Downregulation of NF-κB and PCNA in the regulatory pathways of apoptosis by cyclooxygenase-2 inhibitors in experimental lung cancer

Non-steroidal anti-inflammatory drugs (NSAIDs) are emerging as novel chemopreventive agents against a variety of cancers owing to their capability in blocking the tumor development by cellular proliferation, angiogenesis and by promoting apoptosis. The present study further explored the comparative role of a traditional NSAID, indomethacin and a newly developed coxib, etoricoxib against 9,10-dimethylbenz(a)anthracene (DMBA)-induced lung carcinogenesis in rats. Morphological and histological analysis revealed the occurrence of tumors and lesions along with constricted alveolar spaces in the DMBA treated animals which were largely corrected both by indomethacin and etoricoxib. COX-1 was found to be uniformly expressed in all the groups while COX-2 levels were raised prominently in the DMBA treated animals. Proliferation, as studied by PCNA expression was found to be markedly increased in the DMBA group as compared to the others. Increased NF-κB expression in the DMBA group was found to correct with the co-administration of NSAIDs. Also, fluorescent co-staining of the isolated lung cells revealed a significantly decreased apoptosis and altered mitochondrial membrane potential. In conclusion, these parameters indicate to the chemopreventive action of the two NSAIDs studied in lung cancer and as their mechanism of action suggests, can be achievable both by COX-dependent and COX-independent pathways.     

Chemopreventive effects of NSAIDs as inhibitors of cyclooxygenase-2 and inducers of apoptosis in experimental lung carcinogenesis

Roles of cyclooxygenase (COX) enzyme and intrinsic pathway of apoptosis have been explored for the chemopreventive effects of non-steroidal anti-inflammatory drugs (NSAIDs) on 9,10-dimethyl benz(a)anthracene (DMBA)-induced lung cancer in rat model. 16 weeks after the administration of DMBA, morphological analysis revealed the occurrences of tumours and lesions, which were regressed considerably with the co-administration of indomethacin and etoricoxib, the two NSAIDs under investigation. DMBA group was marked by hyperplasia and dysplasia as observed by histological examination, and these features were corrected to a large extent by the two NSAIDs. Elevated levels of COX-2 were seen in the DMBA group, the enzyme responsible for prostaglandin synthesis during inflammation and cancer, whilst the expression of the constitutive isoform, COX-1, was equally expressed in all the groups. Apoptosis was quantified by studying the activities of apaf-1, caspase-9, and 3 by immunofluorescence and western blots. Their activities were found to diminish in the DMBA-treated animals as compared to the other groups. Fluorescent co-staining of the isolated broncho-alveolar lavage cells showed reduced number of apoptotic cells in the DMBA group, indicating decrease in apoptosis after carcinogen administration. The present results thus suggest that the mechanism of cancer chemoprevention of NSAIDs may include the suppression of COX-2 and the induction of apoptosis.     

Quinine Inhibits Mitochondrial ATP-regulated Potassium Channel from Bovine Heart

The mitochondrial ATP-regulated potassium (mitoK_ATP) channel has been suggested as trigger and effector in myocardial ischemic preconditioning. However, molecular and pharmacological properties of the mitoK_ATP channel remain unclear. In the present study, single-channel activity was measured after reconstitution of the inner mitochondrial membrane from bovine ventricular myocardium into bilayer lipid membrane. After incorporation, a potassium-selective current was recorded with mean conductance of 103 ± 9 pS in symmetrical 150 mM KCl. Single-channel activity of this reconstituted protein showed properties of the mitoK_ATP channel: it was blocked by 500 μM ATP/Mg, activated by the potassium-channel opener diazoxide at 30 μM, inhibited by 50 μM glibenclamide or 150 μM 5-hydroxydecanoic acid, and was not affected by the plasma membrane ATP-regulated potassium-channel blocker HMR1098 at 100 μM. We observed that the mitoK_ATP channel was blocked by quinine in the micromolar concentration range. The inhibition by quinine was additionally verified with the use of ⁸⁶Rb⁺ flux experiments and submitochondrial particles. Quinine inhibited binding of the sulfonylurea derivative [³H]glibenclamide to the inner mitochondrial membrane. We conclude that quinine inhibits the cardiac mitoK_ATP channel by acting on the mitochondrial sulfonylurea receptor.(P. Bednarczyk and A. Kicińska) These authors contributed equally to this work.This revised version was published online in August 2005 with a corrected cover date.     

