Characteristics of Rhizosphere Microbiome, Soil Chemical Properties, and Plant Biomass and Nutrients in Citrus reticulata cv. Shatangju Exposed to Increasing Soil Cu Levels.

The prolonged utilization of copper (Cu)-containing fungicides results in Cu accumulation and affects soil ecological health. Thus, a pot experiment was conducted using Citrus reticulata cv. Shatangju with five Cu levels (38, 108, 178, 318, and 388 mg kg-1) to evaluate the impacts of the soil microbial processes, chemistry properties, and citrus growth. These results revealed that, with the soil Cu levels increased, the soil total Cu (TCu), available Cu (ACu), organic matter (SOM), available potassium (AK), and pH increased while the soil available phosphorus (AP) and alkali-hydrolyzable nitrogen (AN) decreased. Moreover, the soil extracellular enzyme activities related to C and P metabolism decreased while the enzymes related to N metabolism increased, and the expression of soil genes involved in C, N, and P cycling was regulated. Moreover, it was observed that tolerant microorganisms (e.g., p_Proteobacteria, p_Actinobacteria, g_Lysobacter, g_Sphingobium, f_Aspergillaceae, and g_Penicillium) were enriched but sensitive taxa (p_Myxococcota) were suppressed in the citrus rhizosphere. The citrus biomass was mainly positively correlated with soil AN and AP; plant N and P were mainly positively correlated with soil AP, AN, and acid phosphatase (ACP); and plant K was mainly negatively related with soil ß-glucosidase (ßG) and positively related with the soil fungal Shannon index. The dominant bacterial taxa p_Actinobacteriota presented positively correlated with the plant biomass and plant N, P, and K and was negatively correlated with plant Cu. The dominant fungal taxa p_Ascomycota was positively related to plant Cu but negatively with the plant biomass and plant N, P, and K. Notably, arbuscular mycorrhizal fungi (p_Glomeromycota) were positively related with plant P below soil Cu 108 mg kg-1, and pathogenic fungi (p_Mortierellomycota) was negatively correlated with plant K above soil Cu 178 mg kg-1. These findings provided a new perspective on soil microbes and chemistry properties and the healthy development of the citrus industry at increasing soil Cu levels.

Drug-Target Interaction Prediction Based on an Interactive Inference Network.

Drug-target interactions underlie the actions of chemical substances in medicine. Moreover, drug repurposing can expand use profiles while reducing costs and development time by exploiting potential multi-functional pharmacological properties based upon additional target interactions. Nonetheless, drug repurposing relies on the accurate identification and validation of drug-target interactions (DTIs). In this study, a novel drug-target interaction prediction model was developed. The model, based on an interactive inference network, contains embedding, encoding, interaction, feature extraction, and output layers. In addition, this study used Morgan and PubChem molecular fingerprints as additional information for drug encoding. The interaction layer in our model simulates the drug-target interaction process, which assists in understanding the interaction by representing the interaction space. Our method achieves high levels of predictive performance, as well as interpretability of drug-target interactions. Additionally, we predicted and validated 22 Alzheimer's disease-related targets, suggesting our model is robust and effective and thus may be beneficial for drug repurposing.

Combined analysis of the microbiome and metabolome to reveal the characteristics of saliva from different diets: a comparison among vegans, seafood-based omnivores, and red meat (beef and lamb) omnivores.

