Ferritin Heavy-like subunit is involved in the innate immune defense of the red swamp crayfish Procambarus clarkii.

Iron-binding proteins, known as ferritins, play pivotal roles in immunological response, detoxification, and iron storage. Despite their significance to organisms, little is known about how they affect the immunological system of the red swamp crayfish (Procambarus clarkii). In our previous research, one ferritin subunit was completely discovered as an H-like subunit (PcFeH) from P. clarkii. The full-length cDNA of PcFerH is 1779 bp, including a 5'-UTR (untranslated region, UTR) of 89 bp, 3'-UTR (untranslated region, UTR) of 1180 bp and an ORF (open reading frame, ORF) of 510 bp encoding a polypeptide of 169 amino acids that contains a signal peptide and a Ferritin domain. The deduced PcFerH protein sequence has highly identity with other crayfish. PcFerH protein's estimated tertiary structure is quite comparable to animal structure. The PcFerH is close to Cherax quadricarinatus, according to phylogenetic analysis. All the organs examined showed widespread expression of PcFerH mRNA, with the ovary exhibiting the highest levels of expression. Additionally, in crayfish muscles, intestines, and gills, the mRNA transcript of PcFerH was noticeably up-regulated, after LPS and Poly I:C challenge. The expression of downstream genes in the immunological signaling system was suppressed when the PcFerH gene was knocked down. All of these findings suggested that PcFerH played a vital role in regulating the expression of downstream effectors in the immunological signaling pathway of crayfish.

Immunotherapy for Pulmonary Arterial Hypertension: From the Pathogenesis to Clinical Management.

Pulmonary hypertension (PH) is a progressive cardiovascular disease, which may lead to severe cardiopulmonary dysfunction. As one of the main PH disease groups, pulmonary artery hypertension (PAH) is characterized by pulmonary vascular remodeling and right ventricular dysfunction. Increased pulmonary artery resistance consequently causes right heart failure, which is the major reason for morbidity and mortality in this disease. Although various treatment strategies have been available, the poor clinical prognosis of patients with PAH reminds us that further studies of the pathological mechanism of PAH are still needed. Inflammation has been elucidated as relevant to the initiation and progression of PAH, and plays a crucial and functional role in vascular remodeling. Many immune cells and cytokines have been demonstrated to be involved in the pulmonary vascular lesions in PAH patients, with the activation of downstream signaling pathways related to inflammation. Consistently, this influence has been found to correlate with the progression and clinical outcome of PAH, indicating that immunity and inflammation may have significant potential in PAH therapy. Therefore, we reviewed the pathogenesis of inflammation and immunity in PAH development, focusing on the potential targets and clinical application of anti-inflammatory and immunosuppressive therapy.

Discovery of Covalent MLKL PROTAC Degraders via Optimization of a Theophylline Derivative Ligand for Treating Necroptosis.

Mixed lineage kinase domain-like pseudokinase (MLKL) initiates necroptosis and could serve as a therapeutic target related to a series of human diseases. Proteolysis-targeting chimeras (PROTACs) are useful tools for degrading pathological proteins and blocking disease processes. Using computer-aided modeling and molecular dynamics simulations, we developed a series of covalent MLKL PROTACs by linking and optimizing a theophylline derivative that covalently targets MLKL. Via structure-activity relationship studies, MP-11 was identified as a potent MLKL PROTAC degrader. Furthermore, MP-11 showed lower toxicity than the original MLKL ligand, exhibiting nanomolar-scale antinecroptotic activity on human cell lines. Xenograft model studies showed that MP-11 effectively degraded MLKL in vivo. Importantly, our study demonstrates that the covalent binding strategy is an effective approach for designing MLKL-targeting PROTACs, serving as a model for developing PROTACs to treat future necroptosis-related human diseases.

Carnosic acid, a novel food-source AT1R antagonist and its anti-hypertension mechanism.

