Extracellular Processes Research Articles

P-523 rs2277698 Single nucleotides polymorphisms in tissue inhibitors of metalloproteinases 2 (TIMP2) affected clinical outcomes of women undergoing in vitro fertilization

Abstract Study question Is there any association between clinical outcomes of women undergoing in vitro fertilization (IVF) and the polymorphic variants of matrix metalloproteinases (MMPs) or tissue inhibitors of metalloproteinases (TIMPs)? Summary answer Our findings suggested that clinical outcomes of women undergoing IVF were associated with rs2277698 SNPs in TIMP2. What is known already The extracellular matrix proteolytic machinery is one of the most important mechanisms for regulating trophoblast invasion. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), which are involved in the extracellular matrix proteolysis process, have the ability to regulate the degradation of the extracellular matrix and play a critical role in trophoblast invasion. Study design, size, duration A prospective study was composed of 1014 women undergoing their first fresh IVF cycle with non-donor from January 2014 to December 2015. Participants/materials, setting, methods The present study was carried out in Lee Women’s Hospital, Taiwan. DNA was extracted from the peripheral blood of all participants, and the SNPs were genotyped by real-time polymerase chain reaction. The effects of the following three single nucleotide polymorphisms (SNPs) on IVF outcomes were explored: TIMP1 (rs4898 C/T), TIMP2 (rs2277698 C/T) and MMP2 (rs243865 C/T). The SNP genotype, correlation with clinical pregnancy, embryo implantation, abortion and live birth rates of IVF were analyzed. Main results and the role of chance In the analysis of 1014 patients attempting their first cycle of IVF, the genotypes frequencies of the study population showed a statistically significant association between TIMP2 (rs2277698 C/T) and clinical outcome. For TIMP2 polymorphisms, women with wild genotype (CC) hadhigher clinical pregnancy rate (34.8% v.s 28.0%; p = 0.032), embryo implantation rate (19.4% v.s 15.4%; p = 0.009), live birth rate (29.8% v.s 22.1%; p = 0.006), and lower abortion rate (7.0% v.s 17.1%; p = 0.005) compared women with CT/TT genotype. On the other hand, the clinical outcomes in different TIMP1 or MMP2 gene polymorphisms groups were no statistically significant difference. Limitations, reasons for caution The size number of the study samples. Wider implications of the findings This study showed that the TIMP2 SNP polymorphisms CT/TT was associated with low trend of clinical outcomes. The mechanism of TIMP2 gene T allele affected the outcomes of IVF remains to be determined. Further studies should focus on the mechanism of these associations in a larger, more heterogeneous cohort. Trial registration number CS13194

Weighted Gene Co-Expression Network Analysis Identifies Key Modules and Central Genes Associated With Bovine Subcutaneous Adipose Tissue.

BackgroundFat deposition is an important economic trait in livestock and poultry production. However, the relationship between various genes and signal pathways of fat deposition is still unclear to a large extent. The purpose of this study is to analyze the potential molecular targets and related molecular pathways in bovine subcutaneous adipose tissue.ResultsWe downloaded the GSE116775 microarray dataset from Gene Expression Omnibus (GEO). The weighted gene co-expression network (WGCNA) was used to analyze the gene expression profile, and the key gene modules with the highest correlation with subcutaneous adipose tissue were identified, and the functional enrichment of the key modules was analyzed. Then, the “real” Hub gene was screened by in-module analysis and protein–protein interaction network (PPI), and its expression level in tissue samples and adipocytes was verified. The study showed that a total of nine co-expression modules were identified, and the number of genes in these modules ranged from 101 to 1,509. Among them, the blue module is most closely related to subcutaneous adipose tissue, containing 1,387 genes. These genes were significantly enriched in 10 gene ontologies including extracellular matrix organization, biological adhesion, and collagen metabolic process, and were mainly involved in pathways including ECM-receptor interaction, focal adhesion, cAMP signaling pathway, PI3K-AKT signaling pathway, and regulation of lipolysis in adipocytes. In the PPI network and coexpression network, five genes (CAV1, ITGA5, COL5A1, ABL1, and HSPG2) were identified as “real” Hub genes. Analysis of Hub gene expression by dataset revealed that the expression of these Hub genes was significantly higher in subcutaneous adipose tissue than in other tissues. In addition, real-time fluorescence quantitative PCR (qRT-PCR) analysis based on tissue samples and adipocytes also confirmed the above results.ConclusionIn this study, five key genes related to subcutaneous adipose tissue were discovered, which laid a foundation for further study of the molecular regulation mechanism of subcutaneous adipose tissue development and adipose deposition.