Introduction: Revealing individual characteristics is supportive for identifying individuals in forensic crime. As saliva is one of the most common biological samples used in crime scenes, it is important to make full use of the rich individual information contained in saliva. The aim of this study was to explore the application of the microbiome in forensic science by analysing differences in the salivary microbiome and metabolome of healthy individuals with different dietary habits. Methods: We performed 16S rDNA sequencing analysis based on oral saliva samples collected from 12 vegetarians, 12 seafood omnivores and 12 beef and lamb omnivores. Non-targeted metabolomics analyses were also performed based on saliva samples from healthy individuals. Results: The results showed that the dominant flora of vegetarians was dominated by Neisseria (belonging to the phylum Proteobacteria), while seafood omnivores and beef and lamb omnivores were dominated by Streptococcus (belonging to the phylum Firmicutes). NDMS-based and cluster analyses showed that vegetarian dieters were significantly differentiated from meat dieters (seafood omnivores and beef and lamb omnivores), which may be related to the fact that high-fiber diets can create a different salivary flora structure. Variants were also detected in salivary metabolic pathways, including positive correlations with Lipid metabolism, Amino acid metabolism, Carbohydrate metabolism, and Nucleotide metabolism in vegetarians, and correlations in seafood omnivores. In order to select salivary microorganisms and metabolic markers that can distinguish different dietary profiles, a random forest classifier model was constructed in this study, and the results showed that individuals with different dietary profiles could be successfully distinguished based on the core genera and metabolites such as Streptococcus, Histidinyl-Valine. Conclusion: Our study provides a supportive basis for the application of salivary polyomics in order to reveal the dietary characteristics of individuals for forensic investigation and crime solving.

Sex differences in the association between metabolic score for insulin resistance and the reversion to normoglycemia in adults with prediabetes: a cohort study.

BACKGROUND: The metabolic score for insulin resistance (MetS-IR) has become a valid indicator to evaluate insulin resistance. Our investigation sought gender differences in the correlation between MetS-IR and the reversion from prediabetes to normoglycemic status. METHODS: This retrospective research, carried out in 32 areas across 11 cities with several centers in China, encompassed 15,423 participants with prediabetes. We employed a Cox proportional hazards regression model to examine the link between MetS-IR and the reversion to normoglycemic status. We also applied cubic spline functions and smooth curve fitting to detect non-linear relationships. Additionally, we embarked on a range of sensitivity analyses. RESULTS: The study included 15,423 participants, with 10,009 males (64.90%) and 5,414 females (35.10%). The average follow-up time was 2.96 ± 0.93 years, and 6,623 individuals (42.94%) reversed normoglycemia. A non-linear correlation was discovered among MetS-IR and reversion to normoglycemic status in men, with a turning point at 55.48. For a one-unit rise in MetS-IR below this point, the chance of reversal to normoglycemic levels declined by 3% (HR = 0.97, 95% CI:0.96-0.97, P < 0.0001). In women, the association was linear, with every unit rise in MetS-IR leading to a 3% reduction in transitioning to normal glycemic levels. (HR = 0.97, 95% CI: 0.97-0.98, p < 0.0001). CONCLUSION: A negative correlation was discovered between MetS-IR and reversion to normoglycemic status in adults with prediabetes. Specifically, a non-linear association was observed for males, while females exhibited a linear correlation.

Skin locations inference and body fluid identification from skin microbial patterns for forensic applications.

Given that microbiological analysis can be an alternative method that overcomes the shortcomings of traditional forensic technology, and skin samples may be the most common source of cases, the analysis of skin microbiome was investigated in this study. High-throughput sequencing targeting the V3-V4 region of 16S rRNA gene was performed to reveal the skin microbiome of healthy individuals in Guangdong Han. The bacterial diversity of the palm, navel, groin and plantar of the same individual was analyzed. The overall classification based on 16S rRNA gene amplicons revealed that the microbial composition of skin samples from different anatomical parts was different, and the dominant bacterial genus of the navel, plantar, groin and palm skin were dominated by Cutibacterium, Staphylococcus, Corynebacterium and Staphylococcus, respectively. PCoA analysis showed that the skin at these four anatomical locations could only be grouped into three clusters. A predictive model based on random forest algorithm showed the potential to accurately distinguish these four anatomical locations, which indicated that specific bacteria with low abundance were the key taxa. In addition, the skin microbiome in this study is significantly different from the dominant microbiome in saliva and vaginal secretions identified in our previous study, and can be distinguished from these two tissue fluids. In conclusion, the present findings on the community and microbial structure details of the human skin may reveal its potential application value in assessing the location of skin samples and the type of body fluids in forensic medicine.

Elevations in presepsin, PCT, hs-CRP, and IL-6 levels predict mortality among septic patients in the ICU.

