Uniform thin ice on ultraflat graphene for high-resolution cryo-EM.

Cryo-electron microscopy (cryo-EM) visualizes the atomic structure of macromolecules that are embedded in vitrified thin ice at their close-to-native state. However, the homogeneity of ice thickness, a key factor to ensure high image quality, is poorly controlled during specimen preparation and has become one of the main challenges for high-resolution cryo-EM. Here we found that the uniformity of thin ice relies on the surface flatness of the supporting film, and developed a method to use ultraflat graphene (UFG) as the support for cryo-EM specimen preparation to achieve better control of vitreous ice thickness. We show that the uniform thin ice on UFG improves the image quality of vitrified specimens. Using such a method we successfully determined the three-dimensional structures of hemoglobin (64 kDa), α-fetoprotein (67 kDa) with no symmetry, and streptavidin (52 kDa) at a resolution of 3.5 Å, 2.6 Å and 2.2 Å, respectively. Furthermore, our results demonstrate the potential of UFG for the fields of cryo-electron tomography and structure-based drug discovery.

Spontaneous Internal Electric Field in Heterojunction Boosts Bifunctional Oxygen Electrocatalysts for Zinc-Air Batteries: Theory, Experiment, and Application.

Heterojunctions are a promising class of materials for high-efficiency bifunctional oxygen electrocatalysts in both oxygen reduction reaction (ORR) and oxygen evolution reaction (OER). However, the conventional theories fail to explain why many catalysts behave differently in ORR and OER, despite a reversible path (* O2 ⇋* OOH⇋* O⇋* OH). This study proposes the electron-/hole-rich catalytic center theory (e/h-CCT) to supplement the existing theories, it suggests that the Fermi level of catalysts determines the direction of electron transfer, which affects the direction of the oxidation/reduction reaction, and the density of states (DOS) near the Fermi level determines the accessibility for injecting electrons and holes. Additionally, heterojunctions with different Fermi levels form electron-/hole-rich catalytic centers near the Fermi levels to promote ORR/OER, respectively. To verify the universality of the e/h-CCT theory, this study reveals the randomly synthesized heterostructural Fe3 N-FeN0.0324 (Fex N@PC with DFT calculations and electrochemical tests. The results show that the heterostructural F3 N-FeN0.0324 facilitates the catalytic activities for ORR and OER simultaneously by forming an internal electron-/hole-rich interface. The rechargeable ZABs with Fex N@PC cathode display a high open circuit potential of 1.504 V, high power density of 223.67 mW cm-2 , high specific capacity of 766.20 mAh g-1 at 5 mA cm-2 , and excellent stability for over 300 h.

An automated detection of epileptic seizures EEG using CNN classifier based on feature fusion with high accuracy.

BACKGROUND: Epilepsy is a neurological disorder that is usually detected by electroencephalogram (EEG) signals. Since manual examination of epilepsy seizures is a laborious and time-consuming process, lots of automatic epilepsy detection algorithms have been proposed. However, most of the available classification algorithms for epilepsy EEG signals adopted a single feature extraction, in turn to result in low classification accuracy. Although a small account of studies have carried out feature fusion, the computational efficiency is reduced due to too many features, because there are also some poor features that interfere with the classification results. METHODS: In order to solve the above problems, an automatic recognition method of epilepsy EEG signals based on feature fusion and selection is proposed in this paper. Firstly, the Approximate Entropy (ApEn), Fuzzy Entropy (FuzzyEn), Sample Entropy (SampEn), and Standard Deviation (STD) mixed features of the subband obtained by the Discrete Wavelet Transform (DWT) decomposition of EEG signals are extracted. Secondly, the random forest algorithm is used for feature selection. Finally, the Convolutional Neural Network (CNN) is used to classify epilepsy EEG signals. RESULTS: The empirical evaluation of the presented algorithm is performed on the benchmark Bonn EEG datasets and New Delhi datasets. In the interictal and ictal classification tasks of Bonn datasets, the proposed model achieves an accuracy of 99.9%, a sensitivity of 100%, a precision of 99.81%, and a specificity of 99.8%. For the interictal-ictal case of New Delhi datasets, the proposed model achieves a classification accuracy of 100%, a sensitivity of 100%, a specificity of 100%, and a precision of 100%. CONCLUSION: The proposed model can effectively realize the high-precision automatic detection and classification of epilepsy EEG signals. This model can provide high-precision automatic detection capability for clinical epilepsy EEG detection. We hope to provide positive implications for the prediction of seizure EEG.

