Detection of Short-Chain Chlorinated Aliphatic Hydrocarbons through an Engineered Biosensor with Tailored Ligand Specificity.

Short-chain chlorinated aliphatic hydrocarbons (SCAHs), commonly used as industrial reagents and solvents, pose a significant threat to ecosystems and human health as they infiltrate aquatic environments due to extensive usage and accidental spills. Whole-cell biosensors have emerged as cost-effective, rapid, and real-time analytical tools for environmental monitoring and remediation. While the broad ligand specificity of transcriptional factors (TFs) often prohibits the application of such biosensors. Herein, we exploited a semirational transition ligand approach in conjunction with a positive/negative fluorescence-activated cell sorting (FACS) strategy to develop a biosensor based on the TF AlkS, which is highly specific for SCAHs. Furthermore, through promoter-directed evolution, the performance of the biosensor was further enhanced. Mutation in the -10 region of constitutive promoter PalkS resulted in reduced AlkS leakage expression, while mutation in the -10 region of inducible promoter PalkB increased its accessibility to the AlkS-SCAHs complex. This led to an 89% reduction in background fluorescence leakage of the optimized biosensor, M2-463, further enhancing its response to SCAHs. The optimized biosensor was highly sensitive and exhibited a broader dynamic response range with a 150-fold increase in fluorescence output after 1 h of induction. The detection limit (LOD) reached 0.03 ppm, and the average recovery rate of SCAHs in actual water samples ranged from 95.87 to 101.20%. The accuracy and precision of the proposed biosensor were validated using gas chromatography-mass spectrometry (GC-MS), demonstrating the promising application for SCAH detection in an actual environment sample.

Evaluating the Effects of Perinatal Exposures to BPSIP on Hepatic Cholesterol Metabolism in Female and Male Offspring ICR Mice.

BACKGROUND: A broad suite of bisphenol S (BPS) derivatives as alternatives for BPS have been identified in various human biological samples, including 4-hydroxyphenyl 4-isopropoxyphenylsulfone (BPSIP) detected in human umbilical cord plasma and breast milk. However, very little is known about the health outcomes of prenatal BPS derivative exposure to offspring. OBJECTIVES: Our study aimed to investigate the response of hepatic cholesterol metabolism by sex in offspring of dams exposed to BPSIP. METHODS: Pregnant ICR mice were exposed to 5µg/kg body weight (BW)/day of BPSIP, BPS, or E2 through drinking water from gestational day one until the pups were weaned. The concentration of BPSIP, BPS, or E2 in the plasma and liver of pups was determined by liquid chromatography-tandem mass spectrometry. Metabolic phenotypes were recorded, and histopathology was examined for liver impairment. Transcriptome analysis was employed to characterize the distribution and expression patterns of differentially expressed genes across sexes. The metabolic regulation was validated by quantitative real-time PCR, immunohistochemistry, and immunoblotting. The role of estrogen receptors (ERs) in mediating sex-dependent effects was investigated using animal models and liver organoids. RESULTS: Pups of dams exposed to BPSIP showed a higher serum cholesterol level, and liver cholesterol levels were higher in females and lower in males than in the controls. BPSIP concentration in the male liver was 1.22±0.25 ng/g and 0.69±0.27 ng/g in the female liver. Histopathology analysis showed steatosis and lipid deposition in both male and female offspring. Transcriptome and gene expression analyses identified sex-specific differences in cholesterol biosynthesis, absorption, disposal, and efflux between pups of dams exposed to BPSIP and those in controls. In vivo, chromatin immunoprecipitation analysis revealed that the binding of ERα protein to key genes such as Hmgcr, Pcsk9, and Abcg5 was attenuated in BPSIP-exposed females compared to controls, while it was enhanced in males. In vitro, the liver organoid experiments demonstrated that restoration of differential expression induced by BPSIP in key genes, such as Hmgcr, Ldlr, and Cyp7a1, to levels comparable to the controls was only achieved when treated with a combination of ERα agonist and ERß agonist. DISCUSSION: Findings from this study suggest that perinatal exposure to BPSIP disrupted cholesterol metabolism in a sex-specific manner in a mouse model, in which ERα played a crucial role both in vivo and in vitro. Therefore, it is crucial to systematically evaluate BPS derivatives to protect maternal health during pregnancy and prevent the transmission of metabolic disorders across generations. https://doi.org/10.1289/EHP14643.

Bovine serum albumin-based biomimetic gene complexes with specificity facilitate rapid re-endothelialization for anti-restenosis.

