Elucidating the role of the gut microbiota in the physiological effects of dietary fiber.

BACKGROUND: Dietary fiber is an integral part of a healthy diet, but questions remain about the mechanisms that underlie effects and the causal contributions of the gut microbiota. Here, we performed a 6-week exploratory trial in adults with excess weight (BMI: 25-35 kg/m2) to compare the effects of a high-dose (females: 25 g/day; males: 35 g/day) supplement of fermentable corn bran arabinoxylan (AX; n = 15) with that of microbiota-non-accessible microcrystalline cellulose (MCC; n = 16). Obesity-related surrogate endpoints and biomarkers of host-microbiome interactions implicated in the pathophysiology of obesity (trimethylamine N-oxide, gut hormones, cytokines, and measures of intestinal barrier integrity) were assessed. We then determined whether clinical outcomes could be predicted by fecal microbiota features or mechanistic biomarkers. RESULTS: AX enhanced satiety after a meal and decreased homeostatic model assessment of insulin resistance (HOMA-IR), while MCC reduced tumor necrosis factor-α and fecal calprotectin. Machine learning models determined that effects on satiety could be predicted by fecal bacterial taxa that utilized AX, as identified by bioorthogonal non-canonical amino acid tagging. Reductions in HOMA-IR and calprotectin were associated with shifts in fecal bile acids, but correlations were negative, suggesting that the benefits of fiber may not be mediated by their effects on bile acid pools. Biomarkers of host-microbiome interactions often linked to bacterial metabolites derived from fiber fermentation (short-chain fatty acids) were not affected by AX supplementation when compared to non-accessible MCC. CONCLUSION: This study demonstrates the efficacy of purified dietary fibers when used as supplements and suggests that satietogenic effects of AX may be linked to bacterial taxa that ferment the fiber or utilize breakdown products. Other effects are likely microbiome independent. The findings provide a basis for fiber-type specific therapeutic applications and their personalization. TRIAL REGISTRATION: Clinicaltrials.gov, NCT02322112 , registered on July 3, 2015. Video Abstract.

Impact of the COVID-19 pandemic on the oncologic activities (diagnosis, treatment, clinical trials enrollment) of a general hospital in a district with high prevalence of SARS-COV-2 in Italy.

PURPOSE: Little is known about the real impact of the COVID-19 outbreak on the qualitative and quantitative fall-out on the management of cancer patients. Our objective was to provide evidence of the effects of SARS-COV-2 on the management of cancer patients in the real world. METHODS: In a general hospital in a district in Italy with high prevalence of COVID-19 during the first wave, we retrospectively analyzed the data of oncologic activity, namely new cancer diagnosis, types of treatment (intravenous or by mouth), clinical research studies, and drug utilization, and compared the findings with those of 2019, before the pandemic. The data have been summarized in boxplot figures for median and interquartile range. RESULTS: In 2020, a significant reduction in new cancer diagnosis was demonstrated when compared with 2019, with 17.4% fewer cancer diagnoses, 84.5% fewer patients enrolled in clinical trials, a 10.6% reduction in intravenous antitumor treatment, and a 42.7% increase in oral anticancer treatment. CONCLUSION: Our data indicate a significant reduction in cancer diagnosis, antitumor venous treatment, and patients enrolled in clinical research studies in 2020 compared with 2019, although there was a significant increase in oral treatment. These data suggest that the COVID-19 pandemic had a deep impact on the real-world management of cancer patients in a district of Italy with a high prevalence of COVID-19.

Comparison of Four-Dimensional Magnetic Resonance Imaging Analysis of Left Ventricular Fluid Dynamics and Energetics in Ischemic and Restrictive Cardiomyopathies.

