CRISPR-Cas9 screening identifies INTS3 as an anti-apoptotic RNA-binding protein and therapeutic target for colorectal cancer.

Growing evidences indicate that RNA-binding proteins (RBPs) play critical roles in regulating the RNA splicing, polyadenylation, stability, localization, translation, and turnover. Abnormal expression of RBPs can promote tumorigenesis. Here, we performed a CRISPR screen using an RBP pooled CRISPR knockout library and identified 27 potential RBPs with role in supporting colorectal cancer (CRC) survival. We found that the deletion/depletion of INTS3 triggered apoptosis in CRC. The in vitro experiments and RNA sequencing revealed that INTS3 destabilized pro-apoptotic gene transcripts and contributed to the survival of CRC cells. INTS3 loss delayed CRC cells growth in vivo. Furthermore, delivery of DOTAP/cholesterol-mshINTS3 nanoparticles inhibited CRC tumor growth. Collectively, our work highlights the role of INTS3 in supporting CRC survival and provides several novel therapeutic targets for treatment.

ICA/SDF-1α/PBMSCs loaded onto alginate and gelatin cross-linked scaffolds promote damaged cartilage repair.

A three-dimensional alginate-coated scaffold (GAIS) was constructed in the present study to showcase the multidifferentiation potential of peripheral blood mesenchymal stem cells (PBMSCs) and to investigate the role and mechanism by which Icariin (ICA)/stromal cell-derived factor (SDF-1α)/PBMSCs promote damaged articular repair. In addition, the ability of ICA, in combination with SDF-1α, to promote the migration and proliferation of stem cells was validated through the utilization of CCK-8 and migration experiments. The combination of ICA and SDF-1α inhibited the differentiation of PBMSCs into cartilage, as demonstrated by in vivo experiments and histological staining. Both PCR and western blot experiments showed that GAIS could upregulate the expression of particular genes in chondrocytes. In comparison to scaffolds devoid of alginate (G0), PBMSCs seeded into GAIS scaffolds exhibited a greater rate of proliferation, and the conditioned medium derived from scaffolds containing SDF-1α enhanced the capacity for cell migration. Moreover, after a 12-week treatment period, GAIS, when successfully transplanted into osteochondral defects of mice, was found to promote cartilage regeneration and repair. The findings, therefore, demonstrate that GAIS enhanced the in vitro capabilities of PBMSCs, including proliferation, migration, homing and chondrogenic differentiation. In addition, ICA and SDF-1α effectively collaborated to support cartilage formation in vivo. Thus, the ICA/SDF-1α/PBMSC-loaded biodegradable alginate-gelatin scaffolds showcase considerable potential for use in cartilage repair.

Development of an Untargeted LC-MS Metabolomics Method with Postcolumn Infusion for Matrix Effect Monitoring in Plasma and Feces.

Untargeted metabolomics based on reverse phase LC-MS (RPLC-MS) plays a crucial role in biomarker discovery across physiological and disease states. Standardizing the development process of untargeted methods requires paying attention to critical factors that are under discussed or easily overlooked, such as injection parameters, performance assessment, and matrix effect evaluation. In this study, we developed an untargeted metabolomics method for plasma and fecal samples with the optimization and evaluation of these factors. Our results showed that optimizing the reconstitution solvent and sample injection amount was critical for achieving the balance between metabolites coverage and signal linearity. Method validation with representative stable isotopically labeled standards (SILs) provided insights into the analytical performance evaluation of our method. To tackle the issue of the matrix effect, we implemented a postcolumn infusion (PCI) approach to monitor the overall absolute matrix effect (AME) and relative matrix effect (RME). The monitoring revealed distinct AME and RME profiles in plasma and feces. Comparing RME data obtained for SILs through postextraction spiking with those monitored using PCI compounds demonstrated the comparability of these two methods for RME assessment. Therefore, we applied the PCI approach to predict the RME of 305 target compounds covered in our in-house library and found that targets detected in the negative polarity were more vulnerable to the RME, regardless of the sample matrix. Given the value of this PCI approach in identifying the strengths and weaknesses of our method in terms of the matrix effect, we recommend implementing a PCI approach during method development and applying it routinely in untargeted metabolomics.

Logic "AND Gate Circuit"-Based Gasdermin Protein Expressing Nanoplatform Induces Tumor-Specific Pyroptosis to Enhance Cancer Immunotherapy.

