1,25-dihydroxyvitamin D3 augments low-dose PMA-based monocyte-to-macrophage differentiation in THP-1 cells.

The human monocytic THP-1 cell line is the most routinely employed in vitro model for studying monocyte-to-macrophage differentiation. Despite the wide use of this model, differentiation protocols using phorbol 12-myristate-13-acetate (PMA) or 1,25-dihydroxyvitamin D3 (1,25D3) vary drastically between studies. Given that differences in differentiation protocols have the potential to impact the characteristics of the macrophages produced, we aimed to assess the efficacy of three different THP-1 differentiation protocols by assessing changes in morphology and gene- and cell surface macrophage marker expression. THP-1 cells were differentiated with either 5 nM PMA, 10 nM 1,25D3, or a combination thereof, followed by a rest period. The results indicated that all three protocols significantly increased the expression of the macrophage markers, CD11b (p < 0.001) and CD14 (p < 0.010). Despite this, THP-1 cells exposed to 1,25D3 alone did not adopt the morphological and expression characteristics associated with macrophages. PMA was required to produce these characteristics, which were found to be more pronounced in the presence of 1,25D3. Both PMA- and PMA with 1,25D3-differentiated THP-1 cells were capable of M1 and M2 macrophage polarization, though the gene expression of polarization-associated markers was most pronounced in PMA with 1,25D3-differentiated THP-1 cells. Moreover, the combination of PMA with 1,25D3 appeared to support the process of commitment to a particular polarization state.

The Antimicrobial Activity of Human Defensins at Physiological Non-Permeabilizing Concentrations Is Caused by the Inhibition of the Plasma Membrane H+-ATPases.

Human defensins are cysteine-rich peptides (Cys-rich peptides) of the innate immune system. Defensins contain an ancestral structural motif (i.e., γ-core motif) associated with the antimicrobial activity of natural Cys-rich peptides. In this study, low concentrations of human α- and ß-defensins showed microbicidal activity that was not associated with cell membrane permeabilization. The cell death pathway was similar to that previously described for human lactoferrin, also an immunoprotein containing a γ-core motif. The common features were (1) cell death not related to plasma membrane (PM) disruption, (2) the inhibition of microbicidal activity via extracellular potassium, (3) the influence of cellular respiration on microbicidal activity, and (4) the influence of intracellular pH on bactericidal activity. In addition, in yeast, we also observed (1) partial K+-efflux mediated via Tok1p K+-channels, (2) the essential role of mitochondrial ATP synthase in cell death, (3) the increment of intracellular ATP, (4) plasma membrane depolarization, and (5) the inhibition of external acidification mediated via PM Pma1p H+-ATPase. Similar features were also observed with BM2, an antifungal peptide that inhibits Pma1p H+-ATPase, showing that the above coincident characteristics were a consequence of PM H+-ATPase inhibition. These findings suggest, for the first time, that human defensins inhibit PM H+-ATPases at physiological concentrations, and that the subsequent cytosolic acidification is responsible for the in vitro microbicidal activity. This mechanism of action is shared with human lactoferrin and probably other antimicrobial peptides containing γ-core motifs.

Spatial distribution and determinants of Early sexual initiation in Ethiopia.