BACKGROUND: Time-resolved three-directional velocity-encoded (4D flow) magnetic resonance imaging (MRI) enables the quantification of left ventricular (LV) intracavitary fluid dynamics and energetics, providing mechanistic insight into LV dysfunctions. Before becoming a support to diagnosis and patient stratification, this analysis should prove capable of discriminating between clearly different LV derangements. PURPOSE: To investigate the potential of 4D flow in identifying fluid dynamic and energetics derangements in ischemic and restrictive LV cardiomyopathies. STUDY TYPE: Prospective observational study. POPULATION: Ten patients with post-ischemic cardiomyopathy (ICM), 10 patients with cardiac light-chain cardiac amyloidosis (AL-CA), and 10 healthy controls were included. FIELD STRENGTH/SEQUENCE: 1.5 T/balanced steady-state free precession cine and 4D flow sequences. ASSESSMENT: Flow was divided into four components: direct flow (DF), retained inflow, delayed ejection flow, and residual volume (RV). Demographics, LV morphology, flow components, global and regional energetics (volume-normalized kinetic energy [KEV ] and viscous energy loss [ELV ]), and pressure-derived hemodynamic force (HDF) were compared between the three groups. STATISTICAL TESTS: Intergroup differences in flow components were tested by one-way analysis of variance (ANOVA); differences in energetic variables and peak HDF were tested by two-way ANOVA. A P-value of <0.05 was considered significant. RESULTS: ICM patients exhibited the following statistically significant alterations vs. controls: reduced KEV , mostly in the basal region, in systole (-44%) and in diastole (-37%); altered flow components, with reduced DF (-33%) and increased RV (+26%); and reduced basal-apical HDF component on average by 63% at peak systole. AL-CA patients exhibited the following alterations vs. controls: significantly reduced KEV at the E-wave peak in the basal segment (-34%); albeit nonstatistically significant, increased peaks and altered time-course of the HDF basal-apical component in diastole and slightly reduced HDF components in systole. DATA CONCLUSION: The analysis of multiple 4D flow-derived parameters highlighted fluid dynamic alterations associated with systolic and diastolic dysfunctions in ICM and AL-CA patients, respectively. LEVEL OF EVIDENCE: 2 TECHNICAL EFFICACY STAGE: 3.

[Territorial-based management of patients with cancer on active treatment following the Piacenza (north Italy) Model. Results of 4 consecutive years.] / Le cure oncologiche ed ematologiche sul territorio secondo il modello dell'ASL di Piacenza. Rendiconto di 4 anni consecutivi.

INTRODUCTION: The natural history of cancer has radically changed in the last decade. The burden of travel from patient's residence to health care providers is an important issue that can influence access to diagnosis and treatment of cancer; however this issue is quite neglect by the medical community and by the national health system. In addition, community care in the oncology field is actually debated. METHODS: In the district of Piacenza an innovative model to deliver onco-hematologic treatment near the residence of patients was initiated some years ago. The oncologic and hematologic treatments are delivered by specialized nurses under supervision of medical oncologists or hematologists at the 3 community hospital and at 1 house of health in the district of Piacenza. We conducted a retrospective study involving 1,339 cancer patients (CPs) managed and treated near their residence, CPs were on active medical treatment at the oncology and hematology department Azienda sanitaria (ASL) of Piacenza (North Italy). The electronic data base of the antiblastic drug unit (UFA) of the ASL Piacenza, provided: the number of patients treated each year, number of treatments and the accesses to the territorial medical structure each year. The kms saved to reach the nearest territorial structures instead of the oncologic unit of the city hospital, were registered and recorded. RESULTS: During a 4 years period, from January 2017 to December 2020, 1,339 CPs were treated near their residence, 278 in the year 2017, 347 in 2018, 354 in 2019 and 360 in 2020. The total accesses for treatment in 4 years were 10,003: 2,214 in the year 2017, 2,652 in 2018, 2,524 in 2019 and 2,613 in 2020. The mean distance saved for each patient was 937 kms in the year 2017, 891 in 2018, 879 in 2019, 920 in 2020, totally a mean of 3,627 kms in the 4 years. DISCUSSION AND CONCLUSION: We believe that the results of our retrospective study highlight the possibility of treating cancer patients in territorial structures near their residence, with advantages for patients themselves, their caregivers and for the entire community.

Anticoagulation and Vessel Recanalization in Cirrhotic Patients with Splanchnic Vein Thrombosis: A Multidisciplinary "Real Life" Experience.

