Biomolecules. 2023 Apr 15;13(4):678. doi: 10.3390/biom13040678.

ABSTRACT

An increasing number of plant-based herbal treatments, dietary supplements, medical foods and nutraceuticals and their component phytochemicals are used as alternative treatments to prevent or slow the onset and progression of Alzheimer's disease. Their appeal stems from the fact that no current pharmaceutical or medical treatment can accomplish this. While a handful of pharmaceuticals are approved to treat Alzheimer's, none has been shown to prevent, significantly slow or stop the disease. As a result, many see the appeal of alternative plant-based treatments as an option. Here, we show that many phytochemicals proposed or used as Alzheimer's treatments share a common theme: they work via a calmodulin-mediated mode of action. Some phytochemicals bind to and inhibit calmodulin directly while others bind to and regulate calmodulin-binding proteins, including Aβ monomers and BACE1. Phytochemical binding to Aβ monomers can prevent the formation of Aβ oligomers. A limited number of phytochemicals are also known to stimulate calmodulin gene expression. The significance of these interactions to amyloidogenesis in Alzheimer's disease is reviewed.

PMID:37189425 | DOI:10.3390/biom13040678

Biomolecules. 2023 Mar 26;13(4):596. doi: 10.3390/biom13040596.

ABSTRACT

Form I rubiscos evolved in Cyanobacteria ≥ 2.5 billion years ago and are enzymatically unique due to the presence of small subunits (RbcS) capping both ends of an octameric large subunit (RbcL) rubisco assembly to form a hexadecameric (L8S8) holoenzyme. Although RbcS was previously thought to be integral to Form I rubisco stability, the recent discovery of a closely related sister clade of octameric rubiscos (Form I'; L8) demonstrates that the L8 complex can assemble without small subunits (Banda et al. 2020). Rubisco also displays a kinetic isotope effect (KIE) where the 3PG product is depleted in 13C relative to 12C. In Cyanobacteria, only two Form I KIE measurements exist, making interpretation of bacterial carbon isotope data difficult. To aid comparison, we measured in vitro the KIEs of Form I' (Candidatus Promineofilum breve) and Form I (Synechococcus elongatus PCC 6301) rubiscos and found the KIE to be smaller in the L8 rubisco (16.25 ± 1.36‱ vs. 22.42 ± 2.37‱, respectively). Therefore, while small subunits may not be necessary for protein stability, they may affect the KIE. Our findings may provide insight into the function of RbcS and allow more refined interpretation of environmental carbon isotope data.

PMID:37189344 | DOI:10.3390/biom13040596

Front Oncol. 2023 Apr 28;13:1164535. doi: 10.3389/fonc.2023.1164535. eCollection 2023.

ABSTRACT

Heterogeneity describes the differences among cancer cells within and between tumors. It refers to cancer cells describing variations in morphology, transcriptional profiles, metabolism, and metastatic potential. More recently, the field has included the characterization of the tumor immune microenvironment and the depiction of the dynamics underlying the cellular interactions promoting the tumor ecosystem evolution. Heterogeneity has been found in most tumors representing one of the most challenging behaviors in cancer ecosystems. As one of the critical factors impairing the long-term efficacy of solid tumor therapy, heterogeneity leads to tumor resistance, more aggressive metastasizing, and recurrence. We review the role of the main models and the emerging single-cell and spatial genomic technologies in our understanding of tumor heterogeneity, its contribution to lethal cancer outcomes, and the physiological challenges to consider in designing cancer therapies. We highlight how tumor cells dynamically evolve because of the interactions within the tumor immune microenvironment and how to leverage this to unleash immune recognition through immunotherapy. A multidisciplinary approach grounded in novel bioinformatic and computational tools will allow reaching the integrated, multilayered knowledge of tumor heterogeneity required to implement personalized, more efficient therapies urgently required for cancer patients.

