CSB and SMARCAL1 compete for RPA32 at stalled forks and differentially control the fate of stalled forks in BRCA2-deficient cells.

CSB (Cockayne syndrome group B) and SMARCAL1 (SWI/SNF-related, matrix-associated, actin-dependent, regulator of chromatin, subfamily A-like 1) are DNA translocases that belong to the SNF2 helicase family. They both are enriched at stalled replication forks. While SMARCAL1 is recruited by RPA32 to stalled forks, little is known about whether RPA32 also regulates CSB's association with stalled forks. Here, we report that CSB directly interacts with RPA, at least in part via a RPA32C-interacting motif within the N-terminal region of CSB. Modeling of the CSB-RPA32C interaction suggests that CSB binds the RPA32C surface previously shown to be important for binding of UNG2 and SMARCAL1. We show that this interaction is necessary for promoting fork slowing and fork degradation in BRCA2-deficient cells but dispensable for mediating restart of stalled forks. CSB competes with SMARCAL1 for RPA32 at stalled forks and acts non-redundantly with SMARCAL1 to restrain fork progression in response to mild replication stress. In contrast to CSB stimulated restart of stalled forks, SMARCAL1 inhibits restart of stalled forks in BRCA2-deficient cells, likely by suppressing BIR-mediated repair of collapsed forks. Loss of CSB leads to re-sensitization of SMARCAL1-depleted BRCA2-deficient cells to chemodrugs, underscoring a role of CSB in targeted cancer therapy.

Improved Photodynamic Therapy Based on Glutaminase Blockage via Tumor Membrane Coated CB-839/IR-780 Nanoparticles.

Photodynamic therapy (PDT) has promising applications. However, the lethal function of reactive oxygen species (ROS) produced during PDT is typically limited. This restriction is induced by oxygen shortage in the tumor microenvironment due to tumor cell hypermetabolism and reductive chemicals overexpression in tumor tissues. Glutamine (Gln) metabolism is crucial for malignancy development and is closely associated with redox. Herein, a novel nanoparticle (NP) named IRCB@M is constructed to boost PDT through dual effects. This NP simultaneously blocks aerobic respiration and inhibits cellular reduced substances by blocking the Gln metabolic pathway. Within the nanocomplex, a photosensitizer (IR-780) and a glutaminase inhibitor (CB-839) are self-assembled and then encapsulated by cancer cell membranes for homologous targeting. The Gln metabolism intervention relieves hypoxia and decreases the levels of nicotinamide adenine dinucleotide phosphate (NADPH) as well as reduced glutathione (GSH) in vitro and in vivo, which are the dual amplification effects on the IR-780-mediated lethal PDT. The antitumor effects against gastric cancer are ultimately evoked in vivo, thus offering a novel concept for enhancing PDT and other ROS-dependent therapeutic approaches.

Vanadate-based Fe-MOFs as promising negative electrode for hybrid supercapacitor device.

In the supercapacitor field, negative electrodes are mainly concentrated in carbon-based materials, such as activated carbon, carbon nanotubes, graphene, and so forth. However, materials based on metal-organic frameworks (MOFs) as negative active components are relatively rare. Herein, a series of composite materials based on graphene oxide (GO) and vanadate-based Fe-organic frameworks have been prepared by hydrothermal method namely GO/Fe-VO4-BIPY. The deposition amount of polyoxometalate-based metal-organic frameworks (POMOFs) on the surface of graphene is adjusted by changing the content of POMOFs. Through the deposition, it can effectively reduce the accumulation between graphene, and increase the dispersion of POMOFs. As a result, the charge storage performance of the as-obtained materials is greatly improved. Among these materials, GO/Fe-VO4-BIPY-1 has the most prominent performance, with a specific capacitance of 190 F g-1at 0.5 A g-1, which is attributed to the excellent synergistic effect between the Faraday chemical reaction and electric double-layer capacitance. In comparison with pristine Fe-VO4-BIPY, GO/Fe-VO4-BIPY-1 delivers more excellent surface area and therefore exhibits abundant redox reaction sites, achieving better electrochemical performance the best. After assembly with the positive Ni(OH)2electrode, the maximum energy density of 46.84 W h kg-1at a power density of 850 W kg-1is achieved.

