Poly(ADP-ribose) polymerase-1 inhibition protects the brain against systemic inflammation

Poly(ADP-ribose) polymerase-1 (PARP-1) is involved in DNA repair, but its overactivation can induce cell death. Our aim was to investigate the role of PARP-1 in activation of programmed cell death processes in the brain during systemic inflammation.Our data indicated that lipopolysaccharide (1 mg/kg b.w., i.p.)-evoked systemic inflammation enhanced PARP-1 activity in the mouse brain, leading to the lowering of β-NAD⁺ concentration, to translocation of apoptosis inducing factor from mitochondria to the nucleus, and to enhanced lipid peroxidation. Inhibitor of PARP-1, 3-aminobenzamide (30 mg/kg b.w., i.p.), protected the brain against prooxidative and cell death processes, suggesting involvement of PARP-1 in systemic inflammation-related processes in the brain.     

Poly(ADP-ribose) polymerase-1 inhibition protects the brain against systemic inflammation

Poly(ADP-ribose) polymerase-1 (PARP-1) is involved in DNA repair, but its overactivation can induce cell death. Our aim was to investigate the role of PARP-1 in activation of programmed cell death processes in the brain during systemic inflammation.

Our data indicated that lipopolysaccharide (1 mg/kg b.w., i.p.)-evoked systemic inflammation enhanced PARP-1 activity in the mouse brain, leading to the lowering of β-NAD⁺ concentration, to translocation of apoptosis inducing factor from mitochondria to the nucleus, and to enhanced lipid peroxidation. Inhibitor of PARP-1, 3-aminobenzamide (30 mg/kg b.w., i.p.), protected the brain against prooxidative and cell death processes, suggesting involvement of PARP-1 in systemic inflammation-related processes in the brain.     

Poly(ADP-ribose) polymerase-1 is a key mediator of cisplatin-induced kidney inflammation and injury

Cisplatin is a commonly used chemotherapeutic drug, the clinical use of which is limited by the development of dose-dependent nephrotoxicity. Enhanced inflammatory response, oxidative stress, and cell death have been implicated in the development of cisplatin-induced nephropathy; however, the precise mechanisms are elusive. Overactivation of the nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) by oxidative DNA damage under various pathological conditions promotes cell death and up-regulation of key proinflammatory pathways. In this study, using a well-established model of nephropathy, we have explored the role of PARP-1 in cisplatin-induced kidney injury. Genetic deletion or pharmacological inhibition of PARP-1 markedly attenuated the cisplatin-induced histopathological damage, impaired renal function (elevated serum BUN and creatinine levels), and enhanced inflammatory response (leukocyte infiltration; TNF-α, IL-1β, F4/80, adhesion molecules ICAM-1/VCAM-1 expression) and consequent oxidative/nitrative stress (4-HNE, 8-OHdG, and nitrotyrosine content; NOX2/NOX4 expression). PARP inhibition also facilitated the cisplatin-induced death of cancer cells. Thus, PARP activation plays an important role in cisplatin-induced kidney injury, and its pharmacological inhibition may represent a promising approach to preventing the cisplatin-induced nephropathy. This is particularly exciting because several PARP inhibitors alone or in combination with DNA-damaging anticancer agents show considerable promise in clinical trials for treatment of various malignancies (e.g., triple-negative breast cancer).     

Therapeutic effect of a poly(ADP-ribose) polymerase-1 inhibitor on experimental arthritis by downregulating inflammation and Th1 response

Poly(ADP-ribose) polymerase-1 (PARP-1) synthesizes and transfers ADP ribose polymers to target proteins, and regulates DNA repair and genomic integrity maintenance. PARP-1 also plays a crucial role in the progression of the inflammatory response, and its inhibition confers protection in several models of inflammatory disorders. Here, we investigate the impact of a selective PARP-1 inhibitor in experimental arthritis. PARP-1 inhibition with 5-aminoisoquinolinone (AIQ) significantly reduces incidence and severity of established collagen-induced arthritis, completely abrogating joint swelling and destruction of cartilage and bone. The therapeutic effect of AIQ is associated with a striking reduction of the two deleterious components of the disease, i.e. the Th1-driven autoimmune and inflammatory responses. AIQ downregulates the production of various inflammatory cytokines and chemokines, decreases the antigen-specific Th1-cell expansion, and induces the production of the anti-inflammatory cytokine IL-10. Our results provide evidence of the contribution of PARP-1 to the progression of arthritis and identify this protein as a potential therapeutic target for the treatment of rheumatoid arthritis.     