INTRODUCTION: Early sexual initiation has negative health, social, and economic consequences for both women and future generations. The trend of early sexual initiation is increasing globally, leading to higher rates of sexually transmitted diseases and unplanned pregnancies. Ethiopia has been challenged various disasters that makes women vulnerable and position them at heightened risk of early sexual initiation in the last four years. The spatial patterns and factors of early sexual initiation in the post-conflict-post pandemic settings is not well understood. Hence this research aimed at mapping Spatial Patterns and identifying determinant factors in the Post-COVID-Post-Conflict Settings. METHODS: The study was conducted on secondary data from the PMA 2021 cross-sectional survey which conducted nationally from November 2021 to January 2022 which is in the post pandemic and post-war period. Total weighted sample of 6,036 reproductive age women were included in the analysis. ArcGIS Pro and SaTScan software were used to handle spatial analysis. Multilevel logistic regression model was used to estimate the effects of independent variables on early sexual initiation at individual and community level factors. Adjusted odds ratio with the 95% confidence interval was reported to declare the strength and statistical significance of the association. RESULT: The spatial distribution of early sexual initiation was clustered in Ethiopia with a global Moran's I index value of 0.09 and Z-score 6.01 (p-value < 0.001).Significant hotspots were detected in East Gojjam zone of Amhara region, Bale, Arsi, West Hararge, East Wellega and Horo Gudru Wellega zones of Oromia region. The odds of having early sexual initiation was higher in women with primary education (AOR = 1.23, 95%CI: 1.03, 1.47), secondary or above education (AOR = 4.36, 95%CI: 3.49, 5.44), Women aged 26 to 25 (AOR = 1.91, 95%CI: 1.61, 2.26), women aged 36 to 49(AOR = 1.51, 95%CI: 1.24, 1.84). However, there was a significant lower likelihood of early sexual initiation in rural resident women (AOR = 0.53, 95%CI: 0.35, 0.81) and women living in 5 to 7 family size (AOR = 0.79, 95%CI: 0.68, 0.92), and more than 7 members (AOR = 0.63, 95%CI: 0.49, 0.81). CONCLUSIONS: The spatial distribution of early sexual initiation was clustered in Ethiopia. Interventions should be taken to eliminate the observed variation by mobilizing resources to high-risk areas. Policies and interventions targeted to this problem may also take the identified associated factors into account for better results.

Rapid Determination of SARS-CoV-2 Integrity and Infectivity by Using Propidium Monoazide Coupled with Digital Droplet PCR.

SARS-CoV-2 is a highly infectious virus responsible for the COVID-19 pandemic. Therefore, it is important to assess the risk of SARS-CoV-2 infection, especially in persistently positive patients. Rapid discrimination between infectious and non-infectious viruses aids in determining whether prevention, control, and treatment measures are necessary. For this purpose, a method was developed and utilized involving a pre-treatment with 50 µM of propidium monoazide (PMAxx, a DNA intercalant) combined with a digital droplet PCR (ddPCR). The ddPCR method was performed on 40 nasopharyngeal swabs (NPSs) both before and after treatment with PMAxx, revealing a reduction in the viral load at a mean of 0.9 Log copies/mL (SD ± 0.6 Log copies/mL). Furthermore, six samples were stratified based on the Ct values of SARS-CoV-2 RNA (Ct < 20, 20 < Ct < 30, Ct > 30) and analyzed to compare the results obtained via a ddPCR with viral isolation and a negative-chain PCR. Of the five samples found positive via a ddPCR after the PMAxx treatment, two of the samples showed the highest post-treatment SARS-CoV-2 loads. The virus was isolated in vitro from both samples and the negative strand chains were detected. In three NPS samples, SARS CoV-2 was present post-treatment at a low level; it was not isolated in vitro, and, when detected, the strand was negative. Our results indicate that the established method is useful for determining whether the SARS-CoV-2 within positive NPS samples is intact and capable of causing infection.

Use of patent term extensions to restore regulatory time for medical devices in the United States.

BACKGROUND: Medical devices can seek patent term extensions (PTEs), which extend market exclusivity to compensate for delays related to clinical trials and regulatory review. Pharmaceutical companies commonly use PTEs, but their use by medical device companies has not been clear. RESEARCH DESIGN AND METHODS: We examined the use of PTEs by medical device companies between 1984 and 2024 using a database published in the Federal Register and a list published by the Patent and Trademark Office. RESULTS: Only 178 medical device submissions were linked to a PTE application. They were mostly concentrated in 116 product codes associated with 15 medical specialties; nearly half were associated with cardiovascular devices. Numbers increased significantly in the past decade. Successful applications restored 987 days on average. CONCLUSIONS: The patent restoration opportunity appears underutilized. It is unclear whether some companies do not recognize the opportunity it promises, or whether it does not meet their needs. Different business features and marketing strategies in device versus pharmaceutical industries may decrease the usefulness of the PTE program for these types of medical products. However, the finding that a small subset of manufacturers operating in competitive markets adopted patent extension strategies more commonly suggests a significant competitive advantage when competition increases.