Abstract 1089: Targeting amino acid uptake in lung squamous cell carcinomas to improve response to targeted therapies

Abstract Lung squamous cell carcinomas (LUSC) are a subtype of non-small cell lung cancer characterized by poor overall survival. LUSC are highly aggressive in part due to their high metabolic activity, with uptake of high levels of glucose supporting their growth and proliferation. The main driver of high rate of glycolysis in LUSC is elevated expression of glucose transporter GLUT1, whose levels and membrane localization are regulated by activated PI3K-AKT-mTOR pathway. The Cancer Genome Atlas revealed that LUSC have high tumor mutational burden with varied genetic profile that includes amplification of receptor tyrosine kinases (RTKs), PI3KCA, and a loss of PTEN. These alterations lead to increased activity of mTOR pathway, one of the major pathways in cancer cells that drives cell proliferation and metabolism. We have recently demonstrated that catalytic mTOR kinase inhibitor TAK228 (MLN128) inhibits glycolysis in LUSC cell lines in vitro, in xenografts of human LUSC lines, in patient derived xenografts (PDXs) and genetically engineered mouse models. However, despite lowered glucose uptake measured by 18F-FDG signal LUSC tumors were able to maintain high levels of proliferation. Tumors were able to adapt to prolonged treatment with TAK228 by relying on glutaminolysis. Increased glutaminolysis was due to increased uptake of glutamine as well as increased levels of glutaminase (GLS), a key enzyme that converts glutamine to glutamate. Inhibiting both mTOR (with TAK228) and GLS (with CB-839) resulted in reduced proliferation index and lower tumor volume. However, while combination therapy halted tumor growth, there was a lack of tumor regression in xenografts and PDXs of LUSC. Therefore, tumors are poised to further adapt to combination therapy through metabolism of additional amino acids beyond glutamine. Here we show that macropinocytosis, a form of endocytosis that allows cancer cells to uptake extracellular fluids and further process solutes and macromolecules in lysosomes leading to the release of amino acids inside the cytoplasm, is upregulated in LUSC tumors that are resistant to TAK228. Macropinocytosis is induced by active RTKs, PI3KCA, Ras, loss of PTEN, and active NRF2 pathway - oncogenic alterations commonly found in LUSC tumors. This suggests that macropinocytosis might be a mechanism that supports amino acid metabolism utilized by LUSC in order to escape mTOR therapy. Basal levels of macropinocytosis in LUSC appear low, however, upon treatment with mTOR inhibitors, lysosomal catabolism of macropinocytosed proteins increases, allowing tumor cells to increase their proliferation. Resistance to targeted therapies is a major hurdle to achieving effective therapies in multiple cancer types. Identifying ways to overcome resistance to targeted therapies would allow for longer therapeutic response for larger number of patients, ultimately leading to longer survival and better quality of life. Citation Format: Milica Momcilovic, David Shackelford. Targeting amino acid uptake in lung squamous cell carcinomas to improve response to targeted therapies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1089.

MnO2@MXene/Carbon Cloth as an Anode for Microbial Fuel Cells

AbstractHydrophobicity of carbon‐based anodes hinders bacterial adhesion and biofilm formation. Herein, MnO2@MXene coated carbon cloth (CC) was designed as an anode for microbial fuel cells (MFCs). The anode integrated the hydrophilicity and conductivity of MXene with the biocompatibility of MnO2. This unique structure promoted bacterial colonisation and biofilm formation on the anode surface. MnO2@MXene enhanced electricity generation performance and wastewater degradation efficiency owing to the synergistic effect of MXene and MnO2. The MFC achieved a short startup time and an effective extracellular electron transfer process. The MnO2@MXene/CC anode ensured a higher power density and efficient decolourisation in MFCs. The MFC device achieved a high maximum power density of 746.3 mW/m2 and a Congo red decolourisation efficiency of 87.6 % at 48 h. This work offers a potential strategy for the effective degradation of wastewater and recovery of electrical energy.

