Pudilan Keyanning mouthwash inhibits dextran-dependent aggregation and biofilm organization of Streptococcus mutans.

AIMS: This research aimed to investigate the inhibitory effects of Pudilan mouthwash (PDL) on Streptococcus mutans (S. mutans) biofilms and identify its chemical components. METHODS AND RESULTS: The impacts of 100% concentrated PDL on S. mutans biofilm were detected by colony-forming unit (CFU) assays, crystal violet staining, confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM), and quantitative real-time PCR (qRT‒PCR). The biocompatibility with human gingival fibroblasts (HGFs) was evaluated by Cell-Counting-Kit-8 (CCK-8) assay. And chemical components were identified by UPLC-HRMS. PBS and 0.12% chlorhexidine were used as negative and positive controls, respectively. Results indicate early 8-h S. mutans biofilms are sensitive to PDL. Additionally, it leads to a decrease in bacterial activities and dextran-dependent aggregation in 24-h S. mutans biofilms. PDL significantly downregulates the gene expression of gtfB/C/D and smc. And 114 components are identified. CONCLUSIONS: PDL has an inhibitory effect on S. mutans and favorable biocompatibility. It has potential to be exploited as a novel anti-biofilm agent.

Genome-Wide Identification of Histone Deacetylases and Their Roles Related with Light Response in Tartary Buckwheat (Fagopyrum tataricum).

Histone deacetylases (HDACs), known as histone acetylation erasers, function crucially in plant growth and development. Although there are abundant reports focusing on HDACs of Arabidopsis and illustrating their important roles, the knowledge of HDAC genes in Tartary buckwheat (Polygonales Polygonaceae Fagopyrum tataricum (L.) Gaertn) is still scarce. In the study, a total of 14 HDAC genes were identified and divided into three main groups: Reduced Potassium Dependency-3/His-52 tone Deacetylase 1 (RPD3/HDA1), Silent Information Regulator 2 (SIR2), and the plant-53 specific HD2. Domain and motif composition analysis showed there were conserved domains and motifs in members from the same subfamilies. The 14 FtHDACs were distributed asymmetrically on 7 chromosomes, with three segmental events and one tandem duplication event identified. The prediction of the cis-element in promoters suggested that FtHDACs probably acted in numerous biological processes including plant growth, development, and response to environmental signals. Furthermore, expression analysis based on RNA-seq data displayed that all FtHDAC genes were universally and distinctly expressed in diverse tissues and fruit development stages. In addition, we found divergent alterations in FtHDACs transcript abundance in response to different light conditions according to RNA-seq and RT-qPCR data, indicating that five FtHDACs might be involved in light response. Our findings could provide fundamental information for the HDAC gene family and supply several targets for future function analysis of FtHDACs related with light response of Tartary buckwheat.

Evaluation of Human Hepatocyte Drug Metabolism Carrying High-Risk or Protection-Associated Liver Disease Genetic Variants.

Metabolic-dysfunction-associated steatotic liver disease (MASLD), which affects 30 million people in the US and is anticipated to reach over 100 million by 2030, places a significant financial strain on the healthcare system. There is presently no FDA-approved treatment for MASLD despite its public health significance and financial burden. Understanding the connection between point mutations, liver enzymes, and MASLD is important for comprehending drug toxicity in healthy or diseased individuals. Multiple genetic variations have been linked to MASLD susceptibility through genome-wide association studies (GWAS), either increasing MASLD risk or protecting against it, such as PNPLA3 rs738409, MBOAT7 rs641738, GCKR rs780094, HSD17B13 rs72613567, and MTARC1 rs2642438. As the impact of genetic variants on the levels of drug-metabolizing cytochrome P450 (CYP) enzymes in human hepatocytes has not been thoroughly investigated, this study aims to describe the analysis of metabolic functions for selected phase I and phase II liver enzymes in human hepatocytes. For this purpose, fresh isolated primary hepatocytes were obtained from healthy liver donors (n = 126), and liquid chromatography-mass spectrometry (LC-MS) was performed. For the cohorts, participants were classified into minor homozygotes and nonminor homozygotes (major homozygotes + heterozygotes) for five gene polymorphisms. For phase I liver enzymes, we found a significant difference in the activity of CYP1A2 in human hepatocytes carrying MBOAT7 (p = 0.011) and of CYP2C8 in human hepatocytes carrying PNPLA3 (p = 0.004). It was also observed that the activity of CYP2C9 was significantly lower in human hepatocytes carrying HSD17B13 (p = 0.001) minor homozygous compared to nonminor homozygous. No significant difference in activity of CYP2E1, CYP2C8, CYP2D6, CYP2E1, CYP3A4, ECOD, FMO, MAO, AO, and CES2 and in any of the phase II liver enzymes between human hepatocytes carrying genetic variants for PNPLA3 rs738409, MBOAT7 rs641738, GCKR rs780094, HSD17B13 rs72613567, and MTARC1 rs2642438 were observed. These findings offer a preliminary assessment of the influence of genetic variations on drug-metabolizing cytochrome P450 (CYP) enzymes in healthy human hepatocytes, which may be useful for future drug discovery investigations.

