Rhomboid protease AarA mediates quorum-sensing in Providencia stuartii by activating TatA of the twin-arginine translocase

The Providencia stuartii AarA protein is a member of the rhomboid family of intramembrane serine proteases and is required for the production of an unknown quorum-sensing molecule. In a screen to identify rhomboid-encoding genes from Proteus mirabilis, tatA was identified as a multicopy suppressor and restored extracellular signal production as well as complementing all other phenotypes of a Prov. stuartii aarA mutant. TatA is a component of the twin-arginine translocase (Tat) protein secretion pathway and likely forms a secretion pore. By contrast, the native tatA gene of Prov. stuartii in multicopy did not suppress an aarA mutation. We find that TatA in Prov. stuartii has a short N-terminal extension that was atypical of TatA proteins from most other bacteria. This extension was proteolytically removed by AarA both in vivo and in vitro. A Prov. stuartii TatA protein missing the first 7 aa restored the ability to rescue the aarA-dependent phenotypes. To verify that loss of the Tat system was responsible for the various phenotypes exhibited by an aarA mutant, a tatC-null allele was constructed. The tatC mutant exhibited the same phenotypes as an aarA mutant and was epistatic to aarA. These data provide a molecular explanation for the requirement of AarA in quorum-sensing and uncover a function for the Tat protein export system in the production of secreted signaling molecules. Finally, TatA represents a validated natural substrate for a prokaryotic rhomboid protease.     

In vivo evaluation of the HeartWare centrifugal ventricular assist device

In this study, long-term (90-day) hemocompatibility and end-organ effects of a centrifugal left ventricular assist device (the Heartware HVAD) were evaluated in 6 healthy sheep. The device was implanted into the left ventricular apex on beating hearts. The outflow graft of each device was anastomosed to the descending aorta. None of the sheep received anticoagulation or antiaggregation medication during the study. Hematologic and biochemical tests of liver and kidney function were performed pre-operatively (baseline) and throughout the study. Data associated with pump function were collected continuously until 90 +/- 1 days of support, at which time the sheep were humanely killed, and the end-organs were examined macroscopically and histopathologically. Hematologic and biochemical test results were within normal limits during the study period. There were no significant complications. Postmortem examination of the explanted organs revealed no evidence of ischemia or infarction, except in 2 sheep, in which small foci of infarction were detected in each of their left kidneys. There was no significant device failure. In all sheep, the pump's inflow and outflow conduits were free of thrombus. During the 90-day study, the HeartWare HVAD showed exceptional hemocompatibility and reliability, both of which are crucial to the clinical success of any implantable left ventricular assist device.     

A Randomized Trial of a Physical Conditioning Program to Enhance the Driving Performance of Older Persons

Background As the number of older drivers increases, concern has been raised about the potential safety implications. Flexibility, coordination, and speed of movement have been associated with older drivers’ on road performance. Objective To determine whether a multicomponent physical conditioning program targeted to axial and extremity flexibility, coordination, and speed of movement could improve driving performance among older drivers. Design Randomized controlled trial with blinded assignment and end point assessment. Participants randomized to intervention underwent graduated exercises; controls received home, environment safety modules. Participants Drivers, 178, age ≥ 70 years with physical, but without substantial visual (acuity 20/40 or better) or cognitive (Mini Mental State Examination score ≥24) impairments were recruited from clinics and community sources. Measurements On-road driving performance assessed by experienced evaluators in dual-brake equipped vehicle in urban, residential, and highway traffic. Performance rated three ways: (1) 36-item scale evaluating driving maneuvers and traffic situations; (2) evaluator’s overall rating; and (3) critical errors committed. Driving performance reassessed at 3 months by evaluator blinded to treatment group. Results Least squares mean change in road test scores at 3 months compared to baseline was 2.43 points higher in intervention than control participants (P = .03). Intervention drivers committed 37% fewer critical errors (P = .08); there were no significant differences in evaluator’s overall ratings (P = .29). No injuries were reported, and complaints of pain were rare. Conclusions This safe, well-tolerated intervention maintained driving performance, while controls declined during the study period. Having interventions that can maintain or enhance driving performance may allow clinician–patient discussions about driving to adopt a more positive tone, rather than focusing on driving limitation or cessation.     

Symptoms, affects, and self-rated health: evidence for a subjective trajectory of health

OBJECTIVE: Self-rated health (SRH) is known to predict mortality and other health outcomes better than objective ratings, suggesting that patients have important knowledge that physicians do not. The study assessed whether SRH reflects changes in internal states, specifically symptoms and affects. METHOD: In an event-sampling study, 54 elders completed a SRH measure, positive and negative affect scale, a symptom checklist, and a pain scale every evening for 8 weeks. Using lagged (time series) hierarchical regression, the authors modeled associations of SRH with previous symptoms, moods, and changes in symptoms and mood. RESULTS: The SRH was highest when symptoms had decreased from the previous day and lowest when symptoms had increased, suggesting that SRH reflects a sense of change. Symptoms and affects contributed independently to SRH. Self-rated health was more sensitive to positive than negative affect and also sensitive to changes of positive but not negative affect. DISCUSSION: Patients may possess a subjective trajectory of health-an awareness of changes in symptoms and affect. This trajectory may constitute an important component of SRH and help to explain its ability to predict health outcomes.     

