Cerliponase alfa decreases Aß load and alters autophagy- related pathways in mouse hippocampal neurons exposed to fAß1-42.

Extracellular aggregation of amyloid-beta (Aß) in the brain plays a central role in the onset and progression of Alzheimer's disease (AD). Moreover, intraneuronal accumulation of Aß via oligomer internalization might play an important role in the progression of AD. Deficient autophagy, which is a lysosomal degradation process, occurs during the early stages of AD. Tripeptidyl peptidase-1 (TPP1) functions as a lysosomal enzyme, and TPP1 gene mutations are associated with type 2 late infantile neuronal ceroid lipofuscinosis (LINCL). Nevertheless, there is little information about the role of TPP1 in the pathogenesis of AD; therefore, the present study aimed to measure the decrease in intraneuronal Aß accumulation by a recombinant analog of the TPP1 enzyme, cerliponase alfa (CER) (Brineura®), and to determine whether autophagy pathways play a role in this decrease. In this study, endogenous Aß accumulation was induced by fAß1-42 (a toxic fragment of full-length Aß) exposure, and mouse hippocampal neuronal cells (HT-22) were treated with CER (human recombinant rhTPP1 1 mg mL-1). Soluble Aß, TPP1, and the proteins involved in autophagy, including mammalian target of rapamycin (p-mTOR/mTOR), p62/sequestosome-1 (p62/SQSTM1), and microtubule-associated protein 1 A/1B-light chain 3 (LC3), were evaluated using western blotting. The sirtuin-1, beclin-1, and Atg5 genes were also studied using RT-PCR. Aß and TPP1 localizations were observed via immunocytochemistry. CER reduced the Aß load in HT-22 cells by inducing TPP1 expression and converting pro-TPP1 into the mature form. Furthermore, exposure to CER and fAß1-42 induced the autophagy-regulatory/related pathways in HT-22 cells and exposure to CER alone increased sirtuin-1 activity. Based on the present findings, we suggest that augmentation of TPP1 with enzyme replacement therapy may be a potential therapeutic option for the treatment of AD.

Utilising primary care electronic health records to deliver the ALABAMA randomised controlled trial of penicillin allergy assessment.

BACKGROUND: Use of electronic health records (EHR) to provide real-world data for research is established, but using EHR to deliver randomised controlled trials (RCTs) more efficiently is less developed. The Allergy AntiBiotics And Microbial resistAnce (ALABAMA) RCT evaluated a penicillin allergy assessment pathway versus usual clinical care in a UK primary care setting. The aim of this paper is to describe how EHRs were used to facilitate efficient delivery of a large-scale randomised trial of a complex intervention embracing efficient participant identification, supporting minimising GP workload, providing accurate post-intervention EHR updates of allergy status, and facilitating participant follow up and outcome data collection. The generalisability of the EHR approach and health economic implications of EHR in clinical trials will be reported in the main ALABAMA trial cost-effectiveness analysis. METHODS: A descriptive account of the adaptation of functionality within SystmOne used to deliver/facilitate multiple trial processes from participant identification to outcome data collection. RESULTS: An ALABAMA organisation group within SystmOne was established which allowed sharing of trial functions/materials developed centrally by the research team. The 'ALABAMA unit' within SystmOne was also created and provided a secure efficient environment to access participants' EHR data. Processes of referring consented participants, allocating them to a trial arm, and assigning specific functions to the intervention arm were developed by adapting tools such as templates, reports, and protocols which were already available in SystmOne as well as pathways to facilitate allergy de-labelling processes and data retrieval for trial outcome analysis. CONCLUSIONS: ALABAMA is one of the first RCTs to utilise SystmOne EHR functionality and data across the RCT delivery, demonstrating feasibility and applicability to other primary care RCTs. TRIAL REGISTRATION: ClinicalTrials.gov: NCT04108637, registered 05/03/2019. ISRCTN: ISRCTN20579216.

Personality interferences in the pathology of breast cancer: a cross-sectional single-center study.