Pyroptosis mediated by gasdermin protein has shown great potential in cancer immunotherapies. However, the low expression of gasdermin proteins and the systemic toxicity of nonspecific pyroptosis limit its clinical application. Here, we designed a synthetic biology strategy to construct a tumor-specific pyroptosis-inducing nanoplatform M-CNP/Mn@pPHS, in which a pyroptosis-inducing plasmid (pPHS) was loaded onto a manganese (Mn)-doped calcium carbonate nanoparticle and wrapped in a tumor-derived cell membrane. M-CNP/Mn@pPHS showed an efficient tumor targeting ability. After its internalization by tumor cells, the degradation of M-CNP/Mn@pPHS in the acidic endosomal environment allowed the efficient endosomal escape of plasmid pPHS. To trigger tumor-specific pyroptosis, pPHS was designed according to the logic "AND gate circuit" strategy, with Hif-1α and Sox4 as two input signals and gasdermin D induced pyroptosis as output signal. Only in cells with high expression of Hif-1α and Sox4 simultaneously will the output signal gasdermin D be expressed. Since Hif-1α and Sox4 are both specifically expressed in tumor cells, M-CNP/Mn@pPHS induces the tumor-specific expression of gasdermin D and thus pyroptosis, triggering an efficient immune response with little systemic toxicity. The Mn2+ released from the nanoplatform further enhanced the antitumor immune response by stimulating the cGAS-STING pathway. Thus, M-CNP/Mn@pPHS efficiently inhibited tumor growth with 79.8% tumor regression in vivo. We demonstrate that this logic "AND gate circuit"-based gasdermin nanoplatform is a promising strategy for inducing tumor-specific pyroptosis with little systemic toxicity.

Current insights into cow's milk allergy in children: Microbiome, metabolome, and immune response-A systematic review.

The increasing prevalence of IgE-mediated cow's milk allergy (CMA) in childhood is a worldwide health concern. There is a growing awareness that the gut microbiome (GM) might play an important role in CMA development. Therefore, treatment with probiotics and prebiotics has gained popularity. This systematic review provides an overview of the alterations of the GM, metabolome, and immune response in CMA children and animal models, including post-treatment modifications. MEDLINE, PubMed, Scopus, and Web of Science were searched for studies on GM in CMA-diagnosed children, published before 1 March 2023. A total of 21 articles (13 on children and 8 on animal models) were included. The studies suggest that the GM, characterized by an enrichment of the Clostridia class and reductions in the Lactobacillales order and Bifidobacterium genus, is associated with CMA in early life. Additionally, reduced levels of short-chain fatty acids (SCFAs) and altered amino acid metabolism were reported in CMA children. Commonly used probiotic strains belong to the Bifidobacterium and Lactobacillus genera. However, only Bifidobacterium levels were consistently upregulated after the intervention, while alterations of other bacteria taxa remain inconclusive. These interventions appear to contribute to the restoration of SCFAs and amino acid metabolism balance. Mouse models indicate that these interventions tend to restore the Th 2/Th 1 balance, increase the Treg response, and/or silence the overall pro- and anti-inflammatory cytokine response. Overall, this systematic review highlights the need for multi-omics-related research in CMA children to gain a mechanistic understanding of this disease and to develop effective treatments and preventive strategies.

Biomimetic-gasdermin-protein-expressing nanoplatform mediates tumor-specific pyroptosis for cancer immunotherapy.

Pyroptosis, mediated by gasdermin proteins, has shown excellent efficacy in facilitating cancer immunotherapy. The strategies commonly used to induce pyroptosis suffer from a lack of tissue specificity, resulting in the nonselective activation of pyroptosis and consequent systemic toxicity. Moreover, pyroptosis activation usually depends on caspase, which can induce inflammation and metabolic disorders. In this study, inspired by the tumor-specific expression of SRY-box transcription factor 4 (Sox4) and matrix metalloproteinase 2 (MMP2), we constructed a doubly regulated plasmid, pGMD, that expresses a biomimetic gasdermin D (GSDMD) protein to induce the caspase-independent pyroptosis of tumor cells. To deliver pGMD to tumor cells, we used a hyaluronic acid (HA)-shelled calcium carbonate nanoplatform, H-CNP@pGMD, which effectively degrades in the acidic endosomal environment, releasing pGMD into the cytoplasm of tumor cells. Upon the initiation of Sox4, biomimetic GSDMD was expressed and cleaved by MMP2 to induce tumor-cell-specific pyroptosis. H-CNP@pGMD effectively inhibited tumor growth and induced strong immune memory effects, preventing tumor recurrence. We demonstrate that H-CNP@pGMD-induced biomimetic GSDMD expression and tumor-specific pyroptosis provide a novel approach to boost cancer immunotherapy.

Crosstalk between colorectal CSCs and immune cells in tumorigenesis, and strategies for targeting colorectal CSCs.