Sucrose selectively regulates Streptococcus mutans polysaccharide by GcrR.

Streptococcus mutans (S. mutans) is the principal etiological agent in cariogenesis because of its ability to metabolize sucrose into extracellular polysaccharides (EPS). The response regulators GcrR and VicR could regulate EPS metabolism, but with opposing regulatory functions. However, the cooperative interactions between gcrR and vicR regulating sucrose-selective EPS metabolism have not been fully elucidated. First, we constructed several dual-mutant strains (vicR + gcrR+, vicR and gcrR overexpression; vicR + gcrR-, vicR overexpression and gcrR deficient; ASvicRgcrR+, vicR low-expression and gcrR overexpression; ASvicRgcrR-, vicR low-expression and gcrR deficient) to clarify gtfB/gtfC expression levels were modulated by gcrR regardless of the vicR gene expression levels. Next, we found gcrR deletion mutant (SmugcrR) displayed obvious auto-aggregation and bacterial cells were densely packed in enriched EPS induced by sucrose. In the contrast, SmugcrR biofilm showed very little carbohydrate-dependent aggregation in the absence of sucrose. The presence of sucrose amplifies the negative regulation of gcrR acting as a 'switch-off'. After sucrose induction, dexA gene expression was significantly enhanced in gcrR overexpression mutant (SmugcrR+). Furthermore, GcrR was shown to directly bind to the promoter region of the dexA gene. Taken together, our results reveal that GcrR interacts with VicR to block EPS biosynthesis via polysaccharide digestion by DexA, and that this process is induced in a sucrose-selective manner. Hence, targeting GcrR is a potential strategy for the management of dental caries.

SARS-CoV-2 infection of African green monkeys results in mild respiratory disease discernible by PET/CT imaging and shedding of infectious virus from both respiratory and gastrointestinal tracts.

Vaccines are urgently needed to combat the global coronavirus disease 2019 (COVID-19) pandemic, and testing of candidate vaccines in an appropriate non-human primate (NHP) model is a critical step in the process. Infection of African green monkeys (AGM) with a low passage human isolate of SARS-CoV-2 by aerosol or mucosal exposure resulted in mild clinical infection with a transient decrease in lung tidal volume. Imaging with human clinical-grade 18F-fluoro-2-deoxy-D-glucose positron emission tomography (18F-FDG PET) co-registered with computed tomography (CT) revealed pulmonary lesions at 4 days post-infection (dpi) that resolved over time. Infectious virus was shed from both respiratory and gastrointestinal (GI) tracts in all animals in a biphasic manner, first between 2-7 dpi followed by a recrudescence at 14-21 dpi. Viral RNA (vRNA) was found throughout both respiratory and gastrointestinal systems at necropsy with higher levels of vRNA found within the GI tract tissues. All animals seroconverted simultaneously for IgM and IgG, which has also been documented in human COVID-19 cases. Young AGM represent an species to study mild/subclinical COVID-19 disease and with possible insights into live virus shedding. Future vaccine evaluation can be performed in AGM with correlates of efficacy being lung lesions by PET/CT, virus shedding, and tissue viral load.

UHRF1 Controls Thymocyte Fate Decisions through the Epigenetic Regulation of EGR1 Expression.

