Network-level architecture and the evolutionary potential of underground metabolism

A central unresolved issue in evolutionary biology is how metabolic innovations emerge. Low-level enzymatic side activities are frequent and can potentially be recruited for new biochemical functions. However, the role of such underground reactions in adaptation toward novel environments has remained largely unknown and out of reach of computational predictions, not least because these issues demand analyses at the level of the entire metabolic network. Here, we provide a comprehensive computational model of the underground metabolism in Escherichia coli. Most underground reactions are not isolated and 45% of them can be fully wired into the existing network and form novel pathways that produce key precursors for cell growth. This observation allowed us to conduct an integrated genome-wide in silico and experimental survey to characterize the evolutionary potential of E. coli to adapt to hundreds of nutrient conditions. We revealed that underground reactions allow growth in new environments when their activity is increased. We estimate that at least ∼20% of the underground reactions that can be connected to the existing network confer a fitness advantage under specific environments. Moreover, our results demonstrate that the genetic basis of evolutionary adaptations via underground metabolism is computationally predictable. The approach used here has potential for various application areas from bioengineering to medical genetics.How do new molecular pathways evolve? In the best-studied molecular networks, small-molecule metabolism, the prevailing paradigm is that new pathways are patched together from preexisting enzymes borrowed from different parts of the network (1–3). Central to this “patchwork” model of pathway evolution is the notion that many enzymes have limited substrate specificities and can catalyze, albeit at low rates, reactions other than those for which they have evolved (also referred to as enzyme promiscuity) (4). These so-called underground (5) or side activities are prevalent (6–8) and were shown to serve as starting points for the evolution of novel functions both in directed evolution experiments (9) and in the diversification of gene families in the wild (7). However, how the underground catalytic repertoire encoded in the genome can generate novelties within the context of the existing metabolic network remains unknown. Do underground reactions remain isolated, or can they potentially be wired into the native network and allow the organism to survive in novel environments? Furthermore, would it be possible to computationally predict the genetic basis of phenotypic evolution based on a detailed knowledge of the organism’s underground metabolism? Answering these questions requires both large-scale data on underground enzyme activities and systems-level approaches to analyze metabolic capabilities. Although systematic detection of underground activities by unbiased high-throughput approaches is not feasible at present, the accumulated knowledge of enzyme biochemistry in the well-studied prokaryote Escherichia coli provides a valuable resource of such nonnative enzyme activities (10). Thus, to explore the architecture of underground metabolism and its evolutionary potential, we compiled a comprehensive set of experimentally reported side activities of E. coli enzymes and integrated these reactions into a global metabolic network reconstruction of the same organism (11). Analysis of this extended network revealed that a substantial fraction of underground reactions can be fully integrated into the existing metabolism and participate in potential pathways that produce key precursors for cell growth. Using metabolic modeling, we then predicted specific environmental conditions under which such biologically relevant underground reactions confer a growth advantage, and hence deliver a phenotypic novelty. Our analyses revealed that the set of known underground reactions has a significant potential both to increase fitness in existing environments and to exploit new nutrient sources. A genome-wide gene overexpression screen across hundreds of carbon sources showed a good agreement with the model’s predictions, which illustrates that the genetic basis of phenotypic novelties can be predicted based on the knowledge of underground metabolism.     

HDR brachytherapy boost using MR-only workflow for intermediate- and high-risk prostate cancer: 8-year results of a pilot study

