Complement Propriety and Conspiracy in Nanomedicine: Perspective and a Hypothesis.

The complement system is the first line of body's defense against intruders and it acts as a functional bridge between innate and adaptive arms of the immune system. This commentary examines the key roles of complement activation in response to nanomedicine administration, including nucleic acid complexes. These comprise beneficial (eg, adjuvanticity) as well as adverse effects (eg, infusion-related reactions). Pigs (and sheep) are often used as predictive models of nanomedicine-mediated infusion-related reactions in humans. The validity of these models in relation to human responses is questioned, and an alternative hypothesis is presented.

Pathophysiology and treatment of typical and atypical hemolytic uremic syndrome.

Hemolytic uremic syndrome is a rare disease, frequently responsible for renal insufficiency in children. Recent findings have led to renewed interest in this pathology. The discovery of new gene mutations in the atypical form of HUS and the experimental data suggesting the involvement of the complement pathway in the typical form, open new perspectives for treatment. This review summarizes the current state of knowledge on both typical and atypical hemolytic uremic syndrome pathophysiology and examines new perspectives for treatment.

Boosting ADCC and CDC activity by Fc engineering and evaluation of antibody effector functions.

In recent years, therapy with monoclonal antibodies has become standard of care in various clinical applications. Despite obvious clinical activity, not all patients respond and benefit from this generally well tolerated treatment option. Therefore, rational optimization of antibody therapy represents a major area of interest in translational research. Animal models and clinical data suggested important roles of Fc-mediated effector mechanisms such as antibody dependent cell-mediated cytotoxicity (ADCC) or complement dependent cytotoxicity (CDC) in antibody therapy. These novel insights into the mechanisms of action mediated by monoclonal antibodies inspired the development of different engineering approaches to enhance/optimize antibodies' effector functions. Fc-engineering approaches by altering the Fc-bound glycosylation profile or by exchanging amino acids in the protein backbone have been intensively studied. Here, advanced and emerging technologies in Fc-engineering resulting in altered ADCC and CDC activity are summarized and experimental strategies to evaluate antibodies' effector functions are discussed.

Role of complement in a murine model of peanut-induced anaphylaxis.

Peanut allergy is severe and persisting from childhood to adulthood. However, there is no effective prophylaxis or treatment for peanut allergy. Little is known to about the molecular process in the pathogenesis of peanuts allergy, especially in innate immunity. Thus we investigated the role of complement activation in murine peanut anaphylaxis. Complement component C3 deposition on peanut extract (PE) was evaluated using sera from wild-type (WT), mannose-binding lectin associated serine protease (MASP)-1/3 deficient, MASP-2 deficient, and C4 deficient mice. Sera from interferon regulatory factor-4 (IRF-4) deficient mice, which lack serum immunoglobulin, were also used. In anaphylaxis study, mice were pretreated with propranolol and a long-acting form of IL-4, and injected with PE. Mice were then assessed for plasma C3a levels and hypothermia shock by ELISA and rectal temperature measurement, respectively. C3 deposition on PE was abolished in immunoglobulin- and C4-deficient sera. No difference in C3 deposition levels were observed among WT, MASP-1/3 deficient and MASP-2 deficient sera. IgM, IgG2b, IgG3, C1q, and ficolin-A deposits were detected on PE. In anaphylaxis study, MASP-1/3 deficient mice showed elevation of plasma C3a levels similar to WT mice. However, they were significantly reduced in C4- and MASP-2-deficient mice compared to WT mice. Consistently, PE-induced anaphylactic shock was prevented in C4 deficient mice and partially in MASP-2 deficient mice. In conclusion, PE activates complement via both the lectin and classical pathways in vivo, and the complement activation contributes to hypothermia shock in mice.

The complement receptor 2/factor H fusion protein TT30 protects paroxysmal nocturnal hemoglobinuria erythrocytes from complement-mediated hemolysis and C3 fragment.

Paroxysmal nocturnal hemoglobinuria (PNH) is characterized by complement-mediated intravascular hemolysis because of the lack from erythrocyte surface of the complement regulators CD55 and CD59, with subsequent uncontrolled continuous spontaneous activation of the complement alternative pathway (CAP), and at times of the complement classic pathway. Here we investigate in an in vitro model the effect on PNH erythrocytes of a novel therapeutic strategy for membrane-targeted delivery of a CAP inhibitor. TT30 is a 65 kDa recombinant human fusion protein consisting of the iC3b/C3d-binding region of complement receptor 2 (CR2) and the inhibitory domain of the CAP regulator factor H (fH). TT30 completely inhibits in a dose-dependent manner hemolysis of PNH erythrocytes in a modified extended acidified serum assay, and also prevents C3 fragment deposition on surviving PNH erythrocytes. The efficacy of TT30 derives from its direct binding to PNH erythrocytes; if binding to the erythrocytes is disrupted, only partial inhibition of hemolysis is mediated by TT30 in solution, which is similar to that produced by the fH moiety of TT30 alone, or by intact human fH. TT30 is a membrane-targeted selective CAP inhibitor that may prevent both intravascular and C3-mediated extravascular hemolysis of PNH erythrocytes and warrants consideration for the treatment of PNH patients.

