The herbal extract EPs® 7630 increases the antimicrobial airway defense through monocyte-dependent induction of IL-22 in T cells.

The phytotherapeutic compound EPs® 7630, an extract manufactured from Pelargonium sidoides roots, is frequently used for the treatment of airway infections. Nevertheless, the knowledge of the mode of action of EPs® 7630 is still sparse. Our study aimed at further elucidating the underlying pharmacological mechanisms by focusing on antimicrobial defense mechanisms of EPs® 7630. While investigating the influence of EPs® 7630 on lymphokine production by PBMCs, we found that EPs® 7630 is a novel inducer of IL-22 and IL-17. This cytokine-inducing effect was most pronounced for IL-22 and clearly dose-dependent starting from 1 µg/ml of the extract. Furthermore, EPs® 7630 pretreatment selectively enhanced the IL-22 and IL-17 production capacity of CD3/28-activated PBMCs while strongly limiting the IFN-γ production capacity of innate lymphoid cells. The relevance of EPs® 7630-induced IL-22 production was proven in vitro and in vivo, where IL-22 provoked a strong increase of the antimicrobial protein S100A9 in lung epithelial cells and pulmonary tissue, respectively. A detailed analysis of IL-22 induction modi revealed no direct influence of EPs® 7630 on the basal or anti-CD3/CD28 antibody-induced IL-22 production by CD4+ memory T cells. In fact, EPs® 7630-induced IL-22 production by CD4+ memory T cells was found to be essentially dependent on soluble mediators (IL-1/IL-23) as well as on direct cellular contact with monocytes. In summary, our study reveals a new immune-modulating function of EPs® 7630 that might confer IL-22 and IL-17-induced protection from bacterial airway infection. KEY MESSAGES: EPs® 7630 selectively strengthens IL-22 and IL-17 production of memory T cells. EPs® 7630 limits the IFN-y production capacity of innate lymphoid cells. EPs® 7630-caused IL-22 production by T cells is essentially dependent on monocytes. IL-22 increase antimicrobial proteins (AMPs) in airway epithelium. EPs® 7630 might protect against airway infection by induction of AMP-inducers.

The Pelargonium sidoides Extract EPs 7630 Drives the Innate Immune Defense by Activating Selected MAP Kinase Pathways in Human Monocytes.

Pelargonium sidoides is a medical herb and respective extracts are used very frequently for the treatment of respiratory tract infections. However, the effects of Pelargonium sidoides and a special extract prepared from its roots (EPs 7630) on human immune cells are not fully understood. Here we demonstrate that EPs 7630 induced a rapid and dose-dependent production of TNF-α, IL-6, and IL-10 by human blood immune cells. This EPs 7630-induced cytokine profile was more pro-inflammatory in comparison with the profile induced by viral or bacterial infection-mimicking agents. The search for EPs 7630 target cells revealed that T-cells did not respond to EPs 7630 stimulation by production of TNF-α, IL-6, or IL-10. Furthermore, pretreatment of T-cells with EPs 7630 did not modulate their TNF-α, IL-6, and IL-10 secretion during subsequent activation. In contrast to lymphocytes, monocytes showed clear intracellular TNF-α staining after EPs 7630 treatment. Accordingly, EPs 7630 predominantly provoked activation of MAP kinases and inhibition of p38 strongly reduced the monocyte TNF-α production. The pretreatment of blood immune cells with EPs 7630 lowered their secretion of TNF-α and IL-10 and caused an IL-6 dominant response during second stimulation with viral or bacterial infection-mimicking agents. In summary, we demonstrate that EPs 7630 activates human monocytes, induces MAP kinase-dependent pro-inflammatory cytokines in these cells, and specifically modulates their production capacity of mediators known to lead to an increase of acute phase protein production in the liver, neutrophil generation in the bone marrow, and the generation of adaptive Th17 and Th22 cells.

Absolute and functional iron deficiency in professional athletes during training and recovery.

