Capture RIC-seq reveals positional rules of PTBP1-associated RNA loops in splicing regulation.

RNA-binding proteins (RBPs) bind at different positions of the pre-mRNA molecules to promote or reduce the usage of a particular exon. Seeking to understand the working principle of these positional effects, we develop a capture RIC-seq (CRIC-seq) method to enrich specific RBP-associated in situ proximal RNA-RNA fragments for deep sequencing. We determine hnRNPA1-, SRSF1-, and PTBP1-associated proximal RNA-RNA contacts and regulatory mechanisms in HeLa cells. Unexpectedly, the 3D RNA map analysis shows that PTBP1-associated loops in individual introns preferentially promote cassette exon splicing by accelerating asymmetric intron removal, whereas the loops spanning across cassette exon primarily repress splicing. These "positional rules" can faithfully predict PTBP1-regulated splicing outcomes. We further demonstrate that cancer-related splicing quantitative trait loci can disrupt RNA loops by reducing PTBP1 binding on pre-mRNAs to cause aberrant splicing in tumors. Our study presents a powerful method for exploring the functions of RBP-associated RNA-RNA proximal contacts in gene regulation and disease.

High-Precision Mapping of Membrane Proteins on Synaptic Vesicles using Spectrally Encoded Super-Resolution Imaging.

The spatial resolution of single-molecule localization microscopy is limited by the photon number of a single switching event because of the difficulty of correlating switching events dispersed in time. Here we overcome this limitation by developing a new class of photoswitching semiconducting polymer dots (Pdots) with structured and highly dispersed single-particle spectra. We imaged the Pdots at the first and the second vibronic emission peaks and used the ratio of peak intensities as a spectral coding. By correlating switching events using the spectral coding and performing 4-9 frame binning, we achieved a 2-3 fold experimental resolution improvement versus conventional superresolution imaging. We applied this method to count and map SV2 and proton ATPase proteins on synaptic vesicles (SVs). The results reveal that these proteins are trafficked and organized with high precision, showing unprecedented level of detail about the composition and structure of SVs.

Intrinsic Blue Fluorescence of 2.0G PAMAM-DCM Polymer Dots and Its Applications for Fe3+ Sensing.

A typical and environment-friendly fluorescent polyamine-amine (PAMAM) features good compatibility and unique surface modification, while it is restricted by a low fluorescence property performance and an unclear fluorescence mechanism. In this work, we prepared blue fluorescent PAMAM polymer dots (PDs) via a simple hydrothermal method based on dichloromethane (DCM) and 2.0G PAMAM. The quantum yield achieved was 32.1%, which was 25 times stronger than that of 2.0G PAMAM due to the lone-pair electron leap of the amine groups, the aggregation of carbonyl groups, as well as the crosslinking induced by DCM inside the PAMAM. In addition, the fluorescent 2.0G PAMAM-DCM PDs show a great Fe3+ sensing property with the detection limit of 56.6 nM, which is much lower than the safety limits (5.36 µM) in drinking water, indicating its great potential for Fe3+ detection in aqueous media.

Capillary-Mediated Single-Cell Dispenser.

Single-cell manipulation, sorting, and dispensing into multiwell plates is useful for single-cell multiomics studies. Here, we develop a single-cell dispenser inspired by electrohydrodynamic jet printing that achieves accurate droplet generation and single-cell sorting and dispensing using fused silica capillary tubing as both the optical detection window and nozzle for droplet dispensing. Parameters that affect droplet dispensing performance-capillary inner and outer diameter, flow rate, applied voltage, and solution properties-were optimized systematically with COMSOL simulations and experimentation. Small (5-10 nL) droplets were obtained by using 100-µm inner diameter and 160-µm outer diameter capillary tubing and allowed efficient encapsulation and dispensing of single cells. We demonstrate an application of this easy-to-assemble single-cell dispenser by sorting and dispensing cells into multiwell plates for single-cell PCR analysis.

Sequential Ensemble-Decision Aliquot Ranking Isolation and Fluorescence In Situ Hybridization Identification of Rare Cells from Blood by Using Concentrated Peripheral Blood Mononuclear Cells.

