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Journal of Biomedical Optics Feb 2024Glioblastoma (GBM) is a rare but deadly form of brain tumor with a low median survival rate of 14.6 months, due to its resistance to treatment. An independent simulation...
SIGNIFICANCE
Glioblastoma (GBM) is a rare but deadly form of brain tumor with a low median survival rate of 14.6 months, due to its resistance to treatment. An independent simulation of the INtraoperative photoDYnamic therapy for GliOblastoma (INDYGO) trial, a clinical trial aiming to treat the GBM resection cavity with photodynamic therapy (PDT) via a laser coupled balloon device, is demonstrated.
AIM
To develop a framework providing increased understanding for the PDT treatment, its parameters, and their impact on the clinical outcome.
APPROACH
We use Monte Carlo radiative transport techniques within a computational brain model containing a GBM to simulate light path and PDT effects. Treatment parameters (laser power, photosensitizer concentration, and irradiation time) are considered, as well as PDT's impact on brain tissue temperature.
RESULTS
The simulation suggests that 39% of post-resection GBM cells are killed at the end of treatment when using the standard INDYGO trial protocol (light fluence = at balloon wall) and assuming an initial photosensitizer concentration of . Increases in treatment time and light power (light fluence = at balloon wall) result in further cell kill but increase brain cell temperature, which potentially affects treatment safety. Increasing the p hotosensitizer concentration produces the most significant increase in cell kill, with 61% of GBM cells killed when doubling concentration to and keeping the treatment time and power the same. According to these simulations, the standard trial protocol is reasonably well optimized with improvements in cell kill difficult to achieve without potentially dangerous increases in temperature. To improve treatment outcome, focus should be placed on improving the photosensitizer.
CONCLUSIONS
With further development and optimization, the simulation could have potential clinical benefit and be used to help plan and optimize intraoperative PDT treatment for GBM.
Topics: Humans; Photosensitizing Agents; Photochemotherapy; Glioblastoma; Brain Neoplasms; Computer Simulation
PubMed: 38322729
DOI: 10.1117/1.JBO.29.2.025001 -
Sensing chemical-induced DNA damage using CRISPR/Cas9-mediated gene-deletion yeast-reporter strains.Applied Microbiology and Biotechnology Feb 2024Microorganism-based genotoxicity assessments are vital for evaluating potential chemical-induced DNA damage. In this study, we developed both chromosomally integrated...
Microorganism-based genotoxicity assessments are vital for evaluating potential chemical-induced DNA damage. In this study, we developed both chromosomally integrated and single-copy plasmid-based reporter assays in budding yeast using a RNR3 promoter-driven luciferase gene. These assays were designed to compare the response to genotoxic chemicals with a pre-established multicopy plasmid-based assay. Despite exhibiting the lowest luciferase activity, the chromosomally integrated reporter assay showed the highest fold induction (i.e., the ratio of luciferase activity in the presence and absence of the chemical) compared with the established plasmid-based assay. Using CRISPR/Cas9 technology, we generated mutants with single- or double-gene deletions, affecting major DNA repair pathways or cell permeability. This enabled us to evaluate reporter gene responses to genotoxicants in a single-copy plasmid-based assay. Elevated background activities were observed in several mutants, such as mag1Δ cells, even without exposure to chemicals. However, substantial luciferase induction was detected in single-deletion mutants following exposure to specific chemicals, including mag1Δ, mms2Δ, and rad59Δ cells treated with methyl methanesulfonate; rad59Δ cells exposed to camptothecin; and mms2Δ and rad10Δ cells treated with mitomycin C (MMC) and cisplatin (CDDP). Notably, mms2Δ/rad10Δ cells treated with MMC or CDDP exhibited significantly enhanced luciferase induction compared with the parent single-deletion mutants, suggesting that postreplication and for nucleotide excision repair processes predominantly contribute to repairing DNA crosslinks. Overall, our findings demonstrate the utility of yeast-based reporter assays employing strains with multiple-deletion mutations in DNA repair genes. These assays serve as valuable tools for investigating DNA repair mechanisms and assessing chemical-induced DNA damage. KEY POINTS: • Responses to genotoxic chemicals were investigated in three types of reporter yeast. • Yeast strains with single- and double-deletions of DNA repair genes were tested. • Two DNA repair pathways predominantly contributed to DNA crosslink repair in yeast.