Lithium Modulates Cancer Cell Growth, Apoptosis, Gene Expression and Cytokine Production in HL-60 Promyelocytic Leukaemia Cells and Their Drug-Resistant Sub-clones

Lithium has been an FDA-approved and preferred drug for the treatment of mood disorders for many years, and cumulative evidence has pointed towards its potential use as an anti-cancer agent. Previous studies in our laboratory have demonstrated that lithium induces apoptotic cell death in HL-60 promyelocytes at concentrations of 10?mM and above. A lithium-tolerant HL-60 sub-clone, resistant to up to 15?mM lithium, was also generated and its growth profile reported. Treatment of cells with lithium resulted in a dose-dependent induction of p53, retinoblastoma (Rb) and bax expression which was accompanied by concomitant inhibition of bcl-2 expression as demonstrated using immunohistochemical microscopy. These results seem to suggest that lithium induced cell death in these cells by inhibiting expression of anti-apoptotic protein, bcl-2, while inducing higher expression of its pro-apoptotic counterparts which include bax. Expression of bax and bcl-2 is also linked to expression of inflammation-regulating cytokines. Using ELISA assays, lithium was demonstrated to induce production of pro-inflammatory cytokines, IL-6 and TNF-??, while inhibiting release of anti-inflammation-related IL-2 and IL-10 in a dose-dependent fashion. Our findings identify a critical function for lithium in modulating pro- versus anti-apoptotic gene expression and pro- versus anti-inflammatory cytokines in vitro and provide a rationale for suggesting a promising role of lithium in regulation of inflammation and cancer growth.     

Regulation of osteoarthritis-associated key mediators by TNFα and IL-10: effects of IL-10 overexpression in human synovial fibroblasts and a synovial cell line

Synovial fibroblasts (SF) contribute to the pathogenesis of osteoarthritis (OA), but the effects of intra-articular cytokines on SF are not completely understood. The aim of this study was to characterize the interplay between tumor necrosis factor (TNF)α and the anti-inflammatory interleukin (IL)-10. Non-immortalized human SF and SF of the human cell line K4IM were stimulated with recombinant TNFα, IL-10, or TNFα?+?IL-10 (10 ng/ml each) for 24 h or transduced with an adenoviral vector overexpressing human IL-10 (hIL-10) and subsequently treated with 10 ng/ml TNFα for 24 h. Effects on the gene expression and protein synthesis of IL-6, IL-10, matrix metalloproteinases (MMP)-1, ?3, type I collagen, β₁-integrin, and CD44 were investigated via real-time detection polymerase chain reaction, immunofluorescence labeling, flow cytometry, and Western blotting. IL-10 release by transduced SF was confirmed with enzyme-linked immunosorbent assay. Both cell populations were activated by TNFα and by TNFα?+?IL-10, increasing their gene expression and protein synthesis of IL-6, IL-10, MMP-1, and MMP?3 and altering the synthesis of type I collagen, β₁-integrin, and CD44. hIL-10 overexpression greatly elevated the gene expression and protein synthesis of IL-10. However, transduction did not significantly affect the gene expression of IL-6, MMP-1, and MMP?3 in SF. The increased expression of pro-inflammatory and catabolic mediators in TNFα-activated SF indicates their role in OA pathogenesis, suggesting they are a potential therapeutic target. Although the vigorousness of the responses of non-immortalized SF and K4IM clearly differ, the K4IM cell line seems to be a suitable model for non-immortalized human SF.     