Isolation and analysis of circulating rare cells is a promising approach for early detection of cancer and other diseases and for prenatal diagnosis. Isolation of rare cells is usually difficult due to their heterogeneity as well as their low abundance in peripheral blood. We previously reported a two-stage ensemble-decision aliquot ranking platform (S-eDAR) for isolating circulating tumor cells from whole blood with high throughput, high recovery rate (>90%), and good purity (>70%), allowing detection of low surface antigen-expressing cancer cells linked to metastasis. However, due to the scarcity of these cells, large sample volumes and large quantities of antibodies were required to isolate sufficient cells for downstream analysis. Here, we drastically increased the number of nucleated cells analyzed by first concentrating peripheral blood mononuclear cells (PBMCs) from whole blood by density gradient centrifugation. The S-eDAR platform was capable of isolating rare cells from concentrated PBMCs (108/mL, equivalent to processing ∼20 mL of whole blood in the 1 mL sample volume used by our instrument) at a high recovery rate (>85%). We then applied the S-eDAR platform for isolating rare fetal nucleated red blood cells (fNRBCs) from concentrated PBMCs spiked with umbilical cord blood cells and confirmed fNRBC recovery by immunostaining and fluorescence in situ hybridization, demonstrating the potential of the S-eDAR system for isolating rare fetal cells from maternal PBMCs to improve noninvasive prenatal diagnosis.

Analysis of culturable airborne fungi in outdoor environments in Tianjin, China.

BACKGROUND: Fungal spores dispersed in the atmosphere may become cause of different pathological conditions and allergies for human beings. A number of studies have been performed to analyze the diversity of airborne fungi in different environments worldwide, and in particular in many urban areas in China. We investigated, for the first time, the diversity, concentration and distribution of airborne fungi in Tianjin city. We sampled 8 outdoor environments, using open plate method, during a whole winter season. Isolated fungi were identified by morphological and molecular analysis. Environmental factors which could influence the airborne fungi concentration (temperature, humidity, wind speed, and air pressure) were monitored and analyzed. The effect of different urban site functions (busy areas with high traffic flow and commercial activities vs. green areas) on airborne fungal diversity was also analyzed. RESULTS: A total of 560 fungal strains, belonging to 110 species and 49 genera of Ascomycota (80 %), Basidiomycota (18 %), and Mucoromycota (2 %) were isolated in this study. The dominant fungal genus was Alternaria (22 %), followed by Cladosporium (18.4 %), Naganishia (14.1 %), Fusarium (5.9 %), Phoma (4.11 %), and Didymella (4.8 %). A fungal concentration ranging from 0 to 3224.13 CFU m- 3 was recorded during the whole study. Permutational multivariate analysis showed that the month was the most influential factor for airborne fungal community structure, probably because it can be regarded as a proxy of environmental variables, followed by wind speed. The two analyzed environments (busy vs. green) had no detectable effect on the air fungal community, which could be related to the relatively small size of parks in Tianjin and/or to the study season. CONCLUSIONS: Our study shed light on the highly diverse community of airborne fungi characterizing the outdoor environments of Tianjin, and clarified the role that different environmental factors played in shaping the analyzed fungal community. The dominant presence of fungi with potential hazardous effect on human health, such as Alternaria, Cladosporium and Naganishia, deserves further attention. Our results may represent a valuable source of information for air quality monitoring, microbial pollution control, and airborne diseases prevention.

MicroRNA-181b-5p modulates tumor necrosis factor-α-induced inflammatory responses by targeting interleukin-6 in cementoblasts.

Tooth cementum is a bone-like mineralized tissue and serves as a microbial barrier against invasion and destruction. Cementum is also responsible for tooth stability and defending pulp from outside stimuli, which is formed by cementoblasts. Although it is crucial for periodontal and periapical diseases, the mechanisms underlying the pathophysiological changes of cementoblasts and their inflammatory responses remain unclear. MiR-181b is found to modulate vascular inflammation and endotoxin tolerance. In this study, miR-181b-5p was downregulated in tumor necrosis factor-α (TNF-α)-stimulated cementoblasts, whereas proinflammatory molecules increased. The mouse periapical lesions have similar results, which imitate an inflammatory environment for cementoblasts in vivo. The bioinformatics analysis and dual luciferase reporter assay suggested that miR-181b-5p targeted interleukin-6 (IL-6). Overexpressing miR-181b-5p negatively regulated IL-6 and proinflammatory chemokine. Western blot analysis and luciferase activity reporter assay verified that miR-181b-5p weakened the NF-κB activity. Hence, miR-181b-5p moderated proinflammatory chemokine production by targeting IL-6 in cementoblasts and NF-κB signaling pathway was involved. Furthermore, miR-181b-5p promoted cementoblast apoptosis, which may enhance the resolution of inflammation. Overall, our data revealed that miR-181b-5p was a negative regulator of TNF-α-induced inflammatory responses in cementoblasts.