A historical look at alcohol abuse trends in army and civilian populations, 1980-1995

OBJECTIVES: The purpose of this study was to compare civilian and Army alcohol-related hospitalization trends and to plot temporal changes in rates relative to alcohol-related legislation and social policies. METHOD: We compared population-based civilian and Army annual hospitalization rates for overall alcohol-related diagnoses and for alcohol-related diagnostic subgroups (1980-1995) and plotted them against civilian and military substance abuse regulations. Civilian data were adjusted to Army age, gender, and race. RESULTS: Although overall civilian and Army alcohol hospitalization rates were similar, alcohol subgroup rates varied. Simultaneous drug and alcohol abuse (polyabuse) rates were higher among civilians (16.6 per 10,000) than Army soldiers (5.1 per 10,000). Army rates for dependent alcohol-related disorders were higher and increased. Army nondependent alcohol disorders tracked with alcohol-related regulations as rates fell 69% between 1985 and 1995. CONCLUSION: Army and civilian alcohol abuse trends vary by abuse type. Without longitudinal, diagnosis-specific subgroup analyses, these trends would not have emerged. Army policies and screening may explain divergent nondependent alcohol abuse and lower polyabuse rates.     

Acquired Immunity to Malaria

Naturally acquired immunity to falciparum malaria protects millions of people routinely exposed to Plasmodium falciparum infection from severe disease and death. There is no clear concept about how this protection works. There is no general agreement about the rate of onset of acquired immunity or what constitutes the key determinants of protection; much less is there a consensus regarding the mechanism(s) of protection. This review summarizes what is understood about naturally acquired and experimentally induced immunity against malaria with the help of evolving insights provided by biotechnology and places these insights in the context of historical, clinical, and epidemiological observations. We advocate that naturally acquired immunity should be appreciated as being virtually 100% effective against severe disease and death among heavily exposed adults. Even the immunity that occurs in exposed infants may exceed 90% effectiveness. The induction of an adult-like immune status among high-risk infants in sub-Saharan Africa would greatly diminish disease and death caused by P. falciparum. The mechanism of naturally acquired immunity that occurs among adults living in areas of hyper- to holoendemicity should be understood with a view toward duplicating such protection in infants and young children in areas of endemicity.     

Curing CNS autoimmune disease with myelin‐reactive Foxp3+ Treg

The potential use of CD4⁺Foxp3⁺ Treg as a cellular therapy for autoimmune disease is of great interest. For clinical translation, the key objective is to reverse established disease. Here we demonstrate that myelin basic protein (MBP)‐reactive CD4⁺CD25⁺ Treg from TCR Tg mice, but not polyclonal (non‐MBP‐reactive) Treg, can transfer efficient protection against MBP‐induced EAE when used either directly from donor mice, or after in vitro expansion. MBP‐reactive Treg transfer also showed some ability to improve recovery from EAE initiated by T cells recognizing a distinct CNS autoantigen, proteolipid protein. Importantly, we also demonstrate for the first time in the context of EAE that in vitro‐expanded naturally occurring MBP‐reactive Treg can prevent disease relapse when given after the onset of clinical EAE. Our study also contains data pertaining to the most effective Treg sub‐population in vivo (CD4⁺CD25⁺CD62L^hi) and shows that their potent suppressive effects reflect stable expression of Foxp3, although not CD25 or CD62L. Our data provide proof of the principle that Treg‐based therapies can cure CNS autoimmune disease, highlight the challenges for clinical translation and open new avenues for assessing how changing immune function via Treg activity might impact on neurodegeneration and repair.     

Potential prognostic marker ubiquitin carboxyl-terminal hydrolase-L1 does not predict patient survival in non-small cell lung carcinoma

Background

Ubiquitin Carboxyl-Terminal Hydrolase-L1 (UCH-L1) is a deubiquitinating enzyme that is highly expressed throughout the central and peripheral nervous system and in cells of the diffuse neuroendocrine system. Aberrant function of UCH-L1 has been associated with neurological disorders such as Parkinson''s disease and Alzheimer''s disease. Moreover, UCH-L1 exhibits a variable expression pattern in cancer, acting either as a tumour suppressor or promoter, depending on the type of cancer. In non-small cell lung carcinoma primary tumour samples, UCH-L1 is highly expressed and is associated with an advanced tumour stage. This suggests UCH-L1 may be involved in oncogenic transformation and tumour invasion in NSCLC. However, the functional significance of UCH-L1 in the progression of NSCLC is unclear. The aim of this study was to investigate the role of UCH-L1 using NSCLC cell line models and to determine if it is clinically relevant as a prognostic marker for advanced stage disease.

Methods

UCH-L1 expression in NSCLC cell lines H838 and H157 was modulated by siRNA-knockdown, and the phenotypic changes were assessed by flow cytometry, haematoxylin & eosin (H&E) staining and poly (ADP-ribose) polymerase (PARP) cleavage. Metastatic potential was measured by the presence of phosphorylated myosin light chain (MLC2). Tumour microarrays were examined immunohistochemically for UCH-L1 expression. Kaplan-Meier curves were generated using UCH-L1 expression levels and patient survival data extracted from Gene Expression Omnibus data files.

Results

Expression of UCH-L1 was decreased by siRNA in both cell lines, resulting in increased cell death in H838 adenocarcinoma cells but not in the H157 squamous cell line. However, metastatic potential was reduced in H157 cells. Immunohistochemical staining of UCH-L1 in patient tumours confirmed it was preferentially expressed in squamous cell carcinoma rather than adenocarcinoma. However the Kaplan-Meier curves generated showed no correlation between UCH-L1 expression levels and patient outcome.

Conclusions

Although UCH-L1 appears to be involved in carcinogenic processes in NSCLC cell lines, the absence of correlation with patient survival indicates that caution is required in the use of UCH-L1 as a potential prognostic marker for advanced stage and metastasis in lung carcinoma.