Nonsteroidal antiinflammatory drugs alter antibiotic susceptibility and expression of virulence-related genes and protein A of Staphylococcus aureus

Background/aim: Nonsteroidal antiinflammatory drugs (NSAIDs) including diclofenac, naproxen, ibuprofen, acetylsalicylic acid, and acetaminophen have been shown to have antimicrobial effects on various microorganisms. The aim of this study was to investigate the antibacterial effects of NSAIDs on Staphylococcus aureus. Materials and methods: Susceptibilities of S. aureus strains to NSAIDs with or without antimicrobials (moxifloxacin, vancomycin, ciprofloxacin, clindamycin, and gentamicin) were determined using the microdilution method and disk diffusion test. Expression levels of genes in the presence of drugs were investigated by real-time quantitative RT-PCR (qRT-PCR), and immunoblotting analysis was performed for staphylococcal protein A (SpA). Results: Our results showed that all NSAIDs were active against S. aureus strains with MIC values ranging from 195 µg/mL to 6250 µg/ mL. NSAIDs increased the antibiotic susceptibility of the strains, and diclofenac was found to be more effective than the other drugs. Drugs showed different effects on expression levels of virulence factor and/or regulatory genes. Immunoblotting analysis of SpA protein was mostly in accordance with qRT-PCR results. Conclusion: The regulatory/virulence factor genes and proteins of S. aureus investigated in this study may be reasonable targets for these drugs, and we suggest that the data may contribute to the field of infection control and antimicrobial resistance.

Low dose of protein A pretreatment can alleviate the inflammatory reaction and the bio-safety was evaluated in vivo.

BACKGROUND: Staphylococcal protein A (SPA) is a protein of Staphylococcus aureus. Up to now, there have been many studies on the biological activities of SPA. Some reported effects of SPA pretreatment on septic shock in mouse models but there is no study which reports the role of SPA pretreatment on the infected incision. METHODS: According to count results, bacterial suspension was set at a density of ∼1.8 × 10(9) colony forming units/mL. BALB/c mice were anesthetized via intraperitoneal injection with pentobarbital sodium. A longitudinal skin incision was made on the medial side of the right thigh. The length of the incision was 5 mm, and the depth was ∼3 mm. The bacterial suspension was gradually dripped and embrocated onto the incision surface to make the wound infection model. Before making the wound infection model for 48 hours and 24 hours, mice were retreated with SPA via intraperitoneal injection. Rats were intraperitoneally injected with SPA 1 mg/kg and the control group was injected with sterile saline to evaluate the biological safety of the best pretreatment dose. RESULTS: A 1-mL bacterial suspension can be utilized to make the wound infection model of BALB/c mouse lower limb. SPA pretreatment can reduce the inflammatory reactions in wound methicillin-resistant Staphylococcus aureus infection mouse model and the best pretreatment dose is 1 mg/kg. Intraperitoneal injection 1 mg/kg SPA does not destroy the functions of the organs. A 1-mg/kg SPA pretreatment can also reduce the inflammatory reactions in wound various bacterial infection mouse models. CONCLUSION: SPA pretreatment can effectively decrease the infected severity of a wound infected by various bacteria in a BALB/c mouse model. The best pretreatment dose is 1 mg/kg, and this dose does not damage organ function in rats up to a point.

Current therapy of the pemphigus group.

Treatment of pemphigus patients is still challenging and, in some cases, conventional therapy with systemic corticosteroids in combination with adjuvant corticosteroid-sparing immunosuppressive drugs is not sufficient to induce clinical remission. More recently, high-dose intravenous immunoglobulins, immunoadsorption, and the monoclonal anti-CD20 antibody, rituximab, have been established as additional successful therapeutic options. This contribution covers both conventional therapies and most current treatment strategies for pemphigus.

Co-opting endogenous immunoglobulin for the regulation of inflammation and osteoclastogenesis in humans and mice.