Thymocyte differentiation is a highly complex process that is accompanied by epigenetic changes. Ubiquitin-like containing PHD ring finger 1 (UHRF1) is a critical epigenetic modifier involved in various cellular processes. In this study, we demonstrated that it is highly expressed in T cell precursors of the thymus. Further, its deficiency results in significantly reduced thymocyte cellularity and thymus size in mice. Through systematic analysis based on single-cell RNA sequencing, we found that UHRF1 deficiency thwarts αß T cell lineage development, whereas biasing γδ T lineage differentiation dampens the progression of immature single-positive cells. UHRF1 deficiency promotes the IL-17 secreting and RORγt expression in γδ T cell, indicating a Tγδ17 phenotype. Further, the analysis of gene-regulatory networks demonstrated that UHRF1 controls the expression of early growth response 1 (EGR1). UHRF1 interacts with DNA methyltransferase 1 (DNMT1) at the CpG promoter region of Egr1 loci and affects the nearby chromatin modifications of H3K9me3 and H3K4me3. Taken together, our results demonstrate that UHRF1 is a key factor that mediates the epigenetic regulation of EGR1 and, consequently, thymocyte fate decisions.

Enterococcus faecalis rnc gene modulates its susceptibility to disinfection agents: a novel approach against biofilm.

BACKGROUND: Enterococcus faecalis (E. faecalis) plays an important role in the failure of root canal treatment and refractory periapical periodontitis. As an important virulence factor of E. faecalis, extracellular polysaccharide (EPS) serves as a matrix to wrap bacteria and form biofilms. The homologous rnc gene, encoding Ribonuclease III, has been reported as a regulator of EPS synthesis. In order to develop novel anti-biofilm targets, we investigated the effects of the rnc gene on the biological characteristics of E. faecalis, and compared the biofilm tolerance towards the typical root canal irrigation agents and traditional Chinese medicine fluid Pudilan. METHODS: E. faecalis rnc gene overexpression (rnc+) and low-expression (rnc-) strains were constructed. The growth curves of E. faecalis ATCC29212, rnc+, and rnc- strains were obtained to study the regulatory effect of the rnc gene on E. faecalis. Scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), and crystal violet staining assays were performed to evaluate the morphology and composition of E. faecalis biofilms. Furthermore, the wild-type and mutant biofilms were treated with 5% sodium hypochlorite (NaOCl), 2% chlorhexidine (CHX), and Pudilan. The residual viabilities of E. faecalis biofilms were evaluated using crystal violet staining and colony counting assays. RESULTS: The results demonstrated that the rnc gene could promote bacterial growth and EPS synthesis, causing the EPS-barren biofilm morphology and low EPS/bacteria ratio. Both the rnc+ and rnc- biofilms showed increased susceptibility to the root canal irrigation agents. The 5% NaOCl group showed the highest biofilm removing effect followed by Pudilan and 2% CHX. The colony counting results showed almost complete removal of bacteria in the 5% NaOCl, 2% CHX, and Chinese medicine agents' groups. CONCLUSIONS: This study concluded that the rnc gene could positively regulate bacterial proliferation, EPS synthesis, and biofilm formation in E. faecalis. The rnc mutation caused an increase in the disinfectant sensitivity of biofilm, indicating a potential anti-biofilm target. In addition, Pudilan exhibited an excellent ability to remove E. faecalis biofilm.

[Mechanism of Qingwei Powder in treatment of periodontitis based on UPLC-Q-TOF-MS, GC-MS, network pharmacology and molecular docking].

The present study explored the mechanism of Qingwei Powder(QP) in the treatment of periodontitis based on chromatography-mass spectrometry and network pharmacology-molecular docking techniques. UPLC-Q-TOF-MS and GC-MS were used to identify the chemical constituents of QP. The active components and targets were screened out through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP), and their targets were predicted using SwissTargetPrediction. Targets related to periodontitis were obtained from GeneCards, OMIM, and DisGeNET. Venn diagram was constructed using Venny 2.1 to obtain the intersection targets. Cytoscape 3.7.2 was used to construct the "chemical component-target-disease" network. The targets were analyzed for Gene Ontology(GO) function and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment by clusterProfiler R, and the "chemical component-target-pathway" network was constructed. The binding activity of the active components to the target proteins was verified by molecular docking. A total of 189 chemical components were obtained by UPLC-Q-TOF-MS and GC-MS, including 39 active components with 180 potential targets related to periodontitis. Target enrichment analysis of the active components yielded 92 KEGG pathways. Twenty KEGG pathways, 34 active components, and 99 targets were involved in the "chemical component-target-pathway" network. Molecular docking verified a good binding ability of the key targets to the key compounds. This study preliminarily indicates that QP is effective in treating periodontitis through multiple components, multiple targets, and multiple pathways, which reflects the complex system of Chinese medicine. This study provides the theoretical foundation for the subsequent research on the material basis and key quality attributes of QP.

