by increased adhesion of labeled murine macrophages to the aortic endothelium (p&lt;0.05). Finally, when labeled and injected in vivo, HFD RBC exhibited a ~3 fold (p&lt;0.05) increase in splenic phagocytosis as compared with chow RBC. Conclusion: HFD profoundly impact RBC biology, inducing oxidative stress, altering membrane phospholipid display, and increasing chemokine release, phagocytosis by macrophages, and macrophage-blood vessel interactions. These novel findings suggest that RBC may play an important role in transducing vascular inflammation, and thus contributing to atherosclerosis, during HFD.

Effects of neuropeptide S on the proliferation of splenic lymphocytes, phagocytosis, and proinflammatory cytokine production of pulmonary alveolar macrophages in the pig

Neuropeptide S (NPS), a newly identified neuropeptide, is involved in many physiological and pathological activities through the NPS receptor (NPSR). Recently, the NPS and NPSR have been detected in peripheral systems of pigs including immune tissues, suggesting that NPS may play an important role in the regulation of immune function. The aim of this study was to demonstrate the presence and function of NPS and NPSR in splenic lymphocytes (SPLs) and pulmonary alveolar macrophages (PAMs) of pigs. By RT-PCR, the expression of NPS and NPSR mRNA was detected in the SPLs and PAMs. NPS immunoreactivity was observed in the membrane and cytoplasm of both SPLs and PAMs. We found that NPS could stimulate the proliferation of SPLs, when NPS was added at concentrations of 0.01, 0.1, 1, 10, 100 and 1000 nM alone or in combination with PHA/LPS in vitro. In macrophages from bronchoalveolar lavage (BAL) fluid of pigs, various doses of NPS (0.01, 0.1, 1, 10, 100 and 1000 nM) up-regulated the phagocytosis of PAMs in comparison to controls. In PAMs, NPS could induce the production of the pro-inflammatory cytokines IL-1β, IL-6 and TNF-α. Taken together, all data suggest that NPS is capable of inducing phagocytosis of non-opsonized E. coli. NPS might act as potent neuroimmunomodulatory factors and affects the maintenance of immune homeostasis.

Increase in Band 3 Density and Aggregation in Hereditary Spherocytosis

ABSTRACTPurpose: Red cells in hereditary spherocytosis are characterized by a reduced surface area/volume ratio. The mechanisms leading to the loss of membrane material and subsequent elimination of the cells have still not been clarified. It was the aim of the present study to analyze band 3 distribution in the red cell membrane and its putative role in red cell elimination. Methods/Results: Immunogold histochemistry was performed to detect band 3 in red cell membranes. Band 3 density and distribution were visualized by electron microscopy. Unsplenectomized spherocytosis patients (n = 12) showed reduced band 3 density and aggregation compared to controls (n = 15) (density: 1.2 ± 0.1 gold particles/μm circumference of red cell membrane vs 1.5 ± 0.07 gold particles/μm, x ± SEM; P < 0.05; aggregation: 0.26 ± 0.02 aggregates/μm vs 0.3 ± 0.02 aggregates/μm). By contrast, band 3 density and aggregation were increased in spherocytosis patients who had undergone splenectomy (density: 2.8 ± 0.1 gold particles/μm vs 2.0 ± 0.1 gold particles/μm; P < 0.05; aggregation: 1.5 ± 0.1 aggregates/μm vs 0.5 ± 0.03 aggregates/μm; P < 0.01). Artificial ageing of red cells from healthy controls (n = 6) led to a significant increase in band 3 aggregation (2.06 ± 0.2 aggregates/μm vs 0.33 ± 0.1 aggregates/μm; PWilcoxon < 0.01) but no change in band 3 density. In hereditary spherocytosis (n = 6), both band 3 density and aggregation increased significantly after artificial ageing of the red cells. The elevated band 3 aggregation was associated with a stimulated erythrophagocytosis in vitro. Conclusion: Band 3 aggregation characterizes the red cells in hereditary spherocytosis. It may be the cause of selective splenic phagocytosis of both spherocytes and senescent erythrocytes.

The Effect of Methadone on the Immune Status of B6C3F1 Mice

The Effect of Methadone on the Immune Status of B6C3F1 Mice. LeVier, D. G., Brown, R. D., Musgrove, D. L., Butterworth, L. F., McCay, J. A., White, K. L., Jr., Fuchs, B. A., Harris, L. S., and Munson, A. E. (1995). Fundam. Appl. Toxicol. 24, 275-284.Previously, morphine has been shown to elevate corticosterone via the hypothalamic-pituitary-adrenal axis and to suppress the immune system. The present investigation sought to determine if the μ-opiate receptor agonist methadone incurred a similar immune suppression in B6C3F1 mice. Serum methadone and corticosterone levels peaked 1 hr following a single subcutaneous injection of 20 mg/kg methadone HCl. Indeed, the rise in corticosterone levels paralleled that of methadone. After a single injection with 20 mg/kg methadone a pharmacokinetic analysis revealed a serum half-life of ≈2 hr. Following five injections of methadone over a 24-hr period (every 6 hr), methadone levels were elevated as would be expected; however, corticosterone levels did not become elevated. This suggests that the ability of methadone to elevate corticosterone becomes uncoupled following repeated dosing, indicative of either a tolerance or an increased catabolic mechanism. Moreover, dosing every 6 hr for 5 days induced an increase in the catabolism of methadone itself. Therefore, all assays were begun 1 hr after subcutaneous administration of methadone HCl, a time at which both methadone and corticosterone serum levels were elevated. The primary IgM antibody response to sheep red blood cells (sRBC) was suppressed when splenocytes were immunized in vitro. In contrast, animals immunized with sRBC and assayed for the primary IgM antibody response 4 days later were not suppressed. The activity of the resident macrophages of the liver and spleen as measured by the uptake of 51 Cr-sRBC was suppressed in a dose-dependent manner. Previously, it has been demonstrated that morphine suppresses hepatic and splenic phagocytic activity through an opiate receptor-mediated pathway that involves the release of corticosterone. It would appear that methadone plays a similar role in the suppression of hepatic and splenic phagocytosis.