Induced Lymphocyte Proliferation Research Articles

1,4-Naphthoquinone, a pro-oxidant, suppresses immune responses via KEAP-1 glutathionylation

Low levels of oxidative stress have been shown to activate Nrf-2, an important anti-inflammatory transcription factor, by us and also by several other investigators. Earlier we showed that pro-oxidants protect normal lymphocytes against radiation injury by activating Nrf-2. In the present study, we have investigated the effect of oxidative stress on immune responses and delineated the underlying mechanism. Hydrogen peroxide, tert-butylhydroquinone and 1,4-naphthoquinone (NQ) inhibited mitogen induced proliferation of lymphocytes. NQ also inhibited mitogen (Concanavalin A) induced cytokine secretion by murine T cells and lipopolysaccharide induced release of cytokines, nitric oxide and cyclooxygenase-2 expression by macrophages. NQ modulated cellular redox by decreasing GSH/GSSG ratio and the immunosuppressive effects of NQ were significantly abrogated by thiol containing antioxidants and not by non-thiol antioxidants. This redox perturbation led to activation of Nrf-2 pathway and inhibition of NF-κB. NQ treatment increased total protein S-thiolation, induced glutathionylation of KEAP-1 protein and decreased IKKβ levels in lymphocytes. Molecular docking studies revealed that NQ can disrupt KEAP-1/Nrf-2 interaction by directly blocking the binding site of Nrf-2 in the KEAP-1 protein. Further, inhibitors of Nrf-2 and HO-1 abrogated the anti-inflammatory effects of NQ. T cells isolated from spleen and gut associated lymphoid tissue of NQ administered mice also showed suppression of NF-κB activation and were hyporesponsive to mitogenic stimulation. These results demonstrate that pro-oxidants modulate inflammatory and immune responses via oxidative stress mediated KEAP-1 glutathionylation and IKKβ degradation.

Evaluation of immune functions in captive immature loggerhead sea turtles (Caretta caretta)

Sea turtles face numerous environmental challenges, such as exposure to chemical pollution and biotoxins, which may contribute to immune system impairment, resulting in increased disease susceptibility. Therefore, a more thorough assessment of the host's immune response and its susceptibility is needed for these threatened and endangered animals. In this study, the innate and acquired immune functions of sixty-five clinically healthy, immature, captive loggerhead sea turtles (Caretta caretta) were assayed using non-lethal blood sample collection. Functional immune assays were developed and/or optimized for this species, including mitogen-induced lymphocyte proliferation, natural killer (NK) cell activity, phagocytosis, and respiratory burst. Peripheral blood mononuclear cells (PBMC) and phagocytes were isolated by density gradient centrifugation on Ficoll–Paque and discontinuous Percoll gradients, respectively. The T lymphocyte mitogens ConA significantly induced lymphocyte proliferation at 1 and 2μg/mL while PHA significantly induced lymphocyte proliferation at 5 and 10μg/mL. The B lymphocyte mitogen LPS significantly induced proliferation at 1μg/mL. Monocytes demonstrated higher phagocytic activity than eosinophils. In addition, monocytes exhibited respiratory burst. Natural killer cell activity was higher against YAC-1 than K-562 target cells. These optimized assays may help to evaluate the integrity of loggerhead sea turtle's immune system upon exposure to environmental contaminants, as well as part of a comprehensive health assessment and monitoring program.

A Brucella Virulence Factor Targets Macrophages to Trigger B-cell Proliferation

Brucella spp. and Trypanosoma cruzi are two intracellular pathogens that have no evolutionary common origins but share a similar lifestyle as they establish chronic infections for which they have to circumvent the host immune response. Both pathogens have a virulence factor (prpA in Brucella and tcPrac in T. cruzi) that induces B-cell proliferation and promotes the establishment of the chronic phase of the infectious process. We show here that, even though PrpA promotes B-cell proliferation, it targets macrophages in vitro and is translocated to the cytoplasm during the intracellular replication phase. We observed that PrpA-treated macrophages induce the secretion of a soluble factor responsible for B-cell proliferation and identified nonmuscular myosin IIA (NMM-IIA) as a receptor required for binding and function of this virulence factor. Finally, we show that the Trypanosoma cruzi homologue of PrpA also targets macrophages to induce B-cell proliferation through the same receptor, indicating that this virulence strategy is conserved between a bacterial and a protozoan pathogen.