BACKGROUND: Iron deficiency (ID) is one of the most important metabolic dysfunctions. Athletic performance depends on oxygen transport and mitochondrial efficiency, thus on optimal iron balance. We hypothesised that physical extremes result in ID in elite athletes and that the short recovery period may be insufficient to allow a lasting replenishment of iron reserves. METHODS: Iron metabolism was examined in 20 elite rowing athletes and 10 professional soccer players at the end of a competitive season, after recuperation and during pre-season training. Absolute ID values were defined as ferritin <30 µg/L, functional ID as ferritin 30-99 µg/L or 100-299 µg/L+transferrin saturation <20%. RESULTS: At the end of season, 27% of all athletes had absolute ID and 70% showed functional ID. Absolute iron depletion was not generally restored after recuperation and observed at all time points in 14% of the athletes. Although athletes with initially low ferritin levels showed a slight increase during recuperation (p<0.09), these increases remained within borderline levels. Furthermore, 10% showed borderline haemoglobin levels, suggestive of mild anaemia, as defined by the World Health Organisation. CONCLUSION: A significant proportion of professional athletes have ID, independent of the training mode. Although recuperation seems to allow a certain recovery of iron storage, particularly in athletes with initially low ferritin levels, this retrieval was insufficient to fully normalise reduced iron levels. Therefore, iron status should be carefully monitored during the various training and competitive periods in elite athletes. An adequate iron supplementation may be needed to maintain balanced iron stores.

Preventive antibacterial therapy in acute ischemic stroke: a randomized controlled trial.

BACKGROUND: Pneumonia is a major risk factor of death after acute stroke. In a mouse model, preventive antibacterial therapy with moxifloxacin not only prevents the development of post-stroke infections, it also reduces mortality, and improves neurological outcome significantly. In this study we investigate whether this approach is effective in stroke patients. METHODS: Preventive ANtibacterial THERapy in acute Ischemic Stroke (PANTHERIS) is a randomized, double-blind, placebo-controlled trial in 80 patients with severe, non-lacunar, ischemic stroke (NIHSS>11) in the middle cerebral artery (MCA) territory. Patients received either intravenous moxifloxacin (400 mg daily) or placebo for 5 days starting within 36 hours after stroke onset. Primary endpoint was infection within 11 days. Secondary endpoints included neurological outcome, survival, development of stroke-induced immunodepression, and induction of bacterial resistance. FINDINGS: On intention-to treat analysis (79 patients), the infection rate at day 11 in the moxifloxacin treated group was 15.4% compared to 32.5% in the placebo treated group (p = 0.114). On per protocol analysis (n = 66), moxifloxacin significantly reduced infection rate from 41.9% to 17.1% (p = 0.032). Stroke associated infections were associated with a lower survival rate. In this study, neurological outcome and survival were not significantly influenced by treatment with moxifloxacin. Frequency of fluoroquinolone resistance in both treatment groups did not differ. On logistic regression analysis, treatment arm as well as the interaction between treatment arm and monocytic HLA-DR expression (a marker for immunodepression) at day 1 after stroke onset was independently and highly predictive for post-stroke infections. INTERPRETATION: PANTHERIS suggests that preventive administration of moxifloxacin is superior in reducing infections after severe non-lacunar ischemic stroke compared to placebo. In addition, the results emphasize the pivotal role of immunodepression in developing post-stroke infections. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN74386719.

Oral high-dose atorvastatin treatment in relapsing-remitting multiple sclerosis.

BACKGROUND: Recent data from animal models of multiple sclerosis (MS) and from a pilot study indicated a possible beneficial impact of statins on MS. METHODOLOGY/PRINCIPAL FINDINGS: Safety, tolerability and effects on disease activity of atorvastatin given alone or in combination with interferon-beta (IFN-beta) were assessed in a phase II open-label baseline-to-treatment trial in relapsing-remitting MS (RRMS). Patients with at least one gadolinium-enhancing lesion (CEL) at screening by magnetic resonance imaging (MRI) were eligible for the study. After a baseline period of 3 monthly MRI scans (months -2 to 0), patients followed a 9-month treatment period on 80 mg atorvastatin daily. The number of CEL in treatment months 6 to 9 compared to baseline served as the primary endpoint. Other MRI-based parameters as well as changes in clinical scores and immune responses served as secondary endpoints. Of 80 RRMS patients screened, 41 were included, among them 16 with IFN-beta comedication. The high dose of 80 mg atorvastatin was well tolerated in the majority of patients, regardless of IFN-beta comedication. Atorvastatin treatment led to a substantial reduction in the number and volume of CEL in two-sided multivariate analysis (p = 0.003 and p = 0.008). A trend towards a significant decrease in number and volume of CEL was also detected in patients with IFN-beta comedication (p = 0.060 and p = 0.062), in contrast to patients without IFN-beta comedication (p = 0.170 and p = 0.140). Immunological investigations showed no suppression in T cell response but a significant increase in IL-10 production. CONCLUSIONS/SIGNIFICANCE: Our data suggest that high-dose atorvastatin treatment in RRMS is safe and well tolerated. Moreover, MRI analysis indicates a possible beneficial effect of atorvastatin, alone or in combination with IFN-beta, on the development of new CEL. Thus, our findings provide a rationale for phase II/III trials, including combination of atorvastatin with already approved immunomodulatory therapy regimens. TRIAL REGISTRATION: ClinicalTrials.gov NCT00616187.