Autophagy induces eosinophil extracellular traps formation and allergic airway inflammation in a murine asthma model

Studies have shown autophagy participation in the immunopathology of inflammatory diseases. However, autophagy role in asthma and in eosinophil extracellular traps (EETs) release is poorly understood. Here, we attempted to investigate the autophagy involvement in EETs release and in lung inflammation in an experimental asthma model. Mice were sensitized with ovalbumin (OVA), followed by OVA challenge. Before the challenge with OVA, mice were treated with an autophagy inhibitor, 3-methyladenine (3-MA). We showed that 3-MA treatment decreases the number of eosinophils, eosinophil peroxidase (EPO) activity, goblet cells hyperplasia, proinflammatory cytokines, and nuclear factor kappa B (NFκB) p65 immunocontent in the lung. Moreover, 3-MA was able to improve oxidative stress, mitochondrial energy metabolism, and Na⁺, K⁺-ATPase activity. We demonstrated that treatment with autophagy inhibitor 3-MA reduced EETs formation in the airway. On the basis of our results, 3-MA treatment can be an interesting alternative for reducing lung inflammation, oxidative stress, mitochondrial damage, and EETs formation in asthma.     

Poly(ADP-ribose) polymerase-1 inhibition: preclinical and clinical development of synthetic lethality

The hereditary forms of breast cancer identified by BRCA1 and BRCA2 genes have a defect in homologous DNA repair and demonstrate a dependence on alternate DNA repair processes by base excision repair, which requires poly(ADP-ribose) polymerase 1 (PARP-1). siRNA and deletion mutations demonstrate that interference with PARP-1 function results in enhanced cell death when the malignancy has a defect in homologous recombination. These findings resulted in a plethora of agents in clinical trials that interfere with DNA repair, and these agents offer the potential of being more selective in their effects than classic chemotherapeutic drugs. An electronic search of the National Library of Medicine for published articles written in English used the terms "PARP inhibitors" and "breast cancer" to find prospective, retrospective and review articles. Additional searches were done for articles dealing with mechanism of action. A total of 152 articles dealing with breast cancer and PARP inhibition were identified. PARP inhibition not only affects nonhomologous repair, but also has several other nongenomic functions. Mutational resistance to these agents was seen in preclinical studies. To date, PARP-1 inhibitors were shown to enhance cytotoxic effects of some chemotherapy agents. This new class of agents may offer more therapeutic specificity by exploiting a DNA repair defect seen in some human tumors with initial clinical trials demonstrating antitumor activity. Although PARP inhibitors may offer a therapeutic option for selected malignancies, the long-term effects of these agents have not yet been defined.     

Poly(ADP-ribose) polymerase-1 modulates DNA repair capacity and prevents formation of DNA double strand breaks

Although poly(ADP-ribose) polymerase-1 (PARP-1) has no enzymatic activity involved in DNA damage processing by the base excision repair (BER) pathway, PARP-1 deficient cells are genetically unstable and sensitive to DNA-damaging agents. To explain this paradox, we investigated the impact of PARP-1 on BER in mammalian cells. We reduced cellular PARP-1 protein levels using siRNA, then introduced DNA damage by hydrogen peroxide treatment and examined the repair response. We find that PARP-1 is not involved in recruitment of the major BER proteins to sites of DNA damage. However, we find that PARP-1 protects excessive DNA single strand breaks (SSBs) from converting into DNA double strand breaks (DSBs) thus preserving them for subsequent repair by BER enzymes. This suggests that PARP-1 plays an important role in BER by extending the ability of BER enzymes to process DNA single strand breaks arising directly after mutagen stress or during processing of DNA lesions following extensive DNA damage.     