Cancer immunotherapy has emerged as a promising strategy in the treatment of colorectal cancer, and relapse after tumor immunotherapy has attracted increasing attention. Cancer stem cells (CSCs), a small subset of tumor cells with self-renewal and differentiation capacities, are resistant to traditional therapies such as radiotherapy and chemotherapy. Recently, CSCs have been proven to be the cells driving tumor relapse after immunotherapy. However, the mutual interactions between CSCs and cancer niche immune cells are largely uncharacterized. In this review, we focus on colorectal CSCs, CSC-immune cell interactions and CSC-based immunotherapy. Colorectal CSCs are characterized by robust expression of surface markers such as CD44, CD133 and Lgr5; hyperactivation of stemness-related signaling pathways, such as the Wnt/ß-catenin, Hippo/Yap1, Jak/Stat and Notch pathways; and disordered epigenetic modifications, including DNA methylation, histone modification, chromatin remodeling, and noncoding RNA action. Moreover, colorectal CSCs express abnormal levels of immune-related genes such as MHC and immune checkpoint molecules and mutually interact with cancer niche cells in multiple tumorigenesis-related processes, including tumor initiation, maintenance, metastasis and drug resistance. To date, many therapies targeting CSCs have been evaluated, including monoclonal antibodies, antibody‒drug conjugates, bispecific antibodies, tumor vaccines adoptive cell therapy, and small molecule inhibitors. With the development of CSC-/niche-targeting technology, as well as the integration of multidisciplinary studies, novel therapies that eliminate CSCs and reverse their immunosuppressive microenvironment are expected to be developed for the treatment of solid tumors, including colorectal cancer.

Icariin activates far upstream element binding protein 1 to regulate hypoxia-inducible factor-1α and hypoxia-inducible factor-2α signaling and benefits chondrocytes.

Icariin (ICA) is a typical flavonoid glycoside derived from epimedium plants. It has both anabolic and anti-catabolic effects to improve bone mineral density and reduce bone microstructural degradation. However, the effect and underlying mechanism of ICA on the proliferation and metabolism of chondrocyte and synthesis of extracellular matrix are still unclear. This study aimed to investigate the role and regulation of far upstream element binding protein 1 (FUBP1) in chondrocytes treated with ICA to maintain homeostasis and suppress inflammatory responses. In the study, the effect of ICA on chondrocytes with overexpressed or silenced FUBP1 was detected by the MTS and single-cell cloning methods. The expression of hypoxia-inducible factor-1/2α (HIF-1/2α), FUBP1, matrix metalloproteinase (MMP)9, SRY-box transcription factor 9 (SOX9), and type II collagen (Col2α) in ATDC5 cells, a mouse chondrogenic cell line, treated with ICA was evaluated by immunoblotting. Western blotting revealed 1 µM ICA to have the most significant effect on chondrocytes. Alcian blue staining and colony formation assays showed that the promoting effect of ICA was insignificant in FUBP1-knockdown cells (P > 0.05) but significantly enhanced in FUBP1-overexpressed cells (P < 0.05). Western blot results from FUBP1-knockdown cells treated with or without ICA showed no significant difference in the expression of FUBP1, HIF-1/2α, MMP9, SOX9, and Col2α proteins, whereas the same proteins showed increased expression in FUBP1-overexpressed chondrocytes; moreover, HIF-2α and MMP9 expression was significantly inhibited in FUBP1-knockdown chondrocytes (P < 0.05). In conclusion, as a bioactive monomer of traditional Chinese medicine, ICA is beneficial to chondrocytes.

[Research Advances on Strategies to Promote Homing and Engraftment of Hematopoietic Stem Cells--Review].

The homing and engraftment of hematopoietic stem cells (HSC) into bone marrow is the first critical step for successful clinical hematopoietic stem cell transplantation (HSCT). SDF-1 / CXCR4 is considered to be a very promising target to promote HSC homing. In recent years, with the in-depth research on the HSC homing, a variety of new strategies for promoting HSC homing and engraftment have been explored, such as nuclear hormone receptor, histone deacetylase inhibitor, prostaglandin and metabolic regulation, so as to increase the success rate of HSCT and improve the survival of patients. In this review, the recent research advances in the mechanism of HSC homing and strategies to promote HSC homing and engraftment were summarized and discussed.

mcPGK1-dependent mitochondrial import of PGK1 promotes metabolic reprogramming and self-renewal of liver TICs.

