DNA-Guided Extracellular-Vesicle Metallization with High Catalytic Activity for Accurate Diagnosis of Pulmonary Nodules.

Sensitive and specific analysis of extracellular vesicles (EVs) offers a promising minimally invasive way to identify malignant pulmonary nodules from benign lesions. However, accurate analysis of EVs is subject to free target proteins in blood samples, which compromises the clinical diagnosis value of EVs. Here a DNA-guided extracellular-vesicle metallization (DEVM) strategy is described for ultrasensitive and specific analysis of EV protein biomarkers and classification of pulmonary nodules. The facile DEVM process mainly includes the incorporation of DNA labeled with cholesterol and thiol groups into EV membranes and subsequent deposition of Au3+ and Pt4+ to allow the DNA-functionalized EVs to be encapsulated with AuPt nanoshells. It is found that the synthesized AuPt-metallized EVs possess extrinsic peroxidase-like activity. Utilizing the feature of the catalytic metal nanoshells just growth on the EV membranes, the DEVM method enables multiparametric recognition of target proteins and EV membranes and can produce an amplified colorimetric signal, avoiding the interference of free proteins. By profiling four surface proteins of EVs from 48 patients with pulmonary nodules, the highest area under the receiver operating characteristic curve (0.9983) is obtained. Therefore, this work provides a feasible EVs analysis tool for accurate pulmonary nodules management.

CPA-Cas12a-based lateral flow strip for portable assay of Methicillin-resistant Staphylococcus aureus in clinical sample.

The rapid and accurate identification of methicillin-resistant Staphylococcus aureus at an early antibiotic therapy stage would be benefit to disease diagnosis and antibiotic selection. Herein, we integrated cross-priming amplification (CPA) and CRISPR/Cas 12a (designated as CPA-Cas 12a) systems to establish a sensitive and efficient lateral flow assay to detect methicillin-resistant Staphylococcus aureus. This assay relies on the CPA isothermal nucleic acid amplification strategy which can amplify the DNA extracted from Staphylococcus aureus and accompanying the indiscriminately trans-cleavage process of Cas 12a/CrRNA duplex after recognizing specific sequence. Taking the advantage of reporter and high turnover Cas 12a activity, a dramatic change in response was achieved to produce a significant increase in the analytical sensitivity. The signal conversion and output were realized using a lateral flow strip to achieve field-deployable detection. Furthermore, this bioassay was accommodated with a microfluidic device to realize automatically portable detection. This proposed assay completed within 30 min with the detection limit of 5 CFU mL-1, was verified by testing bacterial suspension and 202 clinical samples. Given the high sensitivity, specificity and efficiency, this colorimetric readout assay through strip could be further promoted to the clinical diagnosis, clinical medication of multidrug-resistant bacteria.

NH4I-Triggered [4 + 2] Annulation of α,ß-Unsaturated Ketoxime Acetates with N-Acetyl Enamides for the Synthesis of Pyridines.

The NH4I-triggered formal [4 + 2] annulation of α,ß-unsaturated ketoxime acetates with N-acetyl enamides has been developed. The current protocol employs electron-rich enamides as C2 synthons and enables the efficient and straightforward construction of polysubstituted pyridines in moderate to good yields based on metal-free systems. The reaction tolerates a wide range of functional groups and represents an alternate route toward the synthesis of pyridine derivatives.

Copper-Catalyzed N-O Cleavage of α,ß-Unsaturated Ketoxime Acetates toward Structurally Diverse Pyridines.

The copper-catalyzed [4 + 2] annulation of α,ß-unsaturated ketoxime acetates with 1,3-dicarbonyl compounds for the synthesis of three classes of structurally diverse pyridines has been developed. This method employs 1,3-dicarbonyl compounds as C2 synthons and enables the synthesis of multifunctionalized pyridines with diverse electron-withdrawing groups in moderate to good yields. The mechanistic investigation suggests that the reactions proceed through an ionic pathway.

Production and characterization of monoclonal antibodies against Toxoplasma gondii ROP18 with strain-specific reactivity.