PMID:37188201 | PMC:PMC10175698 | DOI:10.3389/fonc.2023.1164535

Bio Protoc. 2023 May 5;13(9):e4662. doi: 10.21769/BioProtoc.4662. eCollection 2023 May 5.

ABSTRACT

X-ray computed microtomography (µCT) is a powerful tool to reveal the 3D structure of tissues and organs. Compared with the traditional sectioning, staining, and microscopy image acquisition, it allows a better understanding of the morphology and a precise morphometric analysis. Here, we describe a method for 3D visualization and morphometric analysis by µCT scanning of the embryonic heart of iodine-stained E15.5 mouse embryos.

PMID:37188107 | PMC:PMC10176208 | DOI:10.21769/BioProtoc.4662

iScience. 2023 Apr 20;26(5):106536. doi: 10.1016/j.isci.2023.106536. eCollection 2023 May 19.

ABSTRACT

Exponential accumulation of single-cell transcriptomes poses great challenge for efficient assimilation. Here, we present an approach entitled generative pretraining from transcriptomes (tGPT) for learning feature representation of transcriptomes. tGPT is conceptually simple in that it autoregressive models the ranking of a gene in the context of its preceding neighbors. We developed tGPT with 22.3 million single-cell transcriptomes and used four single-cell datasets to evalutate its performance on single-cell analysis tasks. In addition, we examine its applications on bulk tissues. The single-cell clusters and cell lineage trajectories derived from tGPT are highly aligned with known cell labels and states. The feature patterns of tumor bulk tissues learned by tGPT are associated with a wide range of genomic alteration events, prognosis, and treatment outcome of immunotherapy. tGPT represents a new analytical paradigm for integrating and deciphering massive amounts of transcriptome data and it will facilitate the interpretation and clinical translation of single-cell transcriptomes.

PMID:37187700 | PMC:PMC10176267 | DOI:10.1016/j.isci.2023.106536

Front Microbiol. 2023 Apr 28;14:1200733. doi: 10.3389/fmicb.2023.1200733. eCollection 2023.

NO ABSTRACT

PMID:37187534 | PMC:PMC10175803 | DOI:10.3389/fmicb.2023.1200733

Int J Biol Macromol. 2023 May 13:124855. doi: 10.1016/j.ijbiomac.2023.124855. Online ahead of print.

ABSTRACT

Functional nanofibrils from globular proteins are usually formed by heating for several hours at pH 2.0, which induces acidic hydrolysis and consecutive self-association. The functional properties of these micro-metre-long anisotropic structures are promising for biodegradable biomaterials and food applications, but their stability at pH > 2.0 is low. The results presented here show that modified β-lactoglobulin can also form nanofibrils by heating at neutral pH without prior acidic hydrolysis; the key is removing covalent disulfide bonds. The aggregation behaviour of various recombinant β-lactoglobulin variants was systemically studied at pH 3.5 and 7.0. The suppression of intra- and intermolecular disulfide bonds by eliminating one to three out of the five cysteines makes the non-covalent interactions more prevalent and allow for structural rearrangement. This stimulated the linear growth of worm-like aggregates. Full elimination of all five cysteines led to the transformation of worm-like aggregates into actual fibril structures (several hundreds of nanometres long) at pH 7.0. This understanding of the role of cysteine in protein-protein interactions will help to identify proteins and protein modifications to form functional aggregates at neutral pH.

PMID:37187417 | DOI:10.1016/j.ijbiomac.2023.124855

EBioMedicine. 2023 May 13;92:104613. doi: 10.1016/j.ebiom.2023.104613. Online ahead of print.

ABSTRACT

BACKGROUND: Gestational age (GA) and associated level of gastrointestinal tract maturation are major factors driving the initial gut microbiota composition in preterm infants. Besides, compared to term infants, premature infants often receive antibiotics to treat infections and probiotics to restore optimal gut microbiota. How GA, antibiotics, and probiotics modulate the microbiota's core characteristics, gut resistome and mobilome, remains nascent.