BACKGROUND AND AIM: Splanchnic vein thrombosis (SVT) is a potentially life-threatening complication of liver cirrhosis. This study aimed to evaluate the impact of a multi-disciplinary approach and early anticoagulation therapy (AT) on bleeding/thrombotic events, recanalization rates and outcome of cirrhotic patients with SVT. METHODS: This is a single-center, registry-based cohort study. Over 17 years, 149 SVT patients were enrolled and prospectively evaluated. Regarding cirrhotic-SVT, a pre-specified algorithm, guiding initial posology of AT and follow-up visits schedule, was performed. Major bleeding (MB), thrombotic events, functional liver scores and all cause-mortality were investigated. Efficacy of AT was evaluated by radiological imaging. RESULTS: In cirrhotic-SVT, the incidence rate of MB was 8.4 per 100 patient-year (95% CI, 3.83-15.97), while the incidence rate of thrombosis was 5.6 per 100 patient-year (95% CI, 2.05-12.2). In incidental SVT treated with AT, MB incidence was 6.5 per 100 patient-year (95% CI: 2.8-12.82), while in symptomatic SVT was 2.2 per 100 patient-year (95% CI: 0.25-8.02). All thrombotic recurrences occurred in incidental SVT (7.7 per 100 patient-years; 95% CI, 3.71-14.26). Overall survival was significantly higher in patients who had at least a partial recanalization (p < 0.01) and partial/total recanalization was independently associated with improved MELD score at multivariate analysis (HR 2.62, 95% CI 1.1-6.47, p = 0.03). CONCLUSION: In cirrhotic SVT patients, partial or total resolution of thrombosis ameliorates liver function and is associated with higher overall survival. A multidisciplinary approach together with radiological follow-up at pre-fixed time improves patient selection and monitoring.

4D flow evaluation of blood non-Newtonian behavior in left ventricle flow analysis.

Blood is generally modeled as a Newtonian fluid, assuming a standard and constant viscosity; however, this assumption may not hold for the highly pulsatile and recirculating intracavitary flow in the left ventricle (LV), hampering the quantification of fluid dynamic indices of potential clinical relevance. Herein, we investigated the effect of three viscosity models on the patient-specific quantification of LV blood energetics, namely on viscous energy loss (EL), from 4D Flow magnetic resonance imaging: I) Newtonian with standard viscosity (3.7 cP), II) Newtonian with subject-specific hematocrit-dependent viscosity, III) non-Newtonian accounting for the effect of hematocrit and shear rate. Analyses were performed on 5 controls and 5 patients with cardiac light-chain amyloidosis. In Model II, viscosity ranged between 3.0 (-19%) and 4.3 cP (+16%), mildly deviating from the standard value. In the non-Newtonian model, this effect was emphasized: viscosity ranged from 3.2 to 6.0 cP, deviating maximally from the standard value in low shear rate (i.e., <100 s-1) regions. This effect reflected on EL quantifications: in particular, as compared to Model I, Model III yielded markedly higher EL values (up to +40%) or markedly lower (down to -21%) for subjects with hematocrit higher than 39.5% and lower than 30%, respectively. Accounting for non-Newtonian blood behavior on a patient-specific basis may enhance the accuracy of intracardiac energetics assessment by 4D Flow, which may be explored as non-invasive index to discriminate between healthy and pathologic LV.

Crypt residing bacteria and proximal colonic carcinogenesis in a mouse model of Lynch syndrome.