Isolating Rare Cells and Circulating Tumor Cells with High Purity by Sequential eDAR.

Isolation and analysis of circulating tumor cells (CTCs) from the blood of patients at risk of metastatic cancers is a promising approach to improving cancer treatment. However, CTC isolation is difficult due to low CTC abundance and heterogeneity. Previously, we reported an ensemble-decision aliquot ranking (eDAR) platform for the rare cell and CTC isolation with high throughput, greater than 90% recovery, and high sensitivity, allowing detection of low surface antigen-expressing cells linked to metastasis. Here we demonstrate a sequential eDAR platform capable of isolating rare cells from whole blood with high purity. This improvement in purity is achieved by using a sequential sorting and flow stretching design in which whole blood is sorted and fluid elements are stretched using herringbone features and the parabolic flow profile being sorted a second time. This platform can be used to collect single CTCs in a multiwell plate for downstream analysis.

A Fluorescence-Activated Single-Droplet Dispenser for High Accuracy Single-Droplet and Single-Cell Sorting and Dispensing.

The ability to sort and dispense droplets accurately is essential to droplet-based single-cell analysis. Here, we describe a fluorescence-activated single-droplet dispenser (FASD) that is analogous to a conventional fluorescence-activated cell sorter, but sorts droplets containing single cells within an oil emulsion. The FASD system uses cytometric detection and electrohydrodynamic actuation-based single-droplet manipulation, allowing droplet isolation and dispensing with high efficiency and accuracy. The system is compatible with multiwell plates and can be integrated with existing microfluidic devices and large-scale screening systems. By enabling sorting based on single-cell reactions such as PCR, this platform will help expand the basis of cell sorting from mainly protein biomarkers to nucleic acid sequences and secreted biomolecules.

A Self-Digitization Dielectrophoretic (SD-DEP) Chip for High-Efficiency Single-Cell Capture, On-Demand Compartmentalization, and Downstream Nucleic Acid Analysis.

The design and fabrication of a self-digitization dielectrophoretic (SD-DEP) chip with simple components for single-cell manipulation and downstream nucleic acid analysis is presented. The device employed the traditional DEP and insulator DEP to create the local electric field that is tailored to approximately the size of single cells, enabling highly efficient single-cell capture. The multistep procedures of cell manipulation, compartmentalization, lysis, and analysis were performed in the integrated microdevice, consuming minimal reagents, minimizing contamination, decreasing lysate dilution, and increasing assay sensitivity. The platform developed here could be a promising and powerful tool in single-cell research for precise medicine.

Dual-Responsive Mechanized Mesoporous Silica Nanoparticles Based on Sulfonatocalixarene Supramolecular Switches.

Mesoporous silica nanoparticles (MSNs) have been functionalized with supramolecular switches, composed of cleavable disulfide bond-containing alkylammonium stalks encircled by water-soluble sulfonatocalix[4,6]arenes, to result in smart mechanized MSNs. Cargo can be encased tightly in the nanopores of these mechanized MSNs in their closed state, but are released efficiently either in response to L-glutathione (GSH), by cleaving the disulfide bonds in the stalks, or in response to pH variation, by turning on the calixarene-based supramolecular switches. The higher concentration of GSH in cancer cell cytosol and the relatively lower pH value of cancer cell lysosome can simultaneously activate the mechanized MSNs, enabling them to release the pre-loaded cargo in place. This sufficient use of the different environment of cancer cells and normal healthy cells can enhance the targeting effect of delivery vehicles and effectively lower the side effects of delivered anti-cancer drugs. In vitro cytotoxicity tests suggest good biocompatibility and low toxicity of these newly developed drug-delivery systems.

Topical Application of Nitrate Ameliorates Skin Fibrosis by Regulating ST2+CD4+ T Cells in Systemic Sclerosis Mouse Model.