IFN-gamma-inducing factor (IL-18) increases allergic sensitization, serum IgE, Th2 cytokines, and airway eosinophilia in a mouse model of allergic asthma

We investigated the effects of IFN-gamma-inducing factor (IL-18) in a ragweed (RW) mouse model of allergic asthma. Administration of IL-18 in conjunction with allergic sensitization and challenge in wild-type, but not IFN-gamma -/- mice, inhibited the bronchoalveolar lavage (BAL) eosinophilia induced by RW challenge, and increased serum levels of RW-specific IgG2a and production of IFN-gamma from splenocytes cultured with RW, indicating a critical role for IFN-gamma in mediating these effects. Paradoxically, the same treatment schedule in WT mice increased serum levels of RW-specific IgE and IgG1, and production of IL-4 and IL-5 from splenocytes cultured with RW. When the effects of the same IL-18 treatment schedule were allowed to mature for 3 wk, the inhibition of lung eosinophil recruitment was replaced by augmentation of lung eosinophil recruitment. In another experiment, IL-18 administered only with allergic sensitization increased BAL eosinophilia and lung expression of IL-5 and IFN-gamma, while IL-18 administered only with RW challenge decreased BAL eosinophilia and increased lung IFN-gamma expression, while lung expression of IL-5 remained unchanged. IL-18 administered without RW or adjuvant to naive mice increased total serum IgE levels. Finally, intrapulmonary administrations of IL-18 plus RW in naive mice dramatically increased Th2 cytokine production, IgE levels, eosinophil recruitment, and airway mucus, demonstrating induction of allergic sensitization. This is the first report demonstrating that IL-18 promotes a Th2 phenotype in vivo, and potently induces allergic sensitization. These results suggest that IL-18 may contribute to the pathogenesis of allergic asthma.     

Poly(ADP-ribose) polymerase-2 is a lipid-modulated modulator of muscular lipid homeostasis

There is a growing body of evidence that poly(ADP-ribose) polymerase-2 (PARP2), although originally described as a DNA repair protein, has a widespread role as a metabolic regulator. We show that the ablation of PARP2 induced characteristic changes in the lipidome. The silencing of PARP2 induced the expression of sterol regulatory element-binding protein-1 and -2 and initiated de novo cholesterol biosynthesis in skeletal muscle. Increased muscular cholesterol was shunted to muscular biosynthesis of dihydrotestosterone, an anabolic steroid. Thus, skeletal muscle fibers in PARP2^?/? mice were stronger compared to those of their wild-type littermates. In addition, we detected changes in the dynamics of the cell membrane, suggesting that lipidome changes also affect the biophysical characteristics of the cell membrane. In in silico and wet chemistry studies, we identified lipid species that can decrease the expression of PARP2 and potentially phenocopy the genetic abruption of PARP2, including artificial steroids. In view of these observations, we propose a new role for PARP2 as a lipid-modulated regulator of lipid metabolism.     

Poly(ADP-ribose) polymerase-1 dimerizes at a 5' recessed DNA end in vitro: a fluorescence study

Activation of poly(ADP-ribose) polymerase-1 (PARP-1) is an immediate cellular reaction to DNA strand breakage as induced by alkylating agents, ionizing radiation, or oxidants. The resulting formation of protein-bound poly(ADP-ribose) facilitates survival of proliferating cells under conditions of DNA damage probably via its contribution to DNA base excision repair. In this study, we investigated the association of the amino-terminal DNA binding domain of human PARP-1 (hPARP-1 DBD) with a 5' recessed oligonucleotide mimicking a telomeric DNA end. We used the fluorescence of the Trp residues naturally occurring in the zinc finger domain of hPARP-1 DBD. Fluorescence intensity and fluorescence anisotropy measurements consistently show that the binding stoichiometry is two proteins per DNA molecule. hPARP-1 was found to bind the 5' recessed DNA end with a binding constant of approximately 10(14) M(-2) if a cooperative binding model is assumed. These results indicate that hPARP-1 DBD dimerizes during binding to the DNA target site. A footprint experiment shows that hPARP-1 DBD is asymmetrically positioned at the junction between the double-stranded and the single-stranded telomeric repeat. The largest contribution to the stability of the complex is given by nonionic interactions. Moreover, time-resolved fluorescence measurements are in line with the involvement of one Trp residue in the stacking interaction with DNA bases. Taken together, our data open new perspectives for interpretation of the selective binding of hPARP-1 to the junction between double- and single-stranded DNA.     