METHODS: We analysed metagenomic data from a longitudinal observational study in six Norwegian neonatal intensive care units to describe the bacterial microbiota of infants of varying GA and receiving different treatments. The cohort consisted of probiotic-supplemented and antibiotic-exposed extremely preterm infants (n = 29), antibiotic-exposed very preterm (n = 25), antibiotic-unexposed very preterm (n = 8), and antibiotic-unexposed full-term (n = 10) infants. The stool samples were collected on days of life 7, 28, 120, and 365, and DNA extraction was followed by shotgun metagenome sequencing and bioinformatical analysis.

FINDINGS: The top predictors of microbiota maturation were hospitalisation length and GA. Probiotic administration rendered the gut microbiota and resistome of extremely preterm infants more alike to term infants on day 7 and ameliorated GA-driven loss of microbiota interconnectivity and stability. GA, hospitalisation, and both microbiota-modifying treatments (antibiotics and probiotics) contributed to an elevated carriage of mobile genetic elements in preterm infants compared to term controls. Finally, Escherichia coli was associated with the highest number of antibiotic-resistance genes, followed by Klebsiella pneumoniae and Klebsiella aerogenes.

INTERPRETATION: Prolonged hospitalisation, antibiotics, and probiotic intervention contribute to dynamic alterations in resistome and mobilome, gut microbiota characteristics relevant to infection risk.

FUNDING: Odd-Berg Group, Northern Norway Regional Health Authority.

PMID:37187112 | DOI:10.1016/j.ebiom.2023.104613

Nat Plants. 2023 May 15. doi: 10.1038/s41477-023-01419-8. Online ahead of print.

ABSTRACT

Plant specialized metabolites modulate developmental and ecological functions and comprise many therapeutic and other high-value compounds. However, the mechanisms determining their cell-specific expression remain unknown. Here we describe the transcriptional regulatory network that underlies cell-specific biosynthesis of triterpenes in Arabidopsis thaliana root tips. Expression of thalianol and marneral biosynthesis pathway genes depends on the phytohormone jasmonate and is limited to outer tissues. We show that this is promoted by the activity of redundant bHLH-type transcription factors from two distinct clades and coactivated by homeodomain factors. Conversely, the DOF-type transcription factor DAG1 and other regulators prevent expression of the triterpene pathway genes in inner tissues. We thus show how precise expression of triterpene biosynthesis genes is determined by a robust network of transactivators, coactivators and counteracting repressors.

PMID:37188853 | DOI:10.1038/s41477-023-01419-8

Nat Microbiol. 2023 May 15. doi: 10.1038/s41564-023-01383-1. Online ahead of print.

ABSTRACT

Norovirus (NoV) is the leading global cause of viral gastroenteritis. Young children bear the highest burden of disease and play a key role in viral transmission throughout the population. However, which host factors contribute to age-associated variability in NoV severity and shedding are not well-defined. The murine NoV (MNoV) strain CR6 causes persistent infection in adult mice and targets intestinal tuft cells. Here we find that natural transmission of CR6 from infected dams occurred only in juvenile mice. Direct oral CR6 inoculation of wild-type neonatal mice led to accumulation of viral RNA in the ileum and prolonged shedding in the stool that was replication-independent. This viral exposure induced both innate and adaptive immune responses including interferon-stimulated gene expression and MNoV-specific antibody responses. Interestingly, viral uptake depended on passive ileal absorption of luminal virus, a process blocked by cortisone acetate administration, which prevented ileal viral RNA accumulation. Neonates lacking interferon signalling in haematopoietic cells were susceptible to productive infection, viral dissemination and lethality, which depended on the canonical MNoV receptor CD300LF. Together, our findings reveal developmentally associated aspects of persistent MNoV infection, including distinct tissue and cellular tropism, mechanisms of interferon regulation and severity of infection in the absence of interferon signalling. These emphasize the importance of defining viral pathogenesis phenotypes across the developmental spectrum and highlight passive viral uptake as an important contributor to enteric infections in early life.