PICK1 inhibits the E3 ubiquitin ligase activity of Parkin and reduces its neuronal protective effect.

Parkin functions as a multipurpose E3 ubiquitin ligase, and Parkin loss of function is associated with both sporadic and familial Parkinson's disease (PD). We report that the Bin/Amphiphysin/Rvs (BAR) domain of protein interacting with PRKCA1 (PICK1) bound to the really interesting new gene 1 (RING1) domain of Parkin and potently inhibited the E3 ligase activity of Parkin by disrupting its interaction with UbcH7. Parkin translocated to damaged mitochondria and led to their degradation in neurons, whereas PICK1 robustly inhibited this process. PICK1 also impaired the protective function of Parkin against stresses in SH-SY5Y cells and neurons. The protein levels of several Parkin substrates were reduced in young PICK1-knockout mice, and these mice were resistant to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-mediated toxicity. Taken together, the results indicate that PICK1 is a potent inhibitor of Parkin, and the reduction of PICK1 enhances the protective effect of Parkin.

The UV absorption of graphene oxide is size-dependent: possible calibration pitfalls.

Graphene oxide (GO) is often quantified via its UV absorption, typically at around 230 nm. This is convenient but the effect of the size of GO on the accuracy of this method has been ignored so far. The authors report that the molar absorbance of GO is size-dependent. Data are presented on the absorbance of small (hydrodynamic diameter 1 µm), medium sized (1.5 µm), and large (2.2 µm) GO particles at wavelengths of 210, 230 and 250 nm. In general, linear relationship and good regression fits are obtained, but with different slope depending on size even at the same wavelength. This implies that using the UV absorption-based calibration may cause significant errors in GO quantification. Ultimately, this leads to incorrect dosages and faulty conclusions. This may also explain a variety of inconsistent results obtained in previous biological applications of GO. Graphical abstract The size of graphene oxide (GO) determines its UV absorption and the UV absorption-based calibration (GO-s, GO-m and GO-l represent the GO with small, medium and large size).

ICA1L forms BAR-domain complexes with PICK1 and is crucial for acrosome formation in spermiogenesis.

Mutations in the Pick1 gene cause globozoospermia, a male infertility disorder, in both mice and humans. PICK1 is crucial for vesicle trafficking, and its deficiency in sperm cells leads to abnormal vesicle trafficking from the Golgi to the acrosome. This eventually disrupts acrosome formation and leads to male infertility. Here, we identified ICA1L, which has sequence similarities to ICA69 (also known as ICA1), as a new BAR-domain binding partner of PICK1. ICA1L is expressed in testes and brain, and is the major binding partner for PICK1 in testes. ICA1L and PICK1 are highly expressed in spermatids and trafficked together at different stages of spermiogenesis. ICA1L-knockout mice were generated by CRISPR-Cas technology. PICK1 expression was reduced by 80% in the testes of male mice lacking ICA1L. Sperm from ICA1L-knockout mice had abnormalities in the acrosome, nucleus and mitochondrial sheath formation. Both total and mobile sperm numbers were reduced, and about half of the remaining sperm had the characteristics of globozoospermia. These defects ultimately resulted in reduced fertility of male ICA1L-knockout mice, and ICA69/ICA1L-double knockout male mice were sterile.

Collective effects of SNPs on transgenerational inheritance in Caenorhabditis elegans and budding yeast.