Function of s100 Protein in Coronary Atherosclerosis

Atherosclerosis is a complex disease whose pathogenesis includes endothelial activation, accumulation of lipids in the subendothelium, formation of foam cells, fat bands and formation of atherosclerotic plaque. These complex mechanisms involve different cell populations in the intimate sub-endothelium, and the S-100 protein family plays a role in a number of extracellular and intracellular processes during the development of atherosclerotic lesions. The aim of this study was to determine the phenotypic characteristics of smooth muscle cells and the consequent expression of S100 protein in atherosclerotic altered coronary arteries in advanced stages of atherosclerosis. 19 samples of right atherosclerotic coronary arteries in stages of fibro atheroma (type V lesion) and complicated lesions (type VI lesion) have been analyzed. According to the standard protocol, the following primary antibodies have been used in the immunohistochemical analysis: a-smooth muscle actin (α-SMA), vimentin and S-100 protein. All analyzed samples have been in advanced stages of atherosclerosis, fibro atheroma (stage V lesions) and complicated lesions (type VI lesions). Most of them have had the structure of a complicated lesion with atheroma or fibro atheroma as a basis, subsequently complicated by disruption (subtype VI a), hemorrhage (subtype VI b) or thrombosis (subtype VI c), as well as by the presence of several complications on the same sample. Marked hypocellularity is present in the subendothelium of plaques. Cell population at plaque margins is characterized by immunoreactivity to α-SMA, vimentin, and S100 protein. Some of these cells accumulate lipids and look like foam cells. In the cell population at the margins of the plaques, smooth muscle cells of the synthetic phenotype are present, some of which accumulate lipids and demonstrate S100 immunoreactivity. Summarizing numerous literature data and our results, we could assume that smooth muscle cells, due to their synthetic and proliferative activity in the earlier stages of pathogenesis, as well as the consequent expression of S100 protein, could accumulate lipids in the earlier stages of atherosclerosis which, in advanced stages analyzed in this study, result in immunoreactivity of foam cells of smooth muscle origin to S100 protein.

Corrosion behaviors of 2205 duplex stainless steel in biotic and abiotic NaCl solutions

Biofilms on metal surfaces are important in microbiologically influenced corrosion processes. In this study, the corrosion behaviors of 2205 duplex stainless steel (DSS) in 3.5% biotic and abiotic NaCl solutions were explored with scanning Kelvin probe force microscope (SKPFM), electrochemical techniques, and X-ray photoelectron spectroscope (XPS). The results of SKPFM showed that the development process of sulfate-reducing bacteria (SRB) biofilms on 2205DSS was a slow and dynamic process and SRB preferentially attached to the ferrite phase. In the bacterial medium, the pittings of 2205DSS were narrower and deeper and the occlusion degree of pittings was aggravated. XPS and electrochemical measurement results showed that 2205DSS passivation film was sulfureted and that the corrosion current density was increased significantly by SRB metabolism. The voltaic potential measurement results indicated that the biofilm increased the potential difference on the steel surface. In the logarithmic and stable growth phases of SRB, the corrosion mechanism of 2205DSS was dominated by cathode depolarization. In the decline phase of SRB, starved SRB was more corrosive and the corrosion mechanism of 2205DSS was dominated by the extracellular electron transfer process, in which electrons were directly provided by ferrite.

Interplay between arsenic and selenium biomineralization in Shewanella sp. O23S

Bacteria play crucial roles in the biogeochemical cycle of arsenic (As) and selenium (Se) as these elements are metabolized via detoxification, energy generation (anaerobic respiration) and biosynthesis (e.g. selenocysteine) strategies. To date, arsenic and selenium biomineralization in bacteria were studied separately. In this study, the anaerobic metabolism of As and Se in Shewanella sp. O23S was investigated separately and mixed, with an emphasis put on the biomineralization products of this process. Multiple analytical techniques including ICP-MS, TEM-EDS, XRD, Micro-Raman, spectrophotometry and surface charge (zeta potential) were employed. Shewanella sp. O23S is capable of reducing selenate (SeO42−) and selenite (SeO32−) to red Se(-S)0, and arsenate (AsO43−) to arsenite (AsO33−). The release of H2S from cysteine led to the precipitation of AsS minerals: nanorod AsS and granular As2S3. When As and Se oxyanions were mixed, both As–S and Se(-S)0 biominerals were synthesized. All biominerals were extracellular, amorphous and presented a negative surface charge (−24 to −38 mV). Kinetic analysis indicated the following reduction yields: SeO32− (90%), AsO43− (60%), and SeO42− (<10%). The mix of SeO32− with AsO43− led to a decrease in As removal to 30%, while Se reduction yield was unaffected (88%). Interestingly, SeO42− incubated with AsO43− boosted the Se removal (71%). The exclusive extracellular formation of As and Se biominerals might indicate an extracellular respiratory process characteristic of various Shewanella species and strains. This is the first study documenting a complex interplay between As and Se oxyanions: selenite decreased arsenate reduction, whereas arsenate stimulated selenate reduction. Further investigation needs to clarify whether Shewanella sp. O23S employs multi-substrate respiratory enzymes or separate, high affinity enzymes for As and Se oxyanion respiration.