Construction and Validation of Novel Ferroptosis-related Risk Score Signature and Prognostic Prediction Nomogram for Patients with Colorectal Cancer.

Background: Colorectal cancer (CRC) has a high morbidity and mortality. Ferroptosis is a phenomenon in which metabolism and cell death are closely related. The role of ferroptosis-related genes in the progression of CRC is still not clear. Therefore, we screened and validated the ferroptosis-related genes which could determine the prevalence, risk and prognosis of patients with CRC. Methods: We firstly screened differentially expressed ferroptosis-related genes by The Cancer Genome Atlas (TCGA) database. Then, these genes were used to construct a risk-score model using the least absolute shrinkage and selection operator (LASSO) regression algorithm. The function and prognosis of the ferroptosis-related genes were confirmed using multi-omics analysis. The gene expression results were validated using publicly available databases and qPCR. We also used publicly available data and ferroptosis-related genes to construct a prognostic prediction nomogram. Results: A total of 24 differential expressed genes associated with ferroptosis were screened in this study. A three-gene risk score model was then established based on these 24 genes and GPX3, CDKN2A and SLC7A11 were selected. The significant prognostic value of this novel three-gene signature was also assessed. Furthermore, we conducted RT-qPCR analysis on cell lines and tissues, and validated the high expression of CDKN2A, GPX3 and low expression of SLC7A11 in CRC cells. The observed mRNA expression of GPX3, CDKN2A and SLC7A11 was consistent with the predicted outcomes. Besides, eight variables including selected ferroptosis related genes were included to establish the prognostic prediction nomogram for patients with CRC. The calibration plots showed favorable consistency between the prediction of the nomogram and actual observations. Also, the time-dependent AUC (>0.7) indicated satisfactory discriminative ability of the nomogram. Conclusions: The present study constructed and validated a novel ferroptosis-related three-gene risk score signature and a prognostic prediction nomogram for patients with CRC. Also, we screened and validated the ferroptosis-related genes GPX3, CDKN2A, and SLC7A11 which could serve as novel biomarkers for patients with CRC.

NETosis-Inspired Cell Surface-Constrained Framework Nucleic Acids Traps (FNATs) for Cascaded Extracellular Recognition and Cellular Behavior Modulation.

Upon pathogenic stimulation, activated neutrophils release nuclear DNA into the extracellular environment, forming web-like DNA structures known as neutrophil extracellular traps (NETs), which capture and kill bacteria, fungi, and cancer cells. This phenomenon is commonly referred to as NETosis. Inspired by this, we introduce a cell surface-constrained web-like framework nucleic acids traps (FNATs) with programmable extracellular recognition capability and cellular behavior modulation. This approach facilitates dynamic key chemical signaling molecule recognition such as adenosine triphosphate (ATP), which is elevated in the extracellular microenvironment, and triggers FNA self-assembly. This, in turn, leads to in situ tightly interwoven FNAs formation on the cell surface, thereby inhibiting target cell migration. Furthermore, it activates a photosensitizer-capturing switch, chlorin e6 (Ce6), and induces cell self-destruction. This cascade platform provides new potential tools for visualizing dynamic extracellular activities and manipulating cellular behaviors using programmable in situ self-assembling DNA molecular devices.

Nanomaterials in tumor immunotherapy: new strategies and challenges.

Tumor immunotherapy exerts its anti-tumor effects by stimulating and enhancing immune responses of the body. It has become another important modality of anti-tumor therapy with significant clinical efficacy and advantages compared to chemotherapy, radiotherapy and targeted therapy. Although various kinds of tumor immunotherapeutic drugs have emerged, the challenges faced in the delivery of these drugs, such as poor tumor permeability and low tumor cell uptake rate, had prevented their widespread application. Recently, nanomaterials had emerged as a means for treatment of different diseases due to their targeting properties, biocompatibility and functionalities. Moreover, nanomaterials possess various characteristics that overcome the defects of traditional tumor immunotherapy, such as large drug loading capacity, precise tumor targeting and easy modification, thus leading to their wide application in tumor immunotherapy. There are two main classes of novel nanoparticles mentioned in this review: organic (polymeric nanomaterials, liposomes and lipid nanoparticles) and inorganic (non-metallic nanomaterials and metallic nanomaterials). Besides, the fabrication method for nanoparticles, Nanoemulsions, was also introduced. In summary, this review article mainly discussed the research progress of tumor immunotherapy based on nanomaterials in the past few years and offers a theoretical basis for exploring novel tumor immunotherapy strategies in the future.