Colorectal cancer is a multifactorial disease involving inherited DNA mutations, environmental factors, gut inflammation and intestinal microbiota. Certain germline mutations within the DNA mismatch repair system are associated with Lynch syndrome tumors including right-sided colorectal cancer with mucinous phenotype and presence of an inflammatory infiltrate. Such tumors are more often associated with bacterial biofilms, which may contribute to disease onset and progression. Inflammatory bowel diseases are also associated with colorectal cancer and intestinal dysbiosis. Herein we addressed the question, whether inflammation can aggravate colorectal cancer development under mismatch repair deficiency. MSH2loxP/loxP Vill-cre mice were crossed into the IL-10-/- background to study the importance of inflammation and mucosal bacteria as a driver of tumorigenesis in a Lynch syndrome mouse model. An increase in large bowel tumorigenesis was found in double knockout mice both under conventional housing and under specific pathogen-free conditions. This increase was mostly due to the development of proximal tumors, a hotspot for tumorigenesis in Lynch syndrome, and was associated with a higher degree of inflammation. Additionally, bacterial invasion into the mucus of tumor crypts was observed in the proximal tumors. Inflammation shifted fecal and mucosal microbiota composition and was associated with enrichment in Escherichia-Shigella as well as Akkermansia, Bacteroides and Parabacteroides genera in fecal samples. Tumor-bearing double knockout mice showed a similar enrichment for Escherichia-Shigella and Parabacteroides. Lactobacilli, Lachnospiraceae and Muribaculaceae family members were depleted upon inflammation. In summary, chronic inflammation aggravates colonic tumorigenesis under mismatch repair deficiency and is associated with a shift in microbiota composition.

Long-Term Consumption of Anthocyanin-Rich Fruit Juice: Impact on Gut Microbiota and Antioxidant Markers in Lymphocytes of Healthy Males.

Polyphenols are considered protective against diseases associated with oxidative stress. Short-term intake of an anthocyanin-rich fruit juice resulted in significantly reduced deoxyribonucleic acid (DNA) strand-breaks in peripheral blood lymphocytes (PBLs) and affected antioxidant markers in healthy volunteers. Consequently, effects of long-term consumption of fruit juice are of particular interest. In focus was the impact on nuclear factor erythroid 2 (NFE2)-related factor 2 (Nrf2), the Nrf2-regulated genes NAD(P)H quinone oxidoreductase 1 (NQO-1) and heme oxygenase 1 (HO-1) as well as effects on the gut microbiota. In a nine-week placebo-controlled intervention trial with 57 healthy male volunteers, consumption of anthocyanin-rich juice significantly increased NQO-1 and HO-1 transcript levels in PBLs compared to a placebo beverage as measured by real-time polymerase chain reaction (PCR). Three Nrf2-promotor single nucleotide polymorphisms (SNPs), analyzed by pyrosequencing, indicated an association between individual Nrf2 transcript levels and genotype. Moreover, the Nrf2 genotype appeared to correlate with the presence of specific microbial organisms identified by 16S-PCR and classified as Spirochaetaceae. Furthermore, the microbial community was significantly affected by the duration of juice consumption and intake of juice itself. Taken together, long-term consumption of anthocyanin-rich fruit juice affected Nrf2-dependent transcription in PBLs, indicating systemic effects. Individual Nrf2 genotypes may influence the antioxidant response, thus requiring consideration in future intervention studies focusing on the Nrf2 pathway. Anthocyanin-rich fruit juice had an extensive impact on the gut microbiota.

Prevalence study on potential drug-drug interaction in cancer patients in Piacenza hospital's Onco-Haematology department.

Background Cancer patients can be a human model of potential drug interactions. Usually they receive a large number of different medications, including antineoplastic agents, drugs for comorbid illness and medication for supportive care, however information about these interactions are fragmented and poor. Objective We assessed a prospective study to evaluate the prevalence of drug interaction among patients hospitalized in the Onco-Haematology department, Hospital of Piacenza. Methods Data on drugs administered for cancer, comorbidities, or supportive care were collected from different computerized prescription software in use in the department; we compared them with a database to focus on the co-administration of drugs. A literature review was performed to identify major potential drug interaction and to classify them by level of severity and by strengths of scientific evidence. Results In this study 284 cancer patients were enrolled; patients had taken an average of seven drugs on each day of therapy plus chemotherapeutic agents, we identified 67 potential drug interactions. At least 53 patients had one potential drug interaction. Of all potential drug interactions 63 were classified as moderate severity and only four as major. In 55 cases chemotherapeutic agents were involved in possible interactions with supportive care drugs, meanwhile in 12 cases the potential drug interactions were between supportive care drugs. Conclusions In our centre, thanks to a computerized prescription software, integrated with caution alarm in case of possible interaction, we had a lower rate of potential drug interactions than the one from literature. It is possible to improve the software integrating the alarm with the potential drug interactions between chemotherapy agents and supportive care drugs.