The rhoptry kinase 18 of Toxoplasma gondii (TgROP18) has been identified as a key virulence factor that allows the parasite to escape from host immune defences and promotes its proliferation in host cells. Although much research is focused on the interaction between host cells and TgROP18, the development of monoclonal antibodies (mAbs) against TgROP18 has not been reported till date. To produce mAbs targeting TgROP18, two hybridomas secreting mAbs against TgROP18, designated as A1 and T2, were generated using cell fusion technology. The subtypes of the A1 and T2 mAbs were identified as IgG3 λ and IgM κ, and peptide scanning revealed that the core sequences of the antigenic epitopes were 180LRAQRRRSELVFE192 and 351NYFLLMMRAEADM363, respectively. The T2 mAb specifically reacted with both T. gondii type I and Chinese I, but not with T. gondii type II, Plasmodium falciparum or Schistosoma japonicum. Finally, the sequences of heavy chain and light chain complementarity-determining regions of T2 were amplified, cloned and characterized, making the modification of the mAb feasible in the future. The development of mAbs against TgROP18 would aid the investigation of the molecular mechanisms underlying the modulation of host cellular functions by TgROP18, and in the development of strategies to diagnose and treat Toxoplasmosis.

Engineering high-performance hairpin stacking circuits for logic gate operation and highly sensitive biosensing assay of microRNA.

Recently, hairpin stacking circuits (HSC) based on toehold-mediated strand displacement have been engineered to detect nucleic acids and proteins. However, the three metastable hairpins in a HSC system can potentially react non-specifically in the absence of a catalyst, limiting its practical application. Here, we developed a unique hairpin design guideline to eliminate circuit leakage of HSC, and the high-performance HSC was successfully implemented on logic gate building and biosensing. We began by analyzing the sources of circuit leakage and optimizing the toehold lengths of hairpins in the HSC system based on the surface plasmon resonance (SPR) technique. Next, a novel strategy of substituting two nucleotides in a specific domain, termed 'loop-domain substitution', was introduced to eliminate leakages. We also systematically altered the positions and numbers of the introduced substitutions to probe their potential contribution to circuit leakage suppression. Through these efforts, the circuit leakage of HSC was significantly reduced. Finally, by designing different DNA input strands, the logic gates could be activated to achieve the output signal. Using miRNA as a model analyte, this strategy could detect miRNA down to pM levels with minimized circuit leakage. We believe these work indicate significant progress in the DNA circuitry.

Toxoplasma gondii inhibits differentiation of C17.2 neural stem cells through Wnt/ß-catenin signaling pathway.

Toxoplasma gondii is a major cause of congenital brain disease. T. gondii infection in the developing fetus frequently results in major neural developmental damage; however, the effects of the parasite infection on the neural stem cells, the key players in fetal brain development, still remain elusive. This study is aiming to explore the role of T. gondii infection on differentiation of neural stem cells (NSCs) and elucidate the underlying molecular mechanisms that regulate the inhibited differentiation of NSCs induced by the infection. Using a differentiation medium, i.e. , DMEM: F12 (1:1 mixture) supplemented with 2% N2, C17.2 neural stem cells (NSCs) were able to differentiate to neurons and astrocytes, respectively evidenced by immunofluorescence staining of differentiation markers including ßIII-tubulin and glial fibrillary acidic protein (GFAP). After 5-day culture in the differentiation medium, the excreted-secreted antigens of T. gondii (Tg-ESAs) significantly down-regulated the protein levels of ßIII-tubulin and GFAP in C17.2 NSCs in a dose-dependent manner. The protein level of ß-catenin in the nucleus of C17.2 cells treated with both wnt3a (a key activator for Wnt/ß-catenin signaling pathway) and Tg-ESAs was significantly lower than that in the cells treated with only wnt3a, but significantly higher than that in the cells treated with only Tg-ESAs. In conclusion, the ESAs of T. gondii RH blocked the differentiation of C17.2 NCSs and downregulated the expression of ß-catenin, an essential component of Wnt/ß-catenin signaling pathway. The findings suggest a new mechanism underlying the neuropathogenesis induced by T. gondii infection, i.e. inhibition of the differentiation of NSCs via blockade of Wnt/ß-catenin signaling pathway, such as downregulation of ß-catenin expression by the parasite ESAs.