Hypertension is the most prevalent non-communicable disease, affecting billions of people worldwide. Discovery and development of natural antihypertensive lead compounds or drugs are important to resolve the limitations of existing antihypertensive drug safety and resistance. This investigation verified that carnosic acid (CA), an important active ingredient of rosemary, an edible spice plant, indicates a significant anti-hypertensive activity in spontaneous hypertension rats by targeting AT1R. Moreover, our research indicated that CA shared a comparable antagonistic mechanism with established synthetic angiotensin II receptor blockers (ARBs), as it occupies the binding sites of Angiotensin II (AngII) at His6 and Pro7 within the AT1R's ligand-binding pocket. Notably, CA exerted better anti-hypertensive activity since it could not break the Asn1113.35-Asn2957.46 hydrogen bond to stabilize the AT1R inactive state. As the first potent AT1R antagonist identified in a natural food source, CA is poised to become a novel anti-hypertensive lead compound, distinguished by its unique skeleton structure different from conventional ARBs. This research lays a valuable theoretical groundwork for the future exploration of CA and rosemary extract in both fundamental studies and clinical applications.

Clinical Characteristics and Diagnosis of Nonaccelerating MDS/MPN-U Patient with Granulocyte Dysplasia.

BACKGROUND: The goal was to improve the clinical cognition of nonaccelerating myelodysplastic/myeloproliferative neoplasms-unclassifiable (MDS/MPN-U) and avoid misdiagnosis or delayed diagnosis. METHODS: The clinical manifestations, laboratory indicators, histopathology, and therapeutic effects of a patient with nonaccelerating MDS/MPN-U were analyzed and the relevant literature was reviewed. RESULTS: Blood routine: white blood cell 98.48 x 109/L, red blood cell 3.20 x 1012/L, basophils 0.42 x 109/L, eosinophils 1.31 x 109/L, hemoglobin 112 g/L, and platelet 113 x 109/L. Blood smears showed granulocytosis and cells at various stages, polylobular granulocytes also can be seen. Bone marrow images show granulocytosis and dysplastic neutrophils, such as binuclear granulocyte, cyclic nuclear granulocyte, nuclear punch, cytoplasm vacuoles, polylobular granulocytes and so on. Bone marrow biopsy: Bone marrow proliferation tumor, combined with cell morphology and molecular biochemistry is recommended. Gene test showed Jak-2 positive, BCR/ABL and MPL negative. Chromosome examination indicated the presence of 46, XY, add (2)(p25), del (12) (p11.2p13)[16]/46, XY. CONCLUSIONS: MDS/MPN-U with granulocytosis and dysplastic neutrophils is rare, mostly in the elderly, and the diagnosis should be made except for other myeloid tumors. Currently, there is no uniform treatment guideline or expert consensus. The treatment options are limited and need to be further confirmed by more studies. MDS/ MPN-U with granulocytosis and dysplastic neutrophils has adverse prognostic factors such as advanced age, increase of bone marrow original cells and related gene mutations. Whether the adverse prognosis is related to specific gene mutations and cytogenetic variation remains to be clarified by more research data.

Integrin-α9 overexpression underlies the niche-independent maintenance of leukemia stem cells in acute myeloid leukemia.

Leukemia stem cells (LSCs) are widely believed to reside in well-characterized bone marrow (BM) niches; however, the capacity of the BM niches to accommodate LSCs is insufficient, and a significant proportion of LSCs are instead maintained in regions outside the BM. The molecular basis for this niche-independent behavior of LSCs remains elusive. Here, we show that integrin-α9 overexpression (ITGA9 OE) plays a pivotal role in the extramedullary maintenance of LSCs by molecularly mimicking the niche-interacting status, through the binding with its soluble ligand, osteopontin (OPN). Retroviral insertional mutagenesis conducted on leukemia-prone Runx-deficient mice identified Itga9 OE as a novel leukemogenic event. Itga9 OE activates Akt and p38MAPK signaling pathways. The elevated Myc expression subsequently enhances ribosomal biogenesis to overcome the cell integrity defect caused by the preexisting Runx alteration. The Itga9-Myc axis, originally discovered in mice, was further confirmed in multiple human acute myeloid leukemia (AML) subtypes, other than RUNX leukemias. In addition, ITGA9 was shown to be a functional LSC marker of the best prognostic value among 14 known LSC markers tested. Notably, the binding of ITGA9 with soluble OPN, a known negative regulator against HSC activation, induced LSC dormancy, while the disruption of ITGA9-soluble OPN interaction caused rapid cell propagation. These findings suggest that the ITGA9 OE increases both actively proliferating leukemia cells and dormant LSCs in a well-balanced manner, thereby maintaining LSCs. The ITGA9 OE would serve as a novel therapeutic target in AML.

The mechanism of interaction between tri-para-cresyl phosphate and human serum protein: A multispectroscopic and in-silico study.