Individual personality refers to the Ego and the interpersonal sector. The Ego corresponds to consciousness and self-esteem, including the capacities for emotional self-regulation, self-control, self-evaluation, and self-direction in relation to personal goals. When neoplastic and psychiatric diseases coexist, a patient's quality of life is significantly impacted. While there are somatic differences in disease progression, how the illness is perceived and mainly experienced depends on personality traits. In this study, we administered the DECAS Personality Inventory (a Romanian-validated instrument based on the Five-Factor model of personality) to a group of 121 patients diagnosed with breast cancer to explore the relationships among their personality traits. Descriptive statistics revealed that the mean T scores for openness, extroversion, and emotional stability were low, while the scores for conscientiousness and agreeableness were at an average level. Our findings suggest that, in the studied group, low levels of emotional stability, extroversion, and openness were unfavorable personality dimensions that should be a primary focus of therapeutic strategies, as they significantly affect the quality of life in patients with breast cancer.

Strongly ROS-Correlated, Time-Dependent, and Selective Antiproliferative Effects of Synthesized Nano Vesicles on BRAF Mutant Melanoma Cells and Their Hyaluronic Acid-Based Hydrogel Formulation.

Cutaneous metastatic melanoma (CMM) is the most aggressive form of skin cancer with a poor prognosis. Drug-induced secondary tumorigenesis and the emergency of drug resistance worsen an already worrying scenario, thus rendering urgent the development of new treatments not dealing with mutable cellular processes. Triphenyl phosphonium salts (TPPSs), in addiction to acting as cytoplasmic membrane disruptors, are reported to be mitochondria-targeting compounds, exerting anticancer effects mainly by damaging their membranes and causing depolarization, impairing mitochondria functions and their DNA, triggering oxidative stress (OS), and priming primarily apoptotic cell death. TPP-based bola amphiphiles are capable of self-forming nanoparticles (NPs) with enhanced biological properties, as commonly observed for nanomaterials. Already employed in several other biomedical applications, the per se selective potent antibacterial effects of a TPP bola amphiphile have only recently been demonstrated on 50 multidrug resistant (MDR) clinical superbugs, as well as its exceptional and selective anticancer properties on sensitive and MDR neuroblastoma cells. Here, aiming at finding new molecules possibly developable as new treatments for counteracting CMM, the effects of this TPP-based bola amphiphile (BPPB) have been investigated against two BRAF mutants CMM cell lines (MeOV and MeTRAV) with excellent results (even IC50 = 49 nM on MeOV after 72 h treatment). With these findings and considering the low cytotoxicity of BPPB against different mammalian non-tumoral cell lines and red blood cells (RBCs, selectivity indexes up to 299 on MeOV after 72 h treatment), the possible future development of BPPB as topical treatment for CMM lesions was presumed. With this aim, a biodegradable hyaluronic acid (HA)-based hydrogel formulation (HA-BPPB-HG) was prepared without using any potentially toxic crosslinking agents simply by dispersing suitable amounts of the two ingredients in water and sonicating under gentle heating. HA-BPPB-HA was completely characterized, with promising outcomes such as high swelling capability, high porosity, and viscous elastic rheological behavior.

Cotranscriptional Folding of a 5' Stem-loop in the Escherichia coli tbpA Riboswitch at Single-nucleotide Resolution.

Transcription elongation is one of the most important processes in the cell. During RNA polymerase elongation, the folding of nascent transcripts plays crucial roles in the genetic decision. Bacterial riboswitches are prime examples of RNA regulators that control gene expression by altering their structure upon metabolite sensing. It was previously revealed that the thiamin pyrophosphate-sensing tbpA riboswitch in Escherichia coli cotranscriptionally adopts three main structures leading to metabolite sensing. Here, using single-molecule FRET, we characterize the transition in which the first nascent structure, a 5' stem-loop, is unfolded during transcription elongation to form the ligand-binding competent structure. Our results suggest that the structural transition occurs in a relatively abrupt manner, i.e., within a 1-2 nucleotide window. Furthermore, a highly dynamic structural exchange is observed, indicating that riboswitch transcripts perform rapid sampling of nascent co-occurring structures. We also observe that the presence of the RNAP stabilizes the 5' stem-loop along the elongation process, consistent with RNAP interacting with the 5' stem-loop. Our study emphasizes the role of early folding stem-loop structures in the cotranscriptional formation of complex RNA molecules involved in genetic regulation.

Human Beta Defensin-2 mRNA and Proteasome Subunit ß Type 8 mRNA Analysis, Useful in Differentiating Skin Biopsies from Atopic Dermatitis and Psoriasis Vulgaris Patients.