Systemic sclerosis (SSc) is characterized by intractable multiorgan fibrosis caused by vascular and immune dysfunction. Currently, effective therapeutic options for patients with SSc are limited. Nitrate, an abundant nutrient in the diet, has been demonstrated to be preventative and therapeutic for several diseases. To determine whether nitrate can slow or reverse SSc progression, topical application of nitrate delivered by dissolving microneedles was used to treat a bleomycin (BLM)-induced dermal fibrosis mouse model. In this study, nitrate considerably attenuated dermal thickness, stiffness, and collagen deposition. Bulk RNA sequencing of skin revealed that Cd4 was a key hub gene in SSc nitrate therapy. Additionally, BLM-induced cytokines and chemokines were inhibited by nitrate, and CD4+ T cells infiltration markedly declined. Il4, Il6, Il13, and Tgfb expression in CD4+ T cells isolated from skin biopsies also significantly decreased. Mechanistically, Il1rl1, a type2 immune response inducer, was markedly repressed in isolated CD4+ T cells and dermal tissues after nitrate treatment. Remarkably, compared with wild type mice, mice lacking Il1rl1 showed impaired transcriptional profiles after intradermal BLM injection. Adoptive transfer of ST2+CD4+ T cells promoted bleomycin-induced Rag2-/- mice dermal fibrosis. Collectively, these findings demonstrate that nitrate targeting ST2+CD4+ T cells is an effective therapeutic option for SSc.

Differences in choroidal responses to near work between myopic children and young adults.

BACKGROUND: Near work is generally considered as a risk factor for myopia onset and progression. This study aimed to investigate the choroidal responses to a brief-period of near work in children and young adults. METHODS: Thirty myopic medical students (aged 18-28 years) and 30 myopic children (aged 8-12 years) participated in this study. The submacular total choroidal area (TCA), luminal area (LA), stromal area (SA), choroidal vascularity index (CVI) and choriocapillaris flow deficit (CcFD), as well as subfoveal choroidal thickness (SFCT) were measured with swept-source optical coherence tomography/optical coherence tomography angiography (SS-OCT/OCTA) before and immediately after 20 min, 40 min, 60 min of near work at a distance of 33 cm. RESULTS: In adults, 20 min of near work induced a significant reduction in SFCT (- 5.1 ± 6.5 µm), LA [(- 19.2 ± 18.6) × 103 µm2], SA [(- 8.2 ± 12.6) × 103 µm2] and TCA [(- 27.4 ± 24.9) × 103 µm2] (all P < 0.01). After 40 min of near work, LA was still reduced [(- 9.4 ± 18.3) × 103 µm2], accompanied with a decreased CVI (- 0.39% ± 0.70%) and an increased CcFD (0.30% ± 0.78%) (all P < 0.05). After 60 min of near work, CVI was still reduced (- 0.28% ± 0.59%), and CcFD was still increased (0.37% ± 0.75%) (all P < 0.05). In children, 20 min of near work induced a significant increase in CcFD (0.55% ± 0.64%), while 60 min of near work induced increases in SA [(7.2 ± 13.0) × 103 µm2] and TCA [(9.7 ± 25.3) × 103 µm2] and a reduction in CVI (- 0.28% ± 0.72%) (all P < 0.05). Children exhibited lower near work-induced LA and TCA reduction than adults, with a mean difference of - 0.86% and - 0.82%, respectively (all P < 0.05). CONCLUSIONS: The temporal characteristics and magnitude of changes of choroidal vascularity and choriocapillaris perfusion during near work was not identical between children and adults. The initial response to near work was observed in choriocapillaris in children, whereas it was observed in the medium- and large-sized vessels in adults. TRIAL REGISTRATION: Clinical Trial Registry (ChiCTR), ChiCTR2000040205. Registered on 25 November 2020, https://www.chictr.org.cn/bin/project/edit?pid=64501 .

Mendelian randomization study reveals a causal relationship between serum iron status and coronary heart disease and related cardiovascular diseases.