Crocodylus siamensis serum and macrophage phagocytic activity.

Antimicrobial activity of sera from many crocodilian species has been recognized. This activity was proposed to be mediated, at least in part, by complement. Due to the fact that complement proteins have different functions in the immune system, they may be involved in phagocytic process of phagocytes. In the present study, the effects of Siamese crocodile serum on phagocytic activity of macrophages as well as the possible involvement of complement in this process were examined. The results showed increases in the phagocytosis of both Escherichia coli and to a lesser extent, Staphylococcus aureus upon incubation of murine macrophage cell line with fresh crocodile serum (FS). Similar to FS, other crocodile blood products, including freeze dried serum (DS) and freeze dried whole blood (DWB) exhibited phagocytosis-enhancing property. However the ability of DWB to enhance phagocytosis was less efficient than that of FS and DS, suggesting that serum factors were involved in this process. Treatment of FS with heat at 56 degrees C for 30 min deteriorated the effect of FS on bacterial uptake of macrophages, suggesting that complement proteins play a role in the modulation of the phagocytic process. Collectively, the results of the present study suggested that crocodile serum enhances the macrophage phagocytic activity through complement activity and, therefore, may be taken as an alternative medicine for supporting the human immune responses.

The complement cascade: Yin-Yang in neuroinflammation--neuro-protection and -degeneration.

The complement cascade has long been recognized to play a key role in inflammatory and degenerative diseases. It is a 'double edged' sword as it is necessary to maintain health, yet can have adverse effects when unregulated, often exacerbating disease. The contrasting effects of complement, depending on whether in a setting of health or disease, is the price paid to achieve flexibility in scope and degree of a protective response for the host from infection and injury. Loss or even decreased efficiency of critical regulatory control mechanisms can result in aggravated inflammation and destruction of self-tissue. The role of the complement cascade is poorly understood in the nervous system and neurological disorders. Novel studies have demonstrated that the expression of complement proteins in brain varies in different cell types and the effects of complement activation in various disease settings appear to differ. Understanding the functioning of this cascade is essential, as it has therapeutic implications. In this review, we will attempt to provide insight into how this complex cascade functions and to identify potential strategic targets for therapeutic intervention in chronic diseases as well as acute injury in the CNS.

Complement-dependent tumor cell lysis triggered by combinations of epidermal growth factor receptor antibodies.

Therapeutic monoclonal antibodies against the epidermal growth factor receptor (EGFR) have advanced the treatment of colon and head and neck cancer, and show great promise for the development of treatments for other solid cancers. Antibodies against EGFR have been shown to act via inhibition of receptor signaling and induction of antibody-dependent cellular cytoxicity. However, complement-dependent cytotoxicity, which is considered one of the most powerful cell killing mechanisms of antibodies, seems inactive for such antibodies. Here, we show a remarkable synergy for EGFR antibodies. Combinations of antibodies against EGFR were identified, which resulted in potent complement activation via the classic pathway and effective lysis of tumor cells. Studies on a large panel of antibodies indicated that the observed synergy is a general mechanism, which can be activated by combining human IgG1 antibodies recognizing different, nonoverlapping epitopes. Our findings show an unexpected quality of therapeutic EGFR antibodies, which may be exploited to develop novel and more effective treatments for solid cancers.

An in vitro examination of the antioxidant and anti-inflammatory properties of buckwheat honey.