Exogenous auxin regulates the growth and development of peach fruit at the expansion stage by mediating multiple-hormone signaling.

BACKGROUND: Fruit expansion stage is crucial to fruit yield and quality formation, and auxin plays a significant role by mediating multi-hormone signals during fruit expansion. However, till now, it is still unclear of the molecular regulatory network during auxin-mediated peach fruit expansion. RESULTS: Here, exogenous NAA application markedly increased IAA content and drastically decreased ABA content at the fruit expansion stage. Correspondingly, NAA mainly induced the auxin biosynthesis gene (1 PpYUCCA) and early auxin-responsive genes (7PpIAA, 3 PpGH3, and 14 PpSAUR); while NAA down-regulated ABA biosynthesis genes (2 PpNCED, 1 PpABA3, and 1 PpAAO3). In addition, many DEGs involved in other plant hormone biosynthesis and signal transduction were significantly enriched after NAA treatment, including 7 JA, 7 CTK, 6 ETH, and 3 GA. Furthermore, we also found that NAA treatment down-regulated most of genes involved in the growth and development of peach fruit, including the cell wall metabolism-related genes (PpEG), sucrose metabolism-related genes (PpSPS), phenylalanine metabolism-related genes (PpPAL, Pp4CL, and PpHCT), and transcription factors (PpNAC, PpMADS-box, PpDof, PpSBP, and PpHB). CONCLUSION: Overall, NAA treatment at the fruit expansion stage could inhibit some metabolism processes involved in the related genes in the growth and development of peach fruit by regulating multiple-hormone signaling networks. These results help reveal the short-term regulatory mechanism of auxin at the fruit expansion stage and provide new insights into the multi-hormone cascade regulatory network of fruit growth and development.

Targeting KK-LC-1 inhibits malignant biological behaviors of triple-negative breast cancer.

BACKGROUND: Cancer/testis antigens (CTAs) participate in the regulation of malignant biological behaviors in breast cancer. However, the function and mechanism of KK-LC-1, a member of the CTA family, in breast cancer are still unclear. METHODS: Bioinformatic tools, immunohistochemistry, and western blotting were utilized to detect the expression of KK-LC-1 in breast cancer and to explore the prognostic effect of KK-LC-1 expression in breast cancer patients. Cell function assays, animal assays, and next-generation sequencing were utilized to explore the function and mechanism of KK-LC-1 in the malignant biological behaviors of triple-negative breast cancer. Small molecular compounds targeting KK-LC-1 were also screened and drug susceptibility testing was performed. RESULTS: KK-LC-1 was significantly highly expressed in triple-negative breast cancer tissues than in normal breast tissues. KK-LC-1 high expression was related to poor survival outcomes in patients with breast cancer. In vitro studies suggested that KK-LC-1 silencing can inhibit triple-negative breast cancer cell proliferation, invasion, migration, and scratch healing ability, increase cell apoptosis ratio, and arrest the cell cycle in the G0-G1 phase. In vivo studies have suggested that KK-LC-1 silencing decreases tumor weight and volume in nude mice. Results showed that KK-CL-1 can regulate the malignant biological behaviors of triple-negative breast cancer via the MAL2/MUC1-C/PI3K/AKT/mTOR pathway. The small-molecule compound Z839878730 had excellent KK-LC-1 targeting ability and cancer cell killing ability. The EC50 value was 9.7 µM for MDA-MB-231 cells and 13.67 µM for MDA-MB-468 cells. Besides, Z839878730 has little tumor-killing effect on human normal mammary epithelial cells MCF10A and can inhibit the malignant biological behaviors of triple-negative breast cancer cells by MAL2/MUC1-C/PI3K/AKT/mTOR pathway. CONCLUSIONS: Our findings suggest that KK-LC-1 may serve as a novel therapeutic target for triple-negative breast cancer. Z839878730, which targets KK-LC-1, presents a new path for breast cancer clinical treatment.