Evolution of the pathogenic mold Aspergillus fumigatus on high copper levels identifies novel resistance genes.

Aspergillus fumigatus is the leading cause of severe mold infections in immunocompromised patients. This common fungus possesses innate attributes that allow it to evade the immune system, including its ability to survive the high copper (Cu) levels in phagosomes. Our previous work has revealed that under high Cu levels, the A. fumigatus transcription factor AceA is activated, inducing the expression of the copper exporter CrpA to expel excess Cu. To identify additional elements in Cu resistance, we evolved A. fumigatus wild-type and mutant ΔaceA or ΔcrpA strains under increasing Cu concentrations. Sequencing of the resultant resistant strains identified both shared and unique evolutionary pathways to resistance. Reintroduction of three of the most common mutations in genes encoding Pma1 (plasma membrane H+-ATPase), Gcs1 (glutamate cysteine-ligase), and Cpa1 (carbamoyl-phosphate synthetase), alone and in combination, into wild-type A. fumigatus confirmed their additive role in conferring Cu resistance. Detailed analysis indicated that the pma1 mutation L424I preserves Pma1 H+-ATPase activity under high Cu concentrations and that the cpa1 mutation A37V confers a survival advantage to conidia in the presence of Cu. Interestingly, simultaneous mutations of all three genes did not alter virulence in infected mice. Our work has identified novel Cu-resistance pathways and provides an evolutionary approach for dissecting the molecular basis of A. fumigatus adaptation to diverse environmental challenges.IMPORTANCEAspergillus fumigatus is the most common mold infecting patients with weakened immunity. Infection is caused by the inhalation of mold spores into the lungs and is often fatal. In healthy individuals, spores are engulfed by lung immune cells and destroyed by a combination of enzymes, oxidants, and high levels of copper. However, the mold can protect itself by pumping out excess copper with specific transporters. Here, we evolved A. fumigatus under high copper levels and identified new genetic mutations that help it resist the toxic effects of copper. We studied how these mutations affect the mold's ability to resist copper and how they impact its ability to cause disease. This is the first such study in a pathogenic mold, and it gives us a better understanding of how it manages to bypass our body's defenses during an infection.

Leukemia risk assessment of exposure to low-levels of benzene based on the linearized multistage model.

Objectives: To assess leukemia risk in occupational populations exposed to low levels of benzene. Methods: Leukemia incidence data from the Chinese Benzene Cohort Study were fitted using the Linearized multistage (LMS) model. Individual benzene exposure levels, urinary S-phenylmercapturic acid (S-PMA) and trans, trans-muconic acid (t, t-MA) were measured among 98 benzene-exposed workers from factories in China. Subjects were categorized into four groups by rounding the quartiles of cumulative benzene concentrations (< 3, 3-5, 5-12, ≥12 mg/m3·year, respectively). The risk of benzene-induced leukemia was assessed using the LMS model, and the results were validated using the EPA model and the Singapore semi-quantitative risk assessment model. Results: The leukemia risks showed a positive correlation with increasing cumulative concentration in the four exposure groups (excess leukemia risks were 4.34, 4.37, 4.44 and 5.52 × 10-4, respectively; Ptrend < 0.0001) indicated by the LMS model. We also found that the estimated leukemia risk using urinary t, t-MA in the LMS model was more similar to those estimated by airborne benzene compared to S-PMA. The leukemia risk estimated by the LMS model was consistent with both the Singapore semi-quantitative risk assessment model at all concentrations and the EPA model at high concentrations (5-12, ≥12 mg/m3·year), while exceeding the EPA model at low concentrations (< 3 and 3-5 mg/m3·year). However, in all four benzene-exposed groups, the leukemia risks estimated by these three models exceeded the lowest acceptable limit for carcinogenic risk set by the EPA at 1 × 10-6. Conclusion: This study demonstrates the utility of the LMS model derived from the Chinese benzene cohort in assessing leukemia risk associated with low-level benzene exposure, and suggests that leukemia risk may occur at cumulative concentrations below 3 mg/m3·year.