This study aimed to investigate whether changes in presepsin, procalcitonin, high-sensitivity C-reactive protein, and interleukin 6 levels predict mortality in septic patients in the intensive care unit. This study enrolled septic patients between November 2020 and December 2021. Levels of presepsin, procalcitonin, high-sensitivity C-reactive protein, and interleukin 6 were measured on the first (PSEP_0, PCT_0, hsCRP_0, IL-6_0) and third days (PSEP_3, PCT_3, hsCRP_3, IL-6_3). Follow-up was performed on days 3, 7, 14, 21, and 28 after enrollment. The outcome was all-cause death. The study included 119 participants, and the mortality was 18.5%. In univariable Cox proportional hazards regression analysis, ΔPSEP (= PSEP_3 - PSEP_0) > 211.49 pg/mL (hazard ratio, 2.70; 95% confidence interval, 1.17-6.22), ΔPCT (= PCT_3 - PCT_0) > -0.13 ng/mL (hazard ratio, 7.31; 95% confidence interval, 2.68-19.80), ΔhsCRP (= hsCRP_3 - hsCRP_0) > -19.29 mg/L (hazard ratio, 6.89; 95% confidence interval, 1.61-29.40), and ΔIL-6 (= IL-6_3 - IL-6_0) > 1.00 pg/mL (hazard ratio, 3.13; 95% confidence interval, 1.35-7.24) indicated an increased risk of mortality. The composite concordance index for alterations in all 4 distinct biomarkers was highest (concordance index, 0.83; 95% confidence interval, 0.76-0.91), suggesting the optimal performance of this panel in mortality prediction. In decision curve analysis, compared with the Acute Physiology and Chronic Health Evaluation II and Sequential (sepsis-related) Organ Failure Assessment scores, the combination of the 4 biomarkers had a larger net benefit. Interestingly, interleukin 6 was predominantly produced by monocytes upon lipopolysaccharide stimulation in peripheral blood mononuclear cells. ΔPSEP, ΔPCT, ΔhsCRP, and ΔIL-6 are reliable biomarkers for predicting mortality in septic patients in the intensive care unit, and their combination has the best performance.

Survey of Pharmaceutical Industry's Best Practices around In Vitro Transporter Assessment and Implications for Drug Development: Considerations from the International Consortium for Innovation and Quality for Pharmaceutical Development Transporter Working Group.

The International Consortium for Innovation and Quality in Pharmaceutical Development Transporter Working Group had a rare opportunity to analyze a crosspharma collation of in vitro data and assay methods for the evaluation of drug transporter substrate and inhibitor potential. Experiments were generally performed in accordance with regulatory guidelines. Discrepancies, such as not considering the impact of preincubation for inhibition and free or measured in vitro drug concentrations, may be due to the retrospective nature of the dataset and analysis. Lipophilicity was a frequent indicator of crosstransport inhibition (P-gp, BCRP, OATP1B, and OCT1), with high molecular weight (MW ≥500 Da) also common for OATP1B and BCRP inhibitors. A high level of overlap in in vitro inhibition across transporters was identified for BCRP, OATP1B1, and MATE1, suggesting that prediction of DDIs for these transporters will be common. In contrast, inhibition of OAT1 did not coincide with inhibition of any other transporter. Neutrals, bases, and compounds with intermediate-high lipophilicity tended to be P-gp and/or BCRP substrates, whereas compounds with MW <500 Da tended to be OAT3 substrates. Interestingly, the majority of in vitro inhibitors were not reported to be followed up with a clinical study by the submitting company, whereas those compounds identified as substrates generally were. Approaches to metabolite testing were generally found to be similar to parent testing, with metabolites generally being equally or less potent than parent compounds. However, examples where metabolites inhibited transporters in vitro were identified, supporting the regulatory requirement for in vitro testing of metabolites to enable integrated clinical DDI risk assessment. SIGNIFICANCE STATEMENT: A diverse dataset showed that transporter inhibition often correlated with lipophilicity and molecular weight (>500 Da). Overlapping transporter inhibition was identified, particularly that inhibition of BCRP, OATP1B1, and MATE1 was frequent if the compound inhibited other transporters. In contrast, inhibition of OAT1 did not correlate with the other drug transporters tested.