Cytokine Degradation by Biofilms of Porphyromonas gingivalis

The aim of this study was to determine whether biofilms of Porphyromonas gingivalis could proteolytically degrade the cytokines interleukin (IL)-1β, IL-6, or IL-1 receptor antagonist (IL-1ra). Biofilms were grown on membrane filters on the surface of Wilkins-Chalgren blood agar. The biofilms were removed from the plates, and solutions containing 2.5 μg/ml of each cytokine were added. Following incubation for up to 4.0 h, supernatants from the biofilms were subjected to SDS-PAGE. The separated proteins were transferred by Western blotting to PVDF membranes and probed with peroxidase-conjugated antibodies recognizing both the intact cytokines and their degradation products. After 2 h, no intact IL-1β, IL-6, or IL-1ra were detectable. Cytokine proteolysis also occurred in the presence of horse serum. These results demonstrate that biofilm-grown P. gingivalis can degrade both pro- and anti-inflammatory cytokines and so may be able to perturb cytokine networks in vivo by eliminating cytokines from the local environment. Received: 12 August 1997 / Accepted: 15 October 1997     

1,8-Cineole (Eucalyptol) Mitigates Inflammation in Amyloid Beta Toxicated PC12 Cells: Relevance to Alzheimer’s Disease

Inflammatory process has a fundamental role in the pathogenesis of Alzheimer’s disease and insoluble amyloid beta deposits and neurofibrillary tangles provide the obvious stimuli for inflammation. The present study demonstrate the effect of pretreatment of 1,8-cineole (Cin) on inflammation induced by Aβ_(25–35) in differentiated PC12 cells. The cells were treated with Cin at different doses for 24 h and then replaced by media containing Aβ_(25–35) for another 24 h. The cell viability was decreased in Aβ_(25–35) treated cells which was significantly restored by Cin pretreatment. Cin successfully reduced the mitochondrial membrane potential, ROS and NO levels in Aβ_(25–35) treated cells. Cin also lowered the levels of proinflammatory cytokines TNF-α, IL-1β and IL-6 in Aβ_(25–35) treated cells. Moreover, Cin also succeeded in lowering the expression of NOS-2, COX-2 and NF-κB. This study suggests the protective effects of Cin on inflammation and provides additional evidence for its potential beneficial use in therapy as an anti-inflammatory agent in neurodegenerative disease.     

New pentadienone oxime ester derivatives: synthesis and anti-inflammatory activity

To develop novel anti-inflammatory agents, a series of new pentadienone oxime ester compounds were designed and synthesized. The structures were determined by IR, ¹H NMR, ^13?C NMR, and HRMS. All compounds have been screened for their anti-inflammatory activity by evaluating their inhibition against LPS-induced nitric oxide (NO) release in RAW 264.7 cell. Among them, compound 5j was found to be one of the most potent compounds in inhibiting NO and IL-6 (IC₅₀ values were 6.66?µM and 5.07?µM, respectively). Preliminary mechanism studies show that title compound 5j could significantly suppress expressions of nitric oxide synthase, COX-2, and NO, IL-6 through Toll-like receptor 4/mitogen-activated protein kinases/NF-κB signalling pathway. These data support further studies to assess rational design of more efficient pentadienone oxime ester derivatives with anti-inflammatory activity in the future.     