PMID:37188813 | DOI:10.1038/s41564-023-01383-1

Oncogene. 2023 May 15. doi: 10.1038/s41388-023-02714-6. Online ahead of print.

ABSTRACT

The DNA damage response (DDR) is intertwined with signaling pathways downstream of oncogenic receptor tyrosine kinases (RTKs). To drive research into the application of targeted therapies as radiosensitizers, a better understanding of this molecular crosstalk is necessary. We present here the characterization of a previously unreported MET RTK phosphosite, Serine 1016 (S1016) that represents a potential DDR-MET interface. MET S1016 phosphorylation increases in response to irradiation and is mainly targeted by DNA-dependent protein kinase (DNA-PK). Phosphoproteomics unveils an impact of the S1016A substitution on the overall long-term cell cycle regulation following DNA damage. Accordingly, the abrogation of this phosphosite strongly perturbs the phosphorylation of proteins involved in the cell cycle and formation of the mitotic spindle, enabling cells to bypass a G2 arrest upon irradiation and leading to the entry into mitosis despite compromised genome integrity. This results in the formation of abnormal mitotic spindles and a lower proliferation rate. Altogether, the current data uncover a novel signaling mechanism through which the DDR uses a growth factor receptor system for regulating and maintaining genome stability.

PMID:37188738 | DOI:10.1038/s41388-023-02714-6

Plant Physiol Biochem. 2023 May 4;200:107743. doi: 10.1016/j.plaphy.2023.107743. Online ahead of print.

ABSTRACT

Ciboria shiraiana is a fungal pathogen and the causal agent of hypertrophy sorosis scleroteniosis (HSS) in mulberry, leading to substantial economic losses in the mulberry fruit-related industry. To obtain HSS resistant resources and investigate the resistance mechanism, the resistances of 14 mulberry varieties were assessed. Morus laevigata Wall. (MLW) varieties showed strong resistance to C. shiraiana, and the pathogen's infection was associated with mulberry fluorescence. Stigmas were identified as the infection site through cutting experiments. Susceptible varieties (S-varieties) displayed secretory droplets on their stigma papillar cell surfaces, while MLWs lacked these secretions. Correlation analysis between the secretion rate and the diseased fruit rate indicated that the differences between resistant varieties (R-varieties) and S-varieties were related to the stigma type. Furthermore, comparative transcriptome analysis was performed on stigma and ovary samples from R- and S-varieties. Compared with the stigma of R-varieties, the key differentially expressed genes (DEGs) with significantly higher expression in S-variety stigmas mainly participated in the fatty acid biosynthetic process. In R-variety stigmas and ovaries, the transcript levels of DEGs involved in defense response, including resistance (R) genes, were significantly higher than that of S-varieties. Overexpression of MlwRPM1-2 and MlwRGA3 enhances resistance to C. shiraiana and Sclerotinia sclerotiorum, but not Botrytis cinerea in tobacco. These findings help us explain the different resistance mechanisms of mulberry to C. shiraiana, and the critical defense genes in R-varieties can be applied to breeding antifungal plant varieties.

PMID:37186979 | DOI:10.1016/j.plaphy.2023.107743

Proc Natl Acad Sci U S A. 2023 May 23;120(21):e2220741120. doi: 10.1073/pnas.2220741120. Epub 2023 May 15.

ABSTRACT

Mammalian orthoreoviruses (reoviruses) serve as potential triggers of celiac disease and have oncolytic properties, making these viruses potential cancer therapeutics. Primary attachment of reovirus to host cells is mainly mediated by the trimeric viral protein, σ1, which engages cell-surface glycans, followed by high-affinity binding to junctional adhesion molecule-A (JAM-A). This multistep process is thought to be accompanied by major conformational changes in σ1, but direct evidence is lacking. By combining biophysical, molecular, and simulation approaches, we define how viral capsid protein mechanics influence virus-binding capacity and infectivity. Single-virus force spectroscopy experiments corroborated by in silico simulations show that GM2 increases the affinity of σ1 for JAM-A by providing a more stable contact interface. We demonstrate that conformational changes in σ1 that lead to an extended rigid conformation also significantly increase avidity for JAM-A. Although its associated lower flexibility impairs multivalent cell attachment, our findings suggest that diminished σ1 flexibility enhances infectivity, indicating that fine-tuning of σ1 conformational changes is required to successfully initiate infection. Understanding properties underlying the nanomechanics of viral attachment proteins offers perspectives in the development of antiviral drugs and improved oncolytic vectors.

