Self-sufficient control of urate homeostasis in mice by a synthetic circuit

Synthetic biology has shown that the metabolic behavior of mammalian cells can be altered by genetic devices such as epigenetic and hysteretic switches, timers and oscillators, biocomputers, hormone systems and heterologous metabolic shunts. To explore the potential of such devices for therapeutic strategies, we designed a synthetic mammalian circuit to maintain uric acid homeostasis in the bloodstream, disturbance of which is associated with tumor lysis syndrome and gout. This synthetic device consists of a modified Deinococcus radiodurans-derived protein that senses uric acids levels and triggers dose-dependent derepression of a secretion-engineered Aspergillus flavus urate oxidase that eliminates uric acid. In urate oxidase-deficient mice, which develop acute hyperuricemia, the synthetic circuit decreased blood urate concentration to stable sub-pathologic levels in a dose-dependent manner and reduced uric acid crystal deposits in the kidney. Synthetic gene-network devices providing self-sufficient control of pathologic metabolites represent molecular prostheses, which may foster advances in future gene- and cell-based therapies.

Eukaryotic translation elongation factor 1A (eEF1A) domain I from S. cerevisiae is required but not sufficient for inter-species complementation

Ethanolamine phosphoglycerol (EPG) is a protein modification attached exclusively to eukaryotic elongation factor 1A (eEF1A). In mammals and plants, EPG is linked to conserved glutamate residues located in eEF1A domains II and III, whereas in the unicellular eukaryote Trypanosoma brucei, only domain III is modified by a single EPG. A biosynthetic precursor of EPG and structural requirements for EPG attachment to T. brucei eEF1A have been reported, but nothing is known about the EPG modifying enzyme(s). By expressing human eEF1A in T. brucei, we now show that EPG attachment to eEF1A is evolutionarily conserved between T. brucei and Homo sapiens. In contrast, S. cerevisiae eEF1A, which has been shown to lack EPG is not modified in T. brucei. Furthermore, we show that eEF1A cannot functionally complement across species when using T. brucei and S. cerevisiae as model organisms. However, functional complementation in yeast can be obtained using eEF1A chimera containing domains II or III from other species. In contrast, yeast domain I is strictly required for functional complementation in S. cerevisiae.

Current hot potatoes in atrial fibrillation ablation

Atrial fibrillation (AF) ablation has evolved to the treatment of choice for patients with drug-resistant and symptomatic AF. Pulmonary vein isolation at the ostial or antral level usually is sufficient for treatment of true paroxysmal AF. For persistent AF ablation, drivers and perpetuators outside of the pulmonary veins are responsible for AF maintenance and have to be targeted to achieve satisfying arrhythmia-free success rate. Both complex fractionated atrial electrogram (CFAE) ablation and linear ablation are added to pulmonary vein isolation for persistent AF ablation. Nevertheless, ablation failure and necessity of repeat ablations are still frequent, especially after persistent AF ablation. Pulmonary vein reconduction is the main reason for arrhythmia recurrence after paroxysmal and to a lesser extent after persistent AF ablation. Failure of persistent AF ablation mostly is a consequence of inadequate trigger ablation, substrate modification or incompletely ablated or reconducting linear lesions. In this review we will discuss these points responsible for AF recurrence after ablation and review current possibilities on how to overcome these limitations.

Effects of sleep fragmentation on the arousability to resistive loading in NREM and REM sleep in normal men

STUDY OBJECTIVE: In healthy subjects, arousability to inspiratory resistive loading is greater during rapid eye movement (REM) sleep compared with non-REM (NREM) sleep but is poorest in REM sleep in patients with sleep apnea. We therefore examined the hypothesis that sleep fragmentation impairs arousability, especially from REM sleep. DESIGN: Two blocks of 3 polysomnographies (separated by at least 1 week) were performed randomly. An inspiratory-loaded night followed either 2 undisturbed control nights (LN(C)) or 2 acoustically fragmented nights (LN(F)) SETTING: Sleep laboratory. PARTICIPANTS: Sixteen healthy men aged 20 to 29 years. INTERVENTIONS: In both loaded nights, an inspiratory resistive load was added via a valved facemask every 2 minutes during sleep and turned off either when arousal occurred or after 2 minutes. MEASUREMENTS AND RESULTS: During LN(F), arousability remained significantly greater in REM sleep (71% aroused within 2 minutes) compared with stage 2 (29%) or stage 3/4 (16%) sleep. After sleep fragmentation, arousability was decreased in stage 2 sleep (LN(F): 29%; LN(C): 38%; p < .05) and low in early REM sleep, increasing across the night (p < .01). In stage 3/4 sleep, neither an attenuation nor a change across the night was seen after sleep fragmentation. CONCLUSIONS: Mild sleep fragmentation is already sufficient to attenuate arousability in stage 2 sleep and to decrease arousability in early, compared with late, REM sleep. This means that sleep fragmentation affects the arousal response to increasing resistance and that the effects are different in stage 2 and REM sleep. The biologic reason for this increase in the arousal response in REM sleep across the night is not clear.

Is restricted femoral navigation sufficient for accuracy of total knee arthroplasty?