The antioxidant and anti-inflammatory effects of abalone intestine digest, Haliotis discus hannai in RAW 264.7 macrophages

Abalone is a marine gastropod and an important fishery and food industrial resource that is massively maricultured in Asia, Africa, Australia, and America. However, the health beneficial effects of abalone have rarely been reported. In the present study, we examine the antioxidant and anti-inflammatory effects of abalone Haliotis discus hannai in macrophage cells. The results showed that abalone intestine digest (AID) has antioxidant activities against lipid peroxidation, ROS stress and DNA damage in H₂O₂-treated RAW264.7 macrophages. In the lipopolysaccharide (LPS)-induced macrophages, AID suppresses LPS-induced production of nitric oxide (NO) via inducible nitric oxide synthase (iNOS) expression in a dose-dependent manner. It also significantly reduced the generation of proinflammatory cytokines, such as interleukin (IL-1??), tumor necrosis factor (TNF)-??, and IL-6. Furthermore, AID significantly suppresses the phosphorylation of mitogen-activated protein kinases (MAPKs) such as ERK, JNK, and p38. These results indicated that AID inhibits oxidative damage by ROS and LPS-induced inflammatory response via blocking the MAPK signaling pathway in murine macrophages. The potent antioxidant and anti-inflammatory effects of abalone intestine as byproducts from fishery manufacturing may suggest the possibility of high valuable utilization and application as a nutraceutical and therapeutic substance.     

Ammonia Attenuates LPS-Induced Upregulation of Pro-Inflammatory Cytokine mRNA in Co-Cultured Astrocytes and Microglia

Hepatic encephalopathy (HE) is associated with cerebral microglia activation. Ammonia, a major toxin of HE, activates microglia in vitro but does not trigger pro-inflammatory cytokine synthesis. In the present study we analysed effects of ammonia on lipopolysaccharide (LPS)-induced upregulation of microglia activation and cytokine mRNA as well as on cytokine secretion in mono-cultured microglia and co-cultured astrocytes and microglia. In mono-cultured microglia LPS (100 ng/ml, 18 h) strongly elevated mRNA levels of the microglia activation marker CD14 and the pro-inflammatory cytokines IL-1α/β, IL-6 and TNF-α. NH₄Cl (5 mmol/l) had no effect on LPS-induced upregulation of CD14, IL-1α/β and IL-6 mRNA but enhanced LPS-induced upregulation of TNF-α mRNA in mono-cultured microglia. In co-cultured astrocytes and microglia, however, LPS-induced upregulation of IL-1α/β, TNF-α, IL-6, CD14 but not of IL-10, IL-12A/B or TGFβ_1?3 mRNA was attenuated by NH₄Cl. LPS-induced upregulation of IL-1α/β, IL-6 and TNF-α was also diminished by the TGR5-ligands allopregnanolone and taurolithocholic acid in mono-cultured microglia. NH₄Cl also attenuated LPS-induced release of MCP-1, IL-6 and IL-10 in mono-cultured microglia. mRNA level of surrogate marker for microglia activation (CD14) and for the anti-inflammatory M2-type microglia (CD163, CXCL1, CXCL2) were also elevated in post mortem brain tissue taken from the fusiforme gyrus of patients with liver cirrhosis and HE. The findings suggest that ammonia attenuates LPS-induced microglia reactivity in an astrocyte-dependent way. One may speculate that these anti-inflammatory effects of ammonia may be triggered by neurosteroids derived from astrocytes and may account for absence of microglia reactivity in cerebral cortex of cirrhotic patients with HE.     

Curcumin protects against acute liver damage in the rat by inhibiting NF-κB,proinflammatory cytokines production and oxidative stress