Re-endothelialization is a critical problem to inhibit postoperative restenosis, and gene delivery exhibits great potential in rapid endothelialization. Unfortunately, the therapeutic effect is enormously limited by inefficient specificity, poor biocompatibility and in vivo stability owing largely to the complicated in vivo environment. Herein, we developed a series of platelet membrane (PM) cloaked gene complexes based on natural bovine serum albumin (BSA) and polyethyleneimine (PEI). The gene complexes aimed to accelerate re-endothelialization for anti-restenosis via pcDNA3.1-VEGF165 (VEGF) plasmid delivery. Based on BSA and PM coating, these gene complexes exhibited good biocompatibility, stability with serum and robust homing to endothelium-injured site inherited from platelets. Besides, they enhanced the expression of VEGF protein by their high internalization and nucleus accumulation efficiency, and also substantially promoted migration and proliferation of vascular endothelial cells. The biological properties were further optimized via altering PEI and PM content. Finally, rapid recovery of endothelium in a carotid artery injured mouse model (79% re-endothelialization compared with model group) was achieved through two weeks' treatment by the PM cloaked gene complexes. High level of expressed VEGF in vivo was also realized by the gene complexes. Moreover, neointimal hyperplasia (IH) was significantly inhibited by the gene complexes according to in vivo study. The results verified the great potential of the PM cloaked gene complexes in re-endothelialization for anti-restenosis. STATEMENT OF SIGNIFICANCE: Rapid re-endothelialization is a major challenge to inhibit postoperative restenosis. Herein, a series of biodegradable and biocompatible platelet membrane (PM) cloaked gene complexes were designed to accelerate re-endothelialization for anti-restenosis via pcDNA3.1-VEGF165 (VEGF) plasmid delivery. The PM cloaked gene complexes provided high VEGF expression in vascular endothelial cells (VECs), rapid migration and proliferation of VECs and robust homing to endothelium-injured site. In a carotid artery injured mouse model, PM cloaked gene complexes significantly promoted VEGF expression in vivo, accelerated re-endothelialization and inhibited neointimal hyperplasia due to their good biocompatibility and superior specificity. Overall, the optimized PM cloaked gene complexes overcomes multiple obstacles in gene delivery for re-endothelialization and can be a promising candidate for gene delivery and therapy of postoperative restenosis.

5-Boronopicolinic acid-functionalized polymeric nanoparticles for targeting drug delivery and enhanced tumor therapy.

Strong specificity for cancer cells is still the main challenge to deliver drugs for the therapy of cancer. Herein, we developed a convenient strategy to prepare a series of 5-boronopicolinic acid (BA) modified tumor-targeting drug delivery systems (T-DDSs) with strong tumor targeting function. An anti-tumor drug of camptothecin (CPT) was encapsulated into poly(lactide-co-glycolide)-g-polyethylenimine (PLGA-PEI) to form drug-loaded nanoparticles (NP/CPT). Then, the surface of NP/CPT was coated by BA with different polymer and BA molar ratios of 1:1, 1:5, 1:10 and 1:20 via electrostatic interaction to obtain T-DDSs with enhanced biocompatibility and specificity for tumor cells. The introduced BA can endow drug-loaded NPs with high targeting ability to tumor cells because of the overexpression of sialic acids (SA) in tumor cells, which possessed strong interaction with BA. Those T-DDSs exhibited good biocompatibility according to the results of MTT assay, hemolysis test and cellular uptake. Moreover, they were capable of decreasing the viability of breast cancer cell line 4T1 and MCF-7 cells with no obvious cytotoxicity for endothelial cells. Especially, T-DDS with 1:20 molar ratio displayed much higher cellular uptake than other groups, and also exhibited highly efficient in vivo anti-tumor effect. The significantly high targeting function and biocompatibility of T-DDSs improved their drug delivery efficiency and achieved good anti-tumor effect. The BA decorated T-DDSs provides a simple and robust strategy for the design and preparation of DDSs with good biocompatibility and strong tumor-specificity to promote drug delivery efficiency.

Delivery of benzoylaconitine using biodegradable nanoparticles to suppress inflammation via regulating NF-κB signaling.