Investigation of effects of farrerol on suppression of murine T lymphocyte activation in vitro and in vivo

Farrerol, a new type of 2,3-dihydro-flavonoid, has been isolated from the leaves of Rhododendron dauricum L. In the present study, we found that farrerol exerted potent immunosuppressive effects on murine T cells both in vitro and in vivo. In vitro, farrerol markedly suppressed concanavalin A (ConA)-induced lymphocyte proliferation, Th1 and Th2 cytokine production, cluster of differentiation 4-positive (CD4(+)) T cell populations, and the ratio of CD4(+)/cluster of differentiation 8-positive (CD8(+)) T cells. Moreover, farrerol significantly inhibited the T cell-mediated delayed-type hypersensitivity (DTH) reaction in vivo. In addition, we investigated signal transduction mechanisms to determine the effects of farrerol by Western blotting. The data revealed that farrerol could downregulate the activation of the nuclear factor κB (NF-қB) and nuclear factor of activated T cells 2 (NFAT2) signal transduction pathways. These findings suggested that farrerol has potential effects on the regulation of the immune system and could be developed as a practicable immunosuppressive compound.

Patients with Idiopathic Pulmonary Fibrosis with Antibodies to Heat Shock Protein 70 Have Poor Prognoses

Diverse autoantibodies are present in most patients with idiopathic pulmonary fibrosis (IPF). We hypothesized that specific autoantibodies may associate with IPF manifestations. To identify clinically relevant, antigen-specific immune responses in patients with IPF. Autoantibodies were detected by immunoblots and ELISA. Intrapulmonary immune processes were evaluated by immunohistochemistry. Anti-heat shock protein 70 (HSP70) IgG was isolated from plasma by immunoaffinity. Flow cytometry was used for leukocyte functional studies. HSP70 was identified as a potential IPF autoantigen in discovery assays. Anti-HSP70 IgG autoantibodies were detected by immunoblots in 3% of 60 control subjects versus 25% of a cross-sectional IPF cohort (n = 122) (P = 0.0004), one-half the patients with IPF who died (P = 0.008), and 70% of those with acute exacerbations (P = 0.0005). Anti-HSP70 autoantibodies in patients with IPF were significantly associated with HLA allele biases, greater subsequent FVC reductions (P = 0.0004), and lesser 1-year survival (40 ± 10% vs. 80 ± 5%; hazard ratio = 4.2; 95% confidence interval, 2.0-8.6; P < 0.0001). HSP70 protein, antigen-antibody complexes, and complement were prevalent in IPF lungs. HSP70 protein was an autoantigen for IPF CD4 T cells, inducing lymphocyte proliferation (P = 0.004) and IL-4 production (P = 0.01). IPF anti-HSP70 autoantibodies activated monocytes (P = 0.009) and increased monocyte IL-8 production (P = 0.049). ELISA confirmed the association between anti-HSP70 autoreactivity and IPF outcome. Anti-HSP70 autoantibodies were also found in patients with other interstitial lung diseases but were not associated with their clinical progression. Patients with IPF with anti-HSP70 autoantibodies have more near-term lung function deterioration and mortality. These findings suggest antigen-specific immunoassays could provide useful clinical information in individual patients with IPF and may have implications for understanding IPF progression.