OBJECTIVE: Hydroxyl radical and hypochlorite anion formed at the wound site from superoxide anion produced by activated polymorphonuclear neutrophils (PMNs) are considered important factors in impaired wound healing. Superoxide anion may also react with nitric oxide produced by macrophages to form peroxynitrite, a third strong oxidant that damages surrounding tissue. In order to select honey for use in wound-healing products, different samples were compared for their capacity to reduce levels of reactive oxygen species (ROS) in vitro. METHOD: Honey samples were tested in assays for inhibition of ROS production by activated human PMNs, antioxidant activity (scavenging of superoxide anion in a cell-free system) and inhibition of human complement (reducing levels of ROS by limiting formation of complement factors that attract and stimulate PMNs). For buckwheat honey (NewYork, US), moisture and free acid content were determined by refractive index measurement and potentiometric titration respectively. Honey constituents other than sugars were investigated by thin layer chromatography, using natural product reagent to detect phenolic compounds. Constituents with antioxidant properties were detected by spraying the chromatogram with DPPH. RESULTS: Although most honey samples were shown to be active, significant differences were observed, with the highly active honey exceeding the activities of samples with minor effects by factors of 4 to 30. Most pronounced activities were found for American buckwheat honey from the state of NewYork. Phenolic constituents of buckwheat honey were shown to have antioxidant activity. CONCLUSION: As buckwheat honey was most effective in reducing ROS levels, it was selected for use in wound-healing products. The major antioxidant properties in buckwheat honey derive from its phenolic constituents, which are present in relatively large amounts. Its phenolic compounds may also exert antibacterial activity, whereas its low pH and high free acid content may assist wound healing.

Interleukin-4 induces lipogenesis in porcine endothelial cells, which in turn is critical for induction of protection against complement-mediated injury.

Interleukin (IL)-4 has been shown to induce protection in porcine vascular endothelial cells (ECs) from killing by human complement. This protection is dependent on the PI3K/Akt signaling pathway. In this study, we investigated mechanisms downstream of Akt and found that activation of the lipid biosynthesis pathway is required for protection from complement in ECs treated with IL-4. Cells incubated with IL-4 for 48 hours contained increased fatty acids and phospholipids but cholesterol was not increased when compared with medium-treated controls. The transcription factor SREBP-1, which regulates fatty acid synthesis, was found to be activated in extracts of ECs incubated with IL-4 for 6 hours. Finally, induction of protection from complement killing with IL-4 was fully prevented by the presence of the SREBP inhibitor 25-OH cholesterol. This study showed that IL-4 induces lipid biosynthesis in porcine ECs through activation of SREBP-1 and that the activation of this pathway is critical for IL-4 to induce protection of porcine ECs from killing by human complement. Further study of these mechanisms may provide new strategies for the prevention of complement-mediated vascular injury as it occurs in xenograft rejection.

Comparison of the effectiveness of the traditional acupuncture point, ST. 36 and Omura's ST.36 Point (True ST. 36) needling on the isokinetic knee extension & flexion strength of young soccer players.

The purpose of this study was to compare the effects of the Traditional acupuncture point ST.36 and 'Omura's ST.36 Point' ("True ST.36") needling on the isokinetic knee extension & flexion strength of young soccer players. The Bi-Digital O-Ring Test (B.D.O.R.T.) of Yoshiaki Omura, M.D.,Sc.D. was used to determine the "True ST.36". Young soccer players (N = 24) between 16-18 years of age (Mean = 16.92 +/- 0.65) were involved in the study. The extension & flexion strengths of dominant legs were measured with Cybex 350 Extremity System isokinetically. The testing velocity was 60 degrees/sec. The peak torque value in Newton meters (Nm) was evaluated. Subjects were tested 3 times. Extension & Flexion 1 (EXT1, FLEX1) without acupuncture application, EXT2 & FLEX2 after application on the traditional acupuncture point, ST.36 and EXT3 & FLEX3 after application onto the 'Omura's New Foot-point' ("True ST.36"). Before each test, subjects warmed up for 10 minutes by cycling on an isokinetic ergometer at 50 RPM, 75 Watts load followed by stretching exercises of lower extremity. Mean EXT1, EXT2, EXT3 values were 196.92 +/- 28.70: 210.00 +/- 23.00; 224.42 +/- 21.70 respectively, where FLEX1, FLEX2, FLEX3 were 140.88 +/- 22.45; 151.13 +/- 21.27; 161.00 +/- 22.23. Comparisons of EXT1-EXT2, EXT1-EXT3, EXT2-EXT3, FLEX1-FLEX2, FLEX1-FLEX3, FLEX2-FLEX3 strength values showed all very high significance (P < 0.001) in favor of 1) Needling on relevant points and 2) Omura's ST.36 Point ("True ST.36"). We conclude that B.D.O.R.T. can help to determine new (True) Acupuncture points and, both points were effective for increasing the isokinetic knee extension & flexion strength of young soccer players very significantly where as Omura's ST.36 Point ("True ST.36") was more effective than Traditional Acupuncture point, ST.36.