The tumor-repressing effect of CYP27A1 on renal cell carcinoma by 27-HC arising from cholesterol metabolism.

As a key approach to mediate cholesterol metabolism, the role of the CYP27A1/27-HC axis in renal cell carcinoma (RCC) remains unclear. Analysis of CYP27A1 expression from public databases and metastatic cases in our center suggested that CYP27A1 was obviously downregulated in RCC tissues, and survival analysis further showed its correlation with favorable clinicopathological features and prognosis. In vitro, up and downregulation of CYP27A1 expression in RCC cell lines could definitely illustrate its anticipation involving apoptosis, proliferation, invasion, migration, and clonality. This could be achieved through upregulation of 27-HC concentration, which mediates the activation of signaling pathways of apoptosis and cell cycle arrest. Further, recovery of CYP27A1 expression could definitely inhibit the proliferation of RCC tumors in vivo. This is the first study to explore the role of the CYP27A1/27-HC axis in RCC. Attempts to maintain the normal function of the axis may be a potential strategy in the treatment of RCC, and the predictive value of CYP27A1 detection on the efficacy of targeted therapy in metastatic RCC is also worthy of attention.

Major Vault Protein Prevents Atherosclerotic Plaque Destabilization by Suppressing Macrophage ASK1-JNK Signaling.

BACKGROUND: Macrophages are implicated in atherosclerotic plaque instability by inflammation and degradation of extracellular matrix. However, the regulatory mechanisms driving these macrophage-associated processes are not well understood. Here, we aimed to identify the plaque destabilization-associated cytokines and signaling pathways in macrophages. METHODS: The atherosclerotic models of myeloid-specific MVP (major vault protein) knockout mice and control mice were generated. Atherosclerotic instability, macrophage inflammatory signaling, and active cytokines released by macrophages were examined in vivo and in vitro by using cellular and molecular biological approaches. RESULTS: MVP deficiency in myeloid cells exacerbated murine plaque instability by increasing production of both MMP (matrix metallopeptidase)-9 and proinflammatory cytokines in artery wall. Mechanistically, expression of MMP-9 was mediated via ASK1 (apoptosis signal-regulating kinase 1)-MKK-4 (mitogen-activated protein kinase kinase 4)-JNK (c-Jun N-terminal kinase) signaling in macrophages. MVP and its α-helical domain could bind with ASK1 and inhibit its dimerization and phosphorylation. A 62 amino acid peptide (MVP-[686-747]) in the α-helical domain of MVP showed a crucial role in preventing macrophage MMP-9 production and plaque instability. CONCLUSIONS: MVP may act as an inhibitor for ASK1-JNK signaling-mediated MMP-9 production in macrophages and, thereby, attenuate unstable plaque formation. Our findings suggest that suppression of macrophage ASK1-JNK signaling may be a useful strategy antagonizing atherosclerotic diseases.

RNAi machinery regulates nutrient metabolism and fluconazole resistance in the pathogenic fungus Cryptococcus deneoformans.

The RNAi machinery has been extensively studied in plant and animal cells for their crucial roles in the regulation of genome function. However, the potential roles of RNAi in controlling fungal growth and development have been poorly studied, especially in the basidiomycetous yeast Cryptococcus deneoformans. To characterize the biological functions of RNAi in the pathogenic fungus, a comparative analysis of mRNA profiles using high-throughput sequencing technology was performed for the wild type and the RNAi mutants of C. deneoformans. The results revealed a clear difference in the expression of genes associated with metabolic processes in the RNAi mutants. Besides, the growth under nutrient-limited conditions was significantly reduced in the ago2Δ mutant, suggesting the essential roles of Ago2 in nutrient metabolism. Further investigations revealed the differentially expressed transporters in the RNAi mutants, in which transporters involved in fluconazole efflux were significantly up-regulated. More importantly, on account of the upregulated transporters, RNAi mutant strains developed resistance to fluconazole. By disrupting AFR1 gene using the 'suicide' CRISPR-Cas9 system, we verified that the upregulated ABC transporter Afr1 in the RNAi mutants contributed to the fluconazole resistance. In summary, our data demonstrate that in C. deneoformans the RNAi pathway participates in nutrient metabolism and plays a role in the repression of fluconazole resistance, which provides a deep insight into RNAi mechanisms in Cryptococcus and brings great hints for the clinical treatment of cryptococcosis.