Antiviral effect of dietary germanium biotite supplementation in pigs experimentally infected with porcine reproductive and respiratory syndrome virus

Germanium biotite (GB) is an aluminosilicate mineral containing 36 ppm germanium. The present study was conducted to better understand the effects of GB on immune responses in a mouse model, and to demonstrate the clearance effects of this mineral against Porcine reproductive and respiratory syndrome virus (PRRSV) in experimentally infected pigs as an initial step towards the development of a feed supplement that would promote immune activity and help prevent diseases. In the mouse model, dietary supplementation with GB enhanced concanavalin A (ConA)-induced lymphocyte proliferation and increased the percentage of CD3+CD8+ T lymphocytes. In pigs experimentally infected with PRRSV, viral titers in lungs and lymphoid tissues from the GB-fed group were significantly decreased compared to those of the control group 12 days post-infection. Corresponding histopathological analyses demonstrated that GB-fed pigs displayed less severe pathological changes associated with PRRSV infection compared to the control group, indicating that GB promotes PRRSV clearance. These antiviral effects in pigs may be related to the ability of GB to increase CD3+CD8+ T lymphocyte production observed in the mice. Hence, this mineral may be an effective feed supplement for increasing immune activity and preventing disease.

Palladium‐induced Th2 cytokine responses reflect skin test reactivity

Recently, a crucial role of Th2 responses in nickel allergic contact dermatitis (ACD) was demonstrated. As palladium allergy is an issue of growing interest, the diagnostic potential of Th2 parameters for palladium sensitization was investigated. Palladium (Na(2) [PdCl(4)])-induced lymphocyte proliferation (LPT), Th1 and Th2 cytokine production were correlated with skin test (ST) reactivity in 16 positive and 21 negative controls. Furthermore, the diagnostic potential of these assays was evaluated using receiver operating characteristics (ROC) analysis. For comparison, same experiments were carried out for nickel (NiSO(4)). Correlation coefficients between palladium ST reactivity and IFN-γ, LPT, IL-5, and IL-13 were 0.34, 0.51, 0.69, and 0.78, and overall test accuracies were 68%, 81%, 89%, and 95%, respectively. Both palladium- and nickel-mediated Th2 responses tightly correlate with ST reactivity, supporting recent findings on the crucial role of Th2 involvement in ACD. Therefore, these assays may have great potential as diagnostic tools for future in vitro sensitization testing.

Cerebellar fastigial nuclear GABAergic projections to the hypothalamus modulate immune function

Our previous work has shown that the cerebellar fastigial nucleus (FN) is involved in modulation of lymphocyte function. Herein, we investigated effect of FN γ-aminobutyric acid (GABA)-ergic projections to the hypothalamus on lymphocytes to understand pathways and mechanisms underlying cerebellar immunomodulation. By injection of Texas red dextran amine (TRDA), an anterograde tracer, into FN, we found that the TRDA-labeled fibers from the FN traveled through the superior cerebellar peduncle (SCP), crossed in decussation of SCP (XSCP), entered the hypothalamus, and primarily terminated in the lateral hypothalamic area (LHA). Further, by injecting Fluoro-Ruby (FR), a retrograde tracer, in LHA, we observed that the FR-stained fibers retrogradely passed through XSCP and reached FN. Among these FR-positive neurons in the FN, there were GABA-immunoreactive cells. We then microinjected vigabatrin, which is an inhibitor of GABA-transaminase (GABA-T) that degrades GABA, bilaterally into FN. The vigabatrin treatment increased both number of GABA-immunoreactive neurons in FN-LHA projections and GABA content in the hypothalamus. Simultaneously, vigabatrin significantly reduced concanavalin A (Con A)-induced lymphocyte proliferation, anti-sheep red blood cell (SRBC) IgM antibody level, and natural killer (NK) cell number and cytotoxicity. In support of these findings, we inhibited GABA synthesis by using 3-mercaptopropionic acid (3-MP), which antagonizes glutamic acid decarboxylase (GAD). We found that the inhibition of GABA synthesis caused changes that were opposite to those when GABA was increased with vigabatrin. These findings show that the cerebellar FN has a direct GABAergic projection to the hypothalamus and that this projection actively participates in modulation of lymphocytes.