Denitrification strategy of Pantoea sp. MFG10 coupled with microbial dissimilatory manganese reduction: Deciphering the physiological response based on extracellular secretion

In this study, the manganese (Mn) reduction-coupled denitrification strategy of dissimilatory Mn reducing bacteria was insightfully investigated. Different parameters (MnO2 level, pH, and temperature) were optimized by kinetic fitting to improve denitrification and Mn reduction effects. The 300 mg L−1 MnO2 addition achieved 98.72% NO3–-N removal in 12 h, which was 54.62% higher than blank group without MnO2. Scale-up studies showed that the metabolic activity of the bacteria was effectively enhanced by the addition of MnO2. Besides the deepening of humification in the system, tryptophan-like protein and polysaccharide as potential electron donor precursors revealed remarkable contributions to the extracellular secretion-dependent denitrification process of DMRB. The effect of EPS on Mn reduction depends mainly on the capture of MnO2 by the LB-EPS layer versus its dissolution in the TB-EPS layer. Ultimately, the EPS possess a dual effect of accelerated denitrification and Mn reduction efficiency due to the enhanced EET process.

MO711: Evaluation of an in Vitro Co-Culture Model for Studying Modulation of Cross-Talk between Endothelial and Mesothelial Cells by Cytoprotective Additives in Peritoneal Dialysis Fluids

Abstract BACKGROUND AND AIMS Peritoneal dialysis (PD) is a life-saving renal replacement therapy. The composition of all currently available peritoneal dialysis fluids (PDF) triggers morphological and functional changes in the peritoneal membrane. Periodic exposure leads to vasculopathy, hypervascularization and diabetes-like damage of vessels, eventually leading to failure of the technique. In vitro and in vivo studies have shown that cytoprotective additives [e.g. dipeptide alanyl-glutamine (AlaGln) or kinase inhibitor lithium chloride (LiCl)] to PDF reduce peritoneal damage in mesothelial (MC) and endothelial (EC) cells. However, it is yet unclear if potential cross talk between cells present in the peritoneal membrane mediates the observed modulation of PD-associated changes. Currently, there is no model system available to study relevant processes in these cells when in close proximity. Here, we aimed to develop a co-culture model for investigating cell-to-cell communication in a PD-relevant setting using cellular proteome and secretome analyses and investigate the effects of cytoprotective additives. METHOD For modelling the peritoneal membrane in vitro, MCs and ECs were co-cultured in transwell plates. MCs were grown in the upper compartment and primary microvascular ECs were grown in the lower compartment. PDF with or without additives, was added to the upper compartment to expose MCs directly, while the ECs below were kept in medium. Cellular proteome and secretome profiles were analysed for both cell types cultured individually or together by a quantitative mass spectrometry approach. Prior to analysis of the secretome, a combinatorial peptide ligand library coupled to beads was employed to enrich low abundant proteins and deplete high abundant proteins. RESULTS Proteome analysis revealed perturbation of major cellular processes including cytoskeleton reorganization, stress response, and regulation of cell death that characterize PDF cytotoxicity. Co-cultured cells yielded differently regulated pathways following PDF exposure compared to individual cultures. Several pathways relevant for PD, such as VEGF signaling in ECs or oxidative stress response in MCs were found only under co-culture conditions. Combined analysis of proteome and secretome showed specific ligand–receptor pairs expressed only under co-culture conditions regulating some of these pathways. Differentially regulated cellular and secreted proteins after PDF exposure were related to processes like angiogenesis, immune response, or and extracellular processes such as integrin signaling. Addition of 10 mM LiCl to PDF led to modulation of VEGF and oxidative stress response pathways in ECs as well as ligand–receptor pairs involved in these pathways, possibly explaining MC–EC crosstalk during PDF exposure of these cells and the cytoprotective effect of LiCl. CONCLUSION Harmful effects of PDF on MCs may also affect ECs, as demonstrated by our data. Interestingly, both cells type react differently when co-cultured compared to individual culture models, showing the importance of models that allow cell type interaction to mimic the in vivo conditions. We identified potential signaling axes between the cell types that could explain pathophysiological changes of the peritoneal membrane during PD treatment. We have also elucidated potential mechanisms by which the cytoprotective additive LiCl may modulate crosstalk between MCs and ECs to counteract adverse PDF effects in the local peritoneal environment. Linking local peritoneal damage with systemic vasculopathy through perturbations of PD-induced cellular crosstalk, may identify therapeutic targets to reduce the cardiovascular risk of PD patients and current limitations of the therapy.