Food-Borne Biotoxin Neutralization in Vivo by Nanobodies: Current Status and Prospects.

Food-borne biotoxins from microbes, plants, or animals contaminate unclean, spoiled, and rotten foods, posing significant health risks. Neutralizing such toxins is vital for human health, especially after food poisoning. Nanobodies (Nbs), a type of single-domain antibodies derived from the genetic cloning of a variable domain of heavy chain antibodies (VHHs) in camels, offer unique advantages in toxin neutralization. Their small size, high stability, and precise binding enable effective neutralization. The use of Nbs in neutralizing food-borne biotoxins offers numerous benefits, and their genetic malleability allows tailored optimization for diverse toxins. As nanotechnology continues to evolve and improve, Nbs are poised to become increasingly efficient and safer tools for toxin neutralization, playing a pivotal role in safeguarding human health and environmental safety. This review not only highlights the efficacy of these agents in neutralizing toxins but also proposes innovative solutions to address their current challenges. It lays a solid foundation for their further development in this crucial field and propels their commercial application, thereby contributing significantly to advancements in this domain.

A novel AP/PA total body irradiation technique using abutting IMRT fields at extended SSD.

PURPOSE: To develop a Total Body Irradiation (TBI) technique using IMRT at extended SSD that can be performed in any size Linac room. METHODS: Patients studied were placed on a platform close to the floor, directly under the gantry with cranial-caudal axis parallel to the gantry rotation plane and at SSD ∼200 cm. Two abutting fields with the same external isocenter at gantry angles of ±21˚, collimator angle of 90˚, and field size of 25 × 40 cm2 are employed for both supine and prone positions. An iterative optimization algorithm was developed to generate a uniform dose at the patient mid-plane with adequate shielding to critical organs such as lungs and kidneys. The technique was validated in both phantom and patient CT images for treatment planning, and dose measurement and QA were performed in phantom. RESULTS: A uniform dose distribution in the mid-plane within ±5% of the prescription dose was reached after a few iterations. This was confirmed with ion-chamber measurements in phantom. The mean dose to lungs and kidneys can be adjusted according to clinical requirements and can be as low as ∼25% of the prescription dose. For a typical prescription dose of 200 cGy/fraction, the total MU was ∼2400/1200 for the superior/inferior field. The overall treatment time for both supine/prone positions was ∼54 min to meet the maximum absorbed dose rate criteria of 15 cGy/min. IMRT QA with portal dosimetry shows excellent agreement. CONCLUSIONS: We have developed a promising TBI technique using abutting IMRT fields at extended SSD. The patient is in a comfortable recumbent position with good reproducibility and less motion during treatment. An additional benefit of this technique is that full 3D dose distribution is available from the TPS with a DVH summary for organs of interest. The technique allows precise sparing of lungs and kidneys and can be executed in any linac room.

CYP7A1 Gene Induction via SHP-Dependent or Independent Mechanisms can Increase the Risk of Drug-Induced Liver Injury Independently or in Synergy with BSEP Inhibition.