PMID:37186838 | DOI:10.1073/pnas.2220741120

ACS Synth Biol. 2023 May 15. doi: 10.1021/acssynbio.2c00618. Online ahead of print.

ABSTRACT

Rhodococcus opacus is a bacterium that has a high tolerance to aromatic compounds and can produce significant amounts of triacylglycerol (TAG). Here, we present iGR1773, the first genome-scale model (GSM) of R. opacus PD630 metabolism based on its genomic sequence and associated data. The model includes 1773 genes, 3025 reactions, and 1956 metabolites, was developed in a reproducible manner using CarveMe, and was evaluated through Metabolic Model tests (MEMOTE). We combine the model with two Constraint-Based Reconstruction and Analysis (COBRA) methods that use transcriptomics data to predict growth rates and fluxes: E-Flux2 and SPOT (Simplified Pearson Correlation with Transcriptomic data). Growth rates are best predicted by E-Flux2. Flux profiles are more accurately predicted by E-Flux2 than flux balance analysis (FBA) and parsimonious FBA (pFBA), when compared to 44 central carbon fluxes measured by 13C-Metabolic Flux Analysis (13C-MFA). Under glucose-fed conditions, E-Flux2 presents an R2 value of 0.54, while predictions based on pFBA had an inferior R2 of 0.28. We attribute this improved performance to the extra activity information provided by the transcriptomics data. For phenol-fed metabolism, in which the substrate first enters the TCA cycle, E-Flux2's flux predictions display a high R2 of 0.96 while pFBA showed an R2 of 0.93. We also show that glucose metabolism and phenol metabolism function with similar relative ATP maintenance costs. These findings demonstrate that iGR1773 can help the metabolic engineering community predict aromatic substrate utilization patterns and perform computational strain design.

PMID:37186551 | DOI:10.1021/acssynbio.2c00618

J Labelled Comp Radiopharm. 2023 Apr 25. doi: 10.1002/jlcr.4026. Online ahead of print.

ABSTRACT

A strategy has been developed for the carbon-14 radiosynthesis of [14 C]-SHP-141, a 4-(7-hydroxycarbamoyl-heptanoyloxy)-benzoic acid methyl ester derivative containing a terminal hydroxamic acid. The synthesis involved four radiochemical transformations. The key step in the radiosynthesis was the conversion of the 7-[14 C]-cyano-heptanoic acid benzyloxyamide [14 C]-4 directly into the carboxylic acid derivative, 7-benzyloxycarbamoyl-[14 C]-heptanoic acid [14 C]-8 using nitrilase-113 biocatalyst. The final step involved deprotection of the benzyloxy group using catalytic hydrogenation to facilitate the release of the hydroxamic acid without cleaving the phenoxy ester. [14 C]-SHP-141 was isolated with a radiochemical purity of 90% and a specific activity of 190 μCi/mg from four radiochemical steps starting from potassium [14 C]-cyanide in a radiochemical yield of 45%.

PMID:37186406 | DOI:10.1002/jlcr.4026

J Med Virol. 2023 Apr;95(4):e28739. doi: 10.1002/jmv.28739.