Absence of surrogate light chain results in spontaneous autoreactive germinal centres expanding V(H)81X-expressing B cells.

Random recombination of antibody heavy- and light-chain genes results in a diverse B-cell receptor (BCR) repertoire including self-reactive BCRs. However, tolerance mechanisms that prevent the development of self-reactive B cells remain incompletely understood. The absence of the surrogate light chain, which assembles with antibody heavy chain forming a pre-BCR, leads to production of antinuclear antibodies (ANAs). Here we show that the naive follicular B-cell pool is enriched for cells expressing prototypic ANA heavy chains in these mice in a non-autoimmune background with a broad antibody repertoire. This results in the spontaneous formation of T-cell-dependent germinal centres that are enriched with B cells expressing prototypic ANA heavy chains. However, peripheral tolerance appears maintained by selection thresholds on cells entering the memory B-cell and plasma cell pools, as exemplified by the exclusion of cells expressing the intrinsically self-reactive V(H)81X from both pools.

Safety and efficacy of granulocyte colony-stimulating factor biosimilars in engraftment after autologous stem cell transplantation for haematological malignancies: a 4-year, single institute experience with different conditioning regimens.

BACKGROUND: Filgrastim biosimilars have recently been introduced into clinical practice. To date biosimilars have demonstrated comparable efficacy and safety as the originator in chemotherapy-induced neutropenia. Published experience in engraftment after autologous stem cell transplantation (ASCT) is limited and concerns relatively few patients. MATERIALS AND METHODS: With the aim of assessing the efficacy and the safety of filgrastim biosimilars in post-ASCT bone marrow recovery, we conducted a single institution, retrospective study in 56 lymphoma and myeloma patients who received filgrastim biosimilars (Tevagrastim(®) and Zarzio(®)) at standard doses from day 5. We compared our results with recently published data on the originator. A cost analysis of each biosimilar was performed. RESULTS: Neutrophil counts recovered in 55 patients. The median number of filgrastim biosimilar vials injected was seven per patient. The median time to neutrophil and platelet recovery was 10 and 12 days, respectively. Twenty-six patients had febrile neutropenia, in half of whom the agent involved was identified. In the cost analysis, the use of Tevagrastim(®) and Zarzio(®) was associated with cost reductions of 56% and of 86%, respectively. DISCUSSION: Despite differences in CD34+ cell counts and time of starting filgrastim, our results in terms of time to engraftment and median number of vials injected are similar to published data. Comparing our results by single conditioning regimen to recent literature data, the time to engraftment and duration of hospitalisation were equivalent. Significant differences were observed in the incidence of febrile neutropenia, perhaps due to different preventive and prophylactic protocols for infections. Although prospective studies should be performed to confirm our results, filgrastim biosimilars were found to be effective and safe in engraftment after ASCT.

An Anticancer Drug Unit for the whole provincial oncologic network of Piacenza: improving safety and savings.

In recent years, the expenditure for cancer care is increased largely due to the increase in cancer prevalence, demographic changes and incorporation into clinical practice of new and expensive drugs. For these reasons, solutions to contain costs are necessaries. The drugs-related expenditure is proportionally higher in oncology than in other medical specialties and overcomes staffing costs for outpatient care. The introduction of additional measures to contain and reduce expenditures such as waste reduction and human resources optimization is highly desirable. On April 2013, we started a day-to-day monitoring of the consumption of drugs and developed an internal protocol for waste minimization, consisting of five measures. A computerized research through Medline, Cancerlit and Embase was performed, applying the words drug waste, cost-containment, Anticancer Drug Unit and stability instructions. Articles and abstracts were also identified by back-referencing from other relevant papers. Selected for the present review were papers published in English without limit of year. The day-to-day monitoring of the consumption of drugs and the internal protocol for waste minimization were able to achieve a saving of 15,700 every month. The projection of an annual cost-saving result of 188.00 corresponds to a recovery of 4 % on the spending for oncologic drugs. Our data show that in a proper structure working according to the standards of quality, safety and sterility, preserving and reusing the drug waste within the limits imposed by the datasheets, it is possible to achieve a cost-containment policy and produce durable benefits.