Drug-induced liver injury (DILI) is a frequent cause of clinical trial failures during drug development. While inhibiting bile salt export pump (BSEP) is a well-documented DILI mechanism, interference with genes related to bile acid (BA) metabolism and transport can further complicate DILI development. Here, the effects of twenty-eight compounds on genes associated with BA metabolism and transport were evaluated, including those with discontinued development or use, boxed warnings, and clean labels for DILI. The study also included rifampicin and omeprazole, pregnane X receptor and aryl hydrocarbon receptor ligands, and four mitogen-activated protein kinase kinase (MEK1/2) inhibitors. BSEP inhibitors with more severe DILI, notably pazopanib and CP-724714, significantly upregulated the expression of 7 alpha-hydroxylase (CYP7A1), independent of small heterodimer partner (SHP) expression. CYP7A1 expression was marginally induced by omeprazole. In contrast, its expression was suppressed by mometasone (10-fold), vinblastine (18-fold), hexachlorophene (2-fold), bosentan (2.1-fold), and rifampin (2-fold). All four MEK1/2 inhibitors that show clinical DILI were not potent BSEP inhibitors but significantly induced CYP7A1 expression, accompanied by a significant SHP gene suppression. Sulfotransferase 2A1 and BSEP were marginally upregulated, but no other genes were altered by the drugs tested. Protein levels of CYP7A1 were increased with the treatment of CYP7A1 inducers and decreased with obeticholic acid, an farnesoid X receptor ligand. CYP7A1 inducers significantly increased bile acid (BA) production in hepatocytes, indicating the overall regulatory effects of BA metabolism. This study demonstrates that CYP7A1 induction via various mechanisms can pose a risk for DILI, independently or in synergy with BSEP inhibition, and it should be evaluated early in drug discovery. SIGNIFICANCE STATEMENT: Kinase inhibitors, pazopanib and CP-724714, inhibit BSEP and induce CYP7A1 expression independent of small heterodimer partner (SHP) expression, leading to increased bile acid (BA) production and demonstrating clinically elevated drug-induced liver toxicity. MEK1/2 inhibitors that show BSEP-independent drug-induced liver injury (DILI) induced the CYP7A1 gene accompanied by SHP suppression. CYP7A1 induction via SHP-dependent or independent mechanisms can pose a risk for DILI, independently or in synergy with BSEP inhibition. Monitoring BA production in hepatocytes can reliably detect the total effects of BA-related gene regulation for de-risking.

Triterpenoids from the hook-bearing stems of Uncaria rhynchophylla.

An unprescribed nortriterpenoid with an aromatic E ring, uncanortriterpenoid A (1), together with fourteen known triterpenoids (2-15), were isolated from the hook-bearing stems of Uncaria rhynchophylla Miq. Based on extensive spectroscopic analyses, the NMR data of 2, 5, and 10 in CD3OD were assigned for the first time, and the wrongly assigned δC of C-27 and C-29 of 2 were revised. Among the known compounds, 7, 13, and 15 were isolated from this species for the first time, and 15 represents the first lanostane triterpenoid bearing an extra methylidene at C-24 for the Rubiaceae family. Additionally, compounds 6 and 14 exhibited moderate ferroptosis inhibitory activity, with an EC50 value of 14.74 ± 0.20 µM for 6 and 23.11 ± 1.31 µM for 14.

A self-supervised learning approach for registration agnostic imaging models with 3D brain CTA.

Deep learning-based neuroimaging pipelines for acute stroke typically rely on image registration, which not only increases computation but also introduces a point of failure. In this paper, we propose a general-purpose contrastive self-supervised learning method that converts a convolutional deep neural network designed for registered images to work on a different input domain, i.e., with unregistered images. This is accomplished by using a self-supervised strategy that does not rely on labels, where the original model acts as a teacher and a new network as a student. Large vessel occlusion (LVO) detection experiments using computed tomographic angiography (CTA) data from 402 CTA patients show the student model achieving competitive LVO detection performance (area under the receiver operating characteristic curve [AUC] = 0.88 vs. AUC = 0.81) compared to the teacher model, even with unregistered images. The student model trained directly on unregistered images using standard supervised learning achieves an AUC = 0.63, highlighting the proposed method's efficacy in adapting models to different pipelines and domains.

Building a Predictive PBPK Model for Human OATP Substrates: a Strategic Framework for Early Evaluation of Clinical Pharmacokinetic Variations Using Pitavastatin as an Example.