Organophosphate flame retardants (OPFRs) pose the significant risks to the environment and human health and have become a serious public health issue. Tricresyl phosphates (TCPs), a group of aryl OPFRs, exhibit neurotoxicity and endocrine disrupting toxicity. However, the binding mechanisms between TCPs and human serum albumin (HSA) remain unknown. In this study, through fluorescence and ultraviolet-visible (UV-vis) absorption spectroscopy, molecular docking and molecular dynamics (MD), tri-para-cresyl phosphate (TpCP) was selected to explore potential interactions between HSA and TCPs. The results of the fluorescence spectroscopy demonstrated that a decrease in the fluorescence intensity of HSA and a blue shift were observed with the increasing concentrations of TpCP. The binding constant (Ka) was 2.575 × 104 L/mol, 4.701 × 104 L/mol, 5.684 × 104 L/mol and 9.482 × 104 L/mol at 293 K, 298 K, 303 K, and 310 K, respectively. The fluorescence process between HSA and TpCP involved a mix of static and dynamic quenching mechanism. The gibbs free energy (ΔG0) of HSA-TpCP system was -24.452 kJ/mol, -25.907 kJ/mol, -27.363 kJ/mol, and - 29.401 kJ/mol at 293 K, 298 K, 303 K, and 310 K, respectively, suggesting that the HSA-TpCP reaction was spontaneous. The enthalpy change (ΔH0) and thermodynamic entropy change (ΔS0) of the HSA-TpCP system were 60.83 kJ/mol and 291.08 J/(mol·>k), respectively, indicating that hydrophobic force was the major driving force in the HSA-TpCP complex. Furthermore, multispectral analysis also revealed that TpCP could alter the microenvironment of tryptophan residue and the secondary structure of HSA and bind with the active site I of HSA. Molecular docking and MD simulations confirmed that TpCP could spontaneously form a stable complex with HSA, which was consistent with the fluorescence experimental results. This study provides novel insights into the mechanisms of underlying the transportation and distribution of OPFRs in humans.

Species sensitivity distribution for nonylphenol: Acute toxicity and ecological risk in Bohai Region.

Nonylphenol (NP), a main byproduct of nonylphenol polyethoxylates (NPEs) degradation, is prevalent across diverse environmental settings. Given its widespread presence, evaluating the ecological risks associated with NP in coastal waters and sediments is essential for the protection of the marine environment. This study evaluates the acute toxicity of NP on ten representative aquatic species from the Bohai Sea, determining the Aquatic Life Criteria (ALC) through two distinct methods. The Criteria Maximum Concentration (CMC) for NP in seawater was established at 12.0 µg/L, with a Predicted No-Effect Concentration (PNEC) for water at 15.2 µg/L and for sediment at 33.3 µg/kg. Additionally, a tiered ecological risk assessment (ERA) of both surface seawater and sediment in the Bohai Sea revealed significant ecological risks at various sediment sites. These results offer crucial insights for assessing the ecological risks to coastal ecosystem and provide foundational data necessary for informed environmental protection and management strategies.

Insights into chlorantraniliprole exposure via activating cytochrome P450-mediated xenobiotic metabolism pathway in the Procambarus clarkii: Identification of P450 genes involved in detoxification.

To investigate the impact of chlorantraniliprole on Procambarus clarkii, acute toxicity tests were performed. Results indicated that 96 h post-exposure to chlorantraniliprole (60 mg/L) led to the separation of the hepatopancreas basement membrane, causing cell swelling, rupture, and vacuolation. Moreover, acid phosphatase (ACP) and alkaline phosphatase (AKP) activities exhibited divergent trends across four concentrations of chlorantraniliprole (0, 30, 60, and 90 mg/L). Hydrogen peroxide (H2O2) and catalase (CAT) levels significantly increased, while total superoxide dismutase (T-SOD) and malonaldehyde (MDA) activities decreased, indicating oxidative stress in the hepatopancreas. A total of 276 differentially expressed genes (DEGs) were identified, with 204 up-regulated and 72 down-regulated. Out of these, 114 DEGs were successfully annotated and classified into 99 pathways, with a primary focus on the cytochrome P450-mediated xenobiotic metabolism pathway. The DEGs enriched in this pathway, along with transcriptome data, were validated using quantitative-polymerase chain reaction. This study enhances the transcriptome database of P. clarkii and provides fundamental insights into its immune defense and antioxidant mechanisms. Additionally, it lays a theoretical foundation for future research on disease prevention in P. clarkii within rice-shrimp culture systems.