(1): Atopic dermatitis and psoriasis vulgaris are chronic, inflammatory diseases. Clinical presentation usually leads to a proper diagnosis, but sometimes neither clinical examination nor histopathological evaluation can be conclusive. Therefore, we aimed to build up a novel diagnostic tool and check it for accuracy. The main objective of our work was to differentiate between healthy skin (C), atopic dermatitis (AD) and psoriasis vulgaris (PV) biopsies on the base of involucrin (IVL) and human ß-defensin-2 (hBD-2) concentrations and their mRNA, as well as mRNA for TPP2 and PSMB8. (2): ELISA for IVL and hBD-2 proteins and Real-time PCR for the relative expression of mRNA for: IVL (IVL mRNA), hBD-2 (hBD-2 mRNA), PSMB8 (PSMB8 mRNA) and TPP2 (TPP2 mRNA), isolated from skin biopsies taken from AD and PV patients and healthy volunteers were performed. (3): hBD-2 mRNA and PSMB8 mRNA correlated with some parameters of clinical assessment of inflammatory disease severity. hBD-2 mRNA expression, exclusively, was sufficient to distinguish inflammatory skin biopsies from the healthy ones. (4): hBD-2 mRNA and PSMB8 mRNA analysis were the most valuable parameters in differentiating AD and PV biopsies.

Tripolyphosphate-functionalized cellulose: A green solution for cadmium contamination.

This study introduces a highly efficient tripolyphosphate -tethered cellulose sorbent for cadmium (Cd2⁺) removal from aqueous solutions. Characterization through FTIR and SEM revealed the material's structural properties. The sorbent achieved 99% Cd2⁺ removal even at a minimal dosage of 0.05 g. Optimal sorption occurred within the pH range of 4-6, influenced by the sorbent's weak acidic functional groups. Rapid kinetics, reaching equilibrium within a minute, and a high sorption capacity (up to 18.03 mg/g at 50 °C) were observed. Langmuir isotherm modeling confirmed monolayer sorption, and thermodynamic studies indicated a spontaneous, endothermic process with increased randomness at the solid-liquid interface. Selectivity studies demonstrated strong Cd2⁺ removal performance in the presence of competing ions, with minimal interference from monovalent ions but notable effects from divalent ions. The sorbent exhibited consistent reusability over multiple cycles. XPS analysis conclusively established an ion exchange mechanism between Cd2⁺ and negatively charged P3O105- groups as the primary removal pathway. This research highlights the potential of TPP-tethered cellulose as a promising sorbent for effective Cd2⁺ remediation.

Artificial Meshed Vessel-Induced Dimensional Breaking Growth of Human Brain Organoids and Multiregional Assembloids.

Brain organoids are widely used to model brain development and diseases. However, a major challenge in their application is the insufficient supply of oxygen and nutrients to the core region, restricting the size and maturation of the organoids. In order to vascularize brain organoids and enhance the nutritional supply to their core areas, two-photon polymerization (TPP) 3D printing is employed to fabricate high-resolution meshed vessels in this study. These vessels made of photoresist with densely distributed micropores with a diameter of 20 µm on the sidewall, are cocultured with brain organoids to facilitate the diffusion of culture medium into the organoids. The vascularized organoids exhibit dimensional breaking growth and enhanced proliferation, reduced hypoxia and apoptosis, suggesting that the 3D-printed meshed vessels partially mimic vascular function to promote the culture of organoids. Furthermore, cortical, striatal and medial ganglionic eminence (MGE) organoids are respectively differentiated to generate Cortico-Striatal-MGE assembloids by 3D-printed vessels. The enhanced migration, projection and excitatory signaling transduction are observed between different brain regional organoids in the assembloids. This study presents an approach using TPP 3D printing to construct vascularized brain organoids and assembloids for enhancing the development and assembly, offering a research model and platform for neurological diseases.

Cellular thermal shift assay: an approach to identify and assess protein target engagement.

INTRODUCTION: A comprehensive and global knowledge of protein target engagement is of vital importance for mechanistic studies and in drug development. Since its initial introduction, the cellular thermal shift assay (CETSA) has proven to be a reliable and flexible technique that can be widely applied to multiple contexts and has profound applications in facilitating the identification and assessment of protein target engagement. AREAS COVERED: This review introduces the principle of CETSA, elaborates on western blot-based CETSA and MS-based thermal proteome profiling (TPP) as well as the major applications and prospects of these approaches. EXPERT OPINION: CETSA primarily evaluates a given ligand binding to a particular target protein in cells and tissues with the protein thermal stabilities analyzed by western blot. When coupling mass spectrometry with CETSA, thermal proteome profiling allows simultaneous proteome-wide experiment that greatly increased the efficiency of target engagement evaluation, and serves as a promising strategy to identify protein targets and off-targets as well as protein-protein interactions to uncover the biological effects. The CETSA approaches have broad applications and potentials in drug development and clinical research.

