Covering: up to 2021Terpenoids are a diverse group of chemicals used in a wide range of industries. Microbial terpenoid production has the potential to displace traditional manufacturing of these ...compounds with renewable processes, but further titre improvements are needed to reach cost competitiveness. This review discusses strategies to increase terpenoid titres in
with a focus on alternative metabolic pathways. Alternative pathways can lead to improved titres by providing higher orthogonality to native metabolism that redirects carbon flux, by avoiding toxic intermediates, by bypassing highly-regulated or bottleneck steps, or by being shorter and thus more efficient and easier to manipulate. The canonical 2-
-methyl-D-erythritol 4-phosphate (MEP) and mevalonate (MVA) pathways are engineered to increase titres, sometimes using homologs from different species to address bottlenecks. Further, alternative terpenoid pathways, including additional entry points into the MEP and MVA pathways, archaeal MVA pathways, and new artificial pathways provide new tools to increase titres. Prenyl diphosphate synthases elongate terpenoid chains, and alternative homologs create orthogonal pathways and increase product diversity. Alternative sources of terpenoid synthases and modifying enzymes can also be better suited for
expression. Mining the growing number of bacterial genomes for new bacterial terpenoid synthases and modifying enzymes identifies enzymes that outperform eukaryotic ones and expand microbial terpenoid production diversity. Terpenoid removal from cells is also crucial in production, and so terpenoid recovery and approaches to handle end-product toxicity increase titres. Combined, these strategies are contributing to current efforts to increase microbial terpenoid production towards commercial feasibility.
Objectives The goal of this study was to evaluate the safety and performance of the NeoChord DS1000 system (NeoChord, Inc., Minneapolis, Minnesota). Background There is an increasing interest in ...transcatheter mitral valve (MV) treatment. The NeoChord DS 1000 system enables off-pump beating heart transapical MV repair with implantation of artificial neo-chordae. Methods Patients with severe mitral regurgitation (MR) due to isolated posterior prolapse were included in this TACT (Transapical Artificial Chordae Tendinae) trial. All patients were scheduled for off-pump transapical implantation of neo-chordae. Results Thirty patients at 7 centers were enrolled. Major adverse events included 1 death due to post-cardiotomy syndrome and concomitant sepsis and 1 minor stroke with the patient fully recovered at the 30-day follow-up visit. Additional patients experienced procedural major adverse events related to a reoperation or conversion to standard of care. Acute procedural success (placement of at least 1 neo-chord and reduction of MR from 3+ or 4+ to ≤2+) was achieved in 26 patients (86.7%). In 4 patients neo-chordae were not placed for technical and/or patient-specific reasons. These patients underwent intraoperative (3 patients) or post-operative (1 patient) standard MV repair. At 30 days, 17 patients maintained an MR grade ≤2+. Four patients who developed recurrent MR were successfully treated with open MV repair during 30-day follow-up. Results improved with experience: durable reduction in MR to ≤2+ at 30 days was achieved in 5 (33.3%) of the first 15 patients and 12 (85.7%) of the last 14 patients. Conclusions Off-pump transapical implantation of artificial chordae to correct MR is technically safe and feasible; however, it yields further potential for improvement of efficacy and durability. (Safety and Performance Study of the NeoChord Device TACT; NCT01777815 )
Peroxisomes house critical metabolic reactions that are essential for seedling development. As seedlings mature, metabolic requirements change, and peroxisomal contents are remodeled. The resident ...peroxisomal protease LON2 is positioned to degrade obsolete or damaged peroxisomal proteins, but data supporting such a role in plants have remained elusive. Arabidopsis thaliana lon2 mutants display defects in peroxisomal metabolism and matrix protein import but appear to degrade matrix proteins normally. To elucidate LON2 functions, we executed a forward-genetic screen for lon2 suppressors, which revealed multiple mutations in key autophagy genes. Disabling core autophagy-related gene (ATG) products prevents autophagy, a process through which cytosolic constituents, including organelles, can be targeted for vacuolar degradation. We found that atg2, atg3, and atg7 mutations suppressed lon2 defects in auxin metabolism and matrix protein processing and rescued the abnormally large size and small number of lon2 peroxisomes. Moreover, analysis of lon2 atg mutants uncovered an apparent role for LON2 in matrix protein turnover. Our data suggest that LON2 facilitates matrix protein degradation during peroxisome content remodeling, provide evidence for the existence of pexophagy in plants, and indicate that peroxisome destruction via autophagy is enhanced when LON2 is absent.