Toxoplasma gondii induce apoptosis of neural stem cells via endoplasmic reticulum stress pathway.

Toxoplasma gondii is a major cause of congenital brain disease; however, the underlying mechanism of neuropathogenesis in brain toxoplasmosis remains elusive. To explore the role of T. gondii in the development of neural stem cells (NSCs), NSCs were isolated from GD14 embryos of ICR mice and were co-cultured with tachyzoites of T. gondii RH strain. We found that apoptosis levels of the NSCs co-cultured with 1×106 RH tachyzoites for 24 and 48 h significantly increased in a dose-dependent manner, as compared with the control. Western blotting analysis displayed that the protein level of C/EBP homologous protein (CHOP) was up-regulated, and caspase-12 and c-Jun N-terminal kinase (JNK) were activated in the NSCs co-cultured with the parasites. Pretreatment with endoplasmic reticulum stress (ERS) inhibitor (TUDCA) and caspase-12 inhibitor (Z-ATAD-FMK) inhibited the expression or activation of the key molecules involved in the ERS-mediated apoptotic pathway, and subsequently decreased the apoptosis levels of the NSCs induced by the T. gondii. The findings here highlight that T. gondii induced apoptosis of the NSCs through the ERS signal pathway via activation of CHOP, caspase-12 and JNK, which may constitute a potential molecular mechanism responsible for the cognitive disturbance in neurological disorders of T. gondii.

ZNF695, A Potential Prognostic Biomarker, Correlates with Immune Infiltrates in Cervical Squamous Cell Carcinoma and Endocervical Adenocarcinoma: Bioinformatic Analysis and Experimental Verification.

BACKGROUND: The role of Zinc Finger Protein 695 (ZNF695) is unclear in cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC). OBJECTIVE: The objective of this study was to conduct a comprehensive analysis and experimental validation of ZNF695 in CESC. METHODS: The study investigated the expression of ZNF695 in both pan-cancer and CESC, utilizing data from The Cancer Genome Atlas (TCGA) database to assess its diagnostic value. The present study investigated the association between ZNF695 expression levels and clinical characteristics, as well as prognosis, in patients with CESC. The study explored potential regulatory networks involving ZNF695, including its association with immune infiltration, immune score, stemness index based on mRNA expression (mRNAsi), and drug sensitivity in CESC. We explored the expression of ZNF695 in CESC single cells. ZNF695 expression was validated using GSE29570. RESULTS: ZNF695 was found to be aberrantly expressed in pan-cancer and CESC. There was a significant correlation observed between an elevated level of ZNF695 expression in patients with CESC and histological grade (p = 0.017). Furthermore, a strong association was found between high ZNF695 expression in CESC patients and poorer overall survival (OS) (HR: 1.87; 95% CI: 1.17-3.00; p = 0.009), Progression-free Survival (PFS) (HR: 1.86; 95% CI: 1.16-2.98; p = 0.010), and Disease-specific Survival (DSS) (HR: 1.98; 95% CI: 1.15-3.42; p = 0.014). The expression of ZNF695 in CESC patients (p = 0.006) was identified as an independent prognostic determinant. ZNF695 was associated with steroid hormone biosynthesis, oxidative phosphorylation, and so on. ZNF695 expression correlated with immune infiltration, immune score, and mRNAsi in CESC. ZNF695 expression significantly and negatively correlated with AICA ribonucleotide, BIX02189, QL-XI-92, STF-62247, and SNX-2112 in CESC. ZNF695 gene was upregulated in CESC tissues and cell lines. ZNF695 was significantly upregulated in the CESC cell lines. CONCLUSION: ZNF695 may be a potential prognostic biomarker and immunotherapeutic target for CESC patients.

Network analysis of perception of exercise benefits/barriers and kinesiophobia among patients with cardiovascular diseases.