Effect of Biophytum sensitivum on cell-mediated immune response in mice.

Effect of Biophytum sensitivum on cell-mediated immune response was studied in normal as well as Ehrlich ascites tumor bearing BALB/c mice. Administration of Biophytum sensitivum significantly enhanced the proliferation of splenocytes, thymocytes and bone marrow cells by stimulating the mitogenic potential of various mitogens such as Lipopolysaccharide (LPS), Concanavalin A (Con A), Phytohaemagglutinin (PHA) and Poke Weed Mitogen (PWM). Natural killer (NK) cell activity was enhanced significantly by Biophytum sensitivum in both the normal (43.6% cell lysis on day 5) and the tumor bearing group (48.2% cell lysis on day 5), and it was found to be earlier than tumor bearing control animals (maximum of 13.4% cell lysis on day 9). Antibody dependent cellular cytotoxicity (ADCC) was also enhanced significantly in both Biophytum treated normal (35% cell lysis on day 7) as well as tumor bearing animals (40.2% cell lysis on day 7) compared to untreated control tumor bearing animals (maximum of 12.3% cell lysis on day 11). An early antibody dependent complement mediated cytotoxicity (ACC) was also observed in the Biophytum treated normal (22.6% cell lysis, on day 15) and tumor bearing animals (26.4% cell lysis, on day 15). Results of our present study suggest the immunomodulatory property of Biophytum sensitivum.

Complement-targeted therapeutics.

The complement system is a central component of innate immunity and bridges the innate to the adaptive immune response. However, it can also turn its destructive capabilities against host cells and is involved in numerous diseases and pathological conditions. Modulation of the complement system has been recognized as a promising strategy in drug discovery, and a large number of therapeutic modalities have been developed. However, successful marketing of complement-targeted drugs has proved to be more difficult than initially expected, and many strategies have been discontinued. The US Food and Drug Administration's approval of the first complement-specific drug, an antibody against complement component C5 (eculizumab; Soliris), in March 2007, was a long-awaited breakthrough in the field. Approval of eculizumab validates the complement system as therapeutic target and might facilitate clinical development of other promising drug candidates.

Anti-complementary activity of ursane-type triterpenoids from Weigela subsessilis.

A new ursane-type triterpenoid, weigelic acid (1), and seven known compounds, ursolic acid (2), ilekudinol A (3), corosolic acid (4), ilekudinol B (5), esculentic acid (6), pomolic acid (7), and asiatic acid (8) were isolated from the leaf and stem of Weigela subsessilis. The structure of the new triterpenoid was established as 1beta,2alpha,3alpha,23-tetrahydroxyurs-12-en-28-oic acid on the basis of spectroscopic analyses. In addition, the isolated compounds were evaluated for their anti-complement activity against the classical pathway of the complement system. Of these, compounds 1-2 and 4-8 exhibited anti-complement activity with IC50 values of 152, 90, 130, 51, 56, 4, and 163 microM, respectively, whereas 3 was inactive. This shows that a carboxylic group of ursane-type triterpenoids seems to play an important role in inhibiting the hemolytic activity of human serum against erythrocytes.

C-reactive-protein-associated increase in myocardial infarct size after ischemia/reperfusion.

C-Reactive protein (CRP), a marker for acute inflammation, is associated with increased risk of cardiovascular events. The mechanism underlying this association is uncertain. An acute inflammatory response was induced in rabbits by subcutaneous injection of croton oil (CO) 1 to 3 days before 30 min of regional myocardial ischemia/180 min of reperfusion. CO treatment increased plasma CRP from below the limit of detection to 2.5 +/- 0.5 mg/dl and was associated with an increase in infarct size expressed as percentage of risk region [32 +/- 6% vehicle controls (n = 7) to 47 +/- 9% CO-treated rabbits (n = 7; P < 0.05]. After 10 min of ischemia and 180 min reperfusion, no infarct was found in controls; however, an infarct of 7 +/- 1% was found in CO-treated rabbits (P < 0.05; CRP, 2.3 +/- 0.4 mg/dl). The CRP-related increase in infarct size was not observed in croton oil-treated, C6-deficient rabbits (n = 5/group), indicating the involvement of complement. In these rabbits, infarct size was 22 +/- 2% (P < 0.05) despite having plasma CRP of 4.3 +/- 0.4 mg/dl. The CRP-associated increase in infarct size was ameliorated by pretreatment with heparin (n = 7; infarct size 33 +/- 3%; CRP, 2.3 +/- 0.3 mg/dl; P < 0.05) or N-acetylheparin (n = 7; infarct size 23 +/- 4%; CRP, 3.1 +/- 0.5 mg/dl; P < 0.05). These observations may explain why increased serum CRP is associated with an augmented risk for cardiovascular events.