Topics: Saccharomyces cerevisiae; CRISPR-Cas Systems; DNA Damage; Mitomycin; Luciferases; DNA
PubMed: 38300351
DOI: 10.1007/s00253-024-13020-w -
Nature Feb 2024Photosystem II (PSII) catalyses the oxidation of water through a four-step cycle of S states (i = 0-4) at the MnCaO cluster, during which an extra oxygen (O6) is...
Photosystem II (PSII) catalyses the oxidation of water through a four-step cycle of S states (i = 0-4) at the MnCaO cluster, during which an extra oxygen (O6) is incorporated at the S state to form a possible dioxygen. Structural changes of the metal cluster and its environment during the S-state transitions have been studied on the microsecond timescale. Here we use pump-probe serial femtosecond crystallography to reveal the structural dynamics of PSII from nanoseconds to milliseconds after illumination with one flash (1F) or two flashes (2F). Y, a tyrosine residue that connects the reaction centre P680 and the MnCaO cluster, showed structural changes on a nanosecond timescale, as did its surrounding amino acid residues and water molecules, reflecting the fast transfer of electrons and protons after flash illumination. Notably, one water molecule emerged in the vicinity of Glu189 of the D1 subunit of PSII (D1-E189), and was bound to the Ca ion on a sub-microsecond timescale after 2F illumination. This water molecule disappeared later with the concomitant increase of O6, suggesting that it is the origin of O6. We also observed concerted movements of water molecules in the O1, O4 and Cl-1 channels and their surrounding amino acid residues to complete the sequence of electron transfer, proton release and substrate water delivery. These results provide crucial insights into the structural dynamics of PSII during S-state transitions as well as O-O bond formation.
Topics: Biocatalysis; Calcium; Crystallography; Electron Transport; Electrons; Manganese; Oxidation-Reduction; Oxygen; Photosystem II Protein Complex; Protons; Time Factors; Tyrosine; Water
PubMed: 38297122
DOI: 10.1038/s41586-023-06987-5 -
Acta Crystallographica. Section D,... Feb 2024Over the last decade, the development of time-resolved serial crystallography (TR-SX) at X-ray free-electron lasers (XFELs) and synchrotrons has allowed researchers to...
Over the last decade, the development of time-resolved serial crystallography (TR-SX) at X-ray free-electron lasers (XFELs) and synchrotrons has allowed researchers to study phenomena occurring in proteins on the femtosecond-to-minute timescale, taking advantage of many technical and methodological breakthroughs. Protein crystals of various sizes are presented to the X-ray beam in either a static or a moving medium. Photoactive proteins were naturally the initial systems to be studied in TR-SX experiments using pump-probe schemes, where the pump is a pulse of visible light. Other reaction initiations through small-molecule diffusion are gaining momentum. Here, selected examples of XFEL and synchrotron time-resolved crystallography studies will be used to highlight the specificities of the various instruments and methods with respect to time resolution, and are compared with cryo-trapping studies.
Topics: Synchrotrons; Crystallography; Crystallography, X-Ray; X-Rays; Proteins; Lasers
PubMed: 38265875
DOI: 10.1107/S2059798323011002 -
Nutrition & Metabolism Jan 2024Bone is continuously produced by osteoblasts and resorbed by osteoclasts to maintain homeostasis. Impaired bone resorption by osteoclasts causes bone diseases such as...
BACKGROUND
Bone is continuously produced by osteoblasts and resorbed by osteoclasts to maintain homeostasis. Impaired bone resorption by osteoclasts causes bone diseases such as osteoporosis and arthritis. Most pharmacological treatment of osteoporosis focuses on inhibiting osteoclast differentiation, often to restore osteoclast/osteoclast balance. However, recent osteoporosis treatments have various side effects. According to a recent study, resveratrol, known as a stilbenoid family, is known to increase bone density, and the osteoclast inhibitory effect was confirmed using oxyresveratrol, a stilbenoid family. Here, we investigated the effect of oxyresveratrol on osteoclast differentiation and an ovariectomized mouse model.
METHODS
Mouse leukemia monocyte/macrophage cell line RAW 264.7 was treated with oxyresveratrol, and cell cytotoxicity was confirmed by measuring MTT assay. Tartrate-resistant acid phosphatase (TRAP), an enzyme marker for osteoclasts, was confirmed by staining. In addition, osteoclast differentiation markers and MAPK-related markers were confirmed at the mRNA level and protein expression. The effect of oxyresveratrol was confirmed using ovariectomized mice. Deoxypyridinoline (DPD) was measured using mouse urine and TRAP activity was observed using serum. Bone mineral density was also measured using Micro-CT.