Liver tumour-initiating cells (TICs) contribute to tumour initiation, metastasis, progression and drug resistance. Metabolic reprogramming is a cancer hallmark and plays vital roles in liver tumorigenesis. However, the role of metabolic reprogramming in TICs remains poorly explored. Here, we identify a mitochondria-encoded circular RNA, termed mcPGK1 (mitochondrial circRNA for translocating phosphoglycerate kinase 1), which is highly expressed in liver TICs. mcPGK1 knockdown impairs liver TIC self-renewal, whereas its overexpression drives liver TIC self-renewal. Mechanistically, mcPGK1 regulates metabolic reprogramming by inhibiting mitochondrial oxidative phosphorylation (OXPHOS) and promoting glycolysis. This alters the intracellular levels of α-ketoglutarate and lactate, which are modulators in Wnt/ß-catenin activation and liver TIC self-renewal. In addition, mcPGK1 promotes PGK1 mitochondrial import via TOM40 interactions, reprogramming metabolism from oxidative phosphorylation to glycolysis through PGK1-PDK1-PDH axis. Our work suggests that mitochondria-encoded circRNAs represent an additional regulatory layer controlling mitochondrial function, metabolic reprogramming and liver TIC self-renewal.

Conjugated linoleic acid alleviates glycolipid metabolic disorders by modulating intestinal microbiota and short-chain fatty acids in obese rats.

Although conjugated linoleic acid (CLA) has been shown to have anti-obesity properties, the effect and mechanism of CLA in alleviating glycolipid metabolism disorders remains unclear. In this work, it was observed that rats fed a high-fat diet (HFD) had lower body weight and body fat levels after 9 weeks of low-dose and high-dose CLA interventions. The results of blood biochemical indices showed that CLA significantly reduced the levels of total cholesterol, triglycerides, fasting blood glucose and insulin. Additionally, high-dose CLA could restore the intestinal microbiota composition, including increasing the relative abundances of short-chain fatty acid (SCFA)-producing microbiota, such as Dubosiella, Faecalibaculum and Bifidobacterium; decreasing the relative abundances of Enterococcus and Ruminococcus_2; and increasing the content of SCFAs in feces and serum. Further analysis showed that high-dose CLA could increase the expression levels of Insr, Irs-2, Akt and Glut4 in the liver tissue of HFD-induced obese rats. Consistently, high dose of CLA could reversibly improve the downregulation of INSR, AKT, PI3K and GLUT4 protein expression caused by HFD and reverse the decline in AKT phosphorylation levels. Correlation clustering analysis with a heatmap showed that the changes in specific microbiota induced by high-dose CLA were correlated with changes in obesity-related indices and gene expression. The molecular docking analysis showed that the molecular docking of SCFAs with the IRS-2, AKT and GLUT4 proteins had high linking activity. The results supported that CLA can alleviate glycolipid metabolic imbalances associated with obesity by altering the intestinal microbiota to induce the production of SCFAs and thereby activate the INSR/IRS-2/AKT/GLUT4 pathway. This study supports CLA may be preferentially used by the intestinal microbiota of the host to promote its health.

Noncoding RNA circBtnl1 suppresses self-renewal of intestinal stem cells via disruption of Atf4 mRNA stability.

Intestinal stem cells (ISCs) at the crypt base are responsible for the regeneration of the intestinal epithelium. However, how ISC self-renewal is regulated still remains unclear. Here we identified a circular RNA, circBtnl1, that is highly expressed in ISCs. Loss of circBtnl1 in mice enhanced ISC self-renewal capacity and epithelial regeneration, without changes in mRNA and protein levels of its parental gene Btnl1. Mechanistically, circBtnl1 and Atf4 mRNA competitively bound the ATP-dependent RNA helicase Ddx3y to impair the stability of Atf4 mRNA in wild-type ISCs. Furthermore, ATF4 activated Sox9 transcription by binding to its promoter via a unique motif, to enhance the self-renewal capacity and epithelial regeneration of ISCs. In contrast, circBtnl1 knockout promoted Atf4 mRNA stability and enhanced ATF4 expression, which caused Sox9 transcription to potentiate ISC stemness. These data indicate that circBtnl1-mediated Atf4 mRNA decay suppresses Sox9 transcription that negatively modulates self-renewal maintenance of ISCs.

Assessment of heavy metals contamination and human health risk assessment of the commonly consumed medicinal herbs in China.