Identification of organic constituents on atmospheric particulate matter in the East Asian background air of free troposphere by GC×GC-TOFMS.

The presence of organic compounds on the particulate matter (PM) or aerosols can arise from the condensation of gaseous organic compounds on the existing aerosols, or from organic precursors to form secondary organic aerosols (SOA) through photochemistry. The objective of this study is to characterize organic constituents on aerosols relevant to their emission sources and the key compounds revealing the evolution of aerosols with the use of a novel analytical technique. A time-of-flight mass spectrometry (TOFMS) coupled with comprehensive two-dimensional gas chromatography (GC×GC) was developed using a flow type of modulator instead of a thermal type as a prelude to field applications without the need for cryogen. The methodology of GC×GC-TOFMS is discussed in this study in detail. Since the coarse PM (PM10-2.5) may exhibit with a relatively high OC content compared to PM2.5, the GC×GC results have been obtained by analyzing PM10 samples collected in parallel with OC/EC analysis of PM2.5 samples at the Lulin Atmospheric Background Station (LABS, 23.47°N, 120.87°E, 2862 m ASL) as the high-mountain background site in East Asia. We found that the organic analytes were in a majority in the range of 12-30 carbon numbers falling in the category of semi-volatile organic compounds (SVOCs) with 43 compounds of alcohol, aldehyde, ketone, and ester varieties if excluding alkanes. Intriguingly, trace amounts of plasticizers and phosphorus flame retardants such as phthalates (PAEs) and triphenyl phosphate (TPP) were also found, likely originating from regions involved in open burning of household solid waste in Southeast Asia or e-waste recycling in southern China and along the long-range transport route. Compounds such as these are unique to the specific sources, demonstrating the wide spread of these hazardous compounds in the environment.

Coincident Thyrotoxic Hypokalemic Periodic Paralysis and Cardiomyopathy.

Thyrotoxic periodic paralysis (TPP) and thyrotoxic cardiomyopathy (TCMP) are potentially lethal complications of thyrotoxicosis that require emergent recognition and management to attenuate significant morbidity and mortality. We present the case of a 23-year-old Asian male with no prior medical history who developed TPP with coincident TCMP, which was successfully managed with antithyroid and heart failure therapies. The clinician should be aware of the diagnosis and treatment of these 2 life-threatening conditions in a hyperthyroid state.

The neuronal ceroid lipofuscinosis type 2 - associated variants: An analysis of alterations in the TPP1 gene and genotype-phenotype correlation in Ukraine.

The neuronal ceroid lipofuscinosis type 2 (CLN2) is a heterogeneous group of neurodegenerative lysosomal storage disorders caused by autosomal recessive inheritance of two pathogenic variants in trans in the TPP1 gene. Classical late-infantile CLN2 disease has a very well-defined natural history. However, a small number of patients with TPP1 enzyme deficiency present a later onset or protracted disease course within this group there are phenotypic variants. Our work aimed to identify pathological variants in the TPP1 gene that conditioned the development of CLN2 disease in Ukrainian patients, to compare these variants with those found in patients from other European and non-European regions, and to make genotype-phenotype associations for this disease. The phenotypes and genotypes of the 48 CLN2-affected individuals belonging to 43 families were profiled through clinical data collection, enzyme analysis, and genotyping. In most patients, genotype and phenotype correlation are in keeping with the data of previous studies. The clinical signs of the disease in patients with new, previously undescribed variants, allowed us to augment existing data about genotype-phenotype correlations for CLN2 disease. The combination of genotype and clinical form of the disease demonstrated that predicting the type and clinical course of the disease based on genotype is very complicated. The data we obtained supplements existing information on genotype-phenotypic correlations in this rare disease, which, in turn, lays the foundation for a personalized approach to the management of this disease.

Mitochondria-Targeted Liposomes for Drug Delivery to Tumor Mitochondria.