Improving the Efficiency of Viability-qPCR with Lactic Acid Enhancer for the Selective Detection of Live Pathogens in Foods.

Pathogenic Escherichia coli are the most prevalent foodborne bacteria, and their accurate detection in food samples is critical for ensuring food safety. Therefore, a quick technique named viability-qPCR (v-qPCR), which is based on the ability of a selective dye, such as propidium monoazide (PMA), to differentiate between alive and dead cells, has been developed. Despite diverse, successful applications, v-qPCR is impaired by some practical limitations, including the ability of PMA to penetrate the outer membrane of dead Gram-negative bacteria. The objective of this study is to evaluate the ability of lactic acid (LA) to improve PMA penetration and, thus, the efficiency of v-qPCR in detecting the live fraction of pathogens. The pre-treatment of E. coli ATCC 8739 cells with 10 mM LA greatly increased PMA penetration into dead cells compared to conventional PMA-qPCR assay, avoiding false positive results. The limit of detection when using LA-PMA qPCR is 1% viable cells in a mixture of dead and alive cells. The optimized LA-PMA qPCR method was reliably able to detect log 2 CFU/mL culturable E. coli in milk spiked with viable and non-viable bacteria. Lactic acid is cheap, has low toxicity, and can be used to improve the efficiency of the v-qPCR assay, which is economically interesting for larger-scale pathogen detection applications intended for food matrices.

Optimization and Testing of a Commercial Viability PCR Protocol to Detect Escherichia coli in Whole Blood.

Bacteremia, specifically if progressed to sepsis, poses a time-sensitive threat to human and animal health. Escherichia coli is a main causative agent of sepsis in humans. The objective was to evaluate a propidium monoazide (PMA)-based viability PCR (vPCR) protocol to detect and quantify live E. coli from whole blood. We optimized the protocol by adding a eukaryotic-specific lysis step prior to PMA exposure, then used spiking experiments to determine the lower limit of detection (LOD) and linear range of quantification. We also compared the vPCR quantification method to standard colony count of spiked inoculum. Lastly, we calculated percent viability in spiked samples containing 50% live cells or 0% live cells. The LOD was 102 CFU/mL for samples containing live cells only and samples with mixed live and heat-killed cells. The linear range of quantification was 102 CFU/mL to 108 CFU/mL (R2 of 0.997) in samples containing only live cells and 103 CFU/mL to 108 CFU/mL (R2 of 0.998) in samples containing live plus heat-killed cells. A Bland-Altman analysis showed that vPCR quantification overestimates compared to standard plate count of the spiked inoculum, with an average bias of 1.85 Log10 CFU/mL across the linear range when only live cells were present in the sample and 1.98 Log10 CFU/mL when live plus heat-killed cells were present. Lastly, percent viability calculations showed an average 89.5% viable cells for samples containing 50% live cells and an average 19.3% for samples containing 0% live cells. In summary, this optimized protocol can detect and quantify viable E. coli in blood in the presence of heat-killed cells. Additionally, the data presented here provide the groundwork for further development of vPCR to detect and quantify live bacteria in blood in clinical settings.

The Ptk2-Pma1 pathway enhances tolerance to terbinafine in Trichophyton rubrum.