To select a drug candidate for clinical development, accurately and promptly predicting human pharmacokinetic (PK) profiles, assessing drug-drug interactions (DDIs), and anticipating potential PK variations in disease populations are crucial steps in drug discovery. The complexity of predicting human PK significantly increases when hepatic transporters are involved in drug clearance (CL) and volume of distribution (Vss). A strategic framework is developed here, utilizing pitavastatin as an example. The framework includes the construction of a monkey physiologically-based PK (PBPK) model, model calibration to obtain scaling factors (SF) of in vitro-in vivo extrapolation (IVIVE) for various clearance parameters, human model development and validation, and assessment of DDIs and PK variations in disease populations. Through incorporating in vitro human parameters and calibrated SFs from the monkey model of 3.45, 0.14, and 1.17 for CLint,active, CLint,passive, and CLint,bile, respectively, and together with the relative fraction transported by individual transporters obtained from in vitro studies and the optimized Ki values for OATP inhibition, the model reasonably captured observed pitavastatin PK profiles, DDIs and PK variations in human subjects carrying genetic polymorphisms, i.e., AUC within 20%. Lastly, when applying the functional reduction based on measured OATP1B biomarkers, the model adequately predicted PK changes in the hepatic impairment population. The present study presents a strategic framework for early-stage drug development, enabling the prediction of PK profiles and assessment of PK variations in scenarios like DDIs, genetic polymorphism, and hepatic impairment-related disease states, specifically focusing on OATP substrates.

Response of salivary microbiome to temporal, environmental, and surface characteristics under in vitro exposure.

The microbiome of saliva stains deposited at crime scenes and in everyday settings is valuable for forensic investigations and environmental ecology. However, the dynamics and applications of microbial communities in these saliva stains have not been fully explored. In this study, we analyzed saliva samples that were exposed to indoor conditions for up to 1 year and to different carriers (cotton, sterile absorbent cotton swab, woolen, dacron) in both indoor and outdoor environments for 1 month using high-throughput sequencing. The analysis of microbial composition and Mfuzz clustering showed that the salivary flora, specifically Streptococcus (cluster7), which was associated with microbial contamination, remained stable over short periods of time. However, prolonged exposure led to significant differences due to the invasion of environmental bacteria such as Pseudomonas and Achromobacter. The growth and colonization of environmental flora were promoted by humidity. The neutral model predictions indicated that the assembly of salivary microbial communities in outdoor environments was significantly influenced by stochastic processes, with environmental characteristics having a greater impact on community change compared to surface characteristics. By incorporating data from previous studies on fecal and vaginal secretion microbiology, we developed RF and XGBoost classification models that achieved high accuracy (>98 %) and AUC (>0.8). Additionally, a RF regression model was created to determine the time since deposition (TsD) of the stains. Time inference models yielded a mean absolute error (MAE) of 7.1 days for stains exposed for 1 year and 14.2 h for stains exposed for 14 days. These findings enhance our understanding of the changes in the microbiome of saliva stains over time, in different environments, and on different surfaces. They also have potential applications in assessing potential microbial contamination, identifying body fluids, and inferring the time of deposition.

Molecular characterization of the grass carp bscl2 gene and its expression response to lipid accumulation, nutritional status, insulin and glucagon.

Bscl2 plays a role in lipid metabolism of mammals, however its role in teleost fish remains unclear. Using the grass carp (Ctenopharyngodon idella) as a model, the bscl2 gene was isolated from the brain and characterized. Thereafter, the tissue distribution of the gene was examined, before expression was analyzed as a function of fasting, refeeding, oral glucose administration and overfeeding. In addition, bscl2 mRNA levels were evaluated in grass carp primary hepatocytes treated with glucagon, insulin, oleic acid, and glucose. Results showed that the cloned bscl2 gene was 1341 bp, encoding 446 amino acids, and was highly expressed in the brain, heart, and gonad. Following oral glucose administration, bscl2 expression increased. Expression of bscl2 decreased in fasted fish but increased following refeeding. Overfeeding, which resulted in elevated lipid accumulation, also stimulated bscl2 expression. In primary hepatocytes, bscl2 levels were increased by glucose, oleic acid, and insulin treatments, and reduced by glucagon treatment. These data suggest that bscl2 may play an important role in nutrient metabolism in teleost fish.

Cloning grass carp (Ctenopharyngodon idella) ccdc3 and its expression affected by nutrition state, insulin, and glucagon.