The special bilayer structure of mitochondrion is a promising therapeutic target in the diagnosis and treatment of diseases such as cancer and metabolic diseases. Nanocarriers such as liposomes modified with mitochondriotropic moieties can be developed to send therapeutic molecules to mitochondria. In this study, DSPE-PEG-TPP polymer conjugate was synthesized and used to prepare mitochondria-targeted liposomes (TPPLs) to improve the therapeutic index of chemotherapeutic agents functioning in mitochondria and reduce their side effects. Doxorubicin (Dox) loaded-TPPL and non-targeted PEGylated liposomes (PPLs) were prepared and compared based on physicochemical properties, morphology, release profile, cellular uptake, mitochondrial localization, and anticancer effects. All formulations were spherically shaped with appropriate size, dispersity, and zeta potential. The stability of the liposomes was favorable for two months at 4 °C. TPPLs localize to mitochondria, whereas PPLs do not. The empty TPPLs and PPLs were not cytotoxic to HCT116 cells. The release kinetics of Dox-loaded liposomes showed that Dox released from TPPLs was higher at pH 5.6 than at pH 7.4, which indicates a higher accumulation of the released drug in the tumor environment. The half-maximal inhibitory concentration of Dox-loaded TPPLs and PPLs was 1.62-fold and 1.17-fold lower than that of free Dox due to sustained drug release, respectively. The reactive oxygen species level was significantly increased when HCT116 cells were treated with Dox-loaded TPPLs. In conclusion, TPPLs may be promising carriers for targeted drug delivery to tumor mitochondria.

Phytopathogenic Fungicidal Activity and Mechanism Approach of Three Kinds of Triphenylphosphonium Salts.

The triphenylphosphonium (TPP) cation has been widely used as a carrier for mitochondria-targeting molecules. We synthesized two commonly employed targeting systems, namely, ω-triphenylphosphonium fatty acids (group 2) and ω-triphenylphosphonium fatty alcohols (group 3), to assess the impact of the TPP module on the biological efficacy of mitochondria-targeting molecules. We evaluated their fungicidal activities against nine plant pathogenic fungi in comparison to alkyl-1-triphenylphosphonium compounds (group 1). All three compound groups exhibited fungicidal activity and displayed a distinct "cut-off effect", which depended on the length of the carbon chain. Specifically, group 1 compounds showed a cut-off point at C10 (compound 1-7), while group 2 and 3 compounds exhibited cut-off points at C15 (compound 2-12) and C14 (compound 3-11), respectively. Notably, group 1 compounds displayed significantly higher fungicidal activity compared to groups 2 and 3. However, group 2 and 3 compounds showed similar activity to each other, although susceptibility may depend on the pathogen tested. Initial investigations into the mechanism of action of the most active compounds suggested that their fungicidal performance may be primarily attributed to their ability to damage the membrane, as well as uncoupling activity and inhibition of fungal respiration. Our findings suggest that the TPP module used in delivery systems as aliphatic acyl or alkoxyl derivatives with carbon chains length < 10 will contribute negligible fungicidal activity to the TPP-conjugate compared to the effect of high level of accumulation in mitochondria due to its mitochondria-targeting ability. These results provide a foundation for utilizing TPP as a promising carrier in the design and development of more effective mitochondria-targeting drugs or pesticides.

Facilitating the use of the target product profile in academic research: a systematic review.

BACKGROUND: The Target Product Profile (TPP) is a tool used in industry to guide development strategies by addressing user needs and fostering effective communication among stakeholders. However, they are not frequently used in academic research, where they may be equally useful. This systematic review aims to extract the features of accessible TPPs, to identify commonalities and facilitate their integration in academic research methodology. METHODS: We searched peer-reviewed papers published in English developing TPPs for different products and health conditions in four biomedical databases. Interrater agreement, computed on random abstract and paper sets (Cohen's Kappa; percentage agreement with zero tolerance) was > 0.91. We interviewed experts from industry contexts to gain insight on the process of TPP development, and extracted general and specific features on TPP use and structure. RESULTS: 138 papers were eligible for data extraction. Of them, 92% (n = 128) developed a new TPP, with 41.3% (n = 57) focusing on therapeutics. The addressed disease categories were diverse; the largest (47.1%, n = 65) was infectious diseases. Only one TPP was identified for several fields, including global priorities like dementia. Our analyses found that 56.5% of papers (n = 78) was authored by academics, and 57.8% of TPPs (n = 80) featured one threshold level of product performance. The number of TPP features varied widely across and within product types (n = 3-44). Common features included purpose/context of use, shelf life for drug stability and validation aspects. Most papers did not describe the methods used to develop the TPP. We identified aspects to be taken into account to build and report TPPs, as a starting point for more focused initiatives guiding use by academics. DISCUSSION: TPPs are used in academic research mostly for infectious diseases and have heterogeneous features. Our extraction of key features and common structures helps to understand the tool and widen its use in academia. This is of particular relevance for areas of notable unmet needs, like dementia. Collaboration between stakeholders is key for innovation. Tools to streamline communication such as TPPs would support the development of products and services in academia as well as industry.

