Clinical grade allogeneic human mesenchymal stem cells restore alveolar fluid clearance in human lungs rejected for transplantation

The lack of suitable donors for all solid-organ transplant programs is exacerbated in lung transplantation by the low utilization of potential donor lungs, due primarily to donor lung injury and dysfunction, including pulmonary edema. The current studies were designed to determine if intravenous clinical-grade human mesenchymal stem (stromal) cells (hMSCs) would be effective in restoring alveolar fluid clearance (AFC) in the human ex vivo lung perfusion model, using lungs that had been deemed unsuitable for transplantation and had been subjected to prolonged ischemic time. The human lungs were perfused with 5% albumin in a balanced electrolyte solution and oxygenated with continuous positive airway pressure. Baseline AFC was measured in the control lobe and if AFC was impaired (defined as

Proteomics on porcine haptoglobin and IgG/IgA show protein species distribution and glycosylation pattern to remain similar in PCV2-SD infection

Haptoglobin (Hp) and immunoglobulins are plasma glycoproteins involved in the immune reaction of the organism after infection and/or inflammation. Porcine circovirus type 2-systemic disease (PCV2-SD), formerly known as postweaning multisystemic wasting syndrome (PMWS), is a globally spread pig disease of great economic impact. PCV2-SD affects the immunological system of pigs causing immunosuppression. The aim of this work was to characterize the Hp protein species of healthy and PCV2-SD affected pigs, as well as the protein backbone and the glycan chain composition of porcine Hp. PCV2-SD affected pigs had an increased overall Hp level, but it did not affect the ratio between Hp species. Glycoproteomic analysis of the Hp β subunits confirmed that porcine Hp is N-glycosylated and, unexpectedly, O-glycosylated, a PTM that is not found on Hp from healthy humans. The glyco-profile of porcine IgG and IgA heavy chains was also characterized; decreased levels of both proteins were found in the investigated group of PCV2-SD affected pigs. Obtained results indicate that no significant changes in the N- and O-glycosylation patterns of these major porcine plasma glycoproteins were detectable between healthy and PCV2-SD affected animals.

Development and characterization of self-assembling nanoparticles using a bio-inspired amphipathic peptide for gene delivery

The design of a non-viral gene delivery vehicle capable of delivering and releasing a functional nucleic acid cargo intracellularly remains a formidable challenge. For systemic gene therapy to be successful a delivery vehicle is required that protects the nucleic acid cargo from enzymatic degradation, extravasates from the vasculature, traverses the cell membrane, disrupts the endosomal vesicles and unloads the cargo at its destination site, namely the nucleus for the purposes of gene delivery. This manuscript reports the extensive investigation of a novel amphipathic peptide composed of repeating RALA units capable of overcoming the biological barriers to gene delivery both in vitro and in vivo. Our data demonstrates the spontaneous self-assembly of cationic DNA-loaded nanoparticles when the peptide is complexed with pDNA. Nanoparticles were < 100 nm, were stable in the presence of serum and were fusogenic in nature, with increased peptide α-helicity at a lower pH. Nanoparticles proved to be non-cytotoxic, readily traversed the plasma membrane of both cancer and fibroblast cell lines and elicited reporter-gene expression following intravenous delivery in vivo. The results of this study indicate that RALA presents an exciting delivery platform for the systemic delivery of nucleic acid therapeutics.

MIC-1 (a multifunctional modulator of dendritic cell phenotype and function) is produced by decidual stromal cells and trophoblasts

Macrophage inhibitory cytokine-1 (MIC-1) is a multifunctional cytokine produced in high amounts by placental tissue. Inhibiting trophoblast invasion and suppressing inflammation through inhibition of macrophage activation, MIC-1 is thought to provide pleiotropic functions in the establishment and maintenance of pregnancy. So far, little is known about the decidual cell subsets producing MIC-1 and the effect of this cytokine on dendritic cells (DCs), which are known to play a distinct role in the development of pro-fetal tolerance in pregnancy.