BACKGROUND: Perception of exercise benefits/barriers and kinesiophobia are important predictors of low exercise behaviors in patients with cardiovascular diseases (CVDs). Little is known about the complex intercorrelations between different components of perception of exercise benefits/barriers and kinesiophobia. OBJECTIVES: To identify the central components of kinesiophobia and to explore the interconnectedness between perception of exercise benefits/barriers and kinesiophobia. METHODS: A total of 258 patients with CVDs were recruited in this study. The Tampa Scale for Kinesiophobia Heart and the Exercise Benefits/Barriers Scale were used to assess kinesiophobia and perception of exercise benefits/barriers. R software was used to visualize the networks and analyze the centrality of the networks. The index "expected influence" and "bridge expected influence" were employed to identify the central components and the bridge components of the networks. RESULTS: In the item network of kinesiophobia, three items ("It is really not safe for a person in my condition to be physically active/exercise", "I cannot do the same things as others because there is a too big risk that I will get heart problems", and "If I tried to be physically active/exercise my heart problem would increase") had the highest expected influence. In the exercise benefits/barriers-kinesiophobia network, the dimension of physical exertion had the highest positive bridge expected influence, while psychological outlook had highest negative value. CONCLUSIONS: The three central components of kinesiophobia and the two bridge components (perception exercise barriers of physical exertion and perception exercise benefits of psychological outlook) should be targeted in specific intervention for relieving kinesiophobia and further promoting exercise behaviors.

Needs and Constraints for Cardiac Rehabilitation Among Patients with Coronary Heart Disease Within a Community-Based Setting: A Study Based on Focus Group Interviews.

Objective: This study aimed to explore the needs and constraints to cardiac rehabilitation (CR) among patients diagnosed with coronary heart disease (CHD) in a community-based setting, and thereby facilitating the implementation of effective CR programs for this population. Methods: Focus group interviews were used as the primary research methodology. A total of 11 community-dwelling individuals diagnosed with CHD were selected from a community hospital to participate in in-depth interviews, aiming to discern and analyze their requirements and constraints experienced concerning medical resources and healthcare agency. The textual data underwent examination using Colaizzi's method of descriptive data analysis. Results: Deficits existed in the perceptions of patients with CHD within a community-based setting about their condition and CR, and in the social support for this disease. Patients expressed expectations for professional guidance during CR, gained an understanding about the beneficial effects of emotional stability on cognitive function. Patients expressed their thoughts and feelings regarding the diversity of physical exercise options. Two main themes and seven sub-themes were identified: (a) "Insufficient CR resources for patients": Lack of awareness about CHD; inadequate knowledge about secondary prevention/CR; insufficient support from family and friends. (b) "Patient CR initiative": Patient self-adjustment; expectation of professional rehabilitation guidance; stable emotions improving cognition; diverse attitudes and awareness of exercise. Conclusion: For more effective CR, community-based medical teams should provide more comprehensive and individualized rehabilitation programs. They should focus on individual variations and preferences of patients, as well as enhance the autonomy of patients and improve their self-care ability through effective empowerment measures.

Advanced Nanoencapsulation-Enabled Ultrasensitive Analysis: Unraveling Tumor Extracellular Vesicle Subpopulations for Differential Diagnosis of Hepatocellular Carcinoma via DNA Cascade Reactions.

Tumor-derived extracellular vesicles (tEVs) hold immense promise as potential biomarkers for the precise diagnosis of hepatocellular carcinoma (HCC). However, their clinical translation is hampered by their inherent characteristics, such as small size and high heterogeneity and complex environment, including non-EV particles and normal cell-derived EVs, which prolong separation procedures and compromise detection accuracy. In this study, we devised a DNA cascade reaction-triggered individual EV nanoencapsulation (DCR-IEVN) strategy to achieve the ultrasensitive and specific detection of tEV subpopulations via routine flow cytometry in a one-pot, one-step fashion. DCR-IEVN enables the direct and selective packaging of multiple tEV subpopulations in clinical serum samples into flower-like particles exceeding 600 nm. This approach bypasses the need for EV isolation, effectively reducing interference from non-EV particles and nontumor EVs. Compared with conventional analytical technologies, DCR-IEVN exhibits superior efficacy in diagnosing HCC owing to its high selectivity for tEVs. Integration of machine learning algorithms with DCR-IEVN resulted in differential diagnosis accuracy of 96.7% for the training cohort (n = 120) and 93.3% for the validation cohort (n = 30), effectively distinguishing HCC, cirrhosis, and healthy donors. This strategy offers a streamlined workflow and rapid assay completion and requires only small-volume serum samples and routine clinical devices, facilitating the clinical translation of tEV-based tumor diagnosis.