Mass spectrometry-based proteomic landscape of rice reveals a post-transcriptional regulatory role of N6-methyladenosine.

Rice is one of the most important staple food and model species in plant biology, yet its quantitative proteomes are largely uncharacterized. Here we quantify the relative protein levels of over 15,000 genes across major rice tissues using a tandem mass tag strategy followed by intensive fractionation and mass spectrometry. We identify tissue-specific and tissue-enriched proteins that are linked to the functional specificity of individual tissues. Proteogenomic comparison of rice and Arabidopsis reveals conserved proteome expression, which differs from mammals in that there is a strong separation of species rather than tissues. Notably, profiling of N6-methyladenosine (m6A) across the rice major tissues shows that m6A at untranslated regions is negatively correlated with protein abundance and contributes to the discordance between RNA and protein levels. We also demonstrate that our data are valuable for identifying novel genes required for regulating m6A methylation. Taken together, this study provides a paradigm for further research into rice proteogenome.

The effect of modified Qiyuan paste on mice with low immunity and sleep deprivation by regulating GABA nerve and immune system.

BACKGROUND: Low immunity and sleep disorders are prevalent suboptimal health conditions in contemporary populations, which render them susceptible to the infiltration of pathogenic factors. LJC, which has a long history in traditional Chinese medicine for nourishing the Yin and blood and calming the mind, is obtained by modifying Qiyuan paste. Dendrobium officinale Kimura et Migo has been shown to improve the immune function in sleep-deprived mice. In this study, based on the traditional Chinese medicine theory, LJC was prepared by adding D. officinale Kimura et Migo to Qiyuan paste decoction. METHODS: Indicators of Yin deficiency syndrome, such as back temperature and grip strength, were measured in each group of mice; furthermore, behavioral tests and pentobarbital sodium-induced sleep tests were performed. An automatic biochemical analyzer, enzyme-linked immunosorbent assay kit, and other methods were used to determine routine blood parameters, serum immunoglobulin (IgG, IgA, and IgM), cont (C3, C4), acid phosphatase (ACP) and lactate dehydrogenase (LDH) levels in the spleen, serum hemolysin, and delayed-type hypersensitivity (DTH) levels. In addition, serum levels of γ-aminobutyric acid (GABA) and glutamate (Glu) were detected using high-performance liquid chromatography (HPLC). Hematoxylin-eosin staining and Nissl staining were used to assess the histological alterations in the hypothalamus tissue. Western blot and immunohistochemistry were used to detect the expressions of the GABA pathway proteins GABRA1, GAD, GAT1, and GABAT1 and those of CD4+ and CD8+ proteins in the thymus and spleen tissues. RESULTS: The findings indicated that LJC prolonged the sleep duration, improved the pathological changes in the hippocampus, effectively upregulated the GABA content in the serum of mice, downregulated the Glu content and Glu/GABA ratio, enhanced the expressions of GABRA1, GAT1, and GAD, and decreased the expression of GABAT1 to assuage sleep disorders. Importantly, LJC alleviated the damage to the thymus and spleen tissues in the model mice and enhanced the activities of ACP and LDH in the spleen of the immunocompromised mice. Moreover, serum hemolysin levels and serum IgG, IgA, and IgM levels increased after LJC administration, which manifested as increased CD4+ content, decreased CD8+ content, and enhanced DTH response. In addition, LJC significantly increased the levels of complement C3 and C4, increased the number of white blood cells and lymphocytes, and decreased the percentage of neutrophils in the blood. CONCLUSIONS: LJC can lead to improvements in immunocompromised mice models with insufficient sleep. The underlying mechanism may involve regulation of the GABA/Glu content and the expression levels of GABA metabolism pathway-related proteins in the brain of mice, enhancing their specific and nonspecific immune functions.

The combination of allicin with domiphen is effective against microbial biofilm formation.