The increasing prevalence of dermatophyte resistance to terbinafine, a key drug in the treatment of dermatophytosis, represents a significant obstacle to treatment. Trichophyton rubrum is the most commonly isolated fungus in dermatophytosis. In T. rubrum, we identified TERG_07844, a gene encoding a previously uncharacterized putative protein kinase, as an ortholog of budding yeast Saccharomyces cerevisiae polyamine transport kinase 2 (Ptk2), and found that T. rubrum Ptk2 (TrPtk2) is involved in terbinafine tolerance. In both T. rubrum and S. cerevisiae, Ptk2 knockout strains were more sensitive to terbinafine compared with the wild types, suggesting that promotion of terbinafine tolerance is a conserved function of fungal Ptk2. Pma1 is activated through phosphorylation by Ptk2 in S. cerevisiae. Overexpression of T. rubrum Pma1 (TrPma1) in T. rubrum Ptk2 knockout strain (ΔTrPtk2) suppressed terbinafine sensitivity, suggesting that the induction of terbinafine tolerance by TrPtk2 is mediated by TrPma1. Furthermore, omeprazole, an inhibitor of plasma membrane proton pump Pma1, increased the terbinafine sensitivity of clinically isolated terbinafine-resistant strains. These findings suggest that, in dermatophytes, the TrPtk2-TrPma1 pathway plays a key role in promoting intrinsic terbinafine tolerance and may serve as a potential target for combinational antifungal therapy against terbinafine-resistant dermatophytes.

N-3-Methylbutyl-benzisoselenazol-3(2H)-one Exerts Antifungal Activity In Vitro and in a Mouse Model of Vulvovaginal Candidiasis.

In the present work, we evaluated the antifungal activities of two novel ebselen analogs, N-allyl-benzisoselenazol-3(2H)-one (N-allyl-bs) and N-3-methylbutylbenzisoselenazol-3(2H)-one (N-3mb-bs). Colorimetric and turbidity assays were performed to determine the minimum inhibitory concentration (MIC) of these compounds in S1 (fluconazole-sensitive) and S2 (fluconazole-resistant) strains of C. albicans. N-3mb-bs was more active than the N-allyl-bs compound. It is noteworthy that the concentration of N-3mb-bs observed to inhibit fungal growth by 50% (18.2 µM) was similar to the concentration observed to inhibit the activity of the yeast plasma membrane H+-ATPase (Pma1p) by 50% (19.6 µM). We next implemented a mouse model of vulvovaginal candidiasis (VVC) using the S1 strain and examined the mouse and yeast proteins present in the vaginal lavage fluid using proteomics. The yeast proteins detected were predominately glycolytic enzymes or virulence factors associated with C. albicans while the mouse proteins present in the lavage fluid included eosinophil peroxidase, desmocollin-1, and gasdermin-A. We then utilized the N-3mb-bs compound (12.5 mg/kg) in the mouse VVC model and observed that it significantly reduced the vaginal fungal burden, histopathological changes in vagina tissue, and expression of myeloperoxidase (MPO). All in all, the present work has identified a potentially promising drug candidate for VVC treatment.

Women alone modern contraceptive use decision making and its correlates, evidence from PMA CS 2021 survey.