The protective effect of Canova homeopathic medicine in cyclophosphamide-treated non-human primates

Background: Canova activates macrophages and indirectly induces lymphocyte proliferation. Here we evaluated the effects of Canova in cyclophosphamide-treated non-human primates.Methods: Twelve Cebus apella were evaluated. Four animals were treated with Canova only. Eight animals were treated with two doses of cyclophosphamide (50mg/kg) and four of these animals received Canova. Body weight, biochemistry and hematologic analyses were performed for 40days. Micronucleus and comet assays were performed for the evaluation of DNA damage.Results: We observed that cyclophosphamide induced abnormal WBC count in all animals. However, the group treated with cyclophosphamide plus Canova presented a higher leukocyte count than that which received only cyclophosphamide. Cyclophosphamide induced micronucleus and DNA damage in all animals. The frequency of these alterations was significantly lower in the Canova group than in the group without this medicine.Conclusions: Our results demonstrated that Canova treatment minimizes cyclophosphamide myelotoxicity in C. apella.

Structural characterization and immunostimulatory activity of a novel linear α-(1 → 6)-D-glucan isolated from Panax ginseng C. A. Meyer

Panax ginseng C. A. Meyer is a well-known plant medicine in the world. Ginseng polysaccharides mainly contain starch-like glucan and pectin. In this paper, a novel glucan WGPA-UH-N1 was purified from ginseng pectin by the treatment of de-esterification and endo-polygalacturonase, followed by the chromatographies on DEAE-Sepharose Fast Flow and Sephadex G-50 column. WGPA-UH-N1 has molecular weight about 17 kDa. WGPA-UH-N1 was determined to be a linear α-(1→6)-D-glucan without side chains by FT-IR, (13)C-NMR, (1)H-NMR, HMQC and HMBC spectra. It is the first time to isolate a linear α-(1→6)-D-glucan from Panax ginseng C. A. Meyer. Immunological activity assays showed that WGPA-UH-N1, although not effective on the phagocytosis of macrophage, could significantly induce lymphocyte proliferation without mitogenic stimuli at 1.0 mg/mL or with LPS at 0.5 mg/mL, also significantly increase NO production at the range of 0.1-1.0 mg/mL in a dose-dependent manner. The immunological activities of WGPA-UH-N1 are different from those of the β-(1→6)-D-glucan (BIWP2) isolated from the fruit bodies of Bulgaria Inquinans (Fries).

Expression of IL‐1Rrp2 by human myelomonocytic cells is unique to DCs and facilitates DC maturation by IL‐1F8 and IL‐1F9

We report for the first time that expression of the novel IL-1 cytokine receptor IL-1Rrp2 (IL-1R6) is unique to DCs within the human myelomonocytic lineage. IL-1Rrp2 was expressed by monocyte-derived dendritic cells (MDDCs) which was dose-dependently increased by IL-4 and correlated with increased numbers of differentiated MDDCs. Human plasmacytoid DCs also express IL-1Rrp2 but the receptor is not expressed by either myeloid DC type 1 (mDC1) or mDC2 cells. We also show that IL-1F8 or IL-1F9 cytokines, which signal through IL-1Rrp2, induce maturation of MDDCs, as measured by increased expression of HLA-DR and CD83 and decreased expression of CD1a. Furthermore, IL-1F8 stimulated increased CD40 and CD80 expression and IL-18 and IL-12 p70 production by MDDCs, which induced proliferation of IFN-γ-producing CD3(+) lymphocytes (indicative of inflammatory Th1 subsets). IL-1F8 and IL-1F2 were equipotent in their ability to stimulate IL-18 secretion from MDDCs but IL-1F8 was not as potent as IL-1F2 in stimulating secretion of IL-12p70 from MDDCs or inducing lymphocyte proliferation Therefore, IL-1Rrp2 expression by some DC subsets may have an important function in the human immune response in vivo via its role in differentiation of inflammatory Th1 lymphocytes.