Objective The aim of this propensity-matched, multicenter study was to compare early clinical and echocardiographic outcomes of patients undergoing transapical aortic valve implantation (TA-TAVI) ...versus patients undergoing sutureless aortic valve replacement (SU-AVR) for severe symptomatic aortic valve stenosis. Methods We reviewed 468 TA-TAVIs performed in 20 centers from April 2008 to May 2011, and 51 SU-AVRs performed in 3 centers from March to September 2011. Based on a propensity score analysis, 2 groups with 38 matched pairs were created. Variables used in the propensity analysis were age, sex, body surface area, New York Heart Association class, logistic EuroSCORE, peripheral vascular disease, chronic obstructive pulmonary disease, aortic valve area, mitral regurgitation, and left ventricular ejection fraction. Results Preoperative characteristics of the 2 groups were comparable. Hospital mortality was 5.3% and 0% in the TA-TAVI and SU-AVR groups, respectively ( P = .49). We did not observe stroke or acute myocardial infarction in the 2 groups. Permanent pacemaker implantation was needed in 2 patients of each group (5.3%, P = 1.0). Dialysis was required in 2 patients (5.3%) in the SU-AVR group and in 1 patient (2.7%) in the TA-TAVI group ( P = 1.0). Predischarge echocardiographic data showed that the incidence of paravalvular leak (at least mild) was greater in the TA-TAVI group (44.7% vs 15.8%, P = .001), but there were no differences in terms of mean transprosthetic gradient (10.3 ± 5 mm Hg vs 11 ± 3.7 mm Hg, P = .59). Conclusions This preliminary experience showed that, in patients at high risk for conventional surgery, SU-AVR is as safe and effective as TA-TAVI and that it is associated with a lower rate of postprocedural paravalvular leak.
Interest in right minithoracotomy mitral valve surgery (MVS) is rapidly growing and, to date, different perfusion strategies and aortic clamping techniques are available. However each approach ...carries specific advantages and drawbacks. This retrospective study analyses our experience in right minithoracotomy MVS with different arterial perfusion and aortic clamping strategies, highlighting the results of a patient tailored approach.
Between March 2009 and March 2014, 460 patients with a full preoperative work-up that included also aortoiliac-femoral axis' screening underwent right minithoracotomy MVS. One hundred and eight were redo cases (23.5%), 63 had aortoiliac atheromatous disease or significant tortuosity (13.7%), and 38 had chronic obstructive pulmonary disease (8.3%). Based on anatomy and comorbidities, each patient was allocated to the most appropriate of 3 approaches: femoral arterial cannulation with endoaortic balloon (P+EB) (247, 53.7%) or with transthoracic clamp (P+XC) (150, 32.6%), and direct aortic cannulation with endoaortic balloon occlusion (C+EB) (63, 13.7%).
No cases of aortic dissection were reported. Early outcome were similar between the 3 groups; no differences were reported in terms of stroke rate (1.7% in the P+EB, 2% in the P+XC, and no cases in the C+EB group; p = NS) and 30-day mortality (2.1% in the P+EB, 2.7% in the P+XC, and 1.6% in the C+EB group; p = NS). Logistic regression showed no influences of arterial perfusion and aortic clamping techniques on 30-day mortality and stroke.
Right minithoracotomy MVS can routinely be performed with favorable outcomes in all comers when perfusion strategies and clamping techniques are carefully selected after proper evaluation of the patient's preoperative characteristics.
Auxin regulates many aspects of plant development, in part, through degradation of the Aux/IAA family of transcriptional repressors. Consequently, stabilizing mutations in several Aux/IAA proteins ...confer reduced auxin responsiveness. However, of the 29 apparent Aux/IAA proteins in
Arabidopsis thaliana
, fewer than half have roles established through mutant analysis. We identified
iaa16
-
1
, a dominant gain-of-function mutation in
IAA16
(
At3g04730
), in a novel screen for reduced root responsiveness to abscisic acid. The
iaa16
-
1
mutation also confers dramatically reduced auxin responses in a variety of assays, markedly restricts growth of adult plants, and abolishes fertility when homozygous. We compared
iaa16
-
1
phenotypes with those of dominant mutants defective in the closely related
IAA7/AXR2
,
IAA14/SLR
, and
IAA17/AXR3
, along with the more distantly related
IAA28
, and found overlapping but distinct patterns of developmental defects. The identification and characterization of
iaa16
-
1
provides a fuller understanding of the IAA7/IAA14/IAA16/IAA17 clade of Aux/IAA proteins and the diverse roles of these repressors in hormone response and plant development.
Key steps of essential metabolic pathways are housed in plant peroxisomes. We conducted a microscopy-based screen for anomalous distribution of peroxisomally targeted fluorescence in Arabidopsis ...thaliana This screen uncovered 34 novel alleles in 15 genes affecting oil body mobilization, fatty acid β-oxidation, the glyoxylate cycle, peroxisome fission, and pexophagy. Partial loss-of-function of lipid-mobilization enzymes conferred peroxisomes clustered around retained oil bodies without other notable defects, suggesting that this microscopy-based approach was sensitive to minor perturbations, and that fatty acid β-oxidation rates in wild type are higher than required for normal growth. We recovered three mutants defective in PECTIN METHYLESTERASE31, revealing an unanticipated role in lipid mobilization for this cytosolic enzyme. Whereas mutations reducing fatty acid import had peroxisomes of wild-type size, mutations impairing fatty acid β-oxidation displayed enlarged peroxisomes, possibly caused by excess fatty acid β-oxidation intermediates in the peroxisome. Several fatty acid β-oxidation mutants also displayed defects in peroxisomal matrix protein import. Impairing fatty acid import reduced the large size of peroxisomes in a mutant defective in the PEROXISOMAL NAD
TRANSPORTER (PXN), supporting the hypothesis that fatty acid accumulation causes pxn peroxisome enlargement. The diverse mutants isolated in this screen will aid future investigations of the roles of β-oxidation and peroxisomal cofactor homeostasis in plant development.