We studied the collective effects of single nucleotide polymorphisms (SNPs) on transgenerational inheritance in Caenorhabditis elegans recombinant inbred advanced intercross lines (RIAILs) and yeast segregants. We divided the RIAILs and segregants into two groups of high and low minor allele content (MAC). RIAILs with higher MAC needed less generations of benzaldehyde training to gain a stable olfactory imprint and showed a greater change from normal after benzaldehyde training. Yeast segregants with higher MAC showed a more dramatic shortening of the lag phase length after ethanol exposure. The short lag phase as acquired by ethanol training was more dramatically lost after recovery in ethanol free medium for the high MAC group. We also found a preferential association between MAC and traits linked with higher number of additive QTLs. These results suggest a role for the collective effects of SNPs in transgenerational inheritance, and may help explain human variations in disease susceptibility.

Cationic liposomes as a drug-free system for efficient anticancer therapy by intracytoplasmic delivery of sodium bicarbonate.

Developing the delivery systems with high therapeutic efficacy and low side effects is of great interest and significance for anticancer therapy. Compared to the high cost in synthesizing new chemotherapeutic drugs, exploring the anticancer potentials of existing chemicals is more convenient and efficient. Sodium bicarbonate (BC), a simple inorganic salt, has shown its tumor inhibition capacity via regulating the acidity of tumor microenvironment. However, the effects of intracytoplasmic BC on tumor growth and the potentials of BC to serve as an anticancer agent are still unknown. Herein, we developed a BC-loaded cationic liposome system (BC-CLP) to deliver BC into the cytosol of cancer cells. The in vitro studies showed that the BC-CLP containing 1% BC (w/v) had a size of 112.9 nm and a zeta potential of 19.1 mV, which reduced the viability of the model cancer cells (human oral squamous cell carcinoma HSC-3 cells) to 13.7%. In contrast, the neutral BC-LP caused less than 50% viability reduction. We further found that BC-CLP released BC directly into cytoplasm via membrane fusion pathway rather than endocytosis, leading to the remarkable increase of cytosolic pH, which may contribute to the anticancer effect of BC-CLP. Our findings indicate that BC-CLP is a potential system for high-efficiency cancer therapy without causing drug-related side effects or resistance.

Cardiovascular Risk Associated With Social Determinants of Health at Individual and Area Levels.

Importance: The benefit of adding social determinants of health (SDOH) when estimating atherosclerotic cardiovascular disease (ASCVD) risk is unclear. Objective: To examine the association of SDOH at both individual and area levels with ASCVD risks, and to assess if adding individual- and area-level SDOH to the pooled cohort equations (PCEs) or the Predicting Risk of CVD Events (PREVENT) equations improves the accuracy of risk estimates. Design, Setting, and Participants: This cohort study included participants data from 4 large US cohort studies. Eligible participants were aged 40 to 79 years without a history of ASCVD. Baseline data were collected from 1995 to 2007; median (IQR) follow-up was 13.0 (9.3-15.0) years. Data were analyzed from September 2023 to February 2024. Exposures: Individual- and area-level education, income, and employment status. Main outcomes and measures: ASCVD was defined as the composite outcome of nonfatal myocardial infarction, death from coronary heart disease, and fatal or nonfatal stroke. Results: A total of 26 316 participants were included (mean [SD] age, 61.0 [9.1] years; 15 494 women [58.9%]; 11 365 Black [43.2%], 703 Chinese American [2.7%], 1278 Hispanic [4.9%], and 12 970 White [49.3%]); 11 764 individuals (44.7%) had at least 1 adverse individual-level SDOH and 10 908 (41.5%) had at least 1 adverse area-level SDOH. A total of 2673 ASCVD events occurred during follow-up. SDOH were associated with increased risk of ASCVD at both the individual and area levels, including for low education (individual: hazard ratio [HR], 1.39 [95% CI, 1.25-1.55]; area: HR, 1.31 [95% CI, 1.20-1.42]), low income (individual: 1.35 [95% CI, 1.25-1.47]; area: HR, 1.28 [95% CI, 1.17-1.40]), and unemployment (individual: HR, 1.61 [95% CI, 1.24-2.10]; area: HR, 1.25 [95% CI, 1.14-1.37]). Adding area-level SDOH alone to the PCEs did not change model discrimination but modestly improved calibration. Furthermore, adding both individual- and area-level SDOH to the PCEs led to a modest improvement in both discrimination and calibration in non-Hispanic Black individuals (change in C index, 0.0051 [95% CI, 0.0011 to 0.0126]; change in scaled integrated Brier score [IBS], 0.396% [95% CI, 0.221% to 0.802%]), and improvement in calibration in White individuals (change in scaled IBS, 0.274% [95% CI, 0.095% to 0.665%]). Adding individual-level SDOH to the PREVENT plus area-level social deprivation index (SDI) equations did not improve discrimination but modestly improved calibration in White participants (change in scaled IBS, 0.182% [95% CI, 0.040% to 0.496%]), Black participants (0.187% [95% CI, 0.039% to 0.501%]), and women (0.289% [95% CI, 0.115% to 0.574%]). Conclusions and Relevance: In this cohort study, both individual- and area-level SDOH were associated with ASCVD risk; adding both individual- and area-level SDOH to the PCEs modestly improved discrimination and calibration for estimating ASCVD risk for Black individuals, and adding individual-level SDOH to PREVENT plus SDI also modestly improved calibration. These findings suggest that both individual- and area-level SDOH may be considered in future development of ASCVD risk assessment tools, particularly among Black individuals.