Roles of the complement system in human neurodegenerative disorders: pro-inflammatory and tissue remodeling activities.

Complement is an important component of the innate immune response with the capacity to recognize and clear infectious challenges that invade the CNS through a damaged blood brain barrier. For instance, the membrane attack complex is involved in cytotoxic and cytolytic activities while other smaller fragments lead to cell activation (chemotaxis) and phagocytosis of the intruders. It is noteworthy that there is a growing body of evidence that uncontrolled complement biosynthesis and activation in the CNS can contribute to exacerbate the neuronal loss in several neurodegenerative disorders. We provide here an insightful review of the double-edged sword activities of the local innate complement system in the CNS and discuss further the potential therapeutic avenues of delivering complement inhibitors to control brain inflammation.

Targeting complement in therapy.

With increasing evidence that complement activation significantly contributes to the pathogenesis of a large number of inflammatory diseases, strategies that interfere with its deleterious action have become a major focus in pharmacological research. Endogenous soluble complement inhibitors (C1 inhibitor, recombinant soluble complement receptor 1, antibodies) blocking key proteins of the cascade reaction, neutralizing the action of the complement-derived anaphylatoxin C5a, or interfering with complement receptor 3 (CR3, CD18/11b)-mediated adhesion of inflammatory cells to the vascular endothelium have successfully been tested in various animal models over the past years. Promising results consequently led to clinical trials. Furthermore, incorporation of membrane-bound complement regulators (decay-accelerating factor (CD55), membrane co-factor protein (CD46), CD59) in transgenic animals has provided a major step forward in protecting xenografts from hyperacute rejection. At the same time, the poor contribution of complement to the antitumor response, which is caused by multiple resistance mechanisms that hamper the efficacy of antibody-based tumor therapy, is increasingly recognized and requires pharmacologic intervention. First attempts have now been made to interfere with the resistance mechanisms, thereby improving complement-mediated tumor cell destruction.

Neutrophils, not complement, mediate the mortality of experimental hemorrhagic pancreatitis.

Chemoactivation of the neutrophil (PMN) via the complement system has been observed in many inflammatory conditions and is thought to play a pathogenic role in acute pancreatitis. This study examined the effects of PMN depletion in experimental hemorrhagic pancreatitis and tested the role played by complement. Severe pancreatitis was induced by a choline-deficient, 0.5% ethionine-supplemented diet in female Institute of Cancer Research (ICR) mice weighing 11-13 g. Neutropenia was induced by an antibody injection. Total complement depletion was achieved by tail vein injections of cobra venom factor (CVF). Serum amylase levels and local pancreatic injury were not significantly modulated by either PMN or complement depletion at 72 hours. Systemic and remote organ injury, assessed by the formation of ascites, hematocrit, and serum alanine aminotransferase levels, was significantly reduced in neutropenic mice but failed to be moderated by complement depletion. In addition, liver and lung myeloperoxidase activity was independent of complement depletion. At 5 days, mortality was zero in PMN-depleted mice. There was no improvement in survival in the CVF-treated group. Neutrophils are important in the systemic injury and mortality of severe pancreatitis. PMN chemoactivation involves mechanisms other than complement.

Prostasomes: current concepts.

BACKGROUND: Prostasomes are membranous vesicles secreted by prostate gland, and they contain large amounts of cholesterol, sphingomyelin, calcium, and several enzymes. Prostasomes are involved in a number of biological functions. At ejaculation, these prostasomes are expelled with prostate secretions and are to be found in the seminal plasma as seminal prostasomes, which facilitate sperm function in various ways. METHODS: In this review, we discuss the structural and functional role of prostasomes, the various enzyme systems associated with these vesicles, and the biological role prostasomes play in male reproduction. RESULTS AND CONCLUSIONS Prostasomes are pluripotent and well-organized organelles secreted by the prostate gland. Prostasomes are ascribed to have many physiologiocal functions, the primary function being enhancement of sperm capacity. The several enzyme systems, small signaling molecules, and neuroendocrine markers associated with prostasomes reveal the complex nature of these vesicles in regulating sperm viability and vitality. The functional significance of these molecules that regulate complex pathways in these small vesicles is still a matter of dogma. Critical evaluation of the biological processes associated with prostasomes might be helpful in modeling new contraceptive agents, improving the techniques of in vitro fertilization, and in furthering our understanding and treatment of male factor infertility.