Activation of Lymphocytes Induced by Bronchial Epithelial Cells with Prolonged RSV Infection

Respiratory syncytial virus (RSV) preferentially infects airway epithelial cells,which might be responsible for susceptibility to asthma; however, the underlying mechanism is not clear. This study determined the activation of lymphocytes and drift of helper T (Th) subsets induced by RSV-infected human bronchial epithelial cells (HBECs) in vitro. HBECs had prolonged infection with RSV, and lymphocytes isolated from human peripheral blood were co-cultured with RSV-infected HBECs. Four groups were established, as follows: lymphocytes (group L); lymphocytes infected with RSV (group RL); co-culture of lymphocytes with non-infected HBECs (group HL); and co-culture of lymphocytes with infected HBECs (group HRL). After co-culture with HBECs for 24 hours, lymphocytes were collected and the following were determined in the 4 groups: cell cycle status; apoptosis rate; and concentrations of IL-4, IFN-γ, and IL-17 in the supernatants. Cell cycle analysis for lymphocytes showed a significant increase in S phase cells, a decrease in G1 phase cells, and a higher apoptosis rate in group HRL compared with the other three groups. In group HRL, the levels of IL-4, IFN-γ, and IL-17 in supernatants were also higher than the other three groups. For further study, lymphocytes were individually treated with supernatants from non-infected and RSV-infected HBECs for 24 h. We showed that supernatants from RSV-infected HBECs induced the differentiation of Th2 and Th17 subsets, and suppressed the differentiation of Treg subsets. Our results showed that HBECs with prolonged RSV infection can induce lymphocyte proliferation and apoptosis, and enhance the release of cytokines by lymphocytes. Moreover, subset drift might be caused by RSV-infected HBECs.

The Mitogen Induced Lymphocyte Proliferation Is Increased After An Adventure Sprint Race

The Mitogen Induced Lymphocyte Proliferation Is Increased After An Adventure Sprint Race

Promoting Effects of Ganoderma lucidum Polysaccharides on B16F10 Cells to Activate Lymphocytes

The immune system in patients with cancer often fails to control tumour growth because of deficient immunogenicity of tumour cells. Ganoderma lucidum polysaccharides (Gl-PS) are believed to have anti-tumour effects by boosting host immune function. Additionally, Gl-PS may have some direct effects on tumour cells in the activation of lymphocytes, thus enhancing the immunogenicity of tumour cells. We tested the effects of Gl-PS in lymphocyte activation by incubating Gl-PS with a tumour cell line deficient in antigen presentation. Our study showed that Gl-PS can promote B16F10 melanoma cells to induce lymphocyte proliferation, CD69 and FasL expression and IFN-γ production, indicating that Gl-PS can improve the nature of B16F10 cells to activate lymphocytes. Furthermore, H-2D(b) [a major histocompatibility (MHC) class I molecule], and B7-1 and B7-2 (two prominent co-stimulatory molecules expressed on B16F10 cells) were enhanced by Gl-PS, suggesting that these molecules may at least partially be involved in the process of Gl-PS on B16F10 cells to activate lymphocytes.

From Pichia anomala killer toxin through killer antibodies to killer peptides for a comprehensive anti-infective strategy