Curcumin, an anti-inflammatory and antioxidant compound, was evaluated for its ability to suppress acute carbon tetrachloride-induced liver damage. Acute hepatotoxicity was induced by oral administration of CCl₄ (4 g/kg, p.o.). Curcumin treatment (200 mg/kg, p.o.) was given before and 2 h after CCl₄ administration. Indicators of necrosis (alanine aminotransferase) and cholestasis (γ-glutamyl transpeptidase and bilirubins) resulted in significant increases after CCl₄ intoxication, but these effects were prevented by curcumin treatment. As an indicator of oxidative stress, GSH was oxidized and the GSH/GSSG ratio decreased significantly by CCl₄, but was preserved within normal values by curcumin. In addition to its antioxidants properties, curcumin is capable of preventing NF-κB activation and therefore to prevent the secretion of proinflammatory cytokines. Therefore, in this study we determined the concentrations of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) mRNA, and NF-κB activation. CCl₄-administered rats depicted significant increases in TNF-α, IL-1β, and IL-6 production, while curcumin remarkably suppressed these mediators of inflammation in liver damage. These results were confirmed by measuring TNF-α, and IL-1β protein production using Western Blot analysis. Accordingly, these proteins were increased by CCl₄ and this effect was abolished by curcumin. Administration of CCl₄ induced the translocation of NF-κB to the nucleus; CCl₄ induced NF-κB DNA binding activity was blocked by curcumin treatment. These findings suggest that curcumin prevents acute liver damage by at least two mechanisms: acting as an antioxidant and by inhibiting NF-κB activation and thus production of proinflammatory cytokines.     

Ingested (oral) thyrotropin releasing factor (TRH) inhibits EAE

BackgroundIngested immunoactive proteins type I IFN, SIRS peptide 1–21, α-MSH, ACTH, SST inhibit clinical attacks and inflammation in acute EAE by decreasing Th1-like cytokines, increasing Th2-like cytokines or increasing T_reg cell frequencies.ObjectiveWe examined whether another protein, thyrotropin releasing factor (TRH), would have similar anti-inflammatory effects in EAE after oral administration.Design/methodsB6 mice were immunized with MOG peptide 35–55 and gavaged with control saline or TRH during ongoing disease. Splenocytes from mock fed or TRH fed mice were adoptively transferred into active MOG peptide 35–55 immunized recipient mice during ongoing disease.ResultsIngested (oral) TRH inhibited ongoing disease and decreased inflammation. Adoptively transferred cells from TRH fed donors protected against actively induced disease and decreased inflammation. In actively fed mice, oral TRH decreased IL-17 and TNF-α cytokines in both the spleen and the CNS. In recipients of donor cells from TRH fed mice there was a reduction of Th1 and Th17 and induction of Th2-like IL-13 cytokines in both the spleen and CNS. Oral TRH decreased clinical score and decreased inflammatory foci in both actively fed and recipients of actively fed mice. There was no significant increase in T_reg cell frequencies in actively fed or recipients of TRH fed donor cells.ConclusionsIngested (orally administered) TRH can inhibit clinical disease, inhibit CNS inflammation by decreasing Th1-like, Th17 and TNF-α cytokines and increasing Th2-like cytokines (IL-13) in the CNS.     

Anti-inflammatory and Anti-apoptotic Effect of Combined Treatment with Methylprednisolone and Amniotic Membrane Mesenchymal Stem Cells After Spinal Cord Injury in Rats

This study was undertaken to investigate the synergistic effects of methylprednisolone (MP) administration and transplantation of amniotic membrane mesenchymal stem cells (AM-MSCs) following T11 spinal cord clip compressive injury in rats. The combination treatment with MP (50 mg/kg) and delayed transplantation of AM-MSCs after rat spinal cord injury, significantly reduced (1) myeloperoxidase activity, (2) the proinflammatory cytokines: tumor necrosis factor-α, interleukin (IL)-1β, IL-6, IL-17, interferon-γ and (3) the cell apoptosis [terminal deoxynucleotidyl transferase, dUTP nick end labeling (TUNEL) staining, and caspase-3, Bax and Bcl-2 expressions]; increased: (1) the levels of the anti-inflammatory cytokines (IL-10 and transforming growth factor-β1) and (2) the survival rate of AM-MSCs in the injury site. The combination therapy significantly ameliorated the recovery of limb function (evaluated by Basso, Beattie and Bresnahan score). Taken together, our results demonstrate that MP in combination with AM-MSCs transplantation is a potential strategy for reducing secondary damage and promoting functional recovery following spinal cord injury.     