Hydrophilic, Clean Graphene for Cell Culture and Cryo-EM Imaging.

The wettability of graphene is critical for numerous applications but is very sensitive to its surface cleanness. Herein, by clarifying the impact of intrinsic contamination, i.e., amorphous carbon, which is formed on the graphene surface during the high-temperature chemical vapor deposition (CVD) process, the hydrophilic nature of clean graphene grown on single-crystal Cu(111) substrate was confirmed by both experimental and theoretical studies, with an average water contact angle of ∼23°. Furthermore, the wettability of as-transferred graphene was proven to be highly dependent on its intrinsic cleanness, because of which the hydrophilic, clean graphene exhibited improved performance when utilized for cell culture and cryoelectron microscopy imaging. This work not only validates the intrinsic hydrophilic nature of graphene but also provides a new insight in developing advanced bioapplications using CVD-grown clean graphene films.

[Dynamic characteristics of brain networks in patients with irritable bowel syndrome based on functional magnetic resonance imaging].

The disorder of brain-gut interaction is an important cause of irritable bowel syndrome (IBS), but the dynamic characteristics of the brain remain unclear. Since there are many shortcomings for evaluating brain dynamic nature in the previous studies, we proposed a new method based on slope calculation by point-by-point analysis of the data from functional magnetic resonance imaging, and detected the abnormalities of brain dynamic changes in IBS patients. The results showed that compared with healthy subjects, there were dynamic changes in the brain for the IBS patients. After correction by false discovery rate (FDR), significant abnormalities were only found in two functional connections of the right posterior cingulate gyrus linked to left middle frontal gyrus, and the right posterior cingulate gyrus linked to left pallidus. The above results of the brain dynamic analysis were totally different from those of the brain static analysis of IBS patients. Our findings provide novel complementary information for illustrating the central nervous mechanism of IBS and may offer a new direction to explore central target for patients with IBS.

The Triticum Mosaic Virus Internal Ribosome Entry Site Relies on a Picornavirus-Like YX-AUG Motif To Designate the Preferred Translation Initiation Site and To Likely Target the 18S rRNA.

Several viruses encode an internal ribosome entry site (IRES) at the 5' end of their RNA, which, unlike most cellular mRNAs, initiates translation in the absence of a 5' m7GpppG cap. Here, we report a uniquely regulated translation enhancer found in the 739-nucelotide (nt) sequence of the Triticum mosaic virus (TriMV) leader sequence that distinguishes the preferred initiation site from a plethora of IRES-encoded AUG triplets. Through deletion mutations of the TriMV 5' untranslated region (UTR), we show that the TriMV 5' UTR encodes a cis-acting picornaviral Y16-X11-AUG-like motif with a 16-nt polypyrimidine CU-tract (Y16), at a precise, 11-nt distance (X11) from the preferred 13th AUG. Phylogenetic analyses indicate that this motif is conserved among potyviral leader sequences with multiple AUGs. Consistent with a broadly conserved mechanism, the motif could be functionally replaced with known picornavirus YX-AUG motifs and is predicted to function as target sites for the 18S rRNA by direct base pairing. Accordingly, mutations that disrupted overall complementarity to the 18S rRNA markedly reduced TriMV IRES activity, as did the delivery of antisense oligonucleotides designed to block YX-AUG accessibility. To our knowledge, this is the first report of a plant viral IRES YX-AUG motif, and our findings suggest that a conserved mechanism regulates translation for multiple economically important plant and animal positive single-stranded RNA viruses.IMPORTANCE Uncapped viral RNAs often rely on their 5' leader sequences to initiate translation, and the Triticum mosaic virus (TriMV) devotes an astonishing 7% of its genome to directing ribosomes to the correct AUG. Here we uncover a novel mechanism by which a TriMV cis-regulatory element controls cap-independent translation. The upstream region of the functional AUG contains a 16-nt polypyrimidine tract located 11 nt from the initiation site. Based on functional redundancy with similar motifs derived from human picornaviruses, the motif is likely to operate by directing ribosome targeting through base pairing with 18S rRNA. Our results provide the first report of a broad-spectrum mechanism regulating translation initiation for both plant- and animal-hosted picornaviruses.