A vaccine for hypertension based on virus-like particles: preclinical efficacy and phase I safety and immunogenicity.

BACKGROUND: Despite the availability of efficacious drugs, the success of treating hypertension is limited by patients' inconsistent drug intake. Immunization against angiotensin II may offer a valuable alternative to conventional drugs for the treatment of hypertension, because vaccines induce relatively long-lasting effects and do not require daily dosing. Here we describe the preclinical development and the phase I clinical trial testing of a virus-like particle (VLP)-based antihypertensive vaccine. METHODS AND RESULTS: An angiotensin II-derived peptide was conjugated to the VLP Qbeta (AngQb). AngQb was highly immunogenic in mice and rats. To test for efficacy, spontaneously hypertensive rats (SHR) were immunized with 400 microg AngQb or VLP alone. Group mean systolic blood pressure (SBP) was reduced by up to 21 mmHg (159 +/- 2 versus 180 +/- 5 mmHg, P < 0.001), and total angiotensin II levels (antibody-bound and free) were increased ninefold (85 +/- 20 versus 9 +/- 1 pmol/l, P = 0.002) compared with VLP controls. SHR treated with the angiotensin-converting enzyme (ACE) inhibitor ramipril (1 mg/kg per day by mouth) reached an SBP of 155 +/- 2 mmHg. Twelve healthy volunteers of a placebo-controlled randomized phase I trial were injected once with 100 microg AngQb. Angiotensin II-specific antibodies were raised in all subjects (100% responder rate) and AngQb was well tolerated. CONCLUSIONS: AngQb reduces blood pressure in SHR to levels obtained with an ACE inhibitor, and is immunogenic and well tolerated in humans. Therefore, vaccination against angiotensin II has the potential to become a useful antihypertensive treatment providing long-lasting effects and improving patient compliance.

Immunologic effector mechanisms of a standardized mistletoe extract on the function of human monocytes and lymphocytes in vitro, ex vivo, and in vivo.

Even though mistletoe extracts have been in clinical use for centuries their exact mode of action is still unknown. Currently, the application scheme for registered preparations is a dose-escalating scheme to thus reduce side effects. In this study, healthy controls and patients were evaluated for their immunologic response to treatment with a standardized mistletoe extract (Iscador). It shows a strong effect as adjuvant that induces TNF-alpha and IL-12, which was partly mediated via CD14. Desensitization of the TNF-alpha response could be shown after repeated application in vitro and in vivo. Furthermore, Iscador induces a specific lymphocyte sensitization upon multiple injections and production of IgG1- and IgG3 -mistletoe antibodies. Remarkably, a systemic bystander effect (heterologous immunity against other recall antigens) was observed after long-term treatment. In conclusion, dose-escalation reduces the monocyte-related clinical side effects. A T-lymphocyte sensitization stimulates mainly a specific Th1 response. The most interesting clinical long-term effect is the bystander stimulation of various memory T cells that might mediate in vivo antitumor and antiinfectious T-cell response under mistletoe-extract immunization.

The effect of micronutrient supplementation on quality-of-life and left ventricular function in elderly patients with chronic heart failure.