"Antibiobodies", antibodies (Abs) with antibiotic activity, internal image of a Pichia anomala killer toxin (PaKT) characterized by microbicidal activity against microorganisms expressing β-glucans cell-wall receptors (PaKTRs), were produced by idiotypic vaccination with a PaKT-neutralizing monoclonal Ab (PaKT-like Abs) or induced by a protein-conjugated β-glucan. Human natural PaKT-like Abs (PaKTAbs) were found in the vaginal fluid of women infected with KT-sensitive microorganisms. Monoclonal and recombinant PaKT-like Abs, and PaKTAbs proved to be protective against experimental candidiasis, cryptococcosis and aspergillosis. A killer decapeptide (KP), synthesized from the sequence of a recombinant PaKT-like Ab or produced in transgenic plants, showed a microbicidal activity in vitro, neutralized by β-glucans, a therapeutic effect in vivo, against experimental mucosal and systemic mycoses, and a prophylactic role in planta, against phytopathogenic microorganisms, respectively. KP showed fungicidal properties against all the defective mutants of a Saccharomyces cerevisiae library, inclusive of strains recognized to be resistant to conventional antifungal drugs. KP inhibited in vitro, ex vivo and/or in vivo HIV-1 and Influenza A virus replication, owing to down-regulation of CCR5 co-receptors, physical block of the gp120-receptor interaction and reduction in the synthesis of glycoproteins, HA and M1 in particular. KP modulated the expression of costimulatory and MHC molecules on murine dendritic cells, improving their capacity to induce lymphocyte proliferation. KP, proven to be devoid of cytotoxicity on human cells, showed self-assembly-releasing hydrogel-like properties, catalyzed by β1,3 glucan. PaKT's biotechnological derivatives may represent the prototypes of novel antifungal vaccines and anti-infective drugs characterized by different mechanisms of action.

New promising Euphorbiaceae extracts with activity in human lymphocytes from primary cell cultures

Context: Euphorbiaceae plants exhibit anti-inflammatory and immunomodulatory properties.Methods: We evaluated the activity of 14 extracts from seven Euphorbiaceae plants on primary immune cell cultures from healthy individuals. Peripheral blood mononuclear cells (PBMC) were exposed to the extracts w/o phytohaemagglutinin A or cycloheximide as agents that induce proliferation or apoptosis in PBMC, respectively.Results: We found that five up to 14 Euphorbiaceae’s extracts had the ability to modulate at least one of the immune parameters evaluated in this study. However, only the latex extracts of Euphorbia cotinifolia and Euphorbia tirucalli strongly induced both proliferation and apoptosis in PBMC. These extracts were further subfractioned by silica gel column chromatography. Two subfractions with enhanced activity in comparison to the crude extracts were obtained. Although these subfractions induced proliferation on both CD3+ and CD3− cells, the most prominent effects were observed in the former subpopulation. Interestingly, the subfraction from E. tirucalli induced lymphocyte proliferation without the need of accessory cells; this ability was not inhibited by the carbohydrates d-galactose and α-Methyl-d-Mannopyranoside.Conclusions: Altogether, these results reveal the presence of novel candidates within the Euphorbia plants to induce proliferation and apoptosis in human lymphocytes, mainly in CD3+ T cells.

HIV-1 Gag p24-Nef fusion peptide induces cellular and humoral immune response in a mouse model

Many Human immunodeficiency virus (HIV) candidate vaccines have been tested in clinical trials, but none was sufficiently effective in the prevention of HIV infection. A HIV vaccine should induce humoral as well as cell-mediated response, the latter including the cytotoxic CD8+ T lymphocyte (CTL) response. In this study, we immunized BALB/c mice with a purified fusion peptide Gag p24-Nef and evaluated immune responses. As for the cellular responses, the adjuvanted fusion peptide induced lymphocyte proliferation, CTL response, and cytokines IFN-gamma and IL-4 in the Th1 pattern. Humoral immune response to the adjuvanted fusion peptide included an increase in IgG antibodies of more IgG2a than IgG1 subtype. These results indicate that the employed HIV-1 peptide construct can elicit both cellular and humoral immune responses in mice. Further studies aimed at memory T cells and other aspects of immune responses are needed before a comprehensive assessment of this candidate vaccine could be provided. epitopes; fusion peptide; HIV-1 p24-Nef; immune response.