On the perils of poor editing: regulation of peptide loading by HLA-DQ and H2-A molecules associated with celiac disease and type 1 diabetes.

This review discusses mechanisms that link allelic variants of major histocompatibility complex (MHC) class II molecules (MHCII) to immune pathology. We focus on HLA (human leukocyte antigen)-DQ (DQ) alleles associated with celiac disease (CD) and type 1 diabetes (T1D) and the role of the murine DQ-like allele, H2-Ag7 (I-Ag7 or Ag7), in murine T1D. MHCII molecules bind peptides, and alleles vary in their peptide-binding specificity. Disease-associated alleles permit binding of disease-inducing peptides, such as gluten-derived, Glu-/Pro-rich gliadin peptides in CD and peptides from islet autoantigens, including insulin, in T1D. In addition, the CD-associated DQ2.5 and DQ8 alleles are unusual in their interactions with factors that regulate their peptide loading, invariant chain (Ii) and HLA-DM (DM). The same alleles, as well as other T1D DQ risk alleles (and Ag7), share nonpolar residues in place of Asp at ß57 and prefer peptides that place acidic side chains in a pocket in the MHCII groove (P9). Antigen-presenting cells from T1D-susceptible mice and humans retain CLIP because of poor DM editing, although underlying mechanisms differ between species. We propose that these effects on peptide presentation make key contributions to CD and T1D pathogenesis.

Measurement of protein synthesis using heavy water labeling and peptide mass spectrometry: Discrimination between major histocompatibility complex allotypes.

Methodological limitations have hampered the use of heavy water ((2)H(2)O), a convenient, universal biosynthetic label, for measuring protein synthesis. Analyses of (2)H-labeled amino acids are sensitive to contamination; labeling of peptides has been measured for a few serum proteins, but this approach awaits full validation. Here we describe a method for quantifying protein synthesis by peptide mass spectrometry (MS) after (2)H(2)O labeling, as applied to various proteins of the major histocompatibility complex (MHC). Human and murine antigen-presenting cells were cultured in medium containing 5% (2)H(2)O; class I and class II MHC proteins were immunoprecipitated, bands were excised, and Ala-/Gly-rich, allele-specific tryptic peptides were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Mass isotopomer distributions were quantified precisely by LC-MS and shifted markedly on (2)H(2)O labeling. Experimental data agreed closely with models obtained by mass isotopomer distribution analysis (MIDA) and were consistent with contributions from Ala, Gly, and other amino acids to labeling. Estimates of fractional protein synthesis from peptides of the same protein were precise and internally consistent. The method was capable of discriminating between MHC isotypes and alleles, applicable to primary cells, and readily extendable to other proteins. It simplifies measurements of protein synthesis, enabling novel applications in physiology, in genotype/phenotype interactions, and potentially in kinetic proteomics.

Therapeutic efficacy of buforin II and rifampin in a rat model of Acinetobacter baumannii sepsis.

OBJECTIVE: To investigate the efficacy of buforin II and rifampin in an experimental rat model of Acinetobacter baumannii sepsis. DESIGN: Prospective, randomized, controlled animal study. SETTING: Research laboratory in a university hospital. SUBJECTS: Adult male Wistar rats. INTERVENTIONS: The animals received intraperitoneally 1 mL saline containing 2 x 10 colony forming units of A. baumannii ATCC 19606 (model i) or the multiresistant strain (model ii). Immediately after bacterial challenge, animals received intravenously a single dose of isotonic sodium chloride solution (control groups C1 and C2), 1 mg/kg of buforin II, 10 mg/kg of rifampin, and 1 mg/kg of buforin II plus 10 mg/kg of rifampin, respectively. MEASUREMENTS AND MAIN RESULTS: Lethality, bacterial growth in blood and tissue burden, endotoxin, interleukin-6, and tumor necrosis factor-alpha concentrations in plasma. Buforin II showed good antimicrobial activity and achieved a significant reduction of plasma endotoxin and cytokines concentration when compared with control and rifampin-treated groups. Combination among buforin II proved to be the most effective treatment in reducing all variables measured. CONCLUSION: In an experimental model, buforin II and rifampin might have a potential role in the treatment of severe infections due to A. baumannii.