The use of theranostic gadolinium-based nanoprobes to improve radiotherapy efficacy

A new efficient type of gadolinium-based theranostic agent (AGuIX®) has recently been developed for MRI-guided radiotherapy (RT). These new particles consist of a polysiloxane network surrounded by a number of gadolinium chelates, usually 10. Owing to their small size (<5 nm), AGuIX typically exhibit biodistributions that are almost ideal for diagnostic and therapeutic purposes. For example, although a significant proportion of these particles accumulate in tumours, the remainder is rapidly eliminated by the renal route. In addition, in the absence of irradiation, the nanoparticles are well tolerated even at very high dose (10 times more than the dose used for mouse treatment). AGuIX particles have been proven to act as efficient radiosensitizers in a large variety of experimental in vitro scenarios, including different radioresistant cell lines, irradiation energies and radiation sources (sensitizing enhancement ratio ranging from 1.1 to 2.5). Pre-clinical studies have also demonstrated the impact of these particles on different heterotopic and orthotopic tumours, with both intratumoural or intravenous injection routes. A significant therapeutical effect has been observed in all contexts. Furthermore, MRI monitoring was proven to efficiently aid in determining a RT protocol and assessing tumour evolution following treatment. The usual theoretical models, based on energy attenuation and macroscopic dose enhancement, cannot account for all the results that have been obtained. Only theoretical models, which take into account the Auger electron cascades that occur between the different atoms constituting the particle and the related high radical concentrations in the vicinity of the particle, provide an explanation for the complex cell damage and death observed.

Rifampicin and sodium fusidate reduces the frequency of methicillin-resistant Staphylococcus aureus (MRSA) isolation in adults with cystic fibrosis and chronic MRSA infection

Nosocomial transmission of methicillin-resistant Staphylococcus aureus (MRSA) to patients with cystic fibrosis (CF) frequently results in chronic respiratory tract carriage. This is an increasing problem, adds to the burden of glycopeptide antibiotic use in hospitals, and represents a relative contraindication to lung transplantation. The aim of this study was to determine whether it is possible to eradicate MRSA with prolonged oral combination antibiotics, and whether this treatment is associated with improved clinical status. Adult CF patients (six male, one female) with chronic MRSA infection were treated for six months with rifampicin and sodium fusidate. Outcome data were examined for six months before treatment, on treatment and after treatment. The patients had a mean age of 29.3 (standard deviation=6.3) years and FEV(1) of 36.1% (standard deviation=12.7) predicted. The mean duration of MRSA isolation was 31 months. MRSA isolates identified in these patients was of the same lineage as the known endemic strain at the hospital when assessed by pulsed-field gel electrophoresis. Five of the seven had no evidence of MRSA during and for at least six months after rifampicin and sodium fusidate. The proportion of sputum samples positive for MRSA was lower during the six months of treatment (0.13) and after treatment (0.19) compared with before treatment (0.85) (P<0.0001). There was a reduction in the number of days of intravenous antibiotics per six months with 20.3+/-17.6 on treatment compared with 50.7 before treatment and 33.0 after treatment (P=0.02). There was no change in lung function. Gastrointestinal side effects occurred in three, but led to therapy cessation in only one patient. Despite the use of antibiotics with anti-staphylococcal activity for treatment of respiratory exacerbation, MRSA infection persists. MRSA can be eradicated from the sputum of patients with CF and chronic MRSA carriage by using rifampicin and sodium fusidate for six months. This finding was associated with a significant reduction in the duration of intravenous antibiotic treatment during therapy.

Burkholderia pseudomallei:another emerging pathogen in cystic fibrosis

BACKGROUND: Burkholderia pseudomallei is an important cause of acute fulminant pneumonia and septicaemia in tropical regions of northern Australia and south east Asia. Subacute and chronic forms of the disease also occur. There have been three recent reports of adults with cystic fibrosis (CF) who presumably acquired B pseudomallei infection during extended vacations or residence in either Thailand or northern Australia.

METHODS: The clinical course, molecular characteristics, serology and response to treatment are described in four adult CF patients infected with B pseudomallei. Polymerase chain reaction (PCR) based methods were used to confirm B pseudomallei and exclude B cepacia complex. Genotyping was performed using randomly amplified polymorphic DNA (RAPD) PCR and pulsed field gel electrophoresis (PFGE).

RESULTS: Four patients are described with a mean duration of infection of 32 months. All but one patient lived in tropical Queensland. Two patients (with the longest duration of infection) deteriorated clinically and one subsequently died of respiratory failure. Both responded to intravenous treatment specifically targeting B pseudomallei. Another patient suffered two severe episodes of acute bronchopneumonia following acquisition of B pseudomallei. Eradication of the organism was not possible in any of the cases. PFGE of a sample isolate from each patient revealed the strains to be unique and RAPD analysis showed retention of the same strain within an individual over time.

CONCLUSIONS: These findings support a potential pathogenic role for B pseudomallei in CF lung disease, producing both chronic infection and possibly acute bronchopneumonia. Identical isolates are retained over time and are unique, consistent with likely environmental acquisition and not person to person spread. B pseudomallei is emerging as a significant pathogen for patients with CF residing and holidaying in the tropics.