As an adipokine, coiled-coil domain-containing 3 (CCDC3) plays multiple physiological roles in fatty liver, lipid metabolism, and abdominal obesity. Grass carp was selected as the experimental animal in this study to investigate the roles of Ccdc3 in teleosts. Results showed that the open reading frame (ORF) of cloned ccdc3 was 831 bp and encoded 276 amino acids. Three N-glycosylation sites and a predicted coiled-coil domain motif were located in the identified Ccdc3. Moreover, a nuclear localization signal (NLS) was contained in the coiled-coil domain motif of the identified Ccdc3. The results on tissue distribution revealed that ccdc3 was highly detected in grass carp fat and brain tissue. In the oral glucose tolerance test (OGTT), the expression of ccdc3 increased remarkably in the brain, hypothalamus, and visceral fat in the glucose treatment group. In the fasting and refeeding experiment, the ccdc3 expression levels were remarkably reduced in the brain, hypothalamus, and visceral fat after 14 days of fasting. In the refeeding group, the ccdc3 expression levels were considerably elevated compared with those in the fasting group. In the induced overfeeding experiment, the ccdc3 expression increased remarkably in the hepatopancreas, brain, and visceral fat tissues. The ccdc3 expression in the primary hepatocytes was remarkably increased with glucose, oleic acid, and insulin treatment. However, ccdc3 expression was markedly decreased with glucagon treatment. In conclusion, these results indicate that Ccdc3 is involved in regulating glucose and lipid metabolism of teleosts.

Toward improved predictions of pharmacokinetics of transported drugs in hepatic impairment: Insights from the extended clearance model.

Hepatic impairment (HI) moderately (<5-fold) affects the systemic exposure (i.e., area under the plasma concentration-time curve [AUC]) of drugs that are substrates of the hepatic sinusoidal organic anion transporting polypeptide (OATP) transporters and are excreted unchanged in the bile and/or urine. However, the effect of HI on their AUC is much greater (>10-fold) for drugs that are also substrates of cytochrome P450 (CYP) 3A enzymes. Using the extended clearance model, through simulations, we identified the ratio of sinusoidal efflux clearance (CL) over the sum of metabolic and biliary CLs as important in predicting the impact of HI on the AUC of dual OATP/CYP3A substrates. Because HI may reduce hepatic CYP3A-mediated CL to a greater extent than biliary efflux CL, the greater the contribution of the former versus the latter, the greater the impact of HI on drug AUC ratio (AUCRHI ). Using physiologically-based pharmacokinetic modeling and simulation, we predicted relatively well the AUCRHI of OATP substrates that are not significantly metabolized (pitavastatin, rosuvastatin, valsartan, and gadoxetic acid). However, there was a trend toward underprediction of the AUCRHI of the dual OATP/CYP3A4 substrates fimasartan and atorvastatin. These predictions improved when the sinusoidal efflux CL of these two drugs was increased in healthy volunteers (i.e., before incorporating the effect of HI), and by modifying the directionality of its modulation by HI (i.e., increase or decrease). To accurately predict the effect of HI on AUC of hepatobiliary cleared drugs it is important to accurately predict all hepatobiliary pathways, including sinusoidal efflux CL.

Lycorine promotes IDH1 acetylation to induce mitochondrial dynamics imbalance in colorectal cancer cells.

Isocitrate dehydrogenase (IDH) 1 and 2, as essential enzymes in energy metabolism, contribute to the survival and drug resistance of a variety of solid tumors, especially for colorectal cancer (CRC). However, the underlying molecular mechanism still remains unclear. In this study, IDH1 was identified as a crucial cellular target of a natural-derived anti-CRC small molecule lycorine, using the unbiased thermal proteome profiling (TPP) strategy. We found that lycorine directly targeted a unique C-terminal domain of IDH1, and disrupted IDH1 interaction with deacetylase sirtuin 1 (SIRT1), thereby significantly promoting IDH1 acetylation modification. Then, lycorine noticeably triggered oxidative stress in CRC cells to cause mitochondrial membranes injury, and subsequently facilitated mitochondrial fission. Specific knockdown of IDH1 or SIRT1 markedly aggrieved lycorine-mediated oxidative stress and mitochondrial fragmentation in CRC cells. Furthermore, the combination of lycorine and sirtuins blocker nicotinamide (NAM) exhibited a synergic therapeutic effect in CRC cells. Collectively, our results reveal that IDH1 may serve as a promising therapeutic target for CRC via pharmacologically driving oxidative stress-dependent mitochondrial dynamics imbalance.