Background: Microorganisms in biofilms are particularly difficult to control because of their increased survival and antibiotic resistance. Allicin and domiphen were employed to inhibit the microbial growth and biofilm formation of Staphylococcus aureus, Escherichia coli, and Candida albicans strains. Methods: Broth microdilution method and checkerboard assay were conducted to determine the efficacy of allicin combined with domiphen against S. aureus, E. coli, and C. albicans. Microbial biofilm formation was measured using the crystal violet staining method and fluorescence microscopy. And the total viable count of the biofilm cells on material surface after the treatment with antimicrobial reagents was calculated with the plate count technique. Results: The two drugs showed synergistic effects against the pathogens with a fractional bactericidal concentration of less than 0.38. The combination of 64 µg/mL allicin with 1 µg/mL domiphen dispersed over 50% of the biofilm mass of S. aureus, E. coli, and C. albicans. In addition, the drug combination reduced the total viable counts of E. coli and C. albicans biofilm cells on stainless steel and polyethylene surfaces by more than 102 CFU/mL. Conclusion: The combination of allicin and domiphen is an effective strategy for efficiently decreasing biofilms formation on various industrial materials surfaces.

Genome-wide DNA methylation in relation to ARID1A deficiency in ovarian clear cell carcinoma.

BACKGROUND: The poor chemo-response and high DNA methylation of ovarian clear cell carcinoma (OCCC) have attracted extensive attentions. Recently, we revealed the mutational landscape of the human kinome and additional cancer-related genes and found deleterious mutations in ARID1A, a component of the SWI/SNF chromatin-remodeling complex, in 46% of OCCC patients. The present study aims to comprehensively investigate whether ARID1A loss and genome-wide DNA methylation are co-regulated in OCCC and identify putative therapeutic targets epigenetically regulated by ARID1A. METHODS: DNA methylation of ARID1Amt/ko and ARID1Awt OCCC tumors and cell lines were analyzed by Infinium MethylationEPIC BeadChip. The clustering of OCCC tumors in relation to clinical and mutational status of tumors were analyzed by hierarchical clustering analysis of genome-wide methylation. GEO expression profiles were used to identify differentially methylated (DM) genes and their expression level in ARID1Amt/ko vs ARID1Awt OCCCs. Combining three pre-ranked GSEAs, pathways and leading-edge genes epigenetically regulated by ARID1A were revealed. The leading-edge genes that passed the in-silico validation and showed consistent ARID1A-related methylation change in tumors and cell lines were regarded as candidate genes and finally verified by bisulfite sequencing and RT-qPCR. RESULTS: Hierarchical clustering analysis of genome-wide methylation showed two clusters of OCCC tumors. Tumor stage, ARID1A/PIK3CA mutations and TP53 mutations were significantly different between the two clusters. ARID1A mutations in OCCC did not cause global DNA methylation changes but were related to DM promoter or gene-body CpG islands of 2004 genes. Three pre-ranked GSEAs collectively revealed the significant enrichment of EZH2- and H3K27me3-related gene-sets by the ARID1A-related DM genes. 13 Leading-edge DM genes extracted from the enriched gene-sets passed the expression-based in-silico validation and showed consistent ARID1A-related methylation change in tumors and cell lines. Bisulfite sequencing and RT-qPCR analysis showed promoter hypermethylation and lower expression of IRX1, TMEM101 and TRIP6 in ARID1Amt compared to ARID1Awt OCCC cells, which was reversed by 5-aza-2'-deoxycytidine treatment. CONCLUSIONS: Our study shows that ARID1A loss is related to the differential methylation of a number of genes in OCCC. ARID1A-dependent DM genes have been identified as key genes of many cancer-related pathways that may provide new candidates for OCCC targeted treatment.

Breaking barriers: assessing the impact of clinical quality improvements on reducing health disparities in hypertension care among Mumbai's urban slums.

The clinical quality improvement initiatives, led by the organisation's Health Equity Working Group (HEWG), aim to support healthcare providers to provide equitable, quality hypertension care worldwide. After coordinating with the India team, we started monitoring the deidentified patient data collected through electronic health records between January and May 2021. After stratifying data by age, sex and residence location, the team found an average of 55.94% of our hypertensive patients control their blood pressure, with an inequity of 11.91% between male and female patients.The objective of this study was to assess the effectiveness of using clinical quality improvement to improve hypertension care in the limited-resourced, mobile healthcare setting in Mumbai slums. We used the model for improvement, developed by Associates in Process Improvement. After 9-month Plan-Do-Study-Act (PDSA) cycles, the average hypertensive patients with controlled blood pressure improved from 55.94% to 89.86% at the endpoint of the initiative. The gender gap reduced significantly from 11.91% to 2.19%. We continued to monitor the blood pressure and found that the average hypertensive patients with controlled blood pressure remained stable at 89.23% and the gender gap slightly increased to 3.14%. Hypertensive patients have 6.43 times higher chance of having controlled blood pressure compared with the preintervention after the 9-month intervention (p<0.001).This paper discusses the efforts to improve hypertension care and reduce health inequities in Mumbai's urban slums. We highlighted the methods used to identify and bridge health inequity gaps and the testing of PDSA cycles to improve care quality and reduce disparities. Our findings have shown that clinical quality improvement initiatives and the PDSA cycle can successfully improve health outcomes and decrease gender disparity in the limited-resource setting.