RESULTS
The polyphenol oxyresveratrol inhibited receptor activator of nuclear factor kappa-Β ligand (RANKL)-induced osteoclast differentiation of RAW 264.7 cells. Furthermore, oxyresveratrol inhibited TRAP activity and actin-ring formation. Moreover, oxyresveratrol suppressed the phosphorylation of the RANKL-induced mitogen-activated protein kinases (MAPKs) p38, JNK, and ERK and significantly reduced the expression of bone differentiation markers (NFATc1, cathepsin K, and TRAP).
CONCLUSION
Oxyresveratrol inhibits osteoclast differentiation via MAPK and increases bone density in ovariectomized rats, suggesting it has therapeutic potential for bone diseases such as osteoporosis. We confirmed the osteoporosis prevention effect of OR in Raw 264.7 cells, and future studies should confirm the effect of OR using rat bone marrow-derived cells.
PubMed: 38243227
DOI: 10.1186/s12986-024-00781-4 -
Journal of Ophthalmology 2023Anti-vascular endothelial growth factor (anti-VEGF) therapy via intravitreal injection is an effective treatment for patients with abnormal ocular neovascularization,...
BACKGROUND
Anti-vascular endothelial growth factor (anti-VEGF) therapy via intravitreal injection is an effective treatment for patients with abnormal ocular neovascularization, such as age-related macular degeneration (AMD) and diabetic macular edema (DME). However, prolonged and frequent anti-VEGF treatment is associated with a risk of local and systemic adverse events, including geographic atrophy, cerebrovascular disease, and death. Furthermore, some patients do not adequately respond to anti-VEGF therapy. Hypoxia-inducible factor (HIF) is a transcription factor that controls the expression of hypoxia-responsive genes involved in angiogenesis, inflammation, and metabolism. The HIF/VEGF pathway plays an important role in neovascularization, and the inhibition of HIF activation could be an effective biomolecular target for neovascular diseases. The demand for disease prevention or treatment using functional foods such as superfoods has increased in recent years. Few reports to date have focused on the antineovascular effects of superfoods in the retinal pigment epithelium (RPE). In light of the growing demand for functional foods, we aimed to find novel HIF inhibitors from superfoods worked in RPE cells, which could be an adjuvant for anti-VEGF therapy.
METHODS
Seven superfoods were examined to identify novel HIF inhibitor candidates using luciferase assay screening. We used the human RPE cell line ARPE-19 and fetal human RPE (fhRPE) to investigate the biomolecular actions of novel HIF inhibitors using quantitative PCR and western blotting.
RESULTS
Under CoCl-induced pseudohypoxic condition and 1% oxygen hypoxic incubation, camu-camu () showed HIF inhibitory effects determined by luciferase assays. Camu-camu downregulated and mRNA expressions in a concentration-dependent manner. Camu-camu also inhibited HIF-1 protein expressions, and its inhibitory effect was greater than that of vitamin C, which is present at high levels in camu-camu.
CONCLUSION
The camu-camu extract suppressed the activation of HIF and VEGF in RPE cells. This could assist anti-VEGF therapy in patients with abnormal ocular neovascularization.
PubMed: 38187496
DOI: 10.1155/2023/6617981 -
Heliyon Jan 2024Tall helophytes such as and often rapidly colonise after rewetting of former agricultural soil and are therefore often the first plants to contribute to the soil...
Comparative flooding tolerance of Typha latifolia and Phalaris arundinacea in wetland restoration: Insights from photosynthetic CO response curves, photobiology and biomass allocation.
Tall helophytes such as and often rapidly colonise after rewetting of former agricultural soil and are therefore often the first plants to contribute to the soil carbon pool. In this study we carried out a mesocosm experiment where these two species grew at three different water levels relative to the soil surface (-15 cm, 0 cm, +15 cm). After eight weeks' growth, measurements of photosynthetic CO-response curves, stomatal conductance and chlorophyll fluorescence of photosystem II were carried out to detect flooding stress. After 10 weeks' growth, the plants were harvested and biomass production, biomass allocation and specific leaf area were determined. had a higher and more stable photosynthetic performance across all water level treatments, which resulted in an overall higher aboveground and belowground production than . In contrast, and decreased by 41 % and 42 %, respectively from drained to flooded conditions with signs of flooding stress as impairment of the photosynthetic apparatus. Moreover, increasing water level resulted in maintenance of aboveground organs for but a decrease in allocation to belowground organs. did not invest in a higher specific leaf area to counter the decreased photosynthesis under flooding. From -15 cm to 0 cm water levels, showed a 68 % reduction in belowground biomass, which has negative implication for carbon retention immediately after rewetting. In contrast, recolonization of is likely to be a suitable contributor to the soil carbon pool due to its stable physiology and high above- and belowground biomass production at all water depths, and also likely under natural water level fluctuations. We showed that even though both species are generally considered wetland plants, they are likely to support considerably different photosynthetic carbon assimilation and soil carbon sequestration rates.