Lipo/TK-CDN/TPP/Y6 nanoparticles inhibit cutaneous melanoma formation.

Stimulation of the innate immune stimulator of interferon genes (STING) pathway has been shown to boost anti-tumour immunity. Nevertheless, the systemic delivery of STING agonists to the tumour presents challenges. Therefore, we designed a cyclic dinucleotide (CDN)-based drug delivery system (DDS) combined photothermal therapy (PTT)/photodynamic therapy (PDT)/immunotherapy for cutaneous melanoma. We coencapsulated a reactive oxygen species (ROS)-responsive prodrug thioketone-linked CDN (TK-CDN), and photoresponsive agents chlorin E6 (Y6) within mitochondria-targeting reagent triphenylphosphonium (TPP)-modified liposomes (Lipo/TK-CDN/TPP/Y6). Lipo/TK-CDN/TPP/Y6 exhibited a photothermal effect similar to Y6, along with a superior cellular uptake rate. Upon endocytosis by B16F10 cells, Lipo/TK-CDN/TPP/Y6 generated large amounts of ROS under laser irradiation for PDT. Mice bearing B16F10 tumours were intravenously injected with Lipo/TK-CDN/TPP/Y6 and exposed to irradiation, resulting in a substantial inhibition of tumour growth. Exploration of the mechanism of anti-tumour action showed that Lipo/TK-CDN/TPP/Y6 had a stronger stimulation of STING activation and anti-tumour immune cell infiltration compared to other groups. Hence, the Lipo/TK-CDN/TPP/Y6 nanoparticles offer great potential as a DDS for targeted and on-demand drug release at tumour sites. These nanoparticles exhibit promise as a candidate for precise and controllable combination therapy in the treatment of tumours.

Weight of evidence evaluation of the metabolism disrupting effects of triphenyl phosphate using an expert knowledge elicitation approach.

Identification of Endocrine-Disrupting Chemicals (EDCs) in a regulatory context requires a high level of evidence. However, lines of evidence (e.g. human, in vivo, in vitro or in silico) are heterogeneous and incomplete for quantifying evidence of the adverse effects and mechanisms involved. To date, for the regulatory appraisal of metabolism-disrupting chemicals (MDCs), no harmonised guidance to assess the weight of evidence has been developed at the EU or international level. To explore how to develop this, we applied a formal Expert Knowledge Elicitation (EKE) approach within the European GOLIATH project. EKE captures expert judgment in a quantitative manner and provides an estimate of uncertainty of the final opinion. As a proof of principle, we selected one suspected MDC -triphenyl phosphate (TPP) - based on its related adverse endpoints (obesity/adipogenicity) relevant to metabolic disruption and a putative Molecular Initiating Event (MIE): activation of peroxisome proliferator activated receptor gamma (PPARγ). We conducted a systematic literature review and assessed the quality of the lines of evidence with two independent groups of experts within GOLIATH, with the objective of categorising the metabolic disruption properties of TPP, by applying an EKE approach. Having followed the entire process separately, both groups arrived at the same conclusion, designating TPP as a "suspected MDC" with an overall quantitative agreement exceeding 85%, indicating robust reproducibility. The EKE method provides to be an important way to bring together scientists with diverse expertise and is recommended for future work in this area.

SLC25A19 is required for NADH homeostasis and mitochondrial respiration.