Distinct oral-associated gastric microbiota and Helicobacter pylori communities for spatial microbial heterogeneity in gastric cancer.

The gastric microbial community plays a fundamental role in gastric cancer (GC), and the two main anatomical subtypes of GC, non-cardia and cardia GC, are associated with different risk factors (Helicobacter pylori for non-cardia GC). To decipher the different microbial spatial communities of GC, we performed a multicenter retrospective analysis to characterize the gastric microbiota in 223 GC patients, including H. pylori-positive or -negative patients, with tumors and paired adjacent normal tissues, using third-generation sequencing. In the independent validation cohort, both dental plaque and GC tumoral tissue samples were collected and sequenced. The prevalence of H. pylori and oral-associated bacteria was verified using fluorescence in situ hybridization (FISH) assays in GC tumoral tissues and matched nontumoral tissues. We found that the vertical distribution of the gastric microbiota, at the upper, middle, and lower third sites of GC, was likely an important factor causing microbial diversity in GC tumor tissues. The oral-associated microbiota cluster, which included Veillonella parvula, Streptococcus oralis, and Prevotella intermedia, was more abundant in the upper third of the GC. However, H. pylori was more abundant in the lower third of the GC and exhibited a significantly high degree of microbial correlation. The oral-associated microbiota module was co-exclusive with H. pylori in the lower third site of the GC tumoral tissue. Importantly, H. pylori-negative GC patients with oral-associated gastric microbiota showed worse overall survival, while the increase in microbial abundance in H. pylori-positive GC patients showed no difference in overall survival. The prevalence of V. parvula in both the dental plaque and GC tissue samples was concordant in the independent validation phase. We showed that the oral-associated species V. parvula and S. oralis were correlated with overall survival. Our study highlights the roles of the oral-associated microbiota in the upper third of the GC. In addition, oral-associated species may serve as noninvasive screening tools for the management of GC and an independent prognostic factor for H. pylori-negative GCs. IMPORTANCE: Our study highlights the roles of the oral-associated microbiota in the upper third of gastric cancer (GC).We showed that the oral-associated species Veillonella parvula and Streptococcus oralis were correlated with overall survival. In addition, oral-associated species may serve as noninvasive screening tools for the management of GC and an independent prognostic factor for Helicobacter pylori-negative GCs.