Rheumatoid arthritis (RA) is a kind of systemic autoimmune disease, and patients with RA usually suffer serious pain, resulting in low quality of life. The development of drug delivery systems (DDSs) provides a valid approach for RA therapy via inhibiting the secretion of inflammatory cytokines from macrophages. As a prevailing drug nanocarrier with distinctive superiority, polymeric nanoparticles (NPs) have attracted much attention in recent years. However, low biocompatibility and limited exploitation of drug with high efficiency are still the main challenges in RA treatment. To overcome the limitations, we prepared a biocompatible copolymer methoxy-poly(ethylene glycol)-poly(lactide-co-glycolide) (mPEG-PLGA). Moreover, benzoylaconitine (BAC) with superior anti-inflammatory effect was selected as model drug. It was isolated from Aconitum kusnezoffii Reichb and encapsulated into mPEG-PLGA NPs (NP/BAC) to increase the bioavailablity of BAC. The NPs exhibited high cytocompatibility for activated macrophages and well compatibility with red blood cells. Furthermore, the anti-inflammatory property of NP/BAC was testified by substantially inhibiting secretion of pro-inflammatory cytokines. The TNF-α and IL-1ß cytokines of NP/BAC group reduced 70 % and 66 % compared with that of activated macrophages. Especially, NP/BAC reduced the overexpression of NF-κB p65 to inhibit NF-κB signaling pathway, which was a critical regulator of inflammatory responses. NP/BAC also showed efficient in vivo anti-inflammatory effect with high ear (69.8 %) and paw (87.1 %) swelling suppressing rate. These results revealed the anti-inflammatory mechanism of NP/BAC and proved it was a suitable DDS to suppress inflammation, providing a promising strategy for RA therapy and research of Aconitum kusnezoffii Reichb.

pH-Zone-refining counter-current chromatography for two new lipo-alkaloids separated from refined alkaline extraction of Kusnezoff monkshood root.

An efficient and refined method for the separation of six aconitine-type alkaloids from the alkaline prepared "Kusnezoff monkshood root" was established. It is the first study that two new lipo-alkaloids were successfully isolated from refined sample by pH-zone-refining counter-current chromatography rather than synthetic method. It was of interest that a great deal of lipo-alkaloids was produced in crude extract from the alkalization of "Kusnezoff monkshood root." A refined sample method was proposed to enrich two types of alkaloids by liquid-liquid extraction, i.e. lipo-alkaloids and monoester-diterpenoid alkaloids. The pH-zone-refining counter-current chromatography was performed with an optimized two-phase solvent system composed of n-hexane-ethyl acetate-methanol-water (3:5:4:5, v/v), where upper organic phase was added to 3 mmol/L triethylamine as a retainer and lower aqueous mobile phase was added to 3 mmol/L hydrochloric acid as an eluter. As a result, six aconitum alkaloids, including two lipo-alkaloids (8-lino-14-benzoylaconine, 8-pal-14-benzoylaconine), three monoester-diterpenoid alkaloids (14-benzoylmesaconine, 14-benzoylaconine, beyzoyldeoxyaconine), and one aconine alkaloid (neoline) were acquired from the plant at the same time. The anti-inflammatory activities of the two new lipo-alkaloids were compared to the six alkaloids in vitro, in cyclo-oxygen-ase-2 inhibition assays. The separation mechanism of six alkaloids by pH-zone-refining counter-current chromatography was illustrated.

Biological Visual Detection for Advanced Photocatalytic Oxidation toward Pesticide Detoxification.

Photocatalytic oxidation treatment is an emerging and fast developed eco-friendly, energy-saving, and efficient advanced oxidation technology for degrading hazardous pesticides. The conventional chemical detection to evaluate the effects for this process depends on the broken chemical structure, only giving residual content and product chemical composition. However, it misses direct visual detection on the toxicity and the quantitative analysis of pesticide detoxification. Here, we develop a novel strategy to combine photocatalytic oxidation with a zebrafish biological model to provide a direct visual detection on the environmental detoxification. The mortality or deformity of zebrafish embryos (ZEs) acts as an indicator. Over the irradiation duration threshold, the mortality of ZEs decreases to 23.3% for pure chlorothalonil (CTL-P) after photocatalytic oxidation treatment for 1 h, and the deformity reduces to 13.3% for commercial CTL (CTL-C) after 30 min and to 3.33% for tetramethylthiuram disulfide (TMTD) after 20 min. The toxicity of CTL-C and TMTD could be completely removed by photocatalytic oxidation treatment and causes no damage to the ZE developmental morphology. Chemical analyses demonstrate the degradation of CTL into inorganic compounds and TMTD into small organic molecules. Among these highlighted heterogeneous photocatalysts (g-C3N4, BiVO4, Ag3PO4, and P25), g-C3N4 exhibits the highest photocatalytic detoxification for CTL-P, CTL-C, and TMTD.