Transcriptome sequencing reveals biological functions of RNAi in C. deneoformans. Nutritional metabolism is deficient due to the AGO2 disruption. RNAi mechanism inhibits fluconazole resistance by regulating the expression of transporters.

Cockayne syndrome group B protein regulates fork restart, fork progression and MRE11-dependent fork degradation in BRCA1/2-deficient cells.

Cockayne syndrome group B (CSB) protein has been implicated in the repair of a variety of DNA lesions that induce replication stress. However, little is known about its role at stalled replication forks. Here, we report that CSB is recruited to stalled forks in a manner dependent upon its T1031 phosphorylation by CDK. While dispensable for MRE11 association with stalled forks in wild-type cells, CSB is required for further accumulation of MRE11 at stalled forks in BRCA1/2-deficient cells. CSB promotes MRE11-mediated fork degradation in BRCA1/2-deficient cells. CSB possesses an intrinsic ATP-dependent fork reversal activity in vitro, which is activated upon removal of its N-terminal region that is known to autoinhibit CSB's ATPase domain. CSB functions similarly to fork reversal factors SMARCAL1, ZRANB3 and HLTF to regulate slowdown in fork progression upon exposure to replication stress, indicative of a role of CSB in fork reversal in vivo. Furthermore, CSB not only acts epistatically with MRE11 to facilitate fork restart but also promotes RAD52-mediated break-induced replication repair of double-strand breaks arising from cleavage of stalled forks by MUS81 in BRCA1/2-deficient cells. Loss of CSB exacerbates chemosensitivity in BRCA1/2-deficient cells, underscoring an important role of CSB in the treatment of cancer lacking functional BRCA1/2.

CSB Regulates Pathway Choice in Response to DNA Replication Stress Induced by Camptothecin.

Topoisomerase inhibitor camptothecin (CPT) induces fork stalling and is highly toxic to proliferating cells. However, how cells respond to CPT-induced fork stalling has not been fully characterized. Here, we report that Cockayne syndrome group B (CSB) protein inhibits PRIMPOL-dependent fork repriming in response to a low dose of CPT. At a high concentration of CPT, CSB is required to promote the restart of DNA replication through MUS81-RAD52-POLD3-dependent break-induced replication (BIR). In the absence of CSB, resumption of DNA synthesis at a high concentration of CPT can occur through POLQ-LIG3-, LIG4-, or PRIMPOL-dependent pathways, which are inhibited, respectively, by RAD51, BRCA1, and BRCA2 proteins. POLQ and LIG3 are core components of alternative end joining (Alt-EJ), whereas LIG4 is a core component of nonhomologous end joining (NHEJ). These results suggest that CSB regulates fork restart pathway choice following high-dosage CPT-induced fork stalling, promoting BIR but inhibiting Alt-EJ, NHEJ, and fork repriming. We find that loss of CSB and BRCA2 is a toxic combination to genomic stability and cell survival at a high concentration of CPT, which is likely due to accumulation of ssDNA gaps, underscoring an important role of CSB in regulating the therapy response in cancers lacking functional BRCA2.

[Mechanism of Sijunzi Decoction in treatment of Alzheimer's disease based on UPLC-Q-TOF-MS, network pharmacology, and experimental verification].

