Riboflavin is an essential vitamin widely used in the food, pharmaceutical, and feed industries. However, the insufficient supply of precursors caused by the imbalance of intracellular metabolic flow ...limits the riboflavin synthesis by industrial strains. Here, we increase riboflavin production by tuning multiple gene expression to balance intracellular metabolic flow. First, we tuned the expression of mCherry and egfp genes within operons by generating libraries of tunable intergenic regions (TIGRs) and confirmed the relative expression of the two reporter genes. The TIGR library can coordinate the expression ratio of reporter genes more than 180 times in Escherichia coli and more than 70 times in Bacillus subtilis. Next, we used this strategy to tune the expression of zwf, ribBA, and ywlf genes within operons through the TIGR library to increase the intracellular precursor pool for riboflavin biosynthesis. Based on the fluorescence characteristics of riboflavin, 96-well plates were used to screen the optimal combination mutants quickly. The best-engineered strain was selected from the library, which produced 2.7 g/L riboflavin, increasing by 64.35% in the shake flask. Finally, the riboflavin titer increased by 59.27% to 11.77 g/L in fed-batch fermentation. The strategy described here will contribute to the industrial production of riboflavin and related products by B. subtilis.
Since residual stresses significantly affect the properties of Ti/CFRP (Carbon Fiber Reinforced Polyimide) laminates, which are inevitably produced in the laminates after preparation owing to the ...different thermal expansion coefficients of the constituents, a finite element model was built in this work to simulate the residual stresses of Ti/CFRP laminates in the cooling process from glass-transition temperature (Tg) to ambient temperature. Distributions of stresses in each ply and interlaminar stresses between the adjacent layers were obtained. Moreover, the free boundary problem, usually leading to the generation of cracks, was observed especially at the edges perpendicular to the fiber direction. Additionally, an analytical approach and two experimental methods (layer removal method and X-ray diffraction technique) were applied to determine the residual stresses. Comparing these results, it was shown that these methods verified each other. The established model and corresponding simulation in our work were effective to describe residual stresses in Ti/CFRP laminates.
Response to post-stroke aphasia language rehabilitation is difficult to anticipate, mainly because few predictors can help identify optimal, individualized treatment options. Imaging techniques, such ...as Voxel-based Lesion Symptom Mapping have been useful in linking specific brain areas to language behavior; however, further development is required to optimize the use of structural and physiological information in guiding individualized treatment for persons with aphasia (PWA). In this study, we will determine if cerebral blood flow (CBF) mapped in patients with chronic strokes can be further used to understand stroke-related factors and behavior.
We collected perfusion MRI data using pseudo-Continuous Arterial Spin Labeling (pCASL) using a single post-labeling delay of 2,200 ms in 14 chronic PWA, along with high-resolution structural MRI to compute maps of tissue damage using Tissue Integrity Gradation via T2w T1w Ratio (TIGR). To quantify the CBF in chronic stroke lesions, we tested at what point spatial smoothing should be applied in the ASL analysis pipeline. We then related CBF to tissue damage, time since stroke, age, sex, and their respective cross-terms to further understand the variability in lesion CBF. Finally, we assessed the feasibility of computing multivariate brain-behavior maps using CBF and compared them to brain-behavior maps extracted with TIGR MRI.
We found that the CBF in chronic stroke lesions is significantly reduced compared to its homologue grey and white matter regions. However, a reliable CBF signal (although smaller than expected) was detected to reveal a negative relationship between CBF and increasing tissue damage. Further, the relationship between the lesion CBF and age, sex, time since stroke, and tissue damage and cross-terms suggested an aging-by-disease interaction. This relationship was strongest when smoothing was applied in the template space. Finally, we show that whole-brain CBF relates to domain-general visuospatial functioning in PWA. The CBF-based brain-behavior maps provide unique and complementary information to structural (lesion-based) brain-behavior maps.
Therefore, CBF can be detected in chronic stroke lesions using a standard pCASL MRI acquisition and is informative at the whole-brain level in identifying stroke rehabilitation targets in PWAs due to its relationship with demographic factors, stroke-related factors, and behavior.