Cytokine and Nitric Oxide Levels in Patients with Sepsis – Temporal Evolvement and Relation to Platelet Mitochondrial Respiratory Function

Background

The levels of nitric oxide (NO) and various cytokines are known to be increased during sepsis. These signaling molecules could potentially act as regulators and underlie the enhancement of mitochondrial function described in the later phase of sepsis. Therefore, we investigated the correlation between observed changes in platelet mitochondrial respiration and a set of pro- and anti-inflammatory cytokines as well as NO plasma levels in patients with sepsis.

Methods and Results

Platelet mitochondrial respiration and levels of TNFα, MCP-1 (monocyte chemotactic protein-1), INFγ (interferon-γ), IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10 and IL-17 and NO were analyzed in 38 patients with severe sepsis or septic shock at three time points during one week following admission to the ICU. Citrate synthase, mitochondrial DNA and cytochrome c were measured as markers of cellular mitochondrial content. All mitochondrial respiratory states increased over the week analyzed (p<0.001). IL-8 levels correlated with maximal mitochondrial respiration on day 6–7 (p = 0.02, r² = 0.22) and was also higher in non-survivors compared to survivors on day 3–4 and day 6–7 (p = 0.03 respectively). Neither NO nor any of the other cytokines measured correlated with respiration or mortality. Cytochrome c levels were decreased at day 1–2 by 24±5% (p = 0.03) and returned towards values of the controls at the last two time points. Citrate synthase activity and mitochondrial DNA levels were similar to controls and remained constant throughout the week.

Conclusions

Out of ten analyzed cytokines and nitric oxide, IL-8 correlated with the observed increase in mitochondrial respiration. This suggests that cytokines as well as NO do not play a prominent role in the regulation of platelet mitochondrial respiration in sepsis. Further, the respiratory increase was not accompanied by an increase in markers of mitochondrial content, suggesting a possible role for post-translational enhancement of mitochondrial respiration rather than augmented mitochondrial mass.     

Alteration in some pro and anti-inflammatory cytokines associated with complete and incomplete gestation cycle of cows

The maternal immune response during pregnancy is regulated by a complex array of pro and anti-inflammatory cytokines which provide optimum conditions for the embryo implantation and survival. The possible role of pro and anti-inflammatory cytokines during the complete gestation cycle of ruminants and the difference in their circadian rhythmicity between successful and unsuccessful pregnancies is still unknown. To study this, blood samples were collected from three groups of cows, pregnant (P), non-pregnant (NP) and aborted (ABORT) cows starting from the day of Artificial Insemination (AI) till calving in P cows and till stages of non pregnancy in other cows. Various pro and anti-inflammatory cytokines were estimated by bovine-specific ELISA test and compared. Successful pregnancies had lower levels of pro-inflammatory cytokines (IL-2, IL-6 and IL-8) but higher anti-inflammatory cytokine (IL-10) mainly during implantation. Significant (p < 0.05) increase in pro-inflammatory cytokines and low IL-10 levels was noticed at abortion and at calving. This study indicates that temporal and spatial aspects of reducing the release of various pro-inflammatory cytokines and maintaining high anti-inflammatory cytokines during pregnancy are essentially required for maintenance of pregnancy. Any disturbance in the cytokine equilibrium may lead to persistent inflammation and pregnancy failure.     

Effect of intravenous vitamin C on cytokine activation and oxidative stress in end-stage renal disease patients receiving intravenous iron sucrose