Extracellular matrix metalloproteinase-9 (MMP-9) is required in female mice for 17β-estradiol enhancement of hippocampal memory consolidation

Hippocampal plasticity and memory are modulated by the potent estrogen 17β-estradiol (E2). Research on the molecular mechanisms of hippocampal E2 signaling has uncovered multiple intracellular pathways that contribute to these effects, but few have questioned the role that extracellular signaling processes may play in E2 action. Modification of the extracellular matrix (ECM) by proteases like matrix metalloproteinase-9 (MMP-9) is critical for activity-dependent remodeling of synapses, and MMP-9 activity is required for hippocampal learning and memory. Yet little is known about the extent to which E2 regulates MMP-9 in the hippocampus, and the influence this interaction may have on hippocampal memory. Here, we examined the effects of hippocampal MMP-9 activity on E2-induced enhancement of spatial and object recognition memory consolidation. Post-training bilateral infusion of an MMP-9 inhibitor into the dorsal hippocampus of ovariectomized female mice blocked the enhancing effects of E2 on object placement and object recognition memory, supporting a role for MMP-9 in estrogenic regulation of memory consolidation. E2 also rapidly increased the activity of dorsal hippocampal MMP-9 without influencing its protein expression, providing further insight into hippocampal E2/MMP-9 interactions. Together, these results provide the first evidence that E2 regulates MMP-9 to modulate hippocampal memory and highlight the need to further study estrogenic regulation of extracellular modification.

Effects of ivacaftor on systemic inflammation and the plasma proteome in people with CF and G551D

BackgroundIvacaftor is a cystic fibrosis transmembrane conductance regulator (CFTR) potentiator for people with CF and the G551D mutation. We aimed to investigate the biology of CFTR modulation and systemic effects of CFTR restoration by examining changes in circulating measurements of inflammation and growth and novel proteins with ivacaftor treatment. MethodsBlood samples from 64 CF subjects with G551D-CFTR were analyzed for inflammatory and growth-related proteins at baseline, 1 and 6 months after ivacaftor initiation. In 30 subjects, plasma was assayed for 1,322 proteins using the SomaScan proteomic platform at baseline and 6 months post-ivacaftor. Correlations with clinical outcomes were assessed. Measurements and Main ResultsSignificant reductions in high mobility group box-1 protein (HMGB-1), calprotectin, serum amyloid A, and granulocyte colony-stimulating factor (G-CSF), and an increase in insulin-like growth factor (IGF-1) occurred 1 month after ivacaftor. This treatment effect was sustained at 6 months for HMGB-1 and calprotectin. Correcting for multiple comparisons in the proteomic analysis, 9 proteins (albumin, afamin, leptin, trypsin, pancreatic stone protein [PSP], pituitary adenylate cyclase-activating polypeptide-38, repulsive guidance molecule A [RGMA], calreticulin, GTPase KRas) changed significantly with ivacaftor. Proteins changing with treatment are involved in lipid digestion and transport and extracellular matrix organization biological processes. Reductions in calprotectin and G-CSF and increases in calreticulin, and RGMA correlated with improved lung function, while increasing IGF-1, leptin and afamin and decreasing PSP correlated with increased weight. ConclusionsIvacaftor led to changes in inflammatory, lipid digestion, and extracellular matrix proteins, lending insights into the extrapulmonary effects of CFTR modulation.