Antitumor and immunomodulating activity of polysaccharides from Enteromorpha intestinalis

Two polysaccharides (WEA and WEB) were isolated from Enteromorpha intestinalis by hot water extraction, anion-exchange, and gel-permeation chromatography. The average molecular weights (Mw) of the two fractions were 72.03 kDa (WEA) and 60.12 kDa (WEB). WEA was composed of Rha, Xyl, Man, and Glc in a molar ratio of 1.39:1.00:0.13:3.23. WEB consisted of Rha, Xyl, Gal, and GlcA (glucuronic acid) in a molar ratio of 7.32:1.00:0.51:1.28. Both polysaccharides could inhibit tumor growth in S180 tumor-bearing mice, and increased the relative spleen and thymus weight. They also increased the expression of tumor necrosis factor-alpha (TNF-α) in serum. WEA and WEB induced lymphocyte proliferation, increased the production of TNF-α in macrophages, and stimulated macrophages to produce nitric oxide dose-dependently through the up-regulation of inducible NO synthase activity. However, no direct cytotoxicity against Sarcoma 180 was investigated in vitro. These results indicate that the antitumor effects of these polysaccharides are associated with immunostimulation.

The residues 35proline and 41cysteine of chicken IL-2 are critical for binding to chicken CD25

The interleukin 2 (IL-2) immunoregulatory cytokine produces its biological effects by binding sequentially to its cell receptor subunits, alpha (CD25), beta (CD122), and common gamma chain (CD132). In this study, we identified the critical amino acid residues of chicken IL-2 (chIL-2) for binding to chicken CD25 (chCD25) by an Ag-capture ELISA, screening of a phage display peptide library, peptide-competitive ELISA and an in vitro T-cell proliferation assay. Specific ligand elution of phage bound to chCD25 and chIL-2 suggested that the P 35T 36C 41T 42Q 43L 46Q 47C 48Y 49L 50G 51 motif within chIL-2 molecule interacts with the S 99F 100C 101G 102M 103P 104Q 105T 106V 107P 108S 111L 112 motif of chCD25 molecule (chCD25 99–112), whereas the peptide competition ELISA assay showed the residues 27KIHLELYTPTETQEC 41 within chIL-2 (chIL-2 27–41) bound to the chCD25 protein. Lymphocyte proliferation and inhibition assays further confirmed that the binding of chIL-2 27–41 to the chCD25 molecule was inhibited by the chCD25 99–112 peptide. Site-specific mutation of the 35P and 41C residues in chIL-2 27–41 resulted in the lack of its ability to induce lymphocyte proliferation and binding to the chCD25 molecule. These findings demonstrate that chIL-2 27–41 and chCD25 99–112 are the binding domains between the chIL-2 and chCD25 molecules, and that the residues P 35 and C 41 within chIL-2 27–41 are the critical sites for its bioactivity and interaction with chCD25.

Molecular Basis of the Anti-Inflammatory Property Exhibited by Cyclo-Pentano Phenanthrenol Isolated from Lippia nodiflora

The objective of this study was to assess the anti-inflammatory potential of the active molecule isolated from Lippia nodiflora and to understand its molecular dynamics in Vitro inflammation models. Human Peripheral Blood Mononuclear Cells were used as models to study mitogen induced lymphocyte proliferation, cytokine mRNA expression (TNF-α, IL-1β and IL-6) and intracellular protein levels of pro-inflammatory mediators (MAPK and NF-κB). The NO release levels, on treatment with the extract and molecule, were correlated with the underlying iNOS mRNA expression in the murine macrophage cell line RAW 264.7. RT-PCR for COX-2, MMP2 and MMP9 were also performed in the cell line. The rat basophilic leukemia cell line RBL-2H3 was used as an in Vitro model for PLA2 activity. Then, 20 μg/ml of Lippia nodiflora crude methanol extract and 10 μg/ml of the purified CPP were used for subsequent studies based on the IC50 values obtained in the proliferation assay. Results demonstrate that the isolated Cyclo-pentano phenanthrenol inhibits TNF-α, IL-1β and IL-6 expression, NO release via iNOS suppression, prostaglandin biosynthesis via PLA2 and COX-2 inhibition and the activation of intracellular targets, MAPK and NF-κB. We conclude, cyclo-pentano phenanthrenol exerts its anti-inflammatory effect via inhibition of MAPK phosphorylation and NF-κB translocation.