BACKGROUND: Women alone contraceptive decisions making has become one of the top burring public health agenda. Despite Contraceptive method options are available and accessible, contraceptive prevalence rate (CPR) in Ethiopia is not far beyond 41%. Evidences showed that the freedom of women to choose the contraceptive method they desired to use is one of the potential determinants for the sluggish pace of increase in contraceptive usage. In this era of sustainable development, determining the level of women own contraceptive use decision making and identifying its correlates is very critical for the ministries and relevant partners' effort in tracking the achievement of Sustainable Development Goal (SDG) 5.2 by providing actionable evidence through informed decision-making with the aim of improving contraceptive uptake; reducing maternal mortality and improve newborn health. METHODS: Nationally representative cross-sectional data from Performance Monitoring for Action (PMA) 2021 was used in this study. The sample was restricted among2446 married women who have been using or most recently used modern contraceptive method. Cell sample size adequacy was checked using a chi-square test. Frequency was computed to characterize the study participants. Multilevel binary logistics regression was used to identify factors associated with women own contraceptive use decision making. The findings were presented in a form of frequencies, percentage and as an odds ratio using 95% confidence interval. A p-value of 0.05 was used to declare significance. RESULTS: This study revealed that higher than one in two women (59.49%; 95% CI: 57.7-61.38%) decide their contraceptive use by themselves. What is more interesting is that 1 in 16 women (6.06%) reported that they did not participated in their contraceptive use decision-making.-. Women aged 20 to 24 years; (AOR: 2.51 (1.04, 4.45)), women who stayed10 and above years in marriage; (AOR: 1.73 (1.08, 2.77)), whose husband and/or partner age is 41 and above years; (AOR: 2.14 (1.06, 4.31)) and those who obtained contraceptive method they desired; (AOR: 2.49 (1.36, 4.57)) had higher odds of deciding their current and/or recent contraceptive use by their own. On the other hand, women mixed feeling if they became pregnant at the time of the survey; (AOR: 0.6 (0.44, 0.91)), women who started using contraceptive at younger age, 19 to 24; (AOR: 0.6 (0.44, 0.81)), those who use long acting and/or permanent method; (AOR: 0.54 (0.41, 0.71)) and those married at younger age, 10 to 19 years; (AOR: 0.28 (0.09, 0.86)) had lower odds of independently deciding their current and/or most recent contraceptive use. CONCLUSION: 59% of women independently decide their contraceptive use which calls up on further improvement to enable each woman to decide by their own, with directing special focus for the 6.06% of women who reported no say in their contraceptive use decision. Activities targeting on enabling women to use the method they preferred, spacing their pregnancy, encouraging women to discuss with their husband on the time and type of contraceptive method they used, advocating and promoting marriage at least to be at the minimum age as indicate by the law and maintain the marriage duration as much as longer are hoped to improve women alone contraceptive use decision making to the fullest.

Unexpected versatile electrical transport behaviors of ferromagnetic nickel films.

Perpendicular magnetic anisotropy (PMA) of magnets is paramount for electrically controlled spintronics due to their intrinsic potentials for higher memory density, scalability, thermal stability and endurance, surpassing an in-plane magnetic anisotropy (IMA). Nickel film is a long-lived fundamental element ferromagnet, yet its electrical transport behavior associated with magnetism has not been comprehensively studied, hindering corresponding spintronic applications exploiting nickel-based compounds. Here, we systematically investigate the highly versatile magnetism and corresponding transport behavior of nickel films. As the thickness reduces within the general thickness regime of a magnet layer for a memory device, the hardness of nickel films' ferromagnetic loop of anomalous Hall effect increases and then decreases, reflecting the magnetic transitions from IMA to PMA and back to IMA. Additionally, the square ferromagnetic loop changes from a hard to a soft one at rising temperatures, indicating a shift from PMA to IMA. Furthermore, we observe a butterfly magnetoresistance resulting from the anisotropic magnetoresistance effect, which evolves in conjunction with the thickness and temperature-dependent magnetic transformations as a complementary support. Our findings unveil the rich magnetic dynamics and most importantly settle down the most useful guiding information for current-driven spintronic applications based on nickel film: The hysteresis loop is squarest for the ∼8 nm-thick nickel film, of highest hardness withRxyr/Rxys∼ 1 and minimumHs-Hc, up to 125 K; otherwise, extra care should be taken for a different thickness or at a higher temperature.

Characterization of extracellular trap production and release by equine neutrophils in response to different stimuli.