Precision Teaching in IV Fluid Prescribing and Patient Safety

OBJECTIVES: Precision Teaching (PT) has been shown to be an effective intervention to assess teaching method effectiveness and evaluate learning outcomes. SAFMEDS (Say All Fast Minute Every Day Shuffled) are a practice/assessment procedure within the PT framework to assist learning and fluency. We explored the effects of a brief intervention with PT, to impart high frequency performance in safe intravenous fluid prescription in a group of final year undergraduate medical students.
METHODS: 133 final year undergraduate medical students completed a multiple choice question (MCQ) test on safe IV fluid prescription at the beginning and end of the study. The control group (n= 76) of students were taught using a current standardized teaching method. Students allocated to the intervention arm of the study were additionally instructed on PT and the use of SAFMEDS. The study group (n = 57) received 50 SAFMEDS cards containing information on the principles of IV fluid prescription scenarios. These students were trained/tested twice per day for 1 minute.
RESULTS: Interim analysis showed that the study group displayed an improvement in fluency and accuracy as the study progressed. There was a statistically significant improvement in MCQ performance for the PT group compared with the control group between the beginning and end of the study (35% vs 15%).
CONCLUSION: These results suggest PT employing SAFMEDS is an effective method for improving fluency, accuracy and patient safety in intravenous fluid prescribing amongst undergraduate medical students.

Seroepidemiology of norovirus-associated travelers' diarrhea

Background: Noroviruses (NoVs) are the most common cause of epidemic gastroenteritis, responsible for at least 50% of all gastroenteritis outbreaks worldwide and were recently identified as a leading cause of travelers' diarrhea (TD) in US and European travelers to Mexico, Guatemala, and India.

Methods: Serum and diarrheic stool samples were collected from 75 US student travelers to Cuernavaca, Mexico, who developed TD. NoV RNA was detected in acute diarrheic stool samples using reverse transcription-polymerase chain reaction (RT-PCR). Serology assays were performed using GI.1 Norwalk virus (NV) and GII.4 Houston virus (HOV) virus-like particles (VLPs) to measure serum levels of immunoglobulin A (IgA) and IgG by dissociation-enhanced lanthanide fluorescent immunoassay (DELFIA); serum IgM was measured by capture enzyme-linked immunosorbent assay (ELISA), and the 50% antibody-blocking titer (BT50 ) was determined by a carbohydrate-blocking assay.

Results: NoV infection was identified in 12 (16%; 9 GI-NoV and 3 GII-NoV) of 75 travelers by either RT-PCR or fourfold or more rise in antibody titer. Significantly more individuals had detectable preexisting IgA antibodies against HOV (62/75, 83%) than against NV (49/75, 65%) (p = 0.025) VLPs. A significant difference was observed between NV- and HOV-specific preexisting IgA antibody levels (p = 0.0037), IgG (p = 0.003), and BT50 (p = <0.0001). None of the NoV-infected TD travelers had BT50  > 200, a level that has been described previously as a possible correlate of protection.

Conclusion: We found that GI-NoVs are commonly associated with TD cases identified in US adults traveling to Mexico, and seroprevalence rates and geometric mean antibody levels to a GI-NoV were lower than to a GII-NoV strain.

Treatment adherence and health outcomes in patients with bronchiectasis

Background: We aimed to determine adherence to inhaled antibiotics, other respiratory medicines and airway clearance and to determine the association between adherence to these treatments and health outcomes (pulmonary exacerbations, lung function and Quality of Life Questionnaire-Bronchiectasis [QOL-B]) in bronchiectasis after 12 months.

Methods: Patients with bronchiectasis prescribed inhaled antibiotics for Pseudomonas aeruginosa infection were recruited into a one-year study. Participants were categorised as " adherent" to medication (medication possession ratio ≥80% using prescription data) or airway clearance (score ≥80% in the Modified Self-Reported Medication-Taking Scale). Pulmonary exacerbations were defined as treatment with a new course of oral or intravenous antibiotics over the one-year study. Spirometry and QOL-B were completed at baseline and 12 months. Associations between adherence to treatment and pulmonary exacerbations, lung function and QOL-B were determined by regression analyses.