[Phenotype and Genotype Analysis in Two Pedigrees with Hereditary Coagulation Factor â ª Deficiency].

OBJECTIVE: To analyze the phenotype and genotype in two pedigrees with hereditary coagulation factor Ⅺ (FⅪ) deficiency, and investigate the molecular mechanisms of FⅪ deficiency. METHODS: Two patients with hereditary coagulation FⅪ deficiency were admitted to Chaozhou Central Hospital in Nov 2014 and Jan 2018. The prothrombin time (PT), activated partial thromboplastin time (APTT), FⅪ activity (FⅪ∶C) and FⅪ antigen (FⅪ∶Ag) were tested for phenotypic diagnosis. All the exons and exon-intron boundaries of FⅪ gene of proband were analyzed by PCR and sequencing. The family members were tested for the mutant site of proband. Then the mRNA of FⅪ in the proband was analyzed with RT-PCR. RESULTS: The proband-1 was a 7-year-old boy, PT was 10.7 s and APTT was 97.4 s (reference range: 9-12.8 s; 24-40 s), FⅪ∶C (0.6%) and FⅪ∶Ag<1% (reference range: 65%-150%; 72.1%-122.3%). The proband-2 was a 30-year-old female, and showed the PT (11.7 s), APTT (71.3 s), FⅪ∶C (0.7%) and FⅪ∶Ag<1%. FⅧ∶C, FⅨ∶C and FⅫ∶C of two proband were within the normal range. DNA sequencing showed that the proband-1 had a combined mutation of c.326-1G>A and c.1107C>A (p.Tyr351X) in exon 10. His grandmother, mother and brother had a heterozygous splicing mutation of c.326-1G>A, his grandmother and father had a homozygous mutation of c.1107C>A. FXI mRNA was undetected in the proband-1. The proband-2 had a homozygous mutation of c.841C>T (p.Gln263X) in exon 8, and this mutation was also found in her father, mother, daughter and son. CONCLUSION: The c.326-1G>A, c.1107C>A(p.Tyr351X) and c.841C>T (p.Gln263X) might be the molecular pathogenesis for two probands with hereditary coagulation factor Ⅺ deficiency.

Superclean Growth of Graphene Using a Cold-Wall Chemical Vapor Deposition Approach.

Chemical vapor deposition (CVD) has become a promising approach for the industrial production of graphene films with appealing controllability and uniformity. However, in the conventional hot-wall CVD system, CVD-derived graphene films suffer from surface contamination originating from the gas-phase reaction during the high-temperature growth. Shown here is that the cold-wall CVD system is capable of suppressing the gas-phase reaction, and achieves the superclean growth of graphene films in a controllable manner. The as-received superclean graphene film, exhibiting improved optical and electrical properties, was proven to be an ideal candidate material used as transparent electrodes and substrate for epitaxial growth. This study provides a new promising choice for industrial production of high-quality graphene films, and the finding about the engineering of the gas-phase reaction, which is usually overlooked, will be instructive for future research on CVD growth of graphene.

[Clinical value of cleaved lymphocytes in assisting the diagnosis of pertussis in children].