This study explores Neutrophil Extracellular Trap (NET) formation in equine neutrophils, which is crucial for eliminating infections and is implicated in various equine inflammatory diseases. We investigated the molecular pathways involved in NET release by equine neutrophils in response to stimuli. We use PMA, A23187, LPS, PAF, OZ, and cytokines, observing NET release in response to PMA, PAF, and A23187. In contrast, LPS, OZ, and the cytokines tested did not induce DNA release or did not consistently induce citrullination of histone 4. Peptidyl-arginine deiminase inhibition completely halted NET release, while NADPH oxidase and mitochondrial reactive oxygen species only played a role in PMA-induced NETs. Neutrophil elastase inhibition modestly affected PAF-induced NET liberation but not in PMA or A23187-induced NET, while myeloperoxidase did not contribute to NET release. We expect to provide a foundation for future investigations into the role of NETs in equine health and disease and the search for potential therapeutic targets.

Identification and quantification of viable Lacticaseibacillus rhamnosus in probiotics using validated PMA-qPCR method.

The identification and quantification of viable bacteria at the species/strain level in compound probiotic products is challenging now. Molecular biology methods, e.g., propidium monoazide (PMA) combination with qPCR, have gained prominence for targeted viable cell counts. This study endeavors to establish a robust PMA-qPCR method for viable Lacticaseibacillus rhamnosus detection and systematically validated key metrics encompassing relative trueness, accuracy, limit of quantification, linear, and range. The inclusivity and exclusivity notably underscored high specificity of the primers for L. rhamnosus, which allowed accurate identification of the target bacteria. Furthermore, the conditions employed for PMA treatment were fully verified by 24 different L. rhamnosus including type strain, commercial strains, etc., confirming its effective discrimination between live and dead bacteria. A standard curve constructed by type strain could apply to commercial strains to convert qPCR Cq values to viable cell numbers. The established PMA-qPCR method was applied to 46 samples including pure cultures, probiotics as food ingredients, and compound probiotic products. Noteworthy is the congruity observed between measured and theoretical values within a 95% confidence interval of the upper and lower limits of agreement, demonstrating the relative trueness of this method. Moreover, accurate results were obtained when viable L. rhamnosus ranging from 103 to 108 CFU/mL. The comprehensive appraisal of PMA-qPCR performances provides potential industrial applications of this new technology in quality control and supervision of probiotic products.

Kilogram-Scale Synthesis of Extremely Small Gadolinium Oxide Nanoparticles as a T1-Weighted Contrast Agent for Magnetic Resonance Imaging.

Magnetic resonance imaging contrast agents are frequently used in clinics to enhance the contrast between diseased and normal tissues. The previously reported poly(acrylic acid) stabilized exceedingly small gadolinium oxide nanoparticles (ES-GdON-PAA) overcame the problems of commercial Gd chelates, but limitations still exist, i.e., high r2/r1 ratio, long blood circulation half-life, and no data for large scale synthesis and formulation optimization. In this study, polymaleic acid (PMA) is found to be an ideal stabilizer to synthesize ES-GdONs. Compared with ES-GdON-PAA, the PMA-stabilized ES-GdON (ES-GdON-PMA) has a lower r2/r1 ratio (2.05, 7.0 T) and a lower blood circulation half-life (37.51 min). The optimized ES-GdON-PMA-9 has an exceedingly small particle size (2.1 nm), excellent water dispersibility, and stability. A facile, efficient, and environmental friendly synthetic method is developed for large-scale synthesis of the ES-GdONs-PMA. The weight of the optimized freeze-dried ES-GdON-PMA-26 synthesized in a 20 L of reactor reaches the kilogram level. The formulation optimization is also finished, and the concentrated ES-GdON-PMA-26 formulation (CGd = 100 mm) after high-pressure steam sterilization possesses eligible physicochemical properties (i.e., pH value, osmolality, viscosity, and density) for investigational new drug application.

Engineering the reductive tricarboxylic acid pathway in Aureobasidium pullulans for enhanced biosynthesis of poly-L-malic acid.