The study identified the blood-entering components of Sijunzi Decoction after gavage administration in rats by UPLC-Q-TOF-MS/MS, and investigated the mechanism of Sijunzi Decoction in treating Alzheimer's disease by virtue of network pharmacology, molecular docking, and experimental verification. The blood-entering components of Sijunzi Decoction were identified based on the mass spectra and data from literature and databases. The potential targets of the above-mentioned blood-entering components in the treatment of Alzheimer's disease were searched against PharmMapper, OMIM, DisGeNET, GeneCards, and TTD. Next, STRING was employed to establish a protein-protein interaction(PPI) network. DAVID was used to perform the Gene Ontology(GO) annotation and the Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment. Cytoscape 3.9.0 was used to carry out visual analysis. AutoDock Vina and PyMOL were used for molecular docking of the blood-entering components with the potential targets. Finally, the phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt) signaling pathway enriched by the KEGG analysis was selected for validation by animal experiments. The results showed that 17 blood-entering components were detected in the serum samples after administration. Among them, poricoic acid B, liquiritigenin, atractylenolide Ⅱ, atractylenolide Ⅲ, ginsenoside Rb_1, and glycyrrhizic acid were the key components of Sijunzi Decoction in treating Alzheimer's disease. HSP90AA1, PPARA, SRC, AR, and ESR1 were the main targets for Sijunzi Decoction to treat Alzheimer's disease. Molecular docking showed that the components bound well with the targets. Therefore, we hypothesized that the mechanism of Sijunzi Decoction in treating Alzheimer's disease may be associated with the PI3K/Akt, cancer treatment, and mitogen-activated protein kinase(MAPK) signaling pathways. The results of animal experiments showed that Sijunzi Decoction significantly attenuated the neuronal damage in the hippocampal dentate gyrus area, increased the neurons, and raised the ratios of p-Akt/Akt and p-PI3K/PI3K in the hippocampus of mice. In conclusion, Sijunzi Decoction may treat Alzheimer's disease by activating the PI3K/Akt signaling pathway. The findings of this study provide a reference for further studies about the mechanism of action and clinical application of Sijunzi Decoction.

[Different processed products of Polygonati Rhizoma treat Alzheimer's disease in rats: urine metabolomics based on UPLC-Q/TOF-MS].

The study investigated the effects of different processed products of Polygonati Rhizoma(black bean-processed Polygonati Rhizoma, BBPR; stewed Polygonati Rhizoma, SPR) on the urinary metabolites in a rat model of Alzheimer's disease(AD). Sixty SPF-grade male SD rats were randomized into a control group, a model group, a donepezil group, a BBPR group, and a SPR group, with twelve rats in each group. Other groups except the control group were administrated with D-galactose injection(100 mg·kg~(-1)) once a day for seven weeks. The control group was administrated with an equal volume of normal saline once a day for seven consecutive weeks. After three weeks of D-galactose injection, bilateral hippocampal Aß_(25-35) injections were performed for modeling. The rats were administrated with corresponding drugs(10 mL·kg~(-1)) by gavage since week 2, and the rats in the model and control group with an equal volume of double distilled water once a day for 35 continuous days. The memory behaviour and pathological changes in the hippocampal tissue were observed. The untargeted metabolites in the urine were detected by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry(UPLC-Q/TOF-MS). Principal component analysis(PCA) and orthogonal partial least square-discriminant analysis(OPLS-DA) were employed to characterize and screen differential metabolites and potential biomarkers, for which the metabolic pathway enrichment analysis was conducted. The results indicated that BBPR and SPR increased the new object recognition index, shortened the escape latency, and increased the times of crossing the platform of AD rats in the Morris water maze test. The results of hematoxylin-eosin(HE) staining showed that the cells in the hippocampal tissue of the drug administration groups were closely arranged. Moreover, the drugs reduced the content of interleukin-6(IL-6, P<0.01) and tumor necrosis factor-α(TNF-α) in the hippocampal tissue, which were more obvious in the BBPR group(P<0.05). After screening, 15 potential biomarkers were identified, involving two metabolic pathways: dicoumarol pathway and piroxicam pathway. BBPR and SPR may alleviate AD by regulating the metabolism of dicoumarol and piroxicam.

[Effect of Erjing Pills on alleviating neuroinflammation of AD rats based on TLR4/NF-κB/NLRP3 pathway and its mechanism].