OBJECTIVE: To study the value of cleaved lymphocytes in peripheral blood smear in assisting the early diagnosis of pertussis. METHODS: Nasopharyngeal swabs and peripheral blood samples were collected from 107 children with pertussis-like disease. PCR-flow fluorescent hybridization was used to detect the nucleic acids of Bordetella pertussis. Based on the detection results, the children were divided into two groups: pertussis (n=52) and non-pertussis (n=55). According to age, the pertussis group was divided into two subgroups: <1 year old (n=42) and ≥1 year old (n=10). According to disease severity, the pertussis group was divided into another two subgroups: mild (n=45) and severe (n=7). An automatic blood cell analyzer was used to determine peripheral blood cell counts. Wright's staining and peroxidase staining were used to observe and count cleaved lymphocytes under a microscope. RESULTS: Cleaved lymphocytes in peripheral blood were round with small cytoplast, less cytoplasm and cleaved or lobulated nuclei. According to the negative peroxidase staining results, these cells were confirmed as lymphocytes. Compared with the non-pertussis group, the pertussis group had significantly higher leukocyte count, lymphocyte percentage, platelet count, and percentage of cleaved lymphocytes (P<0.001). For the children with pertussis, the <1 year old subgroup had significantly higher lymphocyte percentage, platelet count, and percentage of cleaved lymphocytes than the ≥1 year old subgroup (P<0.05). The severe subgroup had slightly higher leukocyte count, lymphocyte percentage, platelet count, and percentage of cleaved lymphocytes than the mild subgroup (P>0.05). CONCLUSIONS: The detection of cleaved lymphocytes combined with peripheral blood cell counts provides a new option for the early diagnosis of pertussis in children.

Bioactive Functionalized Monolayer Graphene for High-Resolution Cryo-Electron Microscopy.

Single-particle cryo-electron microscopy (cryo-EM) has become one of the most essential tools to understand biological mechanisms at molecular level. A major bottleneck in cryo-EM technique is the preparation of good specimens that embed biological macromolecules in a thin layer of vitreous ice. In the canonical cryo-EM specimen preparation method, biological macromolecules tend to be adsorbed to the air-water interface, causing partial denaturation and/or preferential orientations. In this work, we have designed and produced a new type of cryo-EM grids using bioactive-ligand functionalized single-crystalline monolayer graphene membranes as supporting films. The functionalized graphene membrane (FGM) grids exhibit specific binding affinity to histidine (His)-tagged proteins and complexes. In cryo-EM, the FGM grids generate relatively low background for imaging and selectively anchor 20S proteasomes to the supporting film surface, enabling near-atomic-resolution 3D reconstruction of the complex. We envision that the FGM grids could benefit single particle cryo-EM specimen preparation with high reproducibility and robustness, therefore enhancing the efficiency and throughput of high-resolution cryo-EM structural determination.

Copper-Containing Carbon Feedstock for Growing Superclean Graphene.

Chemical vapor deposition (CVD) enables the large-scale growth of high-quality graphene film and exhibits considerable potential for the industrial production of graphene. However, CVD-grown graphene film contains surface contamination, which in turn hinders its potential applications, for example, in electrical and optoelectronic devices and in graphene-membrane-based applications. To solve this issue, we demonstrated a modified gas-phase reaction to achieve the large-scale growth of contamination-free graphene film, i.e., superclean graphene, using a metal-containing molecule, copper(II) acetate, Cu(OAc)2, as the carbon source. During high-temperature CVD, the Cu-containing carbon source significantly increased the Cu content in the gas phase, which in turn suppressed the formation of contamination on the graphene surface by ensuring sufficient decomposition of the carbon feedstock. The as-received graphene with a surface cleanness of about 99% showed enhanced optical and electrical properties. This study opens a new avenue for improving graphene quality with respect to surface cleanness and provides new insight into the mechanism of graphene growth through the gas-phase reaction pathway.

[Novel mutations p.Lys327X and p.Leu424CysfsX8 underlying congenital factor â ª deficiency].