PurposeTo report 8-year clinical outcome with high-dose-rate brachytherapy (HDRBT) boost using MRI-only workflow for intermediate (IR) and high-risk (HR) prostate cancer (PC) patients.Methods and MaterialsFifty-two patients were treated with 46–60 Gy of 3D conformal radiotherapy preceded and/or followed by a single dose of 8–10 Gy MRI-guided HDRBT. Interventions were performed in a 0.35 T MRI scanner. Trajectory planning, navigation, contouring, catheter reconstruction, and dose calculation were exclusively based on MRI images. Biochemical relapse-free- (BRFS), local relapse-free- (LRFS), distant metastasis-free- (DMFS), cancer-specific-(CCS) and overall survival (OS) were analyzed. Late morbidity was scored using the Common Terminology Criteria for Adverse Events (CTCAE 4.0) combined with RTOG (Radiation Therapy Oncology Group) scale for urinary toxicity and rectal urgency (RU) determined by Yeoh.ResultsMedian follow-up time was 107 (range: 19–143) months. The 8-year actuarial rates of BRFS, LRFS, DMFS, CSS and OS were 85.7%, 97%, 97.6%, and 77.6%, respectively. There were no Gr.3 GI side effects. The 8-year actuarial rate of Gr.2 proctitis was 4%. The 8-year cumulative incidence of Gr.3 GU side effects was 8%, including two urinary stenoses (5%) and one cystitis (3%). EPIC urinary and bowel scores did not change significantly over time.ConclusionsMRI-only HDR-BT boost with moderate dose escalation provides excellent 8-year disease control with a favorable toxicity profile for IRPC and HRPC patients. Our results support the clinical importance of MRI across the BT workflow.     

Submental endotracheal intubation in concurrent orthognathic surgery: a technical note.

INTRODUCTION: Achieving the necessary occlusion for orthognathic surgery is not possible with conventional oral intubation since the tube interferes with the occluding teeth. Sometimes nasotracheal intubation is impossible due to developmental malformations requiring repair. Also, the oral or nasotracheal tube may interfere with the operation or may be damaged during the procedure. In 1986, Hernandez Altemir described a method of submental endotracheal intubation. His intentions were to avoid tracheostomy in maxillofacial trauma cases where short-term intermaxillary fixation was required. PATIENTS: Between January 2000 and May 2003, 13 patients were operated on, using submental intubation. Eight of these (three females and five males) had surgery for orthognathic malformations. METHODS: The Hernandez Altemir technique was modified to ease the procedure: a sterile nylon guiding tube and the '222 rule' incision were introduced. Eight cases with concurrent complex orthognathic surgery, using this modified technique are reported in this paper. RESULTS: There were no operative or postoperative complications related to the procedure. CONCLUSION: The technique is easy to use, rapid and free of complications compared to 'alternative' intubation methods (tracheostomy, retromolar location of tube, etc.). Submental scarring is acceptable. It is recommended for orthognathic procedures in selected cases.     

Uniform qualitative electrophysiological changes in postoperative rest tremor.

Ablation and deep brain stimulation (DBS) can treat pharmacologically uncontrollable tremor. Here, we compared the postoperative electrophysiological changes in resting hand tremor after 32 ablations and 12 DBS implantations in patients with severe tremor-dominant idiopathic Parkinson's disease (PD) and essential tremor (ET). Short- and long-term accelerometric data were acquired after surgery and were compared to the preoperative tremor. After effective surgical treatments, significant rest tremor reduction and increase in both frequency and approximate entropy (ApEn) were detected in all PD cases, irrespective of the type and target of intervention. However, the long-term effect of DBS implantation on tremor reduction was significantly better compared to that after ablative treatments. In cases of thalamotomy, the postoperative increase in frequency and ApEn was significantly larger in essential tremor compared to PD, suggesting that the etiology of tremor may influence the size of the similar changes. However, cases where clinical tremor re-emerged 6 to 12 months after the surgery, no change in frequency and ApEn was detected on the second postoperative day, despite an initial tremor reduction and clinical improvement similar to the effective operations. Our results suggest that uniform postoperative changes in rest tremor and the increase in frequency and ApEn could be due to attenuation of pathological oscillators and might be immediate indicators of the effectiveness of neurosurgical treatments relieving tremor.     

Effects of sweet and bitter gustatory stimuli in anorexia nervosa on EEG frequency spectra.