This paper aimed to study the effect of Erjing Pills on the improvement of neuroinflammation of rats with Alzheimer's di-sease(AD) induced by the combination of D-galactose and Aß_(25-35) and its mechanism. SD rats were randomly divided into a sham group, a model control group, a positive drug group(donepezil, 1 mg·kg~(-1)), an Erjing Pills high-dose group(9.0 g·kg~(-1)), and an Erjing Pills low-dose group(4.5 g·kg~(-1)), with 14 rats each group. To establish the rat model of AD, Erjing Pills were intragastrically administrated to rats for 5 weeks after 2 weeks of D-galactose injection. D-galactose was intraperitoneally injected into rats for 3 weeks, and then Aß_(25-35) was injected into the bilateral hippocampus. The new object recognition test was used to evaluate the learning and memory ability of rats after 4 weeks of intragastric administration. Tissues were acquired 24 h after the last administration. The immunofluorescence method was used to detect the activation of microglia in the brain tissue of rats. The positive expressions of Aß_(1-42) and phosphory protein Tau~(404)(p-Tau~(404)) in the CA1 area of the hippocampus were detected by immunohistochemistry. The levels of inflammatory factors interleukin-1ß(IL-1ß), tumor necrosis factor-α(TNF-α), and interleukin-6(IL-6) in the brain tissue were determined by enzyme-linked immunosorbent assay(ELISA). Toll-like receptor 4(TLR4)/nuclear factor kappa B(NF-κB)/nucleotide-binding oligomerization domain-like receptors 3(NLRP3) pathway-associated proteins in the brain tissue were determined by Western blot. The results showed that as compared with the sham group, the new object recognition index of rats in the model control group decreased significantly, the deposition of Aß_(1-42) and p-Tau~(404) positive protein in the hippocampus increased significantly, and the levels of microglia activation increased significantly in the dentate gyrus. The levels of IL-1ß, TNF-α, and IL-6 in the hippocampus of the model control group increased significantly, and the expression levels of TLR4, p-NF-κB p65/NF-κB p65, p-IκBα/IκBα, and NLRP3 proteins in the hippocampus increased significantly. Compared with the model control group, the Erjing Pill groups enhanced the new object recognition index of rats, decreased the deposition of Aß_(1-42) and the expression of p-Tau~(404) positive protein in the hippocampus, inhibited the activation of microglia in the dentate gyrus, reduced the levels of inflammatory factors IL-1ß, TNF-α, and IL-6 in the hippocampus, and down-regulated the expression levels of TLR4, p-NF-κB P65/NF-κB P65, p-IκBα/IκBα, and NLRP3 proteins in the hippocampus. In conclusion, Erjing Pills can improve the learning and memory ability of the rat model of AD presumably by improving the activation of microglia, reducing the expression levels of neuroinflammatory factors IL-1ß, TNF-α, and IL-6, inhibiting the TLR4/NF-κB/NLRP3 neuroinflammation pathway, and decreasing hippocampal deposition of Aß and expression of p-Tau, thereby restoring the hippocampal morphological structure.

Targeting SOST using a small-molecule compound retards breast cancer bone metastasis.

BACKGROUND: Breast cancer metastasis to the bone can be exacerbated by osteoporosis, is associated with poor long-term survival, and has limited therapeutic options. Sclerostin (SOST) is an endogenous inhibitor of bone formation, and an attractive target for treatment of osteoporosis. However, it is unclear whether SOST can be used as a therapeutic target for bone metastases of breast cancer, and whether small molecule compounds that target SOST in breast cancer cells can inhibit breast cancer bone metastasis. METHODS: SOST expression in 442 breast cancer tissues was characterized by immunohistochemistry and statistically analyzed for the association with breast cancer bone metastases. Bone metastatic breast cancer SCP2 cells were induced for SOST silencing or overexpression and their bone metastatic behaviors were tested in vitro and in vivo. To identify potential therapeutics, we screened inhibitors of the interaction of SOST with STAT3 from a small chemical molecule library and tested the inhibitory effects of one inhibitor on breast cancer growth and bone metastasis in vitro and in vivo. RESULTS: We found that up-regulated SOST expression was associated with breast cancer bone metastases and worse survival of breast cancer patients. SOST silencing significantly reduced the bone metastatic capacity of SCP2 cells. SOST interacted with STAT3 to enhance the TGF-ß/KRAS signaling, increasing both tumor growth and bone metastasis. Treatment with one lead candidate, S6, significantly inhibited the growth of breast-cancer organoids and bone metastasis in mice. CONCLUSIONS: Our findings highlight a new class of potential therapeutics for treatment of bone metastasis in breast cancer.

CSB cooperates with SMARCAL1 to maintain telomere stability in ALT cells.