Reticuloendothelial blockade in hemodialysis patients prevents optimal intravenous (IV) iron utilization. Vitamin C has emerged as a potential therapy to improve anemia treatment by enhancing iron mobilization. However, Vitamin C can act as a pro-oxidant in the presence of iron. This was a prospective, open-label, crossover study. Thirteen patients with end-stage renal disease on hemodialysis and four healthy controls were assigned to receive 100?mg of IV iron sucrose (IS) or 100?mg of IV IS co-administered with 300?mg of IV Vitamin C (IS?+?C) in random sequence. Serum samples for IL-1, IL-6, TNF-α and IL-10 and non-transferrin bound iron were obtained at baseline, 45?min and 105?min post study medication administration. Peripheral blood mononuclear cells were isolated at the same time points and stained with fluorescent probes to identify intracellular reactive oxygen species and mitochondrial membrane potential (Δψm) by flow cytometry. Lipid peroxidation was assessed by plasma F2-isoprosatane concentration. Both IS and IS?+?C were associated with increased plasma F2-isoprostanes concentrations post-infusion. Maximal plasma F2-isoprostane concentrations after IS?+?C were significantly elevated from baseline (234?±?0.04 vs. 0.198?±?0.028?ng/mL, p?=?0.02). After IS?+?C, IL-1, IL-6, IL-10, and TNF-alpha were significantly elevated compared to baseline. After IS alone only IL-6 was noted to be elevated. Intracellular production of H(2)O(2) and loss of mitochondrial membrane potential (Δψm) was observed after IS while IS?+?C was associated with increased O (2) (·-) production. Both IS and IS?+?C induced serum cytokine activation accompanied by lipid peroxidation, however, IS?+?C induced higher plasma concentrations of F2-isoprostanes, IL-1, IL-10, and TNF-α post-infusion. Long-term safety studies of IV iron co-administered with Vitamin C are warranted.     

Resolvin E1 and its precursor 18R-HEPE restore mitochondrial function in inflammation

Inflammatory disorders such as sepsis are a major cause of morbidity and mortality. Mitochondrial dysfunction is considered a key factor in the pathogenesis of severe inflammation. In the present study, we aimed to investigate the impact of arachidonic acid, omega-3 (n-3) fatty acids, and n-3-derived lipid mediators 18R-HEPE and resolvin (Rv) E1 on mitochondrial function in experimental inflammation. The results revealed that, in contrast to n-6 and n-3 fatty acids, both 18R-HEPE and RvE1 possess anti-inflammatory and anti-apoptotic properties. Both mediators are able to restore inflammation-induced mitochondrial dysfunction, which is characterized by a decrease in mitochondrial respiration and membrane potential, as well as an imbalance of mitochondrial fission and fusion. Furthermore, inhibition of mitochondrial fission by Mdivi-1 and Dynasore reduces levels of the pro-inflammatory cytokines IL-6 and IL-8. These results suggest a novel functional mechanism for the beneficial effects of RvE1 in inflammatory reactions.     

The effects of highly active antiretroviral therapy on the serum levels of pro-inflammatory and anti-inflammatory cytokines in HIV infected subjects

Background

Cytokines play an important role in controlling the homeostasis of the immune system and infection with Human Immunodeficiency virus (HIV) leads to deregulated production of both pro- and anti-inflammatory cytokines. This study was designed to determine the effects of HIV and Highly Active Antiretroviral Therapy (HAART) on the levels of pro-and anti-inflammatory cytokines in HIV infected subjects.

Method

A total of 50 HIV infected and 50 HIV seronegative control participants were recruited for the study. The HIV infected subjects were recruited before commencement of antiretroviral therapy and were followed up for 12?months. Blood samples were collected at 3 different points: before initiation of therapy, 6?months into therapy and 12?months into therapy. Serum cytokines were analyzed using ELISA method while CD4+ T cells and viral load counts were measured using standard laboratory methods.

Result

The results showed that pro-inflammatory cytokines: Tumour necrosis factor-alpha (TNF-α), Interleukin-6 (IL-6) and anti-inflammatory cytokines Interleukin-4 (IL-4), Interleukin-10 (IL-10) and Transforming growth factor-beta (TGF-β) were significantly elevated in HIV infected subjects before commencement of therapy compared to 6?months and 12?months into therapy (P?<?0.01) and compared to control participants (P?<?0.01). TNF-α, TGF-beta remained significantly elevated even after 12?months of therapy compared to control participants (P?<?0.01), while IL-4, IL-6, and IL-10 showed no significant difference compared to control participants after 12?months of therapy (P?>?0.05). INF-γ was significantly reduced before commencement of therapy and after 12?months of therapy compared to control participants (P?<?0.05) respectively.