Engineered heterologous multi-gene metabolic pathways often suffer from flux imbalance and toxic metabolites, as the production host typically lacks the regulatory mechanisms for the heterologous ...pathway. Here, we first coordinated the expression of all genes of the mevalonate (MEV) pathway from Saccharomyces cerevisiae using the tunable intergenic regions (TIGRs), and then dynamically regulated the TIGR-mediated MEV pathway to prevent the accumulation of toxic metabolites by using IPP/FPP-responsive promoter. After introduction of the dynamically controlled TIGR-mediated MEV pathway into Escherichia coli, the content and concentration of zeaxanthin in shaker flask cultures were 2.0- and 2.1-fold higher, respectively, than those of the strain harboring the statically controlled non-TIGR-mediated MEV pathway. The content and concentration of zeaxanthin in E. coli ZEAX (pZSPgadE-MevTTIGR-MevBTIGRIS-2) reached 722.46mg/L and 23.16mg/g dry cell weight (DCW), respectively, in 5.0L fed-batch fermentation. We also comparatively analyzed the proteomes between E. coli ZEAX and E. coli ZEAX (pZSPgadE-MevTTIGR-MevBTIGRIS-2) to understand the mechanism of zeaxanthin biosynthesis. The results of the comparative proteomes demonstrate that zeaxanthin overproduction may be associated with increased precursor availability, increased NADPH availability, increased ATP availability, oxidative stress response, and increased membrane storage capacity for zeaxanthin due to changes in both cellular shape and membrane composition.
•Dynamic control of the TIGR-mediated MEV pathway by IPP/FPP-responsive promoter.•722.46mg/L and 23.16mg/g DCW of zeaxanthin was produced in 5.0L fed-batch fermentation.•The strain achieved the highest level of zeaxanthin ever reported.•The mechanism of zeaxanthin overproduction was obtained by proteome analysis.•Zeaxanthin production may be related to oxidative stress and cellular shape change.
Metagenomics projects based on shotgun sequencing of populations of micro-organisms yield insight into protein families. We used sequence similarity clustering to explore proteins with a ...comprehensive dataset consisting of sequences from available databases together with 6.12 million proteins predicted from an assembly of 7.7 million Global Ocean Sampling (GOS) sequences. The GOS dataset covers nearly all known prokaryotic protein families. A total of 3,995 medium- and large-sized clusters consisting of only GOS sequences are identified, out of which 1,700 have no detectable homology to known families. The GOS-only clusters contain a higher than expected proportion of sequences of viral origin, thus reflecting a poor sampling of viral diversity until now. Protein domain distributions in the GOS dataset and current protein databases show distinct biases. Several protein domains that were previously categorized as kingdom specific are shown to have GOS examples in other kingdoms. About 6,000 sequences (ORFans) from the literature that heretofore lacked similarity to known proteins have matches in the GOS data. The GOS dataset is also used to improve remote homology detection. Overall, besides nearly doubling the number of current proteins, the predicted GOS proteins also add a great deal of diversity to known protein families and shed light on their evolution. These observations are illustrated using several protein families, including phosphatases, proteases, ultraviolet-irradiation DNA damage repair enzymes, glutamine synthetase, and RuBisCO. The diversity added by GOS data has implications for choosing targets for experimental structure characterization as part of structural genomics efforts. Our analysis indicates that new families are being discovered at a rate that is linear or almost linear with the addition of new sequences, implying that we are still far from discovering all protein families in nature.
Background
In this morphometric study, we describe the anatomy of the TIGR triangle, which is bordered by the tentorial surface of the cerebellum, the internal cerebral vein and vein of Galen ...complex, and the vein of Rosenthal. These structures define the window, or deep keyhole, to access the pineal region in non-midline supracerebellar infratentorial approaches.
Methods
The posterior fossa anatomy of 16 patients was studied in virtual reality (VR), and the TIGR triangles were defined and measured with special attention on its angular orientation in the posterior fossa. The angular expanse of the posterior fossa was measured and recorded as the transverse-sigmoid junction (TSJ) angle. Because a perpendicular corridor through an anatomic aperture provides the best exposure, we studied the starting point along the TSJ angle that offers the best exposure of TIGR.
Results
In the 31 posterior fossa sides included in the study, the perpendicular trajectory through the TIGR triangle was on average 27.13° CI 95% (range: 5.97°–48.53°) from the midline. When comparing the SCIT variants, both the paramedian and lateral approaches provided near-perpendicular trajectory through the TIGR triangle in a majority of specimens. However, the modified paramedian approach, with starting point defined as TSJ angle/3, provided the most perpendicular path through the TIGR triangle.
Conclusion
We studied the size, spatial orientation, and morphology of the TIGR triangle. Our data indicated that the best exposure of TIGR is through a modified paramedian SCIT approach, in which the starting point one third of the way from midline to the TSJ.