Construction of a new type of three-dimensional honeycomb-structure anode in microbial electrochemical systems for energy harvesting and pollutant removal

Electrode materials occupy most of the construction cost of the microbial electrochemical system (MES), and the low mechanical strength and poor electrochemical performance of the commonly used traditional carbon-based materials restrict the promotion and application of this technology. In this study, polymer-based three-dimensional (3D) honeycomb-structure (HS) materials with good mechanical properties were used as supporting materials. Graphene (GR), carbon nanotube (CNT), and polypyrrole (PPy) was separately chosen as a surface conductivity coating layer for preparing MES anodes. The introduction of GR, CNT, and PPy on HS increased surface roughness, hydrophilicity, O and N content, electrochemically active surface area, and decreased charge transfer internal resistance, which promoted the adhesion of microorganisms on their surface and enhanced the extracellular electron transfer process at the electrode/microbe interface. The CNT-HS anode system got the better maximal power density (1700.7 ± 149.0 mW/m2) of the three modified anode systems and 3.60 times that of MES using CC (471.8 ± 27.2 mW/m2) as the anode. The accelerated reactions of the redox species in the outer cell membrane, the promoted electron shuttle secretion, and the enhanced abundance of the tricarboxylic acid cycle-related functional genes in biofilm led to better performance of the CNT-HS anode system. The CNT-HS anode system also exhibited long-term operational stability (>6 months) and a good chemical oxygen demand degradation effect. Furthermore, CNT-HS material exhibited its cost advantage, and its projected cost is estimated to be about $1.8/m2, much lower than the currently used MES anodes ($8.2–548.2/m2). Considering the good mechanical properties, simple preparation process, low manufacturing cost, long-term stability, excellent bio-electrochemical performance, and good pollutant removal ability, HS-based anode has promising potential for high-performance MES in applications.

Fungal dye-decolorizing peroxidase diversity: roles in either intra- or extracellular processes

Fungal dye-decolorizing peroxidases (DyPs) have found applications in the treatment of dye-contaminated industrial wastes or to improve biomass digestibility. Their roles in fungal biology are uncertain, although it has been repeatedly suggested that they could participate in lignin degradation and/or modification. Using a comprehensive set of 162 fully sequenced fungal species, we defined seven distinct fungal DyP clades on basis of a sequence similarity network. Sequences from one of these clades clearly diverged from all others, having on average the lower isoelectric points and hydropathy indices, the highest number of N-glycosylation sites, and N-terminal sequence peptides for secretion. Putative proteins from this clade are absent from brown-rot and ectomycorrhizal species that have lost the capability of degrading lignin enzymatically. They are almost exclusively present in white-rot and other saprotrophic Basidiomycota that digest lignin enzymatically, thus lending support for a specific role of DyPs from this clade in biochemical lignin modification. Additional nearly full-length fungal DyP genes were isolated from the environment by sequence capture by hybridization; they all belonged to the clade of the presumably secreted DyPs and to another related clade. We suggest focusing our attention on the presumably intracellular DyPs from the other clades, which have not been characterized thus far and could represent enzyme proteins with novel catalytic properties.Key points• A fungal DyP phylogeny delineates seven main sequence clades.• Putative extracellular DyPs form a single clade of Basidiomycota sequences.• Extracellular DyPs are associated to white-rot fungi.

Removal effects and potential mechanisms of bisphenol A and 17α-ethynylestradiol by Biogenic Mn oxides generated by Bacillus sp. WH4.

Endocrine disrupting compounds (EDCs), such as bisphenol A (BPA) and 17α-ethynylestradiol (EE2), have increasingly negative effects on human and wildlife health. In this study, the biogenic Mn oxides (BMOs) generated by Bacillus sp. WH4 were characterized, and the removal effects and reaction kinetics of BPA and EE2 by BMOs under different pH values, initial organic concentrations, and dosages of BMOs were discussed. The results showed that the formation of BMOs was extracellular process, and Mn(II) was oxidized to Mn(III) and Mn(IV) with 23.56% and 76.44%, respectively. The degradation processes of BPA and EE2 by BMOs followed first-order reaction kinetics, and the removal effect decreased with increasing initial BPA/EE2 concentrations and increased with increasing dosages of BMOs. However, the removal effect of BPA by BMOs decreased and then increased with increasing pH, while the removal effect of EE2 by BMOs decreased with increasing pH. Under optimal conditions, the removal efficiency of BPA and EE2 exceeded 98.2% and 94.3%, respectively. Additionally, this study showed that BMOs degraded BPA by coupling, oxidative condensation, substitution, and elimination reactions to obtain sixteen intermediate products and EE2 by substitution and elimination reactions to obtain seven intermediate products.