Results: Seventy-five participants were recruited. Thirty-five (53%), 39 (53%) and 31 (41%) participants were adherent to inhaled antibiotics, other respiratory medicines, and airway clearance, respectively. Twelve (16%) participants were adherent to all treatments. Participants who were adherent to inhaled antibiotics had significantly fewer exacerbations compared to non-adherent participants (2.6 vs 4, p = 0.00) and adherence to inhaled antibiotics was independently associated with having fewer pulmonary exacerbations (regression co-efficient = -0.51, 95% CI [-0.81,-0.21], p < 0.001). Adherence to airway clearance was associated with lower QOL-B Treatment Burden (regression co-efficient = -15.46, 95% CI [-26.54, -4.37], p < 0.01) and Respiratory Symptoms domain scores (regression co-efficient = -10.77, 95% CI [-21.45; -0.09], p < 0.05). There were no associations between adherence to other respiratory medicines and any of the outcomes tested. Adherence to treatment was not associated with FEV₁ % predicted.

Conclusions: Treatment adherence is low in bronchiectasis and affects important health outcomes including pulmonary exacerbations. Adherence should be measured as part of bronchiectasis management and future research should evaluate bronchiectasis-specific adherence strategies. 

Lung Clearance Index in cystic fibrosis subjects treated for Pulmonary Exacerbations

Pulmonary exacerbations are important clinical events for cystic fibrosis (CF) patients. Studies assessing the ability of the lung clearance index (LCI) to detect treatment response for pulmonary exacerbations have yielded heterogeneous results. Here, we conduct a retrospective analysis of pooled LCI data to assess treatment with intravenous antibiotics for pulmonary exacerbations and to understand factors explaining the heterogeneous response.

A systematic literature search was performed to identify prospective observational studies. Factors predicting the relative change in LCI and spirometry were evaluated while adjusting for within-study clustering.

Six previously reported studies and one unpublished study, which included 176 pulmonary exacerbations in both paediatric and adult patients, were included. Overall, LCI significantly decreased by 0.40 units (95% CI -0.60 -0.19, p=0.004) or 2.5% following treatment. The relative change in LCI was significantly correlated with the relative change in forced expiratory volume in 1 s (FEV1), but results were discordant in 42.5% of subjects (80 out of 188). Higher (worse) baseline LCI was associated with a greater improvement in LCI (slope: -0.9%, 95% CI -1.0- -0.4%).

LCI response to therapy for pulmonary exacerbations is heterogeneous in CF patients; the overall effect size is small and results are often discordant with FEV1.

siRNA Technology in Kidney Transplantation: Current Status and Future Potential

Kidney transplantation is one of the most common transplantation operations in the world, accounting for up to 50 % of all transplantation surgeries. To curtail the damage to transplanted organs that is caused by ischemia-reperfusion injury and the recipient's immune system, small interfering RNA (siRNA) technology is being explored. Importantly, the kidney as a whole is a preferential site for non-specific systemic delivery of siRNA. To date, most attempts at siRNA-based therapy for transplantation-related conditions have remained at the in vitro stage, with only a few of them being advanced into animal models. Hydrodynamic intravenous injection of naked or carrier-bound siRNAs is currently the most common route for delivery of therapeutic constructs. To our knowledge, no systematic screens for siRNA targets most relevant for kidney transplantation have been attempted so far. A majority of researchers have arrived at one or another target of interest by analyzing current literature that dissects pathological processes taking place in transplanted organs. A majority of the genes that make up the list of 53 siRNA targets that have been tested in transplantation-related models so far belong to either apoptosis- or immune rejection-centered networks. There is an opportunity for therapeutic siRNA combinations that may be delivered within the same delivery vector or injected at the same time and, by targeting more than one pathway, or by hitting the same pathways within two different key points, will augment the effects of each other.

Laser-engineered dissolving microneedle arrays for protein delivery: potential for enhanced intradermal vaccination

OBJECTIVES: We aimed to highlight the utility of novel dissolving microneedle (MN)-based delivery systems for enhanced transdermal protein delivery. Vaccination remains the most accepted and effective approach in offering protection from infectious diseases. In recent years, much interest has focused on the possibility of using minimally invasive MN technologies to replace conventional hypodermic vaccine injections.

METHODS: The focus of this study was exploitation of dissolving MN array devices fabricated from 20% w/w poly(methyl vinyl ether/maleic acid) using a micromoulding technique, for the facilitated delivery of a model antigen, ovalbumin (OVA).

KEY FINDINGS: A series of in-vitro and in-vivo experiments were designed to demonstrate that MN arrays loaded with OVA penetrated the stratum corneum and delivered their payload systemically. The latter was evidenced by the activation of both humoral and cellular inflammatory responses in mice, indicated by the production of immunoglobulins (IgG, IgG1, IgG2a) and inflammatory cytokines, specifically interferon-gamma and interleukin-4. Importantly, the structural integrity of the OVA following incorporation into the MN arrays was maintained.

CONCLUSION: While enhanced manufacturing strategies are required to improve delivery efficiency and reduce waste, dissolving MN are a promising candidate for 'reduced-risk' vaccination and protein delivery strategies.