Interpretation of Protein-Mediated Uptake of Statins by Hepatocytes Is Confounded by the Residual Statin-Protein Complex.

Inclusion of plasma (or plasma proteins) in human hepatocyte uptake studies narrows, but does not close, the gap in in vitro to in vivo extrapolation (IVIVE) of organic anion transporting polypeptide (OATP)-mediated hepatic clearance (CLh) of statins. We have previously shown that this "apparent" protein-mediated uptake effect (PMUE) of statins by OATP1B1-expressing cells, in the presence of 5% human serum albumin (HSA), is mostly an artifact caused by residual statin-HSA complex remaining in the uptake assay. We determined if the same was true with plated human hepatocytes (PHH) and if this artifact can be reduced using suspended human hepatocytes (SHH) and the oil-spin method. We quantified the uptake of a cocktail of five statins by PHH and SHH in the absence and presence of 5% HSA. After terminating the uptake assay, the amount of residual HSA was quantified by quantitative targeted proteomics. For both PHH and SHH, except for atorvastatin and cerivastatin, the increase in total, active, and passive uptake of the statins, in the presence of 5% HSA, was explained by the estimated residual stain-HSA complex. In addition, the increase in active statin uptake by SHH, where present, was marginal (<50%), much smaller than that observed with PHH. Such a marginal increase cannot bridge the gap in IVIVE of CLh of statins. These data disprove the prevailing hypotheses for the in vitro PMUE. A true PMUE should be evaluated using the uptake data corrected for the residual drug-protein complex. SIGNIFICANCE STATEMENT: We show that the apparent protein-mediated uptake (PMUE) of statins by human hepatocytes is largely confounded by residual statin when plated or suspended human hepatocytes are used. Therefore, mechanisms other than PMUE need to be explored to explain the underprediction of the in vivo human hepatic clearance of statins by human hepatocyte uptake assays.

Changes in the microbial community after vaginal fluid exposure in different simulated forensic situations.

If vaginal fluid is found on clothing or on the body of the suspect, it may indicate the occurrence of sexual assault. Therefore, it is important to collect the victim's vaginal fluid at different sites from the suspect. Previous studies have revealed that fresh vaginal fluids can be identified based on 16S rRNA gene sequencing data. However, the influence of environmental factors on the stability of microbial markers must be investigated before being used in forensic practice. We collected vaginal fluid from nine unrelated individuals and placed each individual of vaginal swab on five different substrates. A total of 54 vaginal swabs were analyzed using 16S rRNA on the V3-V4 regions. Then, we constructed a random forest model including the samples of all vaginal fluids in this study and the other four types of body fluids in our previous studies. The alpha diversity of vaginal samples increased after exposure to the substrate environment for 30 days. The dominant vaginal bacteria were Lactobacillus and Gardnerella, which remained relatively stable after exposure, with Lactobacillus being the most abundant in all substrates, while Gardnerella was more abundant in other substrates than in the polyester fiber substrate. Except for bed sheets, Bifidobacterium significantly declined when placed on other substrates. Rhodococcus and Delftia from the substrate environment migrated to the vaginal samples. Rhodococcus was abundant in polyester fibers, and Delftia was abundant in wool substrates, while those environmental bacteria were all in low abundance in bed sheets. Overall, the bed sheet substrates showed a good retention capacity for the dominant flora and could reduce the number of taxa migrated by the environment compared with the other substrates. Both fresh and exposed vaginal samples of the same individuals could mostly be clustered and clearly distinguished from different individuals, showing the potential of individual identification, and the confusion matrix value of body fluid identification for vaginal samples was 1. In summary, vaginal samples placed on the surface of different substrates retained their stability and demonstrated good application potential for individual and body fluid identification.