Comprehensive analysis of TLX2 in pan cancer as a prognostic and immunologic biomarker and validation in ovarian cancer.

T cell leukemia homeobox 2 (TLX2) plays an important role in some tumors. Bioinformatics and experimental validation represent a useful way to explore the mechanisms and functions of TLX2 gene in the cancer disease process from a pan cancer perspective. TLX2 was aberrantly expressed in pan cancer and cell lines and correlated with clinical stage. High TLX2 expression was significantly associated with poor overall survival in COAD, KIRC, OC, and UCS. The greatest frequency of TLX2 alterations in pan cancer was amplification. Alterations of NXF2B, MSLNL, PCGF1, INO80B-WBP1, LBX2-AS1, MRPL53, LBX2, TTC31, WDR54, and WBP1 co-occurred in the TLX2 alteration group. PFS was significantly shorter in the TLX2-altered group (n = 6) compared to the TLX2-unaltered group (n = 400). Methylation levels of TLX2 were high in 17 tumors. TLX2 expression was associated with MSI in seven tumors and TMB in five tumors. TLX2 expression was associated with immune infiltration and immune checkpoint genes. TLX2 may be associated with some pathways and chemoresistance. We constructed a possible competing endogenous RNA (ceRNA) network of LINC01010/miR-146a-5p/TLX2 in OC. TLX2 expression was significantly upregulated in ovarian cancer cell lines compared to ovarian epithelial cell lines. Aberrant expression of TLX2 in pan cancer may promote tumorigenesis and progression through different mechanisms. TLX2 may represent an important therapeutic target for human cancers.

Association of Three SNPs Loci of Kelch-Like-ECH-Associated Protein 1 (Human) with Tuberculosis in Chinese Han Population.

Purpose: Progression from latent tuberculosis infection (LTBI) to pulmonary TB (PTB) was associated with genetic polymorphisms, but there were limited genetic polymorphism data on LTBI and PTB. We aimed at examining the association of KEAP1 gene polymorphisms with PTB and LTBI. Patients and Methods: PTB patients and close contacts of PTB patients were recruited from West China Hospital of Sichuan University. After obtaining the patient's consent, we draw 2-5mL of blood from the patient's peripheral vein. Tag-SNPs of KEAP1 were chosen according to previous studies. The genotyping was done by improved multiplex ligase detection reaction (iMLDR). We used logistic regression to assess the association of SNPs with LTBI/PTB, with sex and age as covariates. Results: A total of 209 PTB patients, 201 LTBI, and 204 HCS were included in the present study. Three Tag-SNPs were included in this study. Significant association was found for KEAP1 rs1048290 between LTBI and HCS. Compared with the KEAP1 rs1048290 CC genotype, genotype GC had an 38% decreased risk for development LTBI (P = 0.043, OR = 0.62, 95% CI: 0.039-0.98). We also found that SNPs in KEAP1 were significantly related to PTB compared to LTBI. Compared with the rs11545829G allele, allele A had an 30% decreased risk for development PTB (P = 0.034, OR = 0.70, 95% CI: 0.51-0.97). We also found the rs11668429 polymorphism was related to PTB. Compared with TT, GT had a significantly increased risk of LTBI developing into PTB (P = 0.041, OR = 1.68, 95% CI: 1.02-2.77). Conclusion: Our study suggested that KEAP1 polymorphisms were significantly related to susceptibility to PTB and LTBI subjects.

Single-Step and Highly Sensitive Imaging of Exosomal PD-L1 through Aptamer-Activated Cascade Primer Exchange Reaction-Generated Branched DNA Nanostructures.