Assessment of Covid-19 vaccine confidence among healthcare personnel in the safety-net sector in the United States and Puerto Rico.

BACKGROUND: This study aimed to assess COVID-19 vaccine confidence among healthcare personnel in the safety net sector of the United States and Puerto Rico. This study aimed to examine the extent to which increased knowledge and positive attitudes toward COVID-19 vaccine safety and efficacy were associated with healthcare workers' COVID-19 vaccination status and their recommendation of the vaccine to all patients. METHODS: Online survey data were collected from health care workers working in Free and Charitable Clinics across the United States and Federally Qualified Health Centers in Puerto Rico. The survey consisted of 62 questions covering various demographic measures and constructs related to healthcare workers' vaccination status, beliefs, and recommendations for COVID-19 vaccination. Statistical analyses, including multivariate analysis, were conducted to identify the factors associated with the COVID-19 vaccine status and recommendations among healthcare personnel. RESULTS: Among the 2273 respondents, 93% reported being vaccinated against COVID-19. The analysis revealed that respondents who believed that COVID-19 vaccines were efficacious and safe were three times more likely to be vaccinated and twice as likely to recommend them to all their patients. Respondents who believed they had received adequate information about COVID-19 vaccination were 10 times more likely to be vaccinated and four times more likely to recommend it to all their patients. CONCLUSIONS: The study results indicate that healthcare workers' confidence in COVID-19 vaccines is closely tied to their level of knowledge, positive beliefs, and attitudes about vaccine safety and efficacy. The study emphasizes the significance of healthcare workers feeling well informed and confident in their knowledge to recommend the vaccine to their patients. These findings have important implications for the development of strategies to boost COVID-19 vaccine confidence among healthcare workers and increase vaccine uptake among patients.

CLK2 mediates IκBα-independent early termination of NF-κB activation by inducing cytoplasmic redistribution and degradation.

Activation of the NF-κB pathway is strictly regulated to prevent excessive inflammatory and immune responses. In a well-known negative feedback model, IκBα-dependent NF-κB termination is a delayed response pattern in the later stage of activation, and the mechanisms mediating the rapid termination of active NF-κB remain unclear. Here, we showed IκBα-independent rapid termination of nuclear NF-κB mediated by CLK2, which negatively regulated active NF-κB by phosphorylating the RelA/p65 subunit of NF-κB at Ser180 in the nucleus to limit its transcriptional activation through degradation and nuclear export. Depletion of CLK2 increased the production of inflammatory cytokines, reduced viral replication and increased the survival of the mice. Mechanistically, CLK2 phosphorylated RelA/p65 at Ser180 in the nucleus, leading to ubiquitin‒proteasome-mediated degradation and cytoplasmic redistribution. Importantly, a CLK2 inhibitor promoted cytokine production, reduced viral replication, and accelerated murine psoriasis. This study revealed an IκBα-independent mechanism of early-stage termination of NF-κB in which phosphorylated Ser180 RelA/p65 turned off posttranslational modifications associated with transcriptional activation, ultimately resulting in the degradation and nuclear export of RelA/p65 to inhibit excessive inflammatory activation. Our findings showed that the phosphorylation of RelA/p65 at Ser180 in the nucleus inhibits early-stage NF-κB activation, thereby mediating the negative regulation of NF-κB.

Efficient synthesis of L-malic acid by malic enzyme biocatalysis with CO2 fixation.