Acute fulminant myocarditis (AFM) may represent a life-threatening event, characterized by rapidly progressive cardiac compromise that ultimately leads to refractory cardiogenic shock or cardiac ...arrest. Venoarterial extracorporeal membrane oxygenation (VA-ECMO) provides effective cardiocirculatory support in this circumstance, but few clinical series are available about early and long-term results. Data from a multicenter study group are reported which analyzed subjects affected by AFM and treated with VA-ECMO during a 5-year period.
From hospital databases, 57 patients with diagnoses of AFM treated with VA-ECMO in the past 5 years were found and analyzed. Mean age was 37.6 ± 11.8 years; 37 patients were women. At VA-ECMO implantation, cardiogenic shock was present in 38 patients, cardiac arrest in 12, and severe hemodynamic instability in 7. A peripheral approach was used with 47 patients, whereas 10 patients had a central implantation or other access.
Mean VA-ECMO support was 9.9 ± 19 days (range, 2 to 24 days). Cardiac recovery with ECMO weaning was achieved in 43 patients (75.5%), major complications were observed in 40 patients (70.1%), and survival to hospital discharge occurred in 41 patients (71.9%). After hospital discharge (median follow-up, 15 months) there were 2 late deaths. The 5-year actual survival was 65.2% ± 7.9%, with recurrent self-recovering myocarditis observed in 2 patients (at 6 and 12 months from the first AFM event), and 1 heart transplantation.
Cardiopulmonary support with VA-ECMO provides an invaluable tool in the treatment of AFM, although major complications may characterize the hospital course. Long-term outcome appears favorable with rare episodes of recurrent myocarditis or cardiac-related events.
Artemisinin, a sesquiterpene lactone produced by
glandular secretory trichomes, is the active ingredient in the most effective treatment for uncomplicated malaria caused by
parasites. Other ...metabolites in
or related species, particularly flavonoids, have been proposed to either act as antimalarials on their own or act synergistically with artemisinin to enhance antimalarial activity. We identified a mutation that disrupts the CHALCONE ISOMERASE 1 (CHI1) enzyme that is responsible for the second committed step of flavonoid biosynthesis. Detailed metabolite profiling revealed that
lacks all major flavonoids but produces wild-type artemisinin levels, making this mutant a useful tool to test the antiplasmodial effects of flavonoids. We used whole-leaf extracts from
and mutant lines impaired in artemisinin production in bioactivity
assays against intraerythrocytic
Dd2. We found that
extracts did not differ from wild-type extracts in antiplasmodial efficacy nor initial rate of cytocidal action. Furthermore, extracts from the
mutant and RNAi lines impaired in amorpha-4,11-diene synthase gene expression, which are both severely compromised in artemisinin biosynthesis but unaffected in flavonoid metabolism, showed very low or no antiplasmodial activity. These results demonstrate that
bioactivity against
of flavonoids is negligible when compared to that of artemisinin.
A variety of metabolic pathways are sequestered in peroxisomes, conserved organelles that are essential for human and plant survival. Peroxin (PEX) proteins generate and maintain peroxisomes. The ...PEX1 ATPase facilitates recycling of the peroxisome matrix protein receptor PEX5 and is the most commonly affected peroxin in human peroxisome biogenesis disorders. Here, we describe the isolation and characterization of, to our knowledge, the first Arabidopsis (Arabidopsis thaliana) pex1 missense alleles: pex1-2 and pex1-3. pex1-2 displayed peroxisome-related defects accompanied by reduced PEX1 and PEX6 levels. These pex1-2 defects were exacerbated by growth at high temperature and ameliorated by growth at low temperature or by PEX6 overexpression, suggesting that PEX1 enhances PEX6 stability and vice versa. pex1-3 conferred embryo lethality when homozygous, confirming that PEX1, like several other Arabidopsis peroxins, is essential for embryogenesis. pex1-3 displayed symptoms of peroxisome dysfunction when heterozygous; this semidominance is consistent with PEX1 forming a heterooligomer with PEX6 that is poisoned by pex1-3 subunits. Blocking autophagy partially rescued PEX1/pex1-3 defects, including the restoration of normal peroxisome size, suggesting that increasing peroxisome abundance can compensate for the deficiencies caused by pex1-3 and that the enlarged peroxisomes visible in PEX1/pex1-3 may represent autophagy intermediates. Overexpressing PEX1 in wild-type plants impaired growth, suggesting that excessive PEX1 can be detrimental. Our genetic, molecular, and physiological data support the heterohexamer model of PEX1-PEX6 function in plants.
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