Background: Growing observational studies have shown that abnormal systemic iron status is associated with Coronary heart disease (CHD). However, these results from observational studies was not entirely consistent.It remains unclear whether this relationship represents causality.It is necessary to explore the causal relationship between iron status and CHD and related cardiovascular diseases (CVD). Objective: We aimed to investigate the potential casual relationship between serum iron status and CHD and related CVD using a two-sample Mendelian randomization (MR) approach. Methods: Genetic statistics for single nucleotide polymorphisms (SNPs) between four iron status parameters were identified in a large-scale genome-wide association study (GWAS) conducted by the Iron Status Genetics organization. Three independent single nucleotide polymorphisms (SNPs) (rs1800562, rs1799945, and rs855791) aligned with four iron status biomarkers were used as instrumental variables. CHD and related CVD genetic statistics We used publicly available summary-level GWAS data. Five different MR methods random effects inverse variance weighting (IVW), MR Egger, weighted median, weighted mode, and Wald ratio were used to explore the causal relationship between serum iron status and CHD and related CVD. Results: In the MR analysis, we found that the causal effect of serum iron (OR = 0.995, 95% CI = 0.992-0.998, p = 0.002) was negatively associated with the odds of coronary atherosclerosis (AS). Transferrin saturation (TS) (OR = 0.885, 95% CI = 0.797-0.982, p = 0.02) was negatively associated with the odds of Myocardial infarction (MI). Conclusion: This MR analysis provides evidence for a causal relationship between whole-body iron status and CHD development. Our study suggests that a high iron status may be associated with a reduced risk of developing CHD.

Aptamers-functionalized nanoscale MOFs for saxitoxin and tetrodotoxin sensing in sea foods through FRET.

Saxitoxin (STX) and tetrodotoxin (TTX) are widely distributed and extremely harmful marine toxins, it is certainly worth to spend effort to develop facile methods to detect them in sea food for human safety. In this work, two nano-sensors were developed by combining with two zirconium fluorescence Nanoscale metal-organic frameworks (NMOFs) with two emissions and TAMRA-labelled aptamers for STX and TTX sensing, respectively. The recognition of STX and TTX by these nano-sensors could change the structure of aptamer, which caused the blue or green emissions from NMOFs (energy donor) decreased while red emission from TAMRA-labelled aptamers (energy acceptor) increased owing to fluorescence resonance energy transfer (FRET) effect. Based on this strategy, NMOFs-Aptasensor 1 and NMOFs-Aptasensor 2 were developed for the ratiometric detection, with detection limits of 1.17 nM and 3.07 nM for STX and TTX, respectively. Moreover, NMOFs-Aptasensors displayed significant stability, pH-independence, selectivity and NMOFs-Aptasensors were successfully applied in shellfish sample for toxin sensing.

Mendelian randomization study reveals a causal relationship between coronary artery disease and cognitive impairment.

Background: Growing evidence suggests that Coronary artery disease (CAD) is associated with cognitive impairment. However, these results from observational studies was not entirely consistent, with some detecting no such association. And it is necessary to explore the causal relationship between CAD and cognitive impairment. Objective: We aimed to explore the potential causal relationship between CAD and cognitive impairment by using bidirectional two-sample mendelian randomization (MR) analyses. Methods: Instrument variants were extracted according to strict selection criteria. And we used publicly available summary-level GWAS data. Five different methods of MR [random-effect inverse-variance weighted (IVW), MR Egger, weighted median, weighted mode and Wald ratio] were used to explore the causal relationship between CAD and cognitive impairment. Results: There was little evidence to support a causal effect of CAD on cognitive impairment in the forward MR analysis. In the reverse MR analyses, We detect causal effects of fluid intelligence score (IVW: ß = -0.12, 95% CI of -0.18 to -0.06, P = 6.8 × 10-5), cognitive performance (IVW: ß = -0.18, 95% CI of -0.28 to -0.08, P = 5.8 × 10-4) and dementia with lewy bodies (IVW: OR = 1.07, 95% CI of 1.04-1.10, P = 1.1 × 10-5) on CAD. Conclusion: This MR analysis provides evidence of a causal association between cognitive impairment and CAD. Our findings highlight the importance of screening for coronary heart disease in patients of cognitive impairment, which might provide new insight into the prevention of CAD. Moreover, our study provides clues for risk factor identification and early prediction of CAD.

Dietary nitrate improves jaw bone remodelling in zoledronate-treated mice.