Conclusion

TNF-α and TGF-β remained significantly elevated even after 12?months of therapy, while IFN-γ remained significantly reduced after 12?months of therapy. Regulating these cytokines which were unresponsive to therapy could serve as a potential measure of therapy for HIV infected subjects. The positive effect of 12?months therapy on IL-4, IL-6 and IL-10 levels can be used to monitor disease prognosis during therapy especially in resource poor setting where regular viral load monitoring is unavailable.

     

Mitochondrial Oscillations and Waves in Cardiac Myocytes: Insights from Computational Models

Periodic cellwide depolarizations of mitochondrial membrane potential (Ψ_M) which are triggered by reactive oxygen species (ROS) and propagated by ROS-induced ROS release (RIRR) have been postulated to contribute to cardiac arrhythmogenesis and injury during ischemia/reperfusion. Two different modes of RIRR have been described: Ψ_M oscillations involving ROS-sensitive mitochondrial inner membrane anion channels (IMAC), and slow depolarization waves related to mitochondrial permeability transition pore (MPTP) opening. In this study, we developed a computational model of mitochondria exhibiting both IMAC-mediated RIRR and MPTP-mediated RIRR, diffusively coupled in a spatially extended network, to study the spatiotemporal dynamics of RIRR on Ψ_M. Our major findings are: 1), as the rate of ROS production increases, mitochondria can exhibit either oscillatory dynamics facilitated by IMAC opening, or bistable dynamics facilitated by MPTP opening; 2), in a diffusively-coupled mitochondrial network, the oscillatory dynamics of IMAC-mediated RIRR results in rapidly propagating (∼25 μm/s) cellwide Ψ_M oscillations, whereas the bistable dynamics of MPTP-mediated RIRR results in slow (0.1-2 μm/s) Ψ_M depolarization waves; and 3), the slow velocity of the MPTP-mediated depolarization wave is related to competition between ROS scavenging systems and ROS diffusion. Our observations provide mechanistic insights into the spatiotemporal dynamics underlying RIRR-induced Ψ_M oscillations and waves observed experimentally in cardiac myocytes.     

The expression analysis of inflammatory and antimicrobial genes in the goldfish (Carassius auratus L.) infected with Trypanosoma carassii

We report results of a comprehensive analysis of inflammatory gene expression during the course of infection of Trypanosoma carassii in the goldfish. We observed significant increases in mRNA levels of genes encoding pro-inflammatory cytokines IFN-γ, TNFα1 and TNFα2; IL-1β-1 and IL-1β-2; IL-12-p35 and IL-12-p40; CCL1; CXCL8, anti-inflammatory cytokines IL-10 and TGFβ and iNOS A and iNOS B, using quantitative PCR. Expression levels and profiles of these cytokines and iNOS isoforms varied in the different tissues (kidney, spleen, liver) of goldfish during the course of T.?carassii infection. The expression of majority of genes that encode pro- and anti-inflammatory cytokines were up-regulated during the acute phase of infection (days 7-21 post-infection). The mRNA levels of these cytokines returned to normal levels or were down-regulated during the elimination phase of infection (days 28-56), with exception of IL-10 in the spleen and liver of infected fish. A parallel up-regulation of IFN-γ and IL-10 mRNA levels were observed in all tissues of infected fish during the acute phase of the infection. The expression of iNOS genes (iNOS A and B) was significantly delayed (day 14?pi) in the kidney, liver and spleen of infected fish. These results provide insights into the interaction between T.?carassii and goldfish, and suggest that Th1/Th2-like responses may be important for controlling T.?carassii infection in the goldfish.