Protective effects of the combination of alpha-helical antimicrobial peptides and rifampicin in three rat models of Pseudomonas aeruginosa infection.

INTRODUCTION: An experimental study has been performed to compare the in vitro activity and the in vivo efficacy of magainin II and cecropin A with or without rifampicin against control and multidrug-resistant Pseudomonas aeruginosa strains. METHODS: In vitro experiments included MIC determinations and synergy studies. For in vivo studies, animals were given an intraperitoneal injection of P. aeruginosa lipopolysaccharide, P. aeruginosa ATCC 27853 and one clinical multiresistant P. aeruginosa strain. Groups of animals received intravenously isotonic sodium chloride solution, 10 mg/kg rifampicin, 1 mg/kg magainin II or 1 mg/kg cecropin A. Two groups of animals received a combined treatment with magainin II + rifampicin or cecropin A + rifampicin at the same dosages as the singly treated groups. In addition, a further group was treated with tazobactam/piperacillin (120 mg/kg). Lethality, bacterial growth in blood and peritoneum, and endotoxin and TNF-alpha concentrations in plasma were evaluated. RESULTS: Combinations of alpha-helical antimicrobial peptides showed in vitro synergistic interaction. Magainin II and cecropin A exerted strong antimicrobial activity and achieved a significant reduction in plasma endotoxin and TNF-alpha concentrations when compared with control and rifampicin-treated groups. Rifampicin exhibited no anti-P. aeruginosa activity and good substantial impact on endotoxin and TNF-alpha plasma concentrations. Combined treatment groups had significant reductions in bacterial count, positive blood cultures and mortality rates when compared with singly treated and control groups. CONCLUSIONS: Our results highlight the potential usefulness of these combinations that provide future therapeutic alternatives in P. aeruginosa infections.

Efficacy of the combination of tachyplesin III and clarithromycin in rat models of Escherichia coli sepsis.

We investigated the efficacy of tachyplesin III and clarithromycin in two experimental rat models of severe gram-negative bacterial infections. Adult male Wistar rats were given either (i) an intraperitoneal injection of 1 mg/kg Escherichia coli 0111:B4 lipopolysaccharide or (ii) 2 x 10(10) CFU of E. coli ATCC 25922. For each model, the animals received isotonic sodium chloride solution, 1 mg/kg tachyplesin III, 50 mg/kg clarithromycin, or 1 mg/kg tachyplesin III combined with 50 mg/kg clarithromycin intraperitoneally. Lethality, bacterial growth in the blood and peritoneum, and the concentrations of endotoxin and tumor necrosis factor alpha (TNF-alpha) in plasma were evaluated. All the compounds reduced the lethality of the infections compared to that for the controls. Tachyplesin III exerted a strong antimicrobial activity and achieved a significant reduction of endotoxin and TNF-alpha concentrations in plasma compared to those of the control and clarithromycin-treated groups. Clarithromycin exhibited no antimicrobial activity but had a good impact on endotoxin and TNF-alpha plasma concentrations. A combination of tachyplesin III and clarithromycin resulted in significant reductions in bacterial counts and proved to be the most-effective treatment in reducing all variables measured.

Censoring of autoreactive B cell development by the pre-B cell receptor.