Bisphosphonate-related osteonecrosis of the jaw (BRONJ) is a serious complication that occurs in patients with osteoporosis or metastatic bone cancer treated with bisphosphonate. There is still no effective treatment and prevention strategy for BRONJ. Inorganic nitrate, which is abundant in green vegetables, has been reported to be protective in multiple diseases. To investigate the effects of dietary nitrate on BRONJ-like lesions in mice, we utilized a well-established mouse BRONJ model, in which tooth extraction was performed. Specifically, 4 mM sodium nitrate was administered in advance through drinking water to assess the short- and long-term effects on BRONJ. Zoledronate injection could induce severe healing inhibition of the tooth extraction socket, while addition of pretreating dietary nitrate could alleviate the inhibition by reducing monocyte necrosis and inflammatory cytokines production. Mechanistically, nitrate intake increased plasma nitric oxide levels, which attenuated necroptosis of monocytes by downregulating lipid and lipid-like molecule metabolism via a RIPK3 dependent pathway. Our findings revealed that dietary nitrate could inhibit monocyte necroptosis in BRONJ, regulate the bone immune microenvironment and promote bone remodelling after injury. This study contributes to the understanding of the immunopathogenesis of zoledronate and supports the feasibility of dietary nitrate for the clinical prevention of BRONJ.

Causal relationship between sleep traits and cognitive impairment: A Mendelian randomization study.

OBJECTIVE: Observational studies had demonstrated a link between sleep disturbances and cognitive decline. Here, we aimed to investigate the causal association between genetically predicted sleep traits and cognitive impairment using Mendelian randomization (MR). METHODS: Using strict criteria, we selected genetic variants from European ancestry Genome-wide association studies (GWAS) from the Sleep Disorders Knowledge Portal and UK Biobank as instrumental variables for several sleep traits, including insomnia, sleep duration, daytime sleepiness, daytime napping, and chronotype. Summary statistics related to cognitive impairment were derived from five different GWAS, including the Social Science Genetic Association Consortium. The role of self-reported sleep trait phenotypes in the etiology of cognitive impairment was explored using inverse-variance weighted (IVW) tests, MR-Egger tests, and weighted medians, and sensitivity analyses were conducted to ensure robustness. RESULTS: In the main IVW analysis, sleep duration (reaction time: ß = -0.05, 95% CI -0.07 to -0.04, p = 1.93×10-12 ), daytime sleepiness (average cortical thickness: ß = -0.12, 95% CI -0.22 to -0.02, p = 0.023), and daytime napping (fluid intelligence: ß = -0.47, 95% CI -0.87 to -0.07, p = 0.021; hippocampal volume in Alzheimer's disease: ß = -0.99, 95% CI -1.64 to -0.35, p = 0.002) were significantly negatively correlated with cognitive performance. However, any effects of insomnia and chronotype on cognitive impairment were not determined. CONCLUSIONS: Our findings highlighted that focusing on sleep behaviors or distinct sleep patterns-particularly sleep duration, daytime sleepiness, and daytime napping, was a promising approach for preventing cognitive impairment. This study also shed light on risk factors for and potential early markers of cognitive impairment risk factors.

Gut microbiota in combination with blood metabolites reveals characteristics of the disease cluster of coronary artery disease and cognitive impairment: a Mendelian randomization study.

Background: The coexistence of coronary artery disease (CAD) and cognitive impairment has become a common clinical phenomenon. However, there is currently limited research on the etiology of this disease cluster, discovery of biomarkers, and identification of precise intervention targets. Methods: We explored the causal connections between gut microbiota, blood metabolites, and the disease cluster of CAD combined with cognitive impairment through two-sample Mendelian randomization (TSMR). Additionally, we determine the gut microbiota and blood metabolites with the strongest causal associations using Bayesian model averaging multivariate Mendelian randomization (MR-BMA) analysis. Furthermore, we will investigate the mediating role of blood metabolites through a two-step Mendelian randomization design. Results: We identified gut microbiota that had significant causal associations with cognitive impairment. Additionally, we also discovered blood metabolites that exhibited significant causal associations with both CAD and cognitive impairment. According to the MR-BMA results, the free cholesterol to total lipids ratio in large very low density lipoprotein (VLDL) was identified as the key blood metabolite significantly associated with CAD. Similarly, the cholesteryl esters to total lipids ratio in small VLDL emerged as the primary blood metabolite with a significant causal association with dementia with lewy bodies (DLB). For the two-step Mendelian randomization analysis, we identified blood metabolites that could potentially mediate the association between genus Butyricicoccus and CAD in the potential causal links. Conclusion: Our study utilized Mendelian randomization (MR) to identify the gut microbiota features and blood metabolites characteristics associated with the disease cluster of CAD combined with cognitive impairment. These findings will provide a meaningful reference for the identification of biomarkers for the disease cluster of CAD combined with cognitive impairment as well as the discovery of targets for intervention to address the problems in the clinic.