Stroke-related tissue damage within lesioned brain areas is topologically non-uniform and has underlying tissue composition changes that may have important implications for rehabilitation. However, ...we know of no uniformly accepted, objective non-invasive methodology to identify pericavitational areas within the chronic stroke lesion. To fill this gap, we propose a novel magnetic resonance imaging (MRI) methodology to objectively quantify the lesion core and surrounding pericavitational perimeter, which we call tissue integrity gradation
T2w T1w ratio (TIGR). TIGR uses standard T1-weighted (T1w) and T2-weighted (T2w) anatomical images routinely collected in the clinical setting. TIGR maps are analyzed with relation to subject-specific gray matter and cerebrospinal fluid thresholds and binned to create a false colormap of tissue damage within the stroke lesion, and these are further categorized into low-, medium-, and high-damage areas. We validate TIGR by showing that the cerebral blood flow within the lesion reduces with greater tissue damage (
= 0.005). We further show that a significant task activity can be detected in pericavitational areas and that medium-damage areas contain a significantly lower magnitude of hemodynamic response function than the adjacent damaged areas (
< 0.0001). We also demonstrate the feasibility of using TIGR maps to extract multivariate brain-behavior relationships (
< 0.05) and show general agreement in location compared to binary lesion, T1w-only, and T2w-only maps but that the extent of brain behavior maps may depend on signal sensitivity as denoted by the sparseness coefficient (
< 0.0001). Finally, we show the feasibility of quantifying TIGR in early and late subacute stroke phases, where higher-damage areas were smaller in size (
= 0.002) and that lesioned voxels transition from lower to higher damage with increasing time post-stroke (
= 0.004). We conclude that TIGR is able to (1) identify tissue damage gradient within the stroke lesion across different post-stroke timepoints and (2) more objectively delineate lesion core from pericavitational areas wherein such areas demonstrate reasonable and expected physiological and functional impairments. Importantly, because T1w and T2w scans are routinely collected in the clinic, TIGR maps can be readily incorporated in clinical settings without additional imaging costs or patient burden to facilitate decision processes related to rehabilitation planning.
The richness and versatility of biological systems make them ideally suited to solve some of the world’s most significant challenges, such as converting cheap, renewable resources into energy-rich ...molecules; producing high-quality, inexpensive drugs to fight disease; and remediating polluted sites. Over the years, significant strides have been made in engineering microorganisms to produce fuels, bulk chemicals, and valuable drugs from inexpensive starting materials; to detect and degrade nerve agents as well as less toxic organic pollutants; and to accumulate metals and reduce radionuclides. The components needed to engineer the chemistry inside a microbial cell are significantly different from those commonly used to overproduce pharmaceutical proteins. Synthetic biology has had and will continue to have a significant impact on the development of these components to engineer cellular metabolism and microbial chassis to host the chemistry. The ready availability of more well-characterized gene expression components and hosts for chemical synthesis, standards for the connection of these components to make larger functioning devices, computer-aided design software, and debugging tools for biological designs will decrease both the time and the support needed to construct these designs. Some of the most important tools for engineering bacterial metabolism and their use for production of the antimalarial drug artemisinin are reviewed.
The aim of this study was to compare seroma production in breast reconstruction with a biological mesh with that of a synthetic mesh, in the same patient. The patients were randomized to biological ...mesh in one breast and synthetical in the other. Twenty‐four breasts were included. The total drain production and the daily drain production were similar in the two groups. After drain removal, there were more seroma aspirations in the biological group. During the exchange to a permanent implant, there was significantly more seroma in the biological group. Seroma formation is different in synthetic and biological meshes.
The aim of this study was to compare inflammatory response and synovial metaplasia in implant-based immediate breast reconstruction with a biological mesh (Veritas
®
) with that of a synthetic mesh ...(TIGR
®
Matrix Surgical Mesh). We hypothesize that the inflammatory response and formation of synovial metaplasia might be different and the rate of capsular contracture therefore different. The patients were recruited from the Gothenburg TIGR
®
/Veritas
®
Study (ClinicalTrials.Gov identifier NCT02985073). All referrals for bilateral immediate breast reconstruction were assessed for inclusions. During the operation, the patients were randomized to which sides the biological and the synthetic mesh were going to be applied. During the implant exchange biopsies were taken. Biopsies were taken from 30 breasts in 15 patients. There seem to be more myofibroblast and neovascularization in the biological meshes than in the synthetic and the collagen fibers seem to be aligned in an irregular pattern with both parallel and vertical fibers. In the synthetic meshes, there were more giant cells and foreign body reaction and the collagen fibers were loosely and well aligned, oriented parallel to the surface of the implant. Synovial metaplasia was seen in the majority of both the biological and the synthetic meshes. The histological patterns in early capsules from biological and synthetic meshes vary considerably. Nonetheless, it is unknown what role different cell types have in capsular formation in the long run and there was no difference in clinical capsular contracture at the clinical follow-up in this study.
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