Elevated replication stress is evident at telomeres of about 10-15% of cancer cells, which maintain their telomeres via a homologous recombination (HR)-based mechanism, referred to as alternative lengthening of telomeres (ALT). How ALT cells resolve replication stress to support their growth remains incompletely characterized. Here, we report that CSB (also known as ERCC6) promotes recruitment of HR repair proteins (MRN, BRCA1, BLM and RPA32) and POLD3 to ALT telomeres, a process that requires the ATPase activity of CSB and is controlled by ATM- and CDK2-dependent phosphorylation. Loss of CSB stimulates telomeric recruitment of MUS81 and SLX4, components of the structure-specific MUS81-EME1-SLX1-SLX4 (MUS-SLX) endonuclease complex, suggesting that CSB restricts MUS-SLX-mediated processing of stalled forks at ALT telomeres. Loss of CSB coupled with depletion of SMARCAL1, a chromatin remodeler implicated in catalyzing regression of stalled forks, synergistically promotes not only telomeric recruitment of MUS81 but also the formation of fragile telomeres, the latter of which is reported to arise from fork stalling. These results altogether suggest that CSB-mediated HR repair and SMARCAL1-mediated fork regression cooperate to prevent stalled forks from being processed into fragile telomeres in ALT cells.

Plasma Exosomal AKR1C3 mRNA Expression Is a Predictive and Prognostic Biomarker in Patients with Metastatic Castration-Resistant Prostate Cancer.

PURPOSE: Aldo-keto reductase family 1 member C3 (AKR1C3) is important in prostate cancer progression, being a potential biomarker in metastatic castration-resistant prostate cancer (mCRPC). Previous explorations of AKR1C3 are mainly based on tissue samples. This study investigates using plasma-based liquid biopsy to validate the prognostic and predictive value of AKR1C3 in patients with mCRPC . MATERIALS AND METHODS: We prospectively recruited 62 patients with mCRPC. All patients received repeated prostate biopsies at the time of mCRPC diagnosis, and immunohistochemistry (IHC) staining was used to detect protein expression of AKR1C3 in the tissues. We took their blood simultaneously and performed digital droplet polymerase chain reaction (ddPCR) to measure expression levels of AKR1C3 in the exosomes. The detected plasma and tissue AKR1C3 expression levels were analyzed for patients' overall survival (OS) and progression-free survival under first-line abiraterone use (ABI-PFS). RESULTS: All other baseline characteristics were balanced between the 2 groups. 15/62 (24.2%) and 25/62 (40.3%) patients showed AKR1C3-EXO positive (≥20 copies/20 µL) and AKR1C3-IHC positive, respectively. Positive AKR1C3-EXO expression was associated with decreased patients' survival [ABI-PFS: 3.9 vs 10.1 months, P < .001; OS: 16.2 vs 32.5 months, P < .001]. AKR1C3-IHC positivity was also correlated with ABI-PFS and OS (P = .010, P = .016). In patients with worse baseline blood tests (including higher alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) level and lower hemoglobin (HB) level), and lower ISUP/WHO group (<4), their OS was significantly worse when showing AKR1C3-EXO positive. CONCLUSION: AKR1C3-EXO is associated with patient prognosis regarding OS and ABI-PFS and can be used as a biomarker in mCRPC.

Mass Fabrication of Capillary Columns Based on Centrifugal Packing.

Packed capillary columns have become the standard front-end separation device for mass spectrometry-based proteomics. The development of simple, fast, and robust capillary column technology, especially that with mass-fabrication capacity, can greatly improve analytical throughput and reproducibility in omics research. In this technical note, we report a centrifugal packing technology, which has the capability to mass fabricate high quality capillary columns with a 2886 columns/day fabrication throughput. The centrifugally packed columns presented significantly improved efficiency (reduced plate height hmin = 1.6, 37%-40% improvement compared with slurry packed columns), advanced kinetic performance limit, and excellent column-to-column reproducibility (2.0% RSD for retention time, 50 columns). Such columns enabled ∼5300 HeLa proteins identified in single-shot proteomic analysis, displaying both intercolumn and inter-run retention time stability (retention time RSD = 0.94% between nine replicates on three columns for probing peptide sequence). The mass-fabrication technology reported in this technical note may support disposable use of high quality chromatographic columns in large-scale bioanalysis.