Exosomal PD-L1 plays a critical role in tumor progress and immunotherapy. However, accurately analyzing exosomal PD-L1 is greatly limited by the small-sized and free-floating nature of exosomes and the few proteins each exosome carries. We described herein a single-step and highly sensitive method, termed aptamer-triggered cascade primer exchange reaction (PER)-generated branched DNA nanostructures, for the quantification and imaging of exosomal PD-L1. The presence of exosomal PD-L1 converted the conformation of the recognition probe, accompanied by the exposure of primer 1. Then, primer 1 actuated the cascade PER, which generated branched DNA nanostructures containing numerous G-quadruplex for binding to thioflavin T (ThT) dye, leading to an amplified fluorescence signal. Profiting from directly growing branched DNA nanostructures on the surface of exosomes, the size of exosomes was enlarged and the movement of exosomes was limited, achieving the imaging of exosomal PD-L1 by conventional optical microscopy in a wash- and label-free fashion. Analyzing exosomal PD-L1 from serum samples of 15 cancer patients and 15 healthy volunteers demonstrated that this simple strategy could distinguish NSCLC patients from healthy donors with high clinical accuracy. Therefore, the developed assay has great potential as a transformative diagnostic toolkit for cancer detection and immunotherapy monitoring.

miR-145-5p exerts anti-tumor effects in diffuse large B-cell lymphoma by regulating S1PR1/STAT3/AKT pathway.

To investigate the molecular mechanism of miR-145-5p in diffuse large B-cell lymphoma (DLBCL) tissues and cells. The tissues from patients with DLBCL were collected for RT-qPCR or immunohistochemistry. Cell viability, proliferation, migration, invasion, the relationship between miR-145-5p and S1PR1, and proteins related pathway were detected using CCK-8, BrdU staining, Transwell assay, dual luciferase report assay, and western blotting, respectively. The results showed that miR-145-5p was down-regulated and positively correlated with the survival of DLBCL patients. Overexpression of miR-145-5p inhibited cell proliferation, migration, and invasion in cell model. miR-145-5p directly targeted S1PR1. miR-145-5p down-regulated S1PR1, p-AKT/AKT, and p-STAT3 expression. The reduction of miR-145-5p-induced cell movement was reversed by S1PR1 overexpression. Moreover, S1PR1-induced addition of cell growth was clearly alleviated in LY294002 or S3I-201 treated cells. S1PR1 was up-regulated in the tissues of DLBCL patients. In conclusion, miR-145-5p regulated DLBCL cell growth and movement through suppressing S1PR1/STAT3/AKT pathway.

Fluorescence spectroscopy and molecular docking analysis of the binding of Lactobacillus acidophilus GIM1.208 ß-glucosidase with quercetin glycosides.

Lactobacillus acidophilus is an important probiotic. The ß-glucosidase produced by L. acidophilus GIM1.208 can transform quercetin glycosides of Rosa roxburghii Tratt to release quercetin and improve the functional activity of raw materials. Understanding the interaction and the characteristics of the two will lay a theoretical foundation for the site-directed transformation and functional application of the catalytic active site of enzymes. In our study, using the heterologously expressed and highly stable, purified L. acidophilus GIM1.208 BGL as the strain, the representative quercetin in ß-glucosidase and Rosa roxburghii Tratt was preliminarily predicted and explored using ultraviolet-visible absorption spectroscopy. Fluorescence spectroscopy combined with molecular docking was used to determine the interaction characteristics of the glycoside substrates, rutin (Rut) and isoquercitrin (Iso). Results from molecular docking showed that Asp159, Arg56, Iso294, Phe292, and Gly25 were the main residues of ß-glucosidase and Rut. Arg56 was found to be the most crucial residue of ß-glucosidase and isoquercitrin; the interaction between Rut and Iso and ß-glucosidase was mainly driven by hydrogen bonding. The combined free energy of ß-glucosidase and Iso was found to be -182.10 kcal/mol, while that of ß-glucosidase and Rut was -32.37 kcal/mol. The results of fluorescence spectroscopy showed that the fluorescence intensity of ß-glucosidase decreased with an increase in Rut and Iso concentrations. This interaction made ß-glucosidase quench endogenous fluorescence, which was static quenching. The binding constants of Rut and Iso with ß-glucosidase were determined to be 0.50×107 and 0.31×107 L/mol, respectively, indicating that rutin had a stronger affinity when interacting with ß-glucosidase. These findings were consistent with our prediction results determined using molecular docking studies.

Target-driven rolling walker based electrochemical biosensor for ultrasensitive detection of circulating tumor DNA using doxorubicin@tetrahedron-Au tags.