The possible differences in processing gustatory stimuli in anorexic patients compared to healthy control subjects was investigated by electrophysiological methods. The electroencephalogram (EEG) was recorded in outpatients treated with anorexia nervosa (AN) and age-matched controls after exposure to sweet (milk chocolate) and bitter (black tea) taste stimuli. Power spectrum analysis was performed on EEG epochs recorded in the above conditions. Compared to controls a significantly higher percent of theta, and lower percent of alpha1 band power was found in anorexic patients, irrespective of the kind of taste effects and hemispheric side. The pattern of activation caused by sweet and bitter stimuli was found to be different in these two groups, possibly indicating altered gustatory processing mechanisms in AN.     

MRI-guided focused ultrasound surgery in the brain: tests in a primate model.

MRI-guided focused ultrasound was tested in the brains of rhesus monkeys. Locations up to 4.8 cm deep were targeted. Focal heating was observed in all cases with MRI-derived temperature imaging. Subthreshold heating was observed at the focus when the ultrasound beam was targeted with low power sonications, and in the ultrasound beam path during high-power exposures. Lethal temperature values and histologically confirmed tissue damage were confined to the focal zone (e.g., not in the ultrasound beam path), except when the focus was close to the bone. In that case, damage to the neighboring brain tissue was observed. Focal lesions were observed on histological examination and, in some cases, in MR images acquired immediately after the ultrasound exposures. The capabilities demonstrated in this study will be of benefit for clinical ultrasound therapies in the brain.     

Quantitative magnetic resonance imaging of brain development in premature and mature newborns

Definition in the living premature infant of the anatomical and temporal characteristics of development of critical brain structures is crucial for insight into the time of greatest vulnerability of such brain structures. We used three-dimensional magnetic resonance imaging (3D MRI) and image-processing algorithms to quantitate total brain volume and total volumes of cerebral gray matter (GM), unmyelinated white matter (WM), myelinated WM, and cerebrospinal fluid (CSF) in 78 premature and mature newborns (postconceptional age, 29–41 weeks). Total brain tissue volume was shown to increase linearly at a rate of 22 ml/wk. Total GM showed a linear increase in relative intracranial volume of approxi mately 1.4% or 15 ml in absolute volume per week. The pronounced increase in total GM reflected primarily a fourfold increase in cortical GM. Unmyelinated WM was found to be the most prominent brain tissue class in the preterm infant younger than 36 weeks of postconceptional age. Although minimal myelinated WM was present in the preterm infant at 29 weeks, between 35 and 41 weeks an abrupt fivefold increase in absolute volume of myelinated WM was documented. Extracerebral and intraventricular CSF was readily quantitated by this technique and found to change minimally. The application of 3D MRI and tissue segmentation to the study of human infant brain from 29 to 41 weeks of postconceptional age has provided new insights into cerebral cortical development and myelination and has for the first time provided means of quantitative assessment in vivo of early human brain development.     

Invited. MRI of laser-induced interstitial thermal injury in an in vivo animal liver model with histologic correlation

Laser-induced interstitial thermal therapy (LITT) is a preferred method of minimally invasive therapy. MRI is a noninvasive method by which to monitor the thermal effects of LITT. To properly control such effects, changes in MRI parameters during and after LITT should be correlated with changes in the tissue. T1-weighted fast spin echo (FSE) MRI (1 image/10 seconds) at 1.5 T monitored LITT in vivo in rabbit liver (n = 6) using an interstitial bare delivery fiber (600-μm diameter; 3.0 W; 1,064 nm; 150 seconds). During laser irradiation, MRI signal intensity decreased around the fiber tip; after irradiation, this hypointensity proved reversible and permanent lesions were evident. The lesions had hyperintense margins that were brighter than surrounding normal tissue (P < .001); the tissue in these bright regions was mapped to tissue necrosis characterized by the presence of thermally damaged ghost red blood cells amid generally normal hepatocytes. T1-FSE identified the spatial extent of the LITT lesions.