IL-27 suppresses Th2 cell development and Th2 cytokines production from polarized Th2 cells: a novel therapeutic way for Th2-mediated allergic inflammation

IL-27 up-regulates Th1 but down-regulates Th2 responses. However, its molecular mechanism and regulatory effects on polarized Th2 cells remain unclear. In this study, we have revealed that IL-27 inhibits Th2 cell development as well as Th2 cytokines production from already polarized Th2 cells by down-regulation of GATA-3 and up-regulation of T-bet expression simultaneously. In vivo daily IL-27 treatment for 1 wk after Leishmania major infection protects BALB/c mice from footpad swelling by diminishing parasite burden via reciprocal regulation of Th1 and Th2 responses. Furthermore, IL-27 stimulation causes marked reduction in the capacity of host mouse to mount a Th2 response against Strongyloides venezuelensis infection. Thus, IL-27-treated mice failed to develop intestinal mastocytosis after S. venezuelensis infection and exhibited a marked delay in parasite expulsion. Finally, intranasal administration of IL-27 inhibits OVA-induced airway hyperresponsiveness and inflammation in OVA-sensitized animals. Thus, IL-27 could provide us with a novel therapeutic way for treating Th2-associated diseases such as bronchial asthma.     

The IL-3/IL-5/GM-CSF common receptor plays a pivotal role in the regulation of Th2 immunity and allergic airway inflammation

The eosinophil is a central effector cell in allergic asthma. Differentiation and function of eosinophils are regulated by the CD4 Th2 cytokines IL-3, IL-5, and GM-CSF, which all signal through a common beta receptor subunit (betac). Recent therapeutic approaches targeting IL-5 alone have not ablated tissue accumulation of eosinophils and have had limited effects on disease progression, suggesting important roles for IL-3 and GM-CSF. By using a mouse model of allergic airways inflammation, we show that allergen-induced expansion and accumulation of eosinophils in the lung are abolished in betac-deficient (betac-/-) mice. Moreover, betac deficiency resulted in inhibition of hallmark features of asthma, including airways hypersensitivity, mucus hypersecretion, and production of Ag-specific IgE. Surprisingly, we also identified a critical role for this receptor in regulating type 2 immunity. Th2 cells in the lung of allergen-challenged betac-/- mice were limited in their ability to proliferate, produce cytokines, and migrate to effector sites, which was attributed to reduced numbers of myeloid dendritic cells in the lung compartment. Thus, the betac plays a critical role in allergen-induced eosinophil expansion and infiltration and is pivotal in regulating molecules that promote both early and late phases of allergic inflammation, representing a novel target for therapy.     

Eosinophilopoiesis in a murine model of allergic airway eosinophilia: involvement of bone marrow IL-5 and IL-5 receptor alpha

The airway inflammation in asthma is dominated by eosinophils. The aim of this study was to elucidate the contribution of newly produced eosinophils in airway allergic inflammation and to determine mechanisms of any enhanced eosinophilopoiesis. OVA-sensitized BALB/c mice were repeatedly exposed to allergen via airway route. Newly produced cells were identified using a thymidine analog, 5-bromo-2'-deoxyuridine, which is incorporated into DNA during mitosis. Identification of IL-5-producing cells in the bone marrow was performed using FACS. Bone marrow CD3+ cells were enriched to evaluate IL-5-protein release in vitro. Anti-IL-5-treatment (TRFK-5) was given either systemically or directly to the airways. IL-5R-bearing cells were localized by immunocytochemistry. Repeated airway allergen exposure caused prominent airway eosinophilia after three to five exposures, and increased the number of immature eosinophils in the bone marrow. Up to 78% of bronchoalveolar lavage (BAL) granulocytes were 5-bromo-2'-deoxyuridine positive. After three allergen exposures, both CD3+ and non-CD3 cells acquired from the bone marrow expressed and released IL-5-protein. Anti-IL-5 given i.p. inhibited both bone marrow and airway eosinophilia. Intranasal administration of anti-IL-5 also reduced BAL eosinophilia, partly via local effects in the airways. Bone marrow cells, but not BAL eosinophils, displayed stainable amounts of the IL-5R alpha-chain. We conclude that the bone marrow is activated by airway allergen exposure, and that newly produced eosinophils contribute to a substantial degree to the airway eosinophilia induced by allergen. Airway allergen exposure increases the number of cells expressing IL-5-protein in the bone marrow. The bone marrow, as well as the lung, are possible targets for anti-IL-5-treatment.     