AIMS: Chronic heart failure (CHF) is a common and leading cause of death in industrialized countries. The potential benefits of micronutrient supplementation in CHF are extensive. Therefore, we examined the influence of long-term multiple micronutrient supplementation on left ventricular (LV) function, levels of pro-inflammatory cytokines, and quality-of-life (QoL) in elderly patients with CHF. METHODS AND RESULTS: Thirty CHF patients [age 75.4 (0.7), mean (SEM), LV ejection fraction (LVEF) < or =35%] were randomized to receive capsules containing a combination of high-dose micronutrients (calcium, magnesium, zinc, copper, selenium, vitamin A, thiamine, riboflavin, vitamin B(6), folate, vitamin B(12), vitamin C, vitamin E, vitamin D, and Coenzyme Q10) or placebo for 9 months in a double-blind fashion. All subjects were on stable optimal medical therapy for at least 3 months before enrolment. At randomization and at study end, tumour necrosis factor-alpha and its soluble receptors TNFR-1 and TNFR-2 were measured and six-minute walk test and QoL were assessed. Cardiac magnetic resonance scanning was performed to evaluate cardiac dimensions and LVEF. Two patients died during follow-up. The remaining patients (14 randomized to placebo and 14 to micronutrients) were well matched for LV function, symptoms, and exercise capacity. At the end of the follow-up period, LV volumes were reduced in the intervention group with no change in the placebo group [-13.1 (17.1)% vs. +3.8 (10.0)%; P<0.05]. LVEF increased by 5.3+/-1.4% in the intervention group and was unchanged in the placebo group (P<0.05). Patients taking micronutrients also had a significant improvement in QoL score between enrolment and study end [+9.5 (1.6)%; P<0.05], whereas those taking placebo had a slight deterioration [-1.1 (0.8)%; P=0.12]. Six-minute walk test and inflammatory cytokine levels remained unchanged in both groups. CONCLUSION: Long-term multiple micronutrient supplementation can improve LV volumes and LVEF and QoL scores in elderly patients with heart failure due to LV systolic dysfunction.

Preventive antibacterial treatment improves the general medical and neurological outcome in a mouse model of stroke.

BACKGROUND AND PURPOSE: Epidemiological studies have demonstrated a high incidence of infections after severe stroke and their prominent role in morbidity and mortality in stroke patients. In a mouse model, it has been shown recently that stroke is coupled with severe and long-lasting immunosuppression, which is responsible for the development of spontaneous systemic infections. Here, we investigated in the same model the effects of preventive antibiotic treatment on survival and functional outcome of experimental stroke. METHODS: Mice were subjected to experimental stroke by occlusion of the middle cerebral artery (MCAO) for 60 minutes. A group of mice received moxifloxacin (6x100 mg/kg body weight every 2 hours over 12 hours) either immediately or 12 hours after MCAO. Control animals received the vector only. Behavior, neurological deficit, fever, survival, and body weight were monitored over 14 days. In a subgroup, infarct volume was measured 4 days after MCAO. Microbiological assessment was based on cultures of lung tissue, blood, and feces of animals 3 days after stroke. For a dose-response study, moxifloxacin was given immediately after MCAO in different doses and at different time points. RESULTS: Microbiological analyses of blood and lung tissue demonstrated high bacterial burden, mainly Escherichia coli, 3 days after stroke. Accordingly, we observed clinical and histological signs of septicemia and pneumonia. Moxifloxacin prevented the development of infections and fever, significantly reduced mortality, and improved neurological outcome. CONCLUSIONS: Preventive antibiotic treatment may be an important new therapeutical approach to improve outcome in patients with severe stroke.

Ex vivo exposure to carbon monoxide prevents hepatic ischemia/reperfusion injury through p38 MAP kinase pathway.

A direct role of carbon monoxide (CO), an effector-signaling molecule during heme oxygenase-1 (HO-1) catalysis of heme, in the protection against hepatic ischemia/reperfusion (I/R) injury needs to be established. This study was designed to determine the effects and downstream mechanisms of CO on cold I/R injury in a clinically relevant isolated perfusion rat liver model. After 24 hours of cold storage, rat livers perfused ex vivo for 2 hours with blood supplemented with CO (300 parts per million) showed significantly decreased portal venous resistance and increased bile production, as compared with control livers perfused with blood devoid of CO. These beneficial effects correlated with improved liver function (serum glutamic oxaloacetic transaminase levels) and diminished histological features of hepatocyte injury (Banff's scores). The CO-mediated cytoprotective effects were nitric oxide synthase- and cyclic guanine monophosphate-independent, but p38 mitogen-activated protein kinase (MAPK)-dependent. Moreover, adjunctive use of zinc protoporphyrin, a competitive HO-1 inhibitor, has shown that exogenous CO could fully substitute for endogenous HO-1 in preventing hepatic I/R insult. This study performed in a clinically relevant ex vivo cold ischemia model is the first to provide the evidence that HO-1-mediated cytoprotection against hepatic I/R injury depends on the generation of, and can be substituted by, exogenous CO. The p38 MAPK signaling pathway represents the key downstream mechanism by which CO prevents the I/R insult. In conclusion, regimens that employ exogenous CO should be revisited, as they may have potential applications in preventing/mitigating I/R injury, and thus expanding the liver donor pool for clinical transplantation.