The malic enzyme (ME) catalyzes the synthesis of L-malic acid (L-MA) from pyruvic acid and CO2 with NADH as the reverse reaction of L-MA decarboxylation. Carboxylation requires excess pyruvic acid, limiting its application. In this study, it was determined that CO2 was the carboxyl donor by parsing the effects of HCO3- and CO2, which provided a basis for improving the L-MA yield. Moreover, the concentration ratio of pyruvic acid to NADH was reduced from 70:1 to 5:1 using CO2 to inhibit decarboxylation and to introduce the ME mutant A464S with a 2-fold lower Km than that of the wild type. Finally, carboxylation was coupled with NADH regeneration, resulting in a maximum L-MA yield of 77 % based on the initial concentration of pyruvic acid. Strategic modifications, including optimal reactant ratios and efficient mutant ME, significantly enhanced L-MA synthesis from CO2, providing a promising approach to the biotransformation process.

The ROCK inhibitor netarsudil in the treatment of corneal endothelial decompensation caused by corneal endotheliitis: A case report and literature review.

Proper hydration and the clarity of the cornea are maintained through the crucial function of the corneal endothelium. Inflammation of the corneal endothelium, known as endotheliitis, can disrupt endothelial function, resulting in alterations to vision. Corneal endotheliitis is characterised by corneal oedema, the presence of keratic precipitates, inflammation within the anterior chamber, and occasionally, limbal injection, neovascularisation, and the concurrent or overlapping presence of uveitis. The aetiology of this condition is diverse, predominantly viral, but it may also be drug-induced, result from bacterial or fungal infections, be associated with systemic diseases and procedures, or remain idiopathic with no identifiable cause. To date, no standardised protocol for the treatment of this ocular disease exists, and in severe cases, corneal transplantation may be required. A 31-year-old male was transferred to our hospital for the management of corneal endothelial decompensation resulting from corneal endotheliitis. Hormonal therapy and antiviral medications proved ineffective, rendering the patient a candidate for corneal transplantation. As a final measure, treatment with the ROCK inhibitor netarsudil was initiated. The patient demonstrated significant improvement in symptoms, and the inflammation was successfully managed after nine months. In this study, a novel approach employing ROCK inhibitor therapy was utilised for the treatment of corneal endotheliitis, leading to marked recovery during patient follow-up. This case report represents the inaugural application of the ROCK inhibitor netarsudil in managing corneal endothelial decompensation attributed to corneal endotheliitis. These findings suggest that this method warrants consideration as a potential novel treatment option for similar conditions.

Cyclodextrin-based ternary supramolecular deep eutectic solvents for efficient extraction and analysis of trace quinolones and sulfonamides in wastewater by adjusting pH.

BACKGROUND: Antibiotics residues can accelerate the growth of drug-resistant bacteria and harm the ecological environment. Under the effect of enrichment and biomagnification, the emergence of drug-resistant pathogenic bacteria may eventually lead to humans being ineffective to drugs in the face of bacterial or fungal disease infections in the future. It is urgent to develop an efficient separation medium and analytical method for simultaneous extraction and determination of antibiotics in the water environment. RESULTS: This work doped 2,6-Di-O-methyl-ß-cyclodextrin, randomly methyl-ß-cyclodextrin, 2-hydroxypropyl-ß-cyclodextrin with thymol:fatty acid respectively to construct non-covalent interaction-dominated pH-responsive ternary supramolecular deep eutectic solvents (SUPRADESs), which can undergo a hydrophilic/hydrophobic transition with aqueous phase to achieve an efficient microextraction. Semi-empirical method illustrated that SUPRADESs have a wide range of hydrogen bond receptor sites. We developed a SUPRADES-based analytical method combined with liquid chromatography-triple quadrupole mass spectrometry for the extraction and determination of trace quinolones and sulfonamides in wastewater. The overall limits of detection of the method were 0.0021-0.0334 ng mL-1 and the limits of quantification were 0.0073-0.1114 ng mL-1. The linearity maintained good in the spiked level of 0.01-100 ng mL-1 (R2 > 0.99). The overall enrichment factors of the method were 157-201 with lower standard deviations (≤8.7). SIGNIFICANCE: The method gave an extraction recovery of 70.1-115.3 % for 28 antibiotics in livestock farming wastewater samples from Zhejiang, China, at trace levels (minimum 0.5 ng mL-1). The results demonstrated that inducing the phase transition between SUPRADES and aqueous phase by adjusting pH for extraction is a novel and efficient pretreatment strategy. To our knowledge, this is the first application of cyclodextrin-based ternary SUPRADESs with pH-responsive reversible hydrophobicity-hydrophilicity transition behavior in wastewater analysis.