Staphylococcal Protein A with Engineered Cysteine: Comparison of Monomeric Content as a Critical Quality Attribute during Intracellular and Extracellular Expression

Background: The introduction of engineered cysteine in staphylococcal protein A (SPA-cys) for site-specific conjugation results in a substantial amount of dimerized SPA due to spontaneous oxidation during its production, leading to inaccessibility and thus rendering it unusable. Monomers are usually recovered from dimers by using reducing agents before conjugation in subsequent steps. However, this leads to low conjugation efficiency and increases overall cost and production time. This study aims to systematically compare and quantify the monomeric and dimeric content of SPA when produced through intracellular and extracellular routes in E. coli. Methods: Purified SPAs with and without cysteine from both intracellular and extracellular processes are compared for their monomeric content and efficiency to conjugate on solid support matrix with and without an additional pre-step of reduction. Results: The monomeric form of SPA-cys, which is a desired key quality attribute, is less than 50% when produced extracellularly. SPA-cys produced through the intracellular production process has high monomeric content (≥85%) and shows higher binding to solid support. Conclusion: The study demonstrates that the intracellular route for production of SPA-cys should be the preferred method, and the release assays for SPA-cys products should include the amount of monomeric content as one of the quality attributes. The abundance of monomeric content enhances the site-specific conjugation efficiency and density of SPA on the resin matrix.

Retinal Transcriptomics Analysis Reveals the Underlying Mechanism of Disturbed Emmetropization Induced by Wavelength Defocus

Purpose Wavelength signals play a vital role in refractive development. This study aimed to explore the retinal transcriptome signature in these processes. Methods Guinea pigs were randomly divided into three groups exposed to white, blue, or green environmental light for eight weeks. Refraction and axial length were evaluated every 4 weeks, and the retinal transcriptome was profiled at 8 weeks. Results Compared with the white group, ocular refraction significantly decreased and ocular axial length significantly extended in the green group whereas these parameters showed opposite trends in the blue group. RNA-sequencing showed that, compared with the white group, 184 and 171 differentially expressed genes (DEGs) were found in the blue and green groups, respectively. Among these DEGs, only 31 overlapped. These two sets of DEGs were enriched in distinct biological processes and pathways. There were 268 DEGs between the blue and green groups, which were primarily enriched in the extracellular matrix, and metabolism, receptor activity, and ion binding processes. In addition, nine human genes, including ECEL1, CHRND, SHBG, PRSS56, OVOL1, RDH5, WNT7B, PEBP4, CA12, were identified to be related to myopia development and wavelength response, indicating the potential role of these genes in human wavelength-induced myopia. Conclusions In this study, we identified retinal targets and pathways involved in the response to wavelength signals in emmetropization.

A multiscale cell-based model of tumor growth for chemotherapy assessment and tumor-targeted therapy through a 3D computational approach.

ObjectivesComputational modeling of biological systems is a powerful tool to clarify diverse processes contributing to cancer. The aim is to clarify the complex biochemical and mechanical interactions between cells, the relevance of intracellular signaling pathways in tumor progression and related events to the cancer treatments, which are largely ignored in previous studies.Materials and MethodsA three‐dimensional multiscale cell‐based model is developed, covering multiple time and spatial scales, including intracellular, cellular, and extracellular processes. The model generates a realistic representation of the processes involved from an implementation of the signaling transduction network.ResultsConsidering a benign tumor development, results are in good agreement with the experimental ones, which identify three different phases in tumor growth. Simulating tumor vascular growth, results predict a highly vascularized tumor morphology in a lobulated form, a consequence of cells' motile behavior. A novel systematic study of chemotherapy intervention, in combination with targeted therapy, is presented to address the capability of the model to evaluate typical clinical protocols. The model also performs a dose comparison study in order to optimize treatment efficacy and surveys the effect of chemotherapy initiation delays and different regimens.ConclusionsResults not only provide detailed insights into tumor progression, but also support suggestions for clinical implementation. This is a major step toward the goal of predicting the effects of not only traditional chemotherapy but also tumor‐targeted therapies.