Comparison of Wild-Type and High-risk PNPLA3 variants in a Human Biomimetic Liver Microphysiology System for Metabolic Dysfunction-associated Steatotic Liver Disease Precision Therapy.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is a worldwide health epidemic with a global occurrence of approximately 30%. The pathogenesis of MASLD is a complex, multisystem disorder driven by multiple factors including genetics, lifestyle, and the environment. Patient heterogeneity presents challenges for developing MASLD therapeutics, creation of patient cohorts for clinical trials and optimization of therapeutic strategies for specific patient cohorts. Implementing pre-clinical experimental models for drug development creates a significant challenge as simple in vitro systems and animal models do not fully recapitulate critical steps in the pathogenesis and the complexity of MASLD progression. To address this, we implemented a precision medicine strategy that couples the use of our liver acinus microphysiology system (LAMPS) constructed with patient-derived primary cells. We investigated the MASLD-associated genetic variant PNPLA3 rs738409 (I148M variant) in primary hepatocytes, as it is associated with MASLD progression. We constructed LAMPS with genotyped wild type and variant PNPLA3 hepatocytes together with key non-parenchymal cells and quantified the reproducibility of the model. We altered media components to mimic blood chemistries, including insulin, glucose, free fatty acids, and immune activating molecules to reflect normal fasting (NF), early metabolic syndrome (EMS) and late metabolic syndrome (LMS) conditions. Finally, we investigated the response to treatment with resmetirom, an approved drug for metabolic syndrome-associated steatohepatitis (MASH), the progressive form of MASLD. This study using primary cells serves as a benchmark for studies using patient biomimetic twins constructed with patient iPSC-derived liver cells using a panel of reproducible metrics. We observed increased steatosis, immune activation, stellate cell activation and secretion of pro-fibrotic markers in the PNPLA3 GG variant compared to wild type CC LAMPS, consistent with the clinical characterization of this variant. We also observed greater resmetirom efficacy in PNPLA3 wild type CC LAMPS compared to the GG variant in multiple MASLD metrics including steatosis, stellate cell activation and the secretion of pro-fibrotic markers. In conclusion, our study demonstrates the capability of the LAMPS platform for the development of MASLD precision therapeutics, enrichment of patient cohorts for clinical trials, and optimization of therapeutic strategies for patient subgroups with different clinical traits and disease stages.

Association Between Young Adult Characteristics and Blood Pressure Trajectories.

BACKGROUND: Blood pressure (BP) trajectories from young adulthood through middle age are associated with cardiovascular risk. We examined the associations of hypertension risk factors with BP trajectories among a large diverse sample. METHODS AND RESULTS: We analyzed data from young adults, aged 18 to 39 years, with untreated BP <140/90 mm Hg at baseline from Kaiser Permanente Southern California (N=355 324). We used latent growth curve models to identify 10-year BP trajectories and to assess the associations between characteristics in young adulthood and BP trajectories. We identified the following 5 distinct systolic BP trajectories, which appeared to be determined mainly by the baseline BP with progressively higher BP at each year: group 1 (lowest BP trajectory, 7.9%), group 2 (26.5%), group 3 (33.0%), group 4 (25.4%), and group 5 (highest BP trajectory, 7.3%). Older age (adjusted odds ratio for 30-39 versus 18-29 years, 1.23 [95% CI, 1.18-1.28]), male sex (13.38 [95% CI, 12.80-13.99]), obesity (body mass index ≥30 versus 18.5-24.9 kg/m2, 14.81 [95% CI, 14.03-15.64]), overweight (body mass index 25-29.9 versus 18.5-24.9 kg/m2, 3.16 [95% CI, 3.00-3.33]), current smoking (1.58 [95% CI, 1.48-1.67]), prediabetes (1.21 [95% CI, 1.13-1.29]), diabetes (1.60 [95% CI, 1.41-1.81]) and high low-density lipoprotein cholesterol (≥160 versus <100 mg/dL, 1.52 [95% CI, 1.37-1.68]) were associated with the highest BP trajectory (group 5) compared with the reference group (group 2). CONCLUSIONS: Traditional hypertension risk factors including smoking, diabetes, and elevated lipids were associated with BP trajectories in young adults, with obesity having the strongest association with the highest BP trajectory group.

Polymorphisms Associated With Metabolic Dysfunction-Associated Steatotic Liver Disease Influence the Progression of End-Stage Liver Disease.