OBJECTIVE: To analyze the phenotype and genetic mutations in a pedigree affected with factor Ⅺ (FⅪ) deficiency. METHODS: Activated partial thromboplastin time (APTT), FⅪ activity (FⅪ:C) and FⅪ antigen (FⅪ:Ag) were determined for the proband and his family members. All exons and exon-intron boundaries of the FⅪ gene of the proband were analyzed by direct sequencing. Suspected mutation was verified in his family members. RESULTS: The proband had APTT of 82.4 s, FⅪ:C of 0.8%, and FⅪ:Ag of <1%. DNA sequencing showed that he has carried c.1033A>T (Lys327X) mutation in exon 10 and c.1325delT (Leu424CysfsX8) mutation in exon 12 of the FⅪ gene. His elder sister, son, daughter, two granddaughters and one grandson were heterozygous carriers of the c.1033A>T mutation, while his older sister and younger brother were heteozygous carriers of the c.1325delT mutation. Analysis using Mutation Taster software showed that both p.Lys327X and p.Leu424CysfsX8 may affect the function of protein and lead to the corresponding disease. CONCLUSION: The novel mutations of Lys327X and Leu424CysfsX8 of the the FⅪ gene probably underlie the pathogenesis of congenital coagulation factor Ⅺ deficiency in this pedigree.

Large-Area Synthesis of Superclean Graphene via Selective Etching of Amorphous Carbon with Carbon Dioxide.

Contamination commonly observed on the graphene surface is detrimental to its excellent properties and strongly hinders its application. It is still a great challenge to produce large-area clean graphene film in a low-cost manner. Herein, we demonstrate a facile and scalable chemical vapor deposition approach to synthesize meter-sized samples of superclean graphene with an average cleanness of 99 %, relying on the weak oxidizing ability of CO2 to etch away the intrinsic contamination, i.e., amorphous carbon. Remarkably, the elimination of amorphous carbon enables a significant reduction of polymer residues in the transfer of graphene films and the fabrication of graphene-based devices and promises strongly enhanced electrical and optical properties of graphene. The facile synthesis of large-area superclean graphene would open the pathway for both fundamental research and industrial applications of graphene, where a clean surface is highly needed.

Ultrafast Broadband Charge Collection from Clean Graphene/CH3NH3PbI3 Interface.

Photocarrier generation in a material, transportation to the material surface, and collection at the electrode interface are of paramount importance in any optoelectronic and photovoltaic device. In the last collection process, ideal performance comprises ultrafast charge collection to enhance current conversion efficiency and broadband collection to enhance energy conversion efficiency. Here, for the first time, we demonstrate ultrafast broadband charge collection achieved simultaneously at the clean graphene/organic-inorganic halide perovskite interface. The clean interface is realized by directly growing perovskite on graphene surface without polymer contamination. The tunable two-color pump-probe spectroscopy, time-resolved photoluminescence spectroscopy, and time-dependent density functional theory all reveal that the clean-interfacial graphene collects band-edge photocarriers of perovskite in an ultrashort time of ∼100 fs, with a current collection efficiency close to 99%. In addition, graphene can extract deep-band hot carriers of perovskite within only ∼50 fs, several orders faster than hot carrier relaxation and cooling in perovskite itself, due to the unique Dirac linear band structure of graphene, indicating a potential high energy conversion efficiency exceeding the Shockley-Queisser limit. Adding other graphene superiority of good transparency, high carrier mobility, and extreme flexibility, clean-interfacial graphene provides an ideal charge collection layer and electrode candidate for future optoelectronic and photovoltaic applications in two dimensions.

Low-Temperature and Rapid Growth of Large Single-Crystalline Graphene with Ethane.