This study investigates heavy metal contamination of commonly consumed medicinal herbs and human health risks to the Chinese population arising from the consumption of herbs that contain potentially harmful elements. Food safety standards for Chinese residents are becoming stricter, and much work in this field needs to be performed. This study examines Co, Ba, Fe, Cr, Mn, Ni, Zn, As, Cd, Pb, Cu, Be, Sb, and Bi concentrations in four regularly consumed Chinese herb species: Radix Paeoniae Alba (RPA), Radix Angelicae Dahuricae (RAD), Rhizoma Atractylodis Macrocephalae (RAM), and Radix Puerariae (RP). A pollution status examination and evaluation of heavy metals in RPA, RAD, RAM, and RP were performed. The human health risk assessment associated with the intake of potentially harmful elements in herbs was calculated in terms of the estimated daily intake (EDI), the target hazard quotient (THQ), the estimated hazard index (HI), and the lifetime cancer risk (CR). The mean single-factor pollution index (PI) showed that in the RPA, RAD, RAM, and RP samples, approximately 10.0%, 10.0%, 30.0%, and 10.0%, respectively, were polluted by Cd. The present study indicated that the pattern of consumption of the studied herbs in China does not seem to suggest an excessive health hazard associated with any of the toxic elements studied.

Cell Multi-Bernoulli (Cell-MB) Sensor Control for Multi-Object Search-While-Tracking (SWT).

Information-driven control can be used to develop intelligent sensors that can optimize their measurement value based on environmental feedback. In object tracking applications, sensor actions are chosen based on the expected reduction in uncertainty also known as information gain. Random finite set (RFS) theory provides a formalism for quantifying and estimating information gain in multi-object tracking problems. However, estimating information gain in these applications remains computationally challenging. This paper presents a new tractable approximation of the RFS expected information gain applicable to sensor control for multi-object search and tracking. Unlike existing RFS approaches, the information gain approximation presented in this paper considers the contributions of non-ideal noisy measurements, missed detections, false alarms, and object appearance/disappearance. The effectiveness of the information-driven sensor control is demonstrated through two multi-vehicle search-while-tracking experiments using real video data from remote terrestrial and satellite sensors.

Intrathecal Dexmedetomidine Combined With Ropivacaine in Cesarean Section: A Prospective Randomized Double-Blind Controlled Study.

Objective: This study aimed to find the best dose of dexmedetomidine in spinal anesthesia for cesarean section. Methods: 120 American Society of Anesthesiologists (ASA) Class I and II parturients undergoing elective cesarean delivery under spinal anesthesia were randomly allocated into four groups treated with intrathecal ropivacaine (12 mg) alone (Group R) or in combination with dexmedetomidine 5 µg (Group RD1), 7.5 µg (Group RD2) and 10 µg (Group RD3). Characteristics of spinal anesthesia, hemodynamic changes, adverse effects, stress reactions and neonatal outcomes were recorded in the four groups. Results: Patients in Group RD1, RD2, and RD3 had significantly longer sustained sensory and motor block time than patients in Group R. All four groups had comparable onset times of sensory and motor block. The time for the level of sensory block to lower to S1 was longer in Group RD1 (411.07 ± 106.66 min), Group RD2 (397.03 ± 125.39 min) and Group RD3 (468.63 ± 116.43 min) than in Group R (273.60 ± 88.34 min) (p < 0.001). The time to recover from motor block to a Bromage score of IV was longer in Group RD1 (353.60.07 ± 137.28 min), Group RD2 (350.57 ± 118.01 min) and Group RD3 (404.67 ± 112.83 min) than in Group R (232.70 ± 93.29) (p < 0.01). The incidence of chills was significantly lower in the Group RD1, RD2, and RD3 than in the Group R (p < 0.001). There was no significant difference in the incidence of adverse effects such as hypotension, bradycardia, nausea, vomiting, hypoxemia and pruritus in the four groups (p > 0.05). There was no statistically significant visceral traction response or fentanyl use in the four groups (p > 0.05). Phenylephrine dosing was significantly higher in Group RD2 and RD3 than in Group R (p < 0.05), and there was no significant difference in phenylephrine dosing between Group RD1 and Group R (p > 0.05). There were no statistical differences in postnatal Apgar scores (1 min, 5 min after birth) (p > 0.05). The postoperative concentrations of ß-endorphin (ß-EP), cortisol (Cor) and tumor necrosis factor-α (TNF-α) in the Group RD1, RD2, and RD3 were lower than that in Group R (p < 0.05). Conclusion: Intrathecal 5µg of dexmedetomidine as an adjuvant to ropivacaine relieved intraoperative chills, did not increase intraoperative and postoperative adverse effects, did not increase the amount of intraoperative vasoconstrictor used, and reduced intraoperative stress reactions as well as prolonged the duration of maternal sensory and motor block, so this dose is appropriate for cesarean section. Clinical Trial Registration: [www.chictr.org.cn/], identifier [ChiCTR2200056052].