OBJECTIVE: Cells of the monocytic lineage play fundamental roles in the regulation of health, ranging from the initiation and resolution of inflammation to bone homeostasis. In rheumatoid arthritis (RA), the inflamed synovium exhibits characteristic infiltration of macrophages along with local osteoclast maturation, which, together, drive chronic inflammation and downstream articular destruction. The aim of this study was to explore an entirely novel route of immunoglobulin-mediated regulation, involving simultaneous suppression of the inflammatory and erosive processes in the synovium. METHODS: Using in vivo and in vitro studies of human cells and a murine model of RA, the ability of staphylococcal protein A (SPA) to interact with and modulate cells of the monocytic lineage was tested. In addition, the efficacy of SPA as a therapeutic agent was evaluated in murine collagen-induced arthritis (CIA). RESULTS: SPA showed a capacity to appropriate circulating IgG, by generating small immunoglobulin complexes that interacted with monocytes, macrophages, and preosteoclasts. Formation of these complexes resulted in Fcγ receptor type I-dependent polarization of macrophages to a regulatory phenotype, rendering them unresponsive to activators such as interferon-γ. The antiinflammatory complexes also had the capacity to directly inhibit differentiation of preosteoclasts into osteoclasts in humans. Moreover, administration of SPA in the early stages of disease substantially alleviated the clinical and histologic erosive features of CIA in mice. CONCLUSION: These findings demonstrate the overarching utility of immunoglobulin complexes for the prevention and treatment of inflammatory diseases. The results shed light on the interface between immunoglobulin complex-mediated pathways, osteoclastogenesis, and associated pathologic processes. Thus, therapeutic agents designed to harness all of these properties may be an effective treatment for arthritis, by targeting both the innate inflammatory response and prodestructive pathways.

Protein A immunoadsorption therapy in the highly sensitized kidney transplant candidates.

BACKGROUND: Sensitization in transplant candidates increases risk of irreversible immunologic injury of graft in the early period postoperatively. Elimination of anti-human leukocyte antigen (HLA) antibodies using protein A immunoadsorption (IA) might benefit these patients. METHODS: Protein A IA was used in 21 patients with high panel reactive antibody (PRA). The patients had IA 1 - 6 times (median 5 times) with the interval period was 2 - 5 days (median 2.5 days). RESULTS: Total 67 IA procedures were carried out smoothly in all patients. IA treatment reduced PRA I (pre (31.4 ± 3.8)% vs. post (24.4 ± 3.4)%, P < 0.01) and II (pre (37.1 ± 4.3)% vs. post (34.1 ± 3.9)%, P < 0.01). However, PRA did not change in some patients after the treatment. The serum immunoglobulin (IgG, IgM and IgA) and complement C3, C4 level were decreased significantly. Hemoglobin and albumin levels were slightly decreased associated with IA procedures. Flu-like symptoms were observed in a few of cases during the procedure but generally mild and transient. CONCLUSION: Protein A IA is capable to efficiently remove serum immunoglobulin and complement, reduce HLA class I and class II PRA in high sensitized transplant candidates, which is likely to benefit the kidney transplantation in these patients.

[Protein scaffolds as alternatives to whole antibodies: from discovery research to clinical development]. / Ingénierie de charpentes protéiques comme alternative aux anticorps : de la recherche au développement clinique.

Recent advances in combinatorial protein engineering have made it possible to develop non-Ig protein scaffolds that can potentially substitute for most whole antibody-associated properties. These protein scaffolds display most of the binding properties associated with the variable domain of antibodies. In theory, many different natural human protein backbones are suitable to be used as recombinant templates for engineering ; in practice however, only a few have yielded the necessary properties to be translated into << druggable biologicals >>. Amongst these properties, potential broad specificities towards any kind of target, ease of production, small size, good tolerability and low immunogenicity are essential. Intellectual property is another key issue. In this review, a particular emphasis will be given to the most validated non-Ig scaffolds that have reached the clinical development phase.

Successful treatment of patients with lipoprotein glomerulopathy by protein A immunoadsorption: a pilot study.

BACKGROUND: No established therapy is available for patients with lipoprotein glomerulopathy (LPG). Protein A immunoadsorption has been proved to be effective in reducing proteinuria in patients with nephrotic syndrome. In this uncontrolled pilot study, we investigated the efficiency of immunoadsorption onto staphylococcal protein A as treatment for LPG. METHODS: Thirteen patients with renal biopsy-proven LPG were treated with staphylococcal protein A immunoadsorption. Immunoadsorption was administered for 10 cycles per session and 10 sessions as a course. A total of 30 l of plasma was regenerated in each course. RESULTS: Single immunoadsorption course led to a rapid decline in proteinuria from 4.01 +/- 3.09 g/24 h to 1.21 +/- 0.97 g/24 h (mean +/- SD) (n = 13, P = 0.001), along with a dramatic decline in apolipoprotein E (apo E) from 9.79 +/- 5.04 mg/dl to 6.20 +/- 2.22 mg/dl (P = 0.004). A repeated renal biopsy (n = 12) showed that intraglomerular lipoprotein thrombi almost disappeared. Six patients were enrolled in the investigation of long-term outcome, and proteinuria returned to baseline levels within 12 months. Four recurrent patients received repeat immunoadsorption treatment; proteinuria decreased from 5.02 +/- 1.85 g/24 h to 1.64 +/- 0.55 g/24 h at the end of the treatment, serum apo E decreased from 14.65 +/- 11.17 mg/dl to 7.90 +/- 1.72 mg/dl. No patients suffered from severe complications. CONCLUSION: Our observations suggest that immunoadsorption onto protein A might be an effective treatment for resolving intraglomerular thrombi and improving nephrotic syndrome in patients with LPG. Further studies are required to define the influence of immunoadsorption on long-term effects in LPG patients.