Mitochondrial transporters facilitate the translocation of metabolites between the cytoplasm and mitochondria and are critical for mitochondrial functional integrity. Although many mitochondrial transporters are associated with metabolic diseases, how they regulate mitochondrial function and their metabolic contributions at the cellular level are largely unknown. Here, we show that mitochondrial thiamine pyrophosphate (TPP) transporter SLC25A19 is required for mitochondrial respiration. SLC25A19 deficiency leads to reduced cell viability, increased integrated stress response (ISR), enhanced glycolysis and elevated cell sensitivity to 2-deoxyglucose (2-DG) treatment. Through a series of biochemical assays, we found that the depletion of mitochondrial NADH is the primary cause of the impaired mitochondrial respiration in SLC25A19 deficient cells. We also showed involvement of SLC25A19 in regulating the enzymatic activities of complexes I and III, the tricarboxylic acid (TCA) cycle, malate-aspartate shuttle and amino acid metabolism. Consistently, addition of idebenone, an analog of coenzyme Q10, restores mitochondrial respiration and cell viability in SLC25A19 deficient cells. Together, our findings provide new insight into the functions of SLC25A19 in mitochondrial and cellular physiology, and suggest that restoring mitochondrial respiration could be a novel strategy for treating SLC25A19-associated disorders.

Ensuring accuracy in the development and application of nucleic acid amplification tests (NAATs) for infectious disease.

Diagnostic tests were heralded as crucial during the Coronavirus disease (COVID-19) pandemic with most of the key methods using bioanalytical approaches that detected larger molecules (RNA, protein antigens or antibodies) rather than conventional clinical biochemical techniques. Nucleic Acid Amplification Tests (NAATs), like the Polymerase Chain Reaction (PCR), and other molecular methods, like sequencing (that often work in combination with NAATs), were essential to the diagnosis and management during COVID-19. This was exemplified both early in the pandemic but also later on, following the emergence of new genetic SARS-CoV-2 variants. The 100 day mission to respond to future pandemic threats highlights the need for effective diagnostics, therapeutics and vaccines. Of the three, diagnostics represents the first opportunity to manage infectious diseases while also being the most poorly supported in terms of the infrastructure needed to demonstrate effectiveness. Where performance targets exist, they are not well served by consensus on how to demonstrate they are being met; this includes analytical factors such as limit of detection (LOD) false positive results as well as how to approach clinical evaluation. The selection of gold standards or use of epidemiological factors such as predictive value, reference ranges or clinical thresholds are seldom correctly considered. The attention placed on molecular diagnostic tests during COVID-19 illustrates important considerations and assumptions on the use of these methods for infectious disease diagnosis and beyond. In this manuscript, we discuss state-of-the-art approaches to diagnostic evaluation and explore how they may be better tailored to diagnostic techniques like NAATs to maximise the impact of these highly versatile bioanalytical tools, both generally and during future outbreaks.

Chitosan-TPP encapsulated quercetin nanoparticles: amplified protection mechanisms unveiled against Ethion-induced developmental toxicity through comprehensive in-vivo and in-silico elucidation.

Aim: The study investigated Ethion-induced developmental toxicity in Wistar albino rats and the potential ameliorative effects of quercetin and nano-quercetin co-administration. Further, In-silico docking of Ethion and quercetin with MCL-1 was conducted. Methodology: Quercetin nanoparticles were synthesized by ionic-gelation method. The encapsulated quercetin nanoparticles were characterized for Zeta size, UV-Vis spectroscopy, encapsulation efficiency, and TEM studies. Male rats were administered Ethion (high/low dose), quercetin, and nano-quercetin alone or in combination for 60 days. Female rats were introduced for mating on the 61st day, and pregnant females were observed for 20 gestational days. On GD 20, rats were sacrificed and evaluated for body/organ weight, reproductive indices, fetal morphology, skeletal, and visceral deformities.In silico binding energies of ethion and quercetin with MCL-1 were determined. Results: Nanoparticle size was 363.2 ± 1.23 nm on day 0 and 385.63 ± 1.53 nm on day 60, with PDI of 0.247 and charge of 22.9 mV. Absorbance maxima were at 374 nm, with encapsulation efficacy of 85.16 ± 0.33%. EHD male crossed females showed decreased body/organ weights, reduced fertility, hematoma, cleft palate, tail curling, and absence of extremity. Nano-quercetin co-administration normalized parameters comparable to controls. Both Ethion and quercetin interacted with MCL-1, with quercetin exhibiting stronger binding energy. Conclusion: Nano-quercetin demonstrated stronger antioxidant properties than quercetin, counteracting ethion-induced maternal/fetal abnormalities.