An audit of antimicrobial treatment of lower respiratory and urinary tract infections in a hospital setting

Objectives: To audit the quality of treatment of lower respiratory tract infections (LRTIs) and urinary tract infections (UTIs) and to identify targets for antibiotic stewardship. Methods: The audit involved collecting data on admitted patients, who were diagnosed with LRTIs or UTIs and subsequently received antibiotic treatment (January 2009-April 2009). Key findings: The percentage adherence rate for hospital antibiotic policy was 68.6% (24/35). Documentation of the CURB-65 score was found in 80% (16/20) of the patients' clinical notes, for which 46.2% (6/13) of patients were treated according to their CURB- 65 score. The percentages of delayed and missed doses for all antibiotics were 21.7% (254/1171) and 8.6% (101/1171), respectively. The percentage of patients switched from intravenous to oral antibiotics in accordance with the policy was 58.5% (31/53). The mean length of stay for patients switched in line with the guidelines was 6.9 days (range: 2-18 days) compared with 13.2 days (range: 4-28 days) for patients treated with intravenous antibiotics >24 h after the intravenous to oral switch criteria were fulfilled; this equates to on average an extra 6.3 days of hospitalisation (p=0.01). Conclusions: The study identified a number of targets for quality improvement including adherence to antibiotic policy, documentation of the CURB-65 score in patients' notes and treating patients accordingly, addressing the issue of missed and delayed doses, and maintaining adherence to the hospital intravenous-to-oral antibiotic switch policy. The findings suggest that the quality of antibiotic prescribing could be improved by measuring and addressing such performance indicators.

Effect of radium-223 dichloride on symptomatic skeletal events in patients with castration-resistant prostate cancer and bone metastases: results from a phase 3, double-blind, randomised trial

BACKGROUND: Bone metastases frequently cause skeletal events in patients with metastatic castration-resistant prostate cancer. Radium-223 dichloride (radium-223) selectively targets bone metastases with high-energy, short-range α-particles. We assessed the effect of radium-223 compared with placebo in patients with castration-resistant prostate cancer and bone metastases.

METHODS: In this phase 3, double-blind, randomised ALSYMPCA trial, we enrolled patients who had symptomatic castration-resistant prostate cancer with two or more bone metastases and no known visceral metastases, who were receiving best standard of care, and had previously either received or were unsuitable for docetaxel. Patients were stratified by previous docetaxel use, baseline total alkaline phosphatase level, and current bisphosphonate use, then randomly assigned (2:1) to receive either six intravenous injections of radium-223 (50 kBq/kg) or matching placebo; one injection was given every 4 weeks. Randomisation was done with an interactive voice response system, taking into account trial stratification factors. Participants and investigators were masked to treatment assignment. The primary endpoint was overall survival, which has been reported previously. Here we report on time to first symptomatic skeletal event, defined as the use of external beam radiation to relieve bone pain, or occurrence of a new symptomatic pathological fracture (vertebral or non-verterbal), or occurence of spinal cord compression, or tumour-related orthopeadic surgical intervention. All events were required to be clinically apparent and were not assessed by periodic radiological review. Statistical analyses of symptomatic skeletal events were based on the intention-to-treat population. The study has been completed and is registered with ClinicalTrials.gov, number NCT00699751.

FINDINGS: Between June 12, 2008, and Feb 1, 2011, 921 patients were enrolled, of whom 614 (67%) were randomly assigned to receive radium-223 and 307 (33%) placebo. Symptomatic skeletal events occurred in 202 (33%) of 614 patients in the radium-223 group and 116 (38%) of 307 patients in the placebo group. Time to first symptomatic skeletal event was longer with radium-223 than with placebo (median 15·6 months [95% CI 13·5-18·0] vs 9·8 months [7·3-23·7]; hazard ratio [HR]=0·66, 95% CI 0·52-0·83; p=0·00037). The risks of external beam radiation therapy for bone pain (HR 0·67, 95% CI 0·53-0·85) and spinal cord compression (HR=0·52, 95% CI 0·29-0·93) were reduced with radium-233 compared with placebo. Radium-223 treatment did not seem to significantly reduce the risk of symptomatic pathological bone fracture (HR 0·62, 95% CI 0·35-1·09), or the need for tumour-related orthopaedic surgical intervention (HR 0·72, 95% CI 0·28-1·82).

INTERPRETATION: Radium-223 should be considered as a treatment option for patients with castration-resistant prostate cancer and symptomatic bone metastases.

FUNDING: Algeta and Bayer HealthCare Pharmaceuticals.