Immunoadsorption in severe C4d-positive acute kidney allograft rejection: a randomized controlled trial.

Antibody-mediated rejection (AMR) frequently causes refractory graft dysfunction. This randomized controlled trial was designed to evaluate whether immunoadsorption (IA) is effective in the treatment of severe C4d-positive AMR. Ten out of 756 kidney allograft recipients were included. Patients were randomly assigned to IA with protein A (N = 5) or no such treatment (N = 5) with the option of IA rescue after 3 weeks. Enrolled recipients were subjected to tacrolimus conversion and, if indicated, 'anti-cellular' treatment. All IA-treated patients responded to treatment. One death unrelated to IA occurred after successful reversal of rejection. Four control subjects remained dialysis-dependent. With the exception of one patient who developed graft necrosis, non-responders were subjected to rescue IA, however, without success. Because of a high graft loss rate in the control group the study was terminated after a first interim analysis. Even though limited by small patient numbers, this trial suggests efficiency of IA in reversing severe AMR.

Options for treating acute bleeds in addition to bypassing agents: extracorporeal immunoadsorption, FVIII/FIX, desmopressin and antifibrinolytics.

Inhibitor patients do not always respond satisfactorily to treatment with bypassing agents, and options to the standard practice are sometimes needed. Temporary inhibitor removal may be achieved using extracorporeal immunoadsorption. This technique uses a column system including either protein A or antihuman IgG. Immunoadsorption may be used as part of an immune tolerance protocol, or in the case of acute bleeds or prior to surgery, thus rendering the patient more responsive to ordinary replacement therapy with factor VIII or factor IX. Desmopressin is a valuable haemostatic agent in many situations and can be especially recommended in mild haemophilia complicated by an inhibitor. Antifibrinolytics are often administered as an adjunct therapy to the treatment protocol and have also been reported to have a direct anti-inhibitor effect.

Treatment of pemphigus vulgaris with protein A immunoadsorption: case report of long-term history showing favorable outcome.

A pemphigus vulgaris (PV) patient with a 14-year history of severe and painful blistering of skin and mucous membranes as well as side effects from corticosteroids and concomitant immunosuppressive drug treatment was managed successfully by protein A immunoadsorption (IA). After 19 sessions of protein A IA, the patient showed remission of PV and healing of mucocutaneous lesions and the skin. The level of the pathogenic autoantibodies to the adhesion proteins desmoglein 1 (Dsg-1) and desmoglein 3 (Dsg-3) measured by ELISA and immunofluorescence microscopy revealed a significant removal of autoantibodies after each IA therapy. There was a weak rebound of anti-Dsg-1 and anti-Dsg-3 antibodies between IA sessions but an overall decrease over the period of IA therapy. This case demonstrates the effective use of protein A IA as an adjuvant and corticosteroid-sparing therapy in severe pemphigus refractory to standard immunosuppressive therapy and underscores the need for careful monitoring of autoantibodies.

Increased ethyl mercury load in protein a immunoadsorption.