Simultaneous high-concentration pyridine removal and denitrification in an electricity assisted bio-photodegradation system

In this study, enhanced degradation of high-concentration pyridine and simultaneous denitrification were achieved in an Electricity assisted bio-photodegradation system (EBPS) with a photoanode zone, a biocathode zone and an aerobic zone. The recombination of photoelectron-hole pairs was significantly suppressed in EBPS, which could not only improve the pyridine oxidation in photoanode zone, but also accelerate the biodegradation process in biocathode zone. NH4+ produced from pyridine degradation could be effectively nitrified into NO2- and NO3- in aerobic zone, which was recirculated into biocathode zone to serve as electron acceptor for accelerating pyridine biodegradation. When influent pyridine concentration was as high as 1250 mg L-1, pyridine was completely degraded without adding any extra electron donors, and complete denitrification was achieved in biocathode zone. Microbial community analysis confirmed the enrichment of the pyridine biodegradation species (Rhodococcus, Hydrogenophaga, Truepera and Thermovirga), denitrification species (Thiobacillus, Thioalkalispira and Rhodococcus) and electroactive species (Thiobacillus, Thermovirga, Hydrogenophaga, Thioalkalispira and Rhodococcus). Furthermore, the mechanism of pyridine metabolism, denitrification and extracellular electron transfer processes in EBPS was investigated, utilizing Phylogenetic Investigation of Communities by Reconstruction of Unobserved States 2 (PICRUSt2). The results showed that the genes involved in pyridine degradation, denitrification and extracellular electron transfer were remarkably enriched in EBPS, indicating the crucial role of photoelectrical stimulation. This study provided a sustainable technical approach for simultaneous removal of pyridine and nitrogen in wastewater.

Phosphoproteomic Analysis of Haemaphysalis longicornis Saliva Reveals the Influential Contributions of Phosphoproteins to Blood-Feeding Success.

Tick saliva, an essential chemical secretion of the tick salivary gland, is indispensable for tick survival owing to the physiological influence it exerts on the host defence mechanisms via the instrumentality of its cocktail of pharmacologically active molecules (proteins and peptides). Much research about tick salivary proteome has been performed, but how most of the individual salivary proteins are utilized by ticks to facilitate blood acquisition and pathogen transmission is not yet fully understood. In addition, the phosphorylation of some proteins plays a decisive role in their function. However, due to the low phosphorylation level of protein, especially for a small amount of protein, it is more difficult to study phosphorylation. Maybe, for this reason, the scarcity of works on the phosphorylated tick salivary proteomes still abound. Here, we performed a phosphoproteomic analysis of Haemaphysalis longicornis tick saliva via TiO2 enrichment and the most advanced Thermo Fisher Orbitrap Exploris 480 mass spectrometer for identification. A total of 262 phosphorylated tick saliva proteins were identified and were subjected to functional annotation/enrichment analysis. Cellular and metabolic process terms accounted for the largest proportion of the saliva proteins, with the participation of these proteins in vital intracellular and extracellular transport-oriented processes such as vesicle-mediated transport, exocytic process, cell adhesion, and movement of cell/subcellular component. “Endocytosis”, “Protein processing in endoplasmic reticulum”, and “Purine metabolism” were the most significantly enriched pathways. The knockdown (RNAi) of Tudor domain-containing protein (TCP), actin-depolymerizing factors (ADF), programmed cell death protein (PD), and serine/threonine-protein kinase (SPK) resulted in the dissociation of collagen fibers and the pilosebaceous unit, increased inflammatory infiltrates/granulocytes (possibly heterophiles), and the depletion of the epithelium. Ticks injected with SPK dsRNA engorged normally but with a change in skin colour (possibly an autoimmune reaction) and the failure to produce eggs pointing to a possible role of SPK in reproduction and host immune modulation. Ticks injected with ADF dsRNA failed to acquire blood, underscoring the role of ADF in facilitating tick feeding. The results of this study showed the presence of phosphorylation in tick saliva and highlight the roles of salivary phosphoproteins in facilitating tick feeding.

Cysteine and related aminothiols in cardiovascular disease, obesity and insulin resistance.

Cysteine, homocysteine, glutathione and cysteinylglycine are metabolically related low molecular weight aminothiols involved in antioxidant defense and in thiol redox control in many cellular and extracellular processes. Alterations in plasma aminothiol profiles are evident in pathologic conditions associated with oxidative stress and accordingly can be considered potential biomarkers. More recently, cysteine has received special attention on this regard and several methodologies for the separation and measurement of cysteine as well as other related aminothiols have been developed and refined. In this review, we provide insight into characteristics, biologic functions, and metabolic interactions of aminothiols (including their involvement in thiol redox status). We highlight cysteine and review methodologic approaches to elucidate its potential role as a risk factor as well as a biomarker in cardiovascular disease, obesity and insulin resistance.