PubMed: 38187246
DOI: 10.1016/j.heliyon.2023.e23657 -
Micromachines Dec 2023The present work is a review of the research on using hydrogels based on natural biodegradable polymers, starch, and gelatin for enzyme immobilization. This review... (Review)
Review
The present work is a review of the research on using hydrogels based on natural biodegradable polymers, starch, and gelatin for enzyme immobilization. This review addresses the main properties of starch and gelatin that make them promising materials in biotechnology for producing enzyme preparations stable during use and storage and insensitive to chemical and physical impacts. The authors summarize their achievements in developing the preparations of enzymes immobilized in starch and gelatin gels and assess their activity, stability, and sensitivity for use as biorecognition elements of enzyme inhibition-based biosensors.
PubMed: 38138386
DOI: 10.3390/mi14122217 -
Journal of Molecular Biology Mar 2024Cryptochromes are a ubiquitously occurring class of photoreceptors. Together with photolyases, they form the Photolyase Cryptochrome Superfamily (PCSf) by sharing a...
Cryptochromes are a ubiquitously occurring class of photoreceptors. Together with photolyases, they form the Photolyase Cryptochrome Superfamily (PCSf) by sharing a common protein architecture and binding mode of the FAD chromophore. Despite these similarities, PCSf members exert different functions. Photolyases repair UV-induced DNA damage by photocatalytically driven electron transfer between FADH and the DNA lesion, whereas cryptochromes are light-dependent signaling molecules and trigger various biological processes by photoconversion of their FAD redox and charge states. Given that most cryptochromes possess a C-terminal extension (CTE) of varying length, the functions of their CTE have not yet been fully elucidated and are hence highly debated. In this study, the role of the CTE was investigated for a novel subclass of the PCSf, the CryP-like cryptochromes, by hydrogen/deuterium exchange and mass-spectrometric analysis. Striking differences in the relative deuterium uptake were observed in different redox states of CryP from the diatom Phaeodactylum tricornutum. Based on these measurements we propose a model for light-triggered conformational changes in CryP-like cryptochromes that differs from other known cryptochrome families like the insect or plant cryptochromes.
Topics: Cryptochromes; Deoxyribodipyrimidine Photo-Lyase; Deuterium; Diatoms; Electron Transport; Protein Domains
PubMed: 38123123
DOI: 10.1016/j.jmb.2023.168408 -
Nature Communications Dec 2023Diatoms are dominant marine algae and contribute around a quarter of global primary productivity, the success of which is largely attributed to their photosynthetic...
Diatoms are dominant marine algae and contribute around a quarter of global primary productivity, the success of which is largely attributed to their photosynthetic capacity aided by specific fucoxanthin chlorophyll-binding proteins (FCPs) to enhance the blue-green light absorption under water. We purified a photosystem II (PSII)-FCPII supercomplex and a trimeric FCP from Cyclotella meneghiniana (Cm) and solved their structures by cryo-electron microscopy (cryo-EM). The structures reveal detailed organizations of monomeric, dimeric and trimeric FCP antennae, as well as distinct assemblies of Lhcx6_1 and dimeric FCPII-H in PSII core. Each Cm-PSII-FCPII monomer contains an Lhcx6_1, an FCP heterodimer and other three FCP monomers, which form an efficient pigment network for harvesting energy. More diadinoxanthins and diatoxanthins are found in FCPs, which may function to quench excess energy. The trimeric FCP contains more chlorophylls c and fucoxanthins. These diversified FCPs and PSII-FCPII provide a structural basis for efficient light energy harvesting, transfer, and dissipation in C. meneghiniana.
Topics: Photosystem II Protein Complex; Diatoms; Cryoelectron Microscopy; Chlorophyll Binding Proteins; Photosynthesis; Light-Harvesting Protein Complexes
PubMed: 38071196
DOI: 10.1038/s41467-023-44055-8