Immunoadsorption is an adsorption technique for extracorporeal removal of circulating autoantibodies in autoimmune diseases. To prevent microbial growth during storage, the protein A columns are primed with thiomersal, which contains toxic ethyl mercury, which may be released during the procedure and potentially begin to accumulate and become toxic. To reduce the thiomersal-related mercury release during immunoadsorption treatment, we introduced a modified rinsing solution containing N-acetylcysteine, which is an avid mercury scavenger. Thirteen patients received 17 protein A immunoadsorption treatments and their venous blood samples were collected immediately before and after each session. The total blood mercury levels were measured by atomic absorption spectrometry, and the ethyl mercury levels by atomic fluorescence spectrometry. Following the manufacturer's recommendations, we used 600 mg of N-acetylcysteine to rinse the mercury from protein-loaded columns before each immunoadsorption treatment. After immunoadsorption, the ethyl mercury levels increased from 0.148 +/- 0.402 ng/g to 2.026 +/- 1.944 ng/g (P < 0.001), and the total blood mercury levels increased from 2.447 +/- 3.065 ng/g to 20.437 +/- 28.603 ng/g (P = 0.02). The post-treatment values of total blood mercury exceeded the upper safety level of 5 ng/g in all 17 immunoadsorption treatments, but no patient developed clinical signs of mercury toxicity. The results of our study showed an increase in total blood mercury and ethyl mercury levels during the immunoadsorption treatments, suggesting mercury release from thiomersal-primed columns despite the addition of N-acetylcysteine to the rinsing solution.

Effect of the B cell superantigen protein A from S. aureus on the early lupus disease of (NZBxNZW) F1 mice.

The (NZBxNZW) F(1) mouse develops a spontaneous autoimmune disease process with striking similarities to human systemic lupus erythematosus (SLE). In female (NZBxNZW) F(1) mice, the production of IgG antinuclear antibodies, including antibodies to double-stranded DNA (dsDNA), is associated with the development of a severe immune complex-mediated glomerulonephritis that results in death from renal failure in virtually all animals by 12 months of age. Since B-1 and marginal zone (MZ) cells represent a potential source of pathogenic antibodies and because B cell superantigens have been demonstrated to reduce B-1 and MZ cells in vivo, we tested the effect of repeated injections of the superantigen protein A (SpA) from S. aureus on the disease of this lupus model. We found that weekly intraperitoneal injections of SpA delay the progression of serum anti-DNA IgG and reduce proteinuria early in young female (NZBxNZW) F(1) mice. This superantigen also induced a specific depression in the numbers of peritoneal B-1 cells, as compared to mice treated with a control protein. These results support the role of B-1 cells in the development of the autoimmune disease in this mouse model and suggest that B cell superantigens may be useful in the management of autoimmune conditions.

Correcting immune imbalance: the use of Prosorba column treatment for immune disorders.

The treatment of selected refractory autoimmune diseases has been complemented by the use of Protein A (Prosorba column) immunoadsorption. US Food and Drug Administration-approved clinical applications include idiopathic thrombocytopenia purpura (ITP) and rheumatoid arthritis (RA). Other common off label uses include thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS). Less common experimental uses in diseases in which efficacy has been reported include autoimmune CNS syndromes, peripheral neuropathies, autoimmune pancytopenia, hemolytic anemia and solid organ transplant rejection. Prosorba column treatment is generally well tolerated but a small proportion of treated patients experience chills, fever, tremor, hypotension and rash. The mechanism of action suggested for the efficacy of the column is the restoration of normal immune balance and normal tolerance. Observations in ITP has suggested that column treatment stimulates a rise in anti-idiotype antibody directed against antiplatelet antibodies, effecting a decrease in pathogenic antiplatelet antibodies and immune complexes.

Protein A immunoadsorption: a novel and effective adjuvant treatment of severe pemphigus.

BACKGROUND: Pemphigus foliaceus (PF) and pemphigus vulgaris (PV) are autoimmune blistering skin diseases usually treated with high-dose systemic corticosteroids and other immunosuppressants that may cause severe side-effects. Plasmapheresis also has been demonstrated to be of benefit in the treatment of pemphigus. In contrast to plasmapheresis, staphylococcal protein A immunoadsorption (PA-IA) specifically removes immunoglobulin from the circulation, allows treatment of larger plasma volumes, and does not require the substitution of plasma components. OBJECTIVES: To determine the effectiveness and side-effects of PA-IA in patients with severe pemphigus. METHODS: Five patients with severe pemphigus (PV, n = 4; PF, n = 1) were treated by PA-IA. Three of these patients had been refractory to various treatment regimens. In addition to PA-IA, methylprednisolone, 0.5 mg x kg-1 body weight day-1 was given initially and subsequently tapered. RESULTS: In all patients, a dramatic clinical improvement was seen within 2 weeks after initiation of therapy. Patients were free of lesions after 3, 4, 4, 10 and 21 weeks of treatment, respectively. Concurrently, autoantibody levels decreased rapidly. CONCLUSIONS: PA-IA is a rational, effective, and safe adjuvant therapy for severe pemphigus and warrants wider use for this indication. A controlled study should compare side-effects and effectiveness of PA-IA with other treatment options for pemphigus.