BACKGROUND AND AIMS: Chronic liver injury that results in cirrhosis and end-stage liver disease (ESLD) causes more than 1 million deaths annually worldwide. Although the impact of genetic factors on the severity of metabolic dysfunction-associated steatotic liver disease (MASLD) and alcohol-related liver disease (ALD) has been previously studied, their contribution to the development of ESLD remains largely unexplored. METHODS: We genotyped 6 MASLD-associated polymorphisms in healthy (n = 123), metabolic dysfunction-associated steatohepatitis (MASH) (n = 145), MASLD-associated ESLD (n = 72), and ALD-associated ESLD (n = 57) cohorts and performed multinomial logistic regression to determine the combined contribution of genetic, demographic, and clinical factors to the progression of ESLD. RESULTS: Distinct sets of factors are associated with the progression to ESLD. The PNPLA3 rs738409:G and TM6SF2 rs58542926:T alleles, body mass index (BMI), age, and female sex were positively associated with progression from a healthy state to MASH. The PNPLA3 rs738409:G allele, age, male sex, and having type 2 diabetes mellitus were positively associated, while BMI was negatively associated with progression from MASH to MASLD-associated ESLD. The PNPLA3 rs738409:G and GCKR rs780094:T alleles, age, and male sex were positively associated, while BMI was negatively associated with progression from a healthy state to ALD-associated ESLD. The findings indicate that the PNPLA3 rs738409:G allele increases susceptibility to ESLD regardless of etiology, the TM6SF2 rs58542926:T allele increases susceptibility to MASH, and the GCKR rs780094:T allele increases susceptibility to ALD-associated ESLD. CONCLUSION: The PNPLA3, TM6SF2, and GCKR minor alleles influence the progression of MASLD-associated or ALD-associated ESLD. Genotyping for these variants in MASLD and ALD patients can enhance risk assessment, prompting early interventions to prevent ESLD.

No evidence for enhanced disease with human polyclonal SARS-CoV-2 antibody in the ferret model.

Since SARS-CoV-2 emerged in late 2019, it spread from China to the rest of the world. An initial concern was the potential for vaccine- or antibody-dependent enhancement (ADE) of disease as had been reported with other coronaviruses. To evaluate this, we first developed a ferret model by exposing ferrets to SARS-CoV-2 by either mucosal inoculation (intranasal/oral/ocular) or inhalation using a small particle aerosol. Mucosal inoculation caused a mild fever and weight loss that resolved quickly; inoculation via either route resulted in virus shedding detected in the nares, throat, and rectum for 7-10 days post-infection. To evaluate the potential for ADE, we then inoculated groups of ferrets intravenously with 0.1, 0.5, or 1 mg/kg doses of a human polyclonal anti-SARS-CoV-2 IgG from hyper-immunized transchromosomic bovines (SAB-185). Twelve hours later, ferrets were challenged by mucosal inoculation with SARS-CoV-2. We found no significant differences in fever, weight loss, or viral shedding after infection between the three antibody groups or the controls. Signs of pathology in the lungs were noted in infected ferrets but no differences were found between control and antibody groups. The results of this study indicate that healthy, young adult ferrets of both sexes are a suitable model of mild COVID-19 and that low doses of specific IgG in SAB-185 are unlikely to enhance the disease caused by SARS-CoV-2.

Penpulimab, an anti-PD-1 antibody, for heavily pretreated metastatic nasopharyngeal carcinoma: a single-arm phase II study.

Penpulimab is an anti-programmed cell death-1 (PD-1) IgG1 antibody with no Fc gamma receptor (FcγR) binding activity, and thus theoretically reduced immune-related adverse events (irAEs) while maintaining efficacy. This single-arm, phase II trial conducted across 20 tertiary care centers in China enrolled adult patients with metastatic nasopharyngeal carcinoma (NPC) who had failed two or more lines of previous systemic chemotherapy. Patients received 200-mg penpulimab intravenously every 2 weeks (4 weeks per cycle) until disease progression or intolerable toxicities. The primary endpoint was objective response rate (ORR) per RECIST (version 1.1), as assessed by an independent radiological review committee. The secondary endpoints included progression-free survival (PFS) and overall survival (OS). One hundred thirty patients were enrolled and 125 were efficacy evaluable. At the data cutoff date (September 28, 2022), 1 patient achieved complete response and 34 patients attained partial response. The ORR was 28.0% (95% CI 20.3-36.7%). The response was durable, with 66.8% still in response at 9 months. Thirty-three patients (26.4%) were still on treatment. The median PFS and OS were 3.6 months (95% CI = 1.9-7.3 months) and 22.8 months (95% CI = 17.1 months to not reached), respectively. Ten (7.6%) patients experienced grade 3 or higher irAEs. Penpulimab has promising anti-tumor activities and acceptable toxicities in heavily pretreated metastatic NPC patients, supporting further clinical development as third-line treatment of metastatic NPC.