ABSTRACT

Supervised machine learning (ML) methods have been used to predict antibody responses elicited by COVID-19 vaccines in a variety of clinical settings. Here, we explored the reliability of a ML approach to predict the presence of detectable neutralizing antibody responses (NtAb) against Omicron BA.2 and BA.4/5 sublineages in the general population. Anti-SARS-CoV-2 receptor-binding domain (RBD) total antibodies were measured by the Elecsys® Anti-SARS-CoV-2 S assay (Roche Diagnostics) in all participants. NtAbs against Omicron BA.2 and BA4/5 were measured using a SARS-CoV-2 S pseudotyped neutralization assay in 100 randomly selected sera. A ML model was built using the variables of age, vaccination (number of doses) and SARS-CoV-2 infection status. The model was trained in a cohort (TC) comprising 931 participants and validated in an external cohort (VC) including 787 individuals. Receiver operating characteristics analysis indicated that an anti-SARS-CoV-2 RBD total antibody threshold of 2300 BAU/mL best discriminated between participants either exhibiting or not detectable Omicron BA.2 and Omicron BA.4/5-Spike targeted NtAb responses (87% and 84% precision, respectively). The ML model correctly classified 88% (793/901) of participants in the TC: 717/749 (95.7%) of those displaying ≥2300 BAU/mL and 76/152 (50%) of those exhibiting antibody levels <2300 BAU/mL. The model performed better in vaccinated participants, either with or without prior SARS-CoV-2 infection. The overall accuracy of the ML model in the VC was comparable. Our ML model, based upon a few easily collected parameters for predicting neutralizing activity against Omicron BA.2 and BA.4/5 (sub)variants circumvents the need to perform not only neutralization assays, but also anti-S serological tests, thus potentially saving costs in the setting of large seroprevalence studies.

PMID:37185857 | DOI:10.1002/jmv.28739

Sci Rep. 2023 Apr 25;13(1):6744. doi: 10.1038/s41598-023-34026-w.

ABSTRACT

Myxofibrosarcoma (MFS) and undifferentiated sarcoma (US) have been considered as tumors of the same lineage based on genetic/epigenetic profiling. Although MFS shows a notably better prognosis than US, there are no clear criteria for distinguishing between them. Here, we examined 85 patients with MFS/US and found that tumors with infiltrative growth patterns tended to have more myxoid areas and higher local recurrence rates but fewer distant metastases and better overall survival. Morphologically characteristic sickle-shaped blood vessels, which tended to have fewer αSMA-positive cells, were also observed in these tumors, compared with normal vessels. Based on the incidence of these sickle-shaped blood vessels, we subdivided conventionally diagnosed US into two groups. This stratification was significantly correlated with metastasis and prognosis. RNA sequencing of 24 tumors (9 MFS and 15 US tumors) demonstrated that the proteasome, NF-kB, and VEGF pathways were differentially regulated among these tumors. Expression levels of KDR and NFATC4, which encode a transcription factor responsible for the neuritin-insulin receptor angiogenic signaling, were elevated in the sickle-shaped blood vessel-rich US tumors. These findings indicate that further analyses may help elucidate the malignant potential of MFS/US tumors as well as the development of therapeutic strategies for such tumors.

PMID:37185612 | DOI:10.1038/s41598-023-34026-w

Mol Omics. 2023 Apr 26. doi: 10.1039/d3mo00026e. Online ahead of print.

ABSTRACT

The infection rate of syphilis continues to rise globally, and the difficulty in diagnosis of neurosyphilis promptly needs to be resolved. More specific and sensitive diagnostic markers for latent syphilis and neurosyphilis should be found. Here the metabolic profiles of 88 cerebrospinal fluid samples from syphilis patients and controls were analyzed by LC/MS-based untargeted metabolomics. In total, 272 metabolites based on 3937 features obtained in ESI- mode and 252 metabolites based on 3799 features in ESI+ mode were identified. The experimental process was evaluated by principal component analysis, partial least squares discriminant analysis, and hierarchical cluster analysis. A clear separation between latent syphilis and neurosyphilis was found. Levels of lipid and linoleic acid metabolites, such as 9-oxo-octadecadienoic acid and 9,10,13-trihydroxyoctadecenoic acid, were increased in syphilis patients. In patients with neurosyphilis, significant changes in levels of 5-hydroxy-L-tryptophan (5-HTP) and acetyl-N-formyl-5-methoxykynurenamine (AFMK) in the tryptophan-kynurenine pathway were also detected. Only one metabolite, theophylline, differed significantly between symptomatic and asymptomatic neurosyphilis patients. Additionally, KEGG analysis revealed significant enrichment of tryptophan metabolism pathways, indicating a high correlation between tryptophan metabolism and syphilis symptoms. Levels of linoleic acid metabolites, 5-HTP, AFMK and theophylline were significantly altered in different patients. The role of these differential metabolites in the development of syphilis is worthy of further exploration. Our results may promote the development of biomarkers for diagnosis of latent syphilis from neurosyphilis, and for that of asymptomatic neurosyphilis from symptomatic neurosyphilis in the future.