Future applications of graphene rely highly on the production of large-area high-quality graphene, especially large single-crystalline graphene, due to the reduction of defects caused by grain boundaries. However, current large single-crystalline graphene growing methodologies are suffering from low growth rate and as a result, industrial graphene production is always confronted by high energy consumption, which is primarily caused by high growth temperature and long growth time. Herein, a new growth condition achieved via ethane being the carbon feedstock to achieve low-temperature yet rapid growth of large single-crystalline graphene is reported. Ethane condition gives a growth rate about four times faster than methane, achieving about 420 µm min-1 for the growth of sub-centimeter graphene single crystals at temperature about 1000 °C. In addition, the temperature threshold to obtain graphene using ethane can be reduced to 750 °C, lower than the general growth temperature threshold (about 1000 °C) with methane on copper foil. Meanwhile ethane always keeps higher graphene growth rate than methane under the same growth temperature. This study demonstrates that ethane is indeed a potential carbon source for efficient growth of large single-crystalline graphene, thus paves the way for graphene in high-end electronical and optoelectronical applications.

Two-Dimensional (C4H9NH3)2PbBr4 Perovskite Crystals for High-Performance Photodetector.

Two-dimensional (2D) layered hybrid perovskites of (RNH3)2PbX4 (R is an alkyl and X is a halide) have been recently synthesized and exhibited rich optical properties including fluorescence and exciton effects. However, few studies on transport and optoelectronic measurements of individual 2D perovskite crystals have been reported, presumably owing to the instability issue during electronic device fabrications. Here we report the first photodetector based on individual 2D (C4H9NH3)2PbBr4 perovskite crystals, built with the protection and top contact of graphene film. Both a high responsivity (∼2100 A/W) and extremely low dark current (∼10-10 A) are achieved with a design of interdigital graphene electrodes. Our study paves the way to build high-performance optoelectronic devices based on the emerging 2D single-crystal perovskite materials.

A Unique 5' Translation Element Discovered in Triticum Mosaic Virus.

UNLABELLED: Several plant viruses encode elements at the 5' end of their RNAs, which, unlike most cellular mRNAs, can initiate translation in the absence of a 5' m7GpppG cap. Here, we describe an exceptionally long (739-nucleotide [nt]) leader sequence in triticum mosaic virus (TriMV), a recently emerged wheat pathogen that belongs to the Potyviridae family of positive-strand RNA viruses. We demonstrate that the TriMV 5' leader drives strong cap-independent translation in both wheat germ extract and oat protoplasts through a novel, noncanonical translation mechanism. Translation preferentially initiates at the 13th start codon within the leader sequence independently of eIF4E but involves eIF4G. We truncated the 5' leader to a 300-nucleotide sequence that drives cap-independent translation from the 5' end. We show that within this sequence, translation activity relies on a stem-loop structure identified at nucleotide positions 469 to 490. The disruption of the stem significantly impairs the function of the 5' untranslated region (UTR) in driving translation and competing against a capped RNA. Additionally, the TriMV 5' UTR can direct translation from an internal position of a bicistronic mRNA, and unlike cap-driven translation, it is unimpaired when the 5' end is blocked by a strong hairpin in a monocistronic reporter. However, the disruption of the identified stem structure eliminates such a translational advantage. Our results reveal a potent and uniquely controlled translation enhancer that may provide new insights into mechanisms of plant virus translational regulation. IMPORTANCE: Many members of the Potyviridae family rely on their 5' end for translation. Here, we show that the 739-nucleotide-long triticum mosaic virus 5' leader bears a powerful translation element with features distinct from those described for other plant viruses. Despite the presence of 12 AUG start codons within the TriMV 5' UTR, translation initiates primarily at the 13th AUG codon. The TriMV 5' UTR is capable of driving cap-independent translation in vitro and in vivo, is independent of eIF4E, and can drive internal translation initiation. A hairpin structure at nucleotide positions 469 to 490 is required for the cap-independent translation and internal translation initiation abilities of the element and plays a role in the ability of the TriMV UTR to compete against a capped RNA in vitro. Our results reveal a novel translation enhancer that may provide new insights into the large diversity of plant virus translation mechanisms.