Inducible costimulator regulates Th2-mediated inflammation, but not Th2 differentiation, in a model of allergic airway disease

A novel costimulatory molecule expressed on activated T cells, inducible costimulator (ICOS), and its ligand, B7-related protein-1 (B7RP-1), were recently identified. ICOS costimulation leads to the induction of Th2 cytokines without augmentation of IL-2 production, suggesting a role for ICOS in Th2 cell differentiation and expansion. In the present study, a soluble form of murine ICOS, ICOS-Ig, was used to block ICOS/B7RP-1 interactions in a Th2 model of allergic airway disease. In this model, mice are sensitized with inactivated Schistosoma mansoni eggs and are subsequently challenged with soluble S. mansoni egg Ag directly in the airways. Treatment of C57BL/6 mice with ICOS-Ig during sensitization and challenge attenuated airway inflammation, as demonstrated by a decrease in cellular infiltration into the lung tissue and airways, as well as by a decrease in local IL-5 production. These inhibitory effects were not due to a lack of T cell priming nor to a defect in Th2 differentiation. In addition, blockade of ICOS/B7RP-1 interactions during ex vivo restimulation of lung Th2 effector cells prevented cytokine production. Thus, blockade of ICOS signaling can significantly reduce airway inflammation without affecting Th2 differentiation in this model of allergic airway disease.     

Poly(ADP-ribose) polymerase inhibition prevents both apoptotic-like delayed neuronal death and necrosis after H(2)O(2) injury

Toxic reactive oxygen species (ROS) such as hydrogen peroxide, nitric oxide, superoxide, and the hydroxyl radical are generated in a variety of neuropathological conditions and cause significant DNA damage. We determined the effects of 3-aminobenzamide (AB), an inhibitor of the DNA repair enzyme poly(ADP-ribose) polymerase (PARP), on cell death in differentiated PC12 cells, a model of sympathetic neurons, after H(2) O(2) injury. Exposure to 0.5 mm H(2) O(2) resulted in a significant decrease in intracellular NAD(H), NADP(H), and ATP levels. This injury resulted in the death of 90% of the cells with significant necrosis early (2 h) after injury and increased apoptosis (12-24 h after injury), as measured by PS exposure and the presence of cytoplasmic oligonucleosomal fragments. Treatment with 2.5 mm AB restored pyridine nucleotide and ATP levels and ameliorated cell death (65% versus 90%) by decreasing the extent of both necrosis and apoptosis. Interestingly, we observed that H(2) O(2) -induced injury caused a delayed cell death exhibiting features of apoptosis but in which caspase-3 like activity was absent. Moreover, pretreatment with AB restored caspase-3-like activity. Our results suggest that apoptosis and necrosis are both triggered by PARP overactivation, and that maintenance of cellular energy levels after injury by inhibiting PARP shifts cell death from necrosis to apoptosis.     

Poly(ADP-ribose) polymerase-1 is a survival factor for radiation-exposed intestinal epithelial stem cells in vivo

Poly(ADP-ribose) polymerase-1 (PARP-1) is a key enzyme mediating the cellular response to DNA strand breaks. It plays a critical role in genomic stability and survival of proliferating cells in culture undergoing DNA damage. Intestinal epithelium is the most proliferative tissue in the mammalian body and its stem cells show extreme sensitivity to low-level genotoxic stress. We investigated the role of PARP-1 in the in vivo damage response of intestinal stem cells in crypts of PARP-1^–/– and control mice following whole-body γ-irradiation (1 Gy). In the PARP-1^–/– mice there was a significant delay during the first 6 h in the transient p53 accumulation in stem cells whereas an increased number of cells were positive for p21^CIP1/WAF1. Either no or only marginal differences were noted in MDM2 expression, apoptosis, induction of or recovery from mitotic blockage, or inhibition of DNA synthesis. We further observed a dose-dependent reduction in crypt survival measured at 4 days post-irradiation in control mice, and this crypt-killing effect was significantly potentiated in PARP-1^–/– mice. Our results thus establish that PARP-1 acts as a survival factor for intestinal stem cells in vivo and suggest a functional link with early p53 and p21^CIP1/WAF1 responses.