PMID:37185577 | DOI:10.1039/d3mo00026e

Biosensors (Basel). 2023 Apr 16;13(4):480. doi: 10.3390/bios13040480.

ABSTRACT

Deep Brain Stimulation (DBS) of the subthalamic nucleus (STN) is a surgical procedure for alleviating motor symptoms of Parkinson's Disease (PD). The pattern of DBS (e.g., the electrode pairs used and the intensity of stimulation) is usually optimized by trial and error based on a subjective evaluation of motor function. We tested the hypotheses that DBS releases glutamate in selected basal ganglia nuclei and that the creation of 6-hydroxydopamine (6-OHDA)-induced nigrostriatal lesions alters glutamate release during DBS in those basal ganglia nuclei. We studied the relationship between a pseudo-random binary sequence of DBS and glutamate levels in the STN itself or in the globus pallidus (GP) in anesthetized, control, and 6-OHDA-treated rats. We characterized the stimulus-response relationships between DBS and glutamate levels using a transfer function estimated using System Identification. Stimulation of the STN elevated glutamate levels in the GP and in the STN. Although the 6-OHDA treatment did not affect glutamate dynamics in the STN during DBS in the STN, the transfer function between DBS in the STN and glutamate levels in the GP was significantly altered by the presence or absence of 6-OHDA-induced lesions. Thus, glutamate responses in the GP in the 6-OHDA-treated animals (but not in the STN) depended on dopaminergic inputs. For this reason, measuring glutamate levels in the GP may provide a useful feedback target in a closed-loop DBS device in patients with PD since the dynamics of glutamate release in the GP during DBS seem to reflect the loss of dopaminergic neurons in the SNc.

PMID:37185555 | DOI:10.3390/bios13040480

Biosensors (Basel). 2023 Apr 6;13(4):463. doi: 10.3390/bios13040463.

ABSTRACT

Detection and quantification of DNA biomarkers relies heavily on the yield and quality of DNA obtained by extraction from different matrices. Although a large number of studies have compared the yields of different extraction methods, the repeatability and intermediate precision of these methods have been largely overlooked. In the present study, five extraction methods were evaluated, using digital PCR, to determine their efficiency in extracting DNA from three different Gram-negative bacteria in sputum samples. The performance of two automated methods (GXT NA and QuickPick genomic DNA extraction kit, using Arrow and KingFisher Duo automated systems, respectively), two manual kit-based methods (QIAamp DNA mini kit; DNeasy UltraClean microbial kit), and one manual non-kit method (CTAB), was assessed. While GXT NA extraction kit and the CTAB method have the highest DNA yield, they did not meet the strict criteria for repeatability, intermediate precision, and measurement uncertainty for all three studied bacteria. However, due to limited clinical samples, a compromise is necessary, and the GXT NA extraction kit was found to be the method of choice. The study also showed that dPCR allowed for accurate determination of extraction method repeatability, which can help standardize molecular diagnostic approaches. Additionally, the determination of absolute copy numbers facilitated the calculation of measurement uncertainty, which was found to be influenced by the DNA extraction method used.

PMID:37185538 | DOI:10.3390/bios13040463