Aureobasidium pullulans produced poly-L-malic acid (PMA) as the main metabolite in fermentation but with relatively low productivity and yield limiting its industrial application. In this study, A. pullulans ZX-10 was engineered to overexpress cytosolic malate dehydrogenase (MDH) and pyruvate carboxylase (PYC) and PMA synthetase (PMS) using a high-copy yeast episomal plasmid with the gpdA promoter from Aspergillus nidulans. Overexpressing endogenous PMS and heterologous MDH and PYC from Aspergillus oryzae respectively increased PMA production by 19 % - 37 % (0.64 - 0.74 g/g vs. 0.54 g/g for wild type) in shake-flask fermentations, demonstrating the importance of the reductive tricarboxylic acid (rTCA) pathway in PMA biosynthesis. A. pullulans co-expressing MDH and PYC produced 96.7 g/L PMA at 0.90 g/L∙h and 0.68 g/g glucose in fed-batch fermentation, which were among the highest yield and productivity reported. The engineered A. pullulans with enhanced rTCA pathway is advantageous and promising for PMA production.

Strong metal support interaction (SMSI) and MoO3 synergistic effect of Pt-based catalysts on the promotion of CO activity and sulfur resistance.

In industrial applications, Pt-based catalysts for CO oxidation have the dual challenges of CO self-poisoning and SO2 toxicity. This study used synthetic Keggin-type H3PMo12O40 (PMA) as the site of Pt, and the Pt-MoO3 produced by decomposition of PMA was anchored to TiO2 to construct the dual-interface structure of Pt-MoO3 and Pt-TiO2, abbreviated as Pt-P&M/TiO2. Pt-0.125P&M/TiO2 with a molar ratio of Pt to PMA of 8:1 showed both good CO oxidation activity and SO2 tolerance. In the CO activity test, the CO complete conversion temperature T100 of Pt-0.125P&M/TiO2 was 113 ℃ (compared with 135 ℃ for Pt/TiO2). In the SO2 resistance test, the conversion efficiency of Pt-0.125P&M/TiO2 at 170 ℃ remained at 60% after 72 h, while that of Pt/TiO2 was only 13%. H2-TPR and XPS tests revealed that lattice oxygen provided by TiO2 and hydroxyl produced by MoO3 increased the CO reaction rate on Pt. According to the DFT theoretical calculation, the electronegative MoO3 attracted the d-orbital electrons of Pt, which reduced the adsorption energy of CO and SO2 from - 4.15 eV and - 2.54 eV to - 3.56 eV and - 1.52 eV, respectively, and further weakened the influence of strong CO adsorption and SO2 poisoning on the catalyst. This work explored the relationship between catalyst structure and catalyst performance and provided a feasible technical idea for the design of high-performance CO catalysts in industrial applications.

Depletion of Gsdma1/2/3 alleviates PMA-induced epidermal hyperplasia by inhibiting the EGFR-Stat3/Akt pathway.

Homeostasis of the skin barrier is essential for maintaining normal skin function. Gasdermin A (GSDMA) is highly expressed in the skin and associated with many skin diseases, such as melanoma and psoriasis. In mice, GSDMA is encoded by three gene homologues, namely Gsdma1, Gsdma2, and Gsdma3. Although Gsdma3 gain-of-function mutations cause hair loss and skin inflammation, Gsdma3-deficient mice do not show any visible phenotypes in skin and hair structures. To explore the physiological function of GSDMA, we generated conventional Gsdma1/2/3 knockout (KO) mice. These mice showed significantly alleviated epidermal hyperplasia and inflammation induced by phorbol 12-myristate 13-acetate (PMA). Furthermore, the alleviation of epidermal hyperplasia depended on the expression of Gsdma1/2/3 specifically in keratinocytes. Mechanistically, Gsdma1/2/3 depletion downregulated epidermal growth factor receptor (EGFR) ligands, leading to the decreased EGFR-Stat3/Akt signalling. These results demonstrate that depletion of Gsdma1/2/3 alleviates PMA-induced epidermal hyperplasia partially by inhibiting the EGFR-Stat3/Akt pathway.