Adaptive evolution of Aspergillus oryzae 3.042 strain and process optimization to reduce the formation of tyrosine crystals in broad bean paste.

Tyrosine crystals occasionally appeared on the broad bean paste surface, which will cause economic losses. This study aimed to eliminate the tyrosine crystals in broad bean paste through decreasing the activities of proteolytic enzymes produced by Aspergillus oryzae and process optimization. Broad bean pastes containing no more than 6.16 mg/g dry material tyrosine showed low possibility to form tyrosine crystals. Using tyrosine as substrate, the A. oryzae 3.042 was adaptively evolved and the tyrosine content in the broad bean paste fermented by the evolved A. oryzae was reduced from 6.49 mg/g dry material to 6.14 mg/g dry material (p < 0.05). When the production process was optimized, the tyrosine content in broad bean paste was further reduced to 5.67 mg/g dry material (p < 0.05). In this condition, no tyrosine crystals were formed in broad bean paste after the 12-month storage while the product quality was not influenced. PRACTICAL APPLICATIONS: Tyrosine crystals were one of the most important factors which negatively influence the quality of traditionally fermented food, including broad bean paste, soybean paste, and sausages. The appearance of tyrosine crystals in these foods will cause economic losses to manufacturers. This study tried to eliminate the appearance of tyrosine crystals through decreasing the activities of proteolytic enzymes produced by Aspergillus oryzae 3.042 and fermentation process optimization. The adoption of modified A. oryzae and optimized fermentation process successfully guaranteed the elimination of tyrosine crystals formation in the production and storage period of broad bean paste. This will not only benefit the broad bean paste manufacturers but also provide guidance for other fermented food producers to deal with tyrosine crystals problem.

A non-target urinary and serum metabolomics strategy reveals therapeutical mechanism of Radix Astragali on adjuvant-induced arthritis rats.

Radix Astragali has been used traditionally in China to treat rheumatoid arthritis (RA) in formulas. In this paper, we conducted a holistic evaluation of Radix Astragali acted on adjuvant-induced arthritis (AIA) rats by urinary and serum metabolomic studies. Histological results and hind paw swelling were used to assess the joint damage, while the levels of IL-1ß, TNF-α, SOD and MDA in serum were used to assess inflammation injury and oxidative stress. Metabolomic study and multivariate statistical analyses were used to investigate the differences between different groups. After processing with multivariate statistical analysis, 13 and 21 potential biomarkers were respectively found in urine and serum when Radix Astragali treatment group compared with model group. The main metabolism pathways in which Radix Astragali affected on AIA rats were tryptophan metabolism, phenylalanine metabolism, citrate cycle metabolism, fatty acid metabolism, vitamin B6 metabolism and so on. The present study demonstrates that urinary and serum metabolomics method could be a potentially powerful tool to understand the holistic therapeutic effect and the mechanisms of herb medicines.

A study on the effective substance of the Wu-tou formula based on the metabonomic method using UPLC-Q-TOF-HDMS.

The Wu-tou formula (WTF) is a Chinese medicine formula which has been applied to treat rheumatic arthritis (RA) and pain of joints for more than a thousand years. In this study, a pharmacodynamics combined urinary metabonomic study using UPLC-Q-TOF-HDMS was performed to assess the holistic efficacy of the Traditional Chinese Medicine (TCM) Wu-tou formula for treating RA in rats. Eighty male Sprague-Dawley rats were randomly divided into eight groups, named as the healthy control group (HG), the model group (AIA), the WTF group and five single herb groups. The treatment groups and the model group were induced for treating rheumatoid arthritis by using complete Freund's adjuvant. Histological results assessed the joint damage and several biochemical parameters such as IL-1ß, TNF-α, SOD and MDA were used to evaluate inflammation injury and oxidative stress. Based on the results, a metabonomic investigation was conducted to study the mechanism of the WTF and single herb treatment groups for treating RA. Multivariate statistical analyses such as PCA and OPLS-DA were used to identify potential biomarkers in urine. As a result, twenty-six potential biomarkers have been found by comparison with the model and the WTF treatment group. The potential biomarkers mainly affect the phenylalanine, tyrosine and tryptophan biosynthesis pathway and the taurine and hypotaurine metabolism pathway. Aconiti Radix Preparata and Ephedrae Herba showed better effects on treating RA from the integrated evaluation by histological results, biochemical parameters and pattern recognition analysis. A comprehensive evaluation of the different therapeutic effects and the mechanism of each herb in the WTF for treating RA was performed in this research.