Antibody diversity occurs randomly as B cells recombine their immunoglobulin (Ig) heavy- and light-chain genes during development. This process inevitably generates reactivity against self structures, and several mechanisms prevent the development of autoreactive B cells. We report here a role for the pre-B cell receptor, composed of Ig heavy and surrogate light chains, in the negative selection of cells expressing Ig heavy chains with the potential to generate autoantibodies. Surrogate light-chain-deficient (SLC-/-) mice harbored elevated levels of antinuclear antibodies (ANAs) in their serum and showed evidence of escape of pre-B cells expressing prototypic autoantibody heavy chains from negative selection, leading to mature autoantibody secreting CD21-CD23- B cells in the periphery. Thus, the pre-B cell receptor appears to censor the development of certain autoantibody-secreting cells and may represent an important factor in multifactorial autoimmune diseases.

RNAIII-inhibiting peptide enhances healing of wounds infected with methicillin-resistant Staphylococcus aureus.

Quorum sensing is a mechanism through which a bacterial population receives input from neighboring cells and elicits an appropriate response to enable survival within the host. Inhibiting quorum sensing by RNAIII-inhibiting peptide (RIP) has been demonstrated as a very effective mode of prevention and therapy for device-associated staphylococcal infections and was tested here for healing of wounds that are otherwise resistant to conventional antibiotics. Wounds, established through the panniculus carnosus of BALB/c mice, were inoculated with 5 x 10(7) CFU of methicillin-resistant Staphylococcus aureus. Mice were treated with Allevyn, RIP-soaked Allevyn (containing 20 microg RIP), daily intraperitoneal teicoplanin (7 mg/kg of body weight), Allevyn and teicoplanin, and RIP-soaked Allevyn and daily intraperitoneal teicoplanin. The main outcome measures were quantitative bacterial culture and histological examination with assessment of microvessel density and of vascular endothelial growth factor (VEGF) expression in tissue sections. Treatment with RIP-soaked Allevyn together with teicoplanin injection greatly reduced the bacterial load to 13 CFU/g (control untreated animals had 10(8) CFU/g bacteria). All other treatments were also significantly effective but only reduced the bacterial load to about 10(3) CFU/ml. Histological examination indicated that only treatment with RIP-soaked Allevyn with teicoplanin injection restored epithelial, granulation, and collagen scores, as well as microvessel density and VEGF expression, to the levels found with uninfected mice. In conclusion, we observed that RIP may be useful for the management of infected wounds and that it could represent an exciting and future alternative to the conventional antibiotics, at present considered the gold-standard treatments for methicillin-resistant S. aureus infections.

Efficacy of LL-37 and granulocyte colony-stimulating factor in a neutropenic murine sepsis due to Pseudomonas aeruginosa.

A promising therapeutic strategy for the management of severe Pseudomonas infection in neutropenic patients may result from the coadministration of colony-stimulating factors (CSFs) that help maintain immune competence and antimicrobial peptides, a novel generation of adjunctive therapeutic agents with antimicrobial and anti-inflammatory properties. A promising peptide with these properties is LL-37, the only member of the cathelicidin family of antimicrobial peptides found in humans. BALB/c male mice were rendered neutropenic by intraperitoneal administration of cyclophosphamide on days -4 and -2 preinfection. Septic shock was induced at time 0 by intraperitoneal injection of 2x10 colony-forming units of P. aeruginosa American Type Culture Collection (ATCC) 27853. All animals were randomized to receive intravenously isotonic sodium chloride solution, 1 mg/kg of LL-37, 20 mg/kg of imipenem, 0.1 mg/kg of granulocyte CSF (G-CSF), 1 mg/kg of LL-37+0.1 mg/kg of G-CSF, or 20 mg/kg of imipenem+0.1 mg/kg of G-CSF. Lethality and bacterial growth in blood, peritoneum, spleen, liver, and kidney were evaluated. All regimens were significantly superior to controls at reducing the mouse lethality rate and bacterial burden in organs. Particularly, the combination between LL-37 and G-CSF was the most effective in protecting neutropenic mice from the onset of sepsis and in vitro significantly reduced the apoptosis of neutrophils. Combination therapy between LL-37 and G-CSF is a promising therapeutic strategy for the management of severe Pseudomonas infection complicated by neutropenia.