A total knee arthroplasty performed with navigation results in more accurate component positioning with fewer outliers. It is not known whether image-based or image-free-systems are preferable and if navigation for only one component leads to equal accuracy in leg alignment than navigation of both components. We evaluated the results of total knee arthroplasties performed with femoral navigation. We studied 90 knees in 88 patients who had conventional total knee arthroplasties, image-based total knee arthroplasties, or total knee arthroplasties with image-free navigation. We compared patients' perioperative times, component alignment accuracy, and short-term outcomes. The total surgical time was longer in the image-based total knee arthroplasty group (109 +/- 7 minutes) compared with the image-free (101 +/- 17 minutes) and conventional total knee arthroplasty groups (87 +/- 20 minutes). The mechanical axis of the leg was within 3 degrees of neutral alignment, although the conventional total knee arthroplasty group showed more (10.6 degrees ) variance than the navigated groups (5.8 degrees and 6.4 degrees , respectively). We found a positive correlation between femoral component malalignment and the total mechanical axis in the conventional group. Our results suggest image-based navigation is not necessary, and image-free femoral navigation may be sufficient for accurate component alignment.

[Vasculitis as a reason of chronic headache]

A 13-year-old girl presented to our emergency with a one week history of fever and skin rash and new onset of chorea for the last three days. There was a long standing history of right predominant headache; followed by personality change, fatigue, arthralgia and weight loss over the last few months. Previous investigations by head CT and ophthalmological examination did not explain the symptoms. Further investigations revealed peri- and pancarditis with aortic insufficiency, a renal involvement with elevated creatinin, protein- and hematuria and a hemolytic anemia. Diagnosis of lupus eythematodes was confirmed by high ANA, anti-dsDNS and Anticardiolipin antibodies. Within the first 48 hours after admission there was significant deterioration with reduced vigilance and dysarthria. MRI of the brain and dopplersonography of cerebral vessels showed a complete thrombosis of the right medial cerebral artery with a small net of collaterals, irregularities of the left cerebral artery due to vasculitis and several subacute leftsided ischemias. Immunosuppressive therapy with high-dose corticosteroids and cyclophosphamid together with antithrombotic therapy induced an improvement of neurologic, renal and cardiac function.

Transmembrane tumor necrosis factor alpha is required for enteropathy and is sufficient to promote parasite expulsion in gastrointestinal helminth infection

To study the specific role of transmembrane tumor necrosis factor (tmTNF) in protective and pathological responses against the gastrointestinal helminth Trichinella spiralis, we compared the immune responses of TNF-alpha/lymphotoxin alpha (LTalpha)(-/-) mice expressing noncleavable transgenic tmTNF to those of TNF-alpha/LTalpha(-/-) and wild-type mice. The susceptibility of TNF-alpha/LTalpha(-/-) mice to T. spiralis infection was associated with impaired induction of a protective Th2 response and the lack of mucosal mastocytosis. Although tmTNF-expressing transgenic (tmTNF-tg) mice also had a reduced Th2 response, the mast cell response was greater than that observed in TNF-alpha/LTalpha(-/-) mice and was sufficient to induce the expulsion of the parasite. T. spiralis infection of tmTNF-tg mice resulted in significant intestinal pathology characterized by villus atrophy and crypt hyperplasia comparable to that induced following the infection of wild-type mice, while pathology in TNF-alpha/LTalpha(-/-) mice was significantly reduced. Our data thus indicate a role for tmTNF in host defense against gastrointestinal helminths and in the accompanying enteropathy. Furthermore, they also demonstrate that TNF-alpha is required for the induction of Th2 immune responses related to infection with gastrointestinal helminth parasites.

Production of interferon-gamma by activated T-cell receptor-alphabeta CD8alphabeta intestinal intraepithelial lymphocytes is required and sufficient for disruption of the intestinal barrier integrity

Maintenance of intestinal epithelial barrier function is of vital importance in preventing uncontrolled influx of antigens and the potentially ensuing inflammatory disorders. Intestinal intraepithelial lymphocytes (IEL) are in intimate contact with epithelial cells and may critically regulate the epithelial barrier integrity. While a preserving impact has been ascribed to the T-cell receptor (TCR)-gammadelta subset of IEL, IEL have also been shown to attenuate the barrier function. The present study sought to clarify the effects of IEL by specifically investigating the influence of the TCR-alphabeta CD8alphabeta and TCR-alphabeta CD8alphaalpha subsets of IEL on the intestinal epithelial barrier integrity. To this end, an in vitro coculture system of the murine intestinal crypt-derived cell-line mIC(cl2) and syngeneic ex vivo isolated IEL was employed. Epithelial integrity was assessed by analysis of transepithelial resistance (TER) and paracellular flux of fluorescein isothiocyanate-conjugated (FITC-) dextran. The TCR-alphabeta CD8alphaalpha IEL and resting TCR-alphabeta CD8alphabeta IEL did not affect TER of mIC(cl2) or flux of FITC-dextran. In contrast, activated TCR-alphabeta CD8alphabeta IEL clearly disrupted the integrity of the mIC(cl2) monolayer. No disrupting effect was seen with activated TCR-alphabeta CD8alphabeta IEL from interferon-gamma knockout mice. These findings demonstrate that secretion of interferon-gamma by activated TCR-alphabeta CD8alphabeta IEL is strictly required and also sufficient for disrupting the intestinal epithelial barrier function.