Recent advances in batch production of transfer-free graphene.

Large-area transfer-free graphene films prepared via chemical vapor deposition have proved appealing for various applications, with exciting examples in electronics, photonics, and optoelectronics. To achieve their commercialisation, batch production is a prerequisite. Nevertheless, the prevailing scalable synthesis strategies that have been reported are still obstructed by production inefficiencies and non-uniformity. There has also been a lack of reviews in this realm. We present herein a comprehensive and timely summary of recent advances in the batch production of transfer-free graphene. Primary issues and promising approaches for improving the graphene growth rate are first addressed, followed by a discussion of the strategies to guarantee in-plane and batch uniformity for graphene grown on planar plates and wafer-scale substrates, with the design of the target equipment to meet productivity requirements. Finally, potential research directions are outlined, aiming to offer insights into guiding the scalable production of transfer-free graphene.

A robust approach for multi-type classification of brain tumor using deep feature fusion.

Brain tumors can be classified into many different types based on their shape, texture, and location. Accurate diagnosis of brain tumor types can help doctors to develop appropriate treatment plans to save patients' lives. Therefore, it is very crucial to improve the accuracy of this classification system for brain tumors to assist doctors in their treatment. We propose a deep feature fusion method based on convolutional neural networks to enhance the accuracy and robustness of brain tumor classification while mitigating the risk of over-fitting. Firstly, the extracted features of three pre-trained models including ResNet101, DenseNet121, and EfficientNetB0 are adjusted to ensure that the shape of extracted features for the three models is the same. Secondly, the three models are fine-tuned to extract features from brain tumor images. Thirdly, pairwise summation of the extracted features is carried out to achieve feature fusion. Finally, classification of brain tumors based on fused features is performed. The public datasets including Figshare (Dataset 1) and Kaggle (Dataset 2) are used to verify the reliability of the proposed method. Experimental results demonstrate that the fusion method of ResNet101 and DenseNet121 features achieves the best performance, which achieves classification accuracy of 99.18 and 97.24% in Figshare dataset and Kaggle dataset, respectively.

Bifidobacterium improves oestrogen-deficiency-induced osteoporosis in mice by modulating intestinal immunity.

Osteoporosis has become one of the major diseases that threaten the health of middle-aged and elderly people, and with the growth of an ageing population, more and more people are affected by osteoporosis these days. In recent years, intestinal flora has been found to affect the host immune system, and an overactive immune system is closely related to bone resorption. Probiotics can effectively improve bone density and strength, reduce bone loss, and improve osteoporosis, but their mechanism of action and relationship with intestinal microbiota are still unclear. In this study, two strains of Bifidobacterium (Bifidobacterium bifidum FL228.1 and Bifidobacterium animalis subsp. Lactis F1-7) that can alleviate intestinal inflammation were screened based on previous experiments. Through the construction of an ovariectomized mouse model, the improvement of osteoporosis by Bifidobacterium was detected, and the influence of Bifidobacterium on intestinal immunity was explored. The results show that Bifidobacterium treatment significantly improved bone mineral density (BMD), bone volume/total volume ratio (BV/TV), and trabecular number (Tb·N), and effectively suppressed bone loss. Furthermore, Bifidobacterium treatment could inhibit the expression of inflammatory cytokines in the gut, alleviate gut inflammation, and thus suppress excessive osteoclast generation. Its mechanism of action includes factors that protect the mucosal barrier, including occludin, ZO-1, claudin-2, and MUC2, and the reduction of pro-inflammatory M1 macrophages. B. bifidum FL228.1 increased the abundance of beneficial bacteria in the colon, including Lactobacillus and Colidextribacter. B. animalis F1-7 increased the abundance of Bifidobacterium and decreased the abundance of Desulfovibrio and Ruminococcus in the colon. These research findings expand our understanding of the gut-bone axis and provide new guidance for the development of probiotic-based therapies for osteoporosis in the future.