In this study, a multiple-legged and highly integrated DNA rolling walker based electrochemical biosensor was developed for ultrasensitive ctDNA analysis through rolling circle amplification (RCA) with doxorubicin@tetrahedron-Au (DOX@TDN-Au) as electrochemical indicator. Upon target-driven RCA, the multiple-legged walker could move along with the predesigned track by strand displacement reactions, resulting in numbers of legs binding irreversibly to iStep probes. The binding of massive legs to iStep probes could effectively impede DOX@TDN-Au tags binding on the surface of sensor and then reached a "signal off" state. Benefiting from the highly amplified efficiency of rolling walker machine and DOX@TDN-Au tags, the established biosensor performed high sensitivity for target detection with a low limit of detection down to 0.29 fM. Moreover, the target ctDNA could hybridize with the ring and capture probe simultaneously, greatly enhancing the specificity of the developed biosensing method. Thus, this biosensing method is a promising tool for detection of ctDNA in the field of clinical diagnostic and tumor progression assessment.

[Effect of moxibustion on clinical symptoms, peripheral inflammatory indexes and T lymphocyte subsets in COVID-19 patients].

OBJECTIVE: To explore the therapeutic effect and the mechanism of the adjuvant treatment with moxibustion on coronavirus disease 2019 (COVID-19). METHODS: A total of 95 patients with COVID-19 were randomly divided into a moxibustion group (45 cases) and a basic treatment group (50 cases). The routine treatment of western medicine was applied in the patients of both groups. In the moxibustion group, on the base of the treatment of western medicine, moxibustion was applied to Dazhui (GV 14), Feishu (BL 13), Qihai (CV 6) and Zusanli (ST 36), once daily and consecutively for 14 days. At the end of treatment courses, clinical symptom scores for cough, asthmatic breathing, chest oppression and short breath, as well as their remission rates were compared between the two groups before and after treatment. Before and after treatment, the white blood cell (WBC) count, the levels of c-reactive protein (CRP) and interleukin-6 (IL-6) and the absolute number of T lymphocyte subsets, i.e. , and of the peripheral blood were compared in the patients between the two groups. The principal component analysis was adopted to analyze the common data extracted from the above 10 clinical indexes variables and comprehensively evaluate the differences in the therapeutic effect of two regimens. RESULTS: The clinical symptom scores were all decreased after treatment in both of the moxibustion group and the basic treatment group as compared with those before treatment (P<0.05). After treatment, the clinical symptom scores of cough, chest oppression and asthmatic breathing in the moxibustion group were lower significantly than those in the basic treatment group (P<0.05) and the remission rates of cough, chest oppression and asthmatic breathing were higher than the basic treatment group (P<0.05). After treatment, WBC count was increased as compared with that before treatment in either group (P<0.05) and the levels of CRP and IL-6 in the moxibustion group were reduced as compared with those before treatment (P<0.05). The reducing range of IL-6 level in the moxibustion group was larger than the basic treatment group (P<0.05). After treatment, the absolute number of , and T lymphocytes was increased as compared with that before treatment in the moxibustion group (P<0.05), and its increase range was larger than the basic treatment group (P<0.05). The difference value was 33.38 for the score of comprehensive evaluation before and after treatment in the moxbustion group, higher obviously than 8.91 in the basic treatment group. CONCLUSION: On the base of the routine treatment with western medicine, moxibustion therapy supplemented relieves the clinical symptoms, reduces the levels of inflammatory indexes, i.e. IL-6 and CRP as well as improves the absolute number of peripheral T lymphocyte subsets. The clinical therapeutic effect of such regimen with moxibustion supplemented is significantly better than the simple routine treatment of western medicine.

CD41-deficient exosomes from non-traumatic femoral head necrosis tissues impair osteogenic differentiation and migration of mesenchymal stem cells.

Non-traumatic osteonecrosis of the femoral head (ONFH) is clinically a devastating and progressive disease without an effective treatment. Mesenchymal stem cells (MSCs) transplantation has been used to treat ONFH in early stage, but the failure rate of this therapy is high due to the reduced osteogenic differentiation and migration of the transplanted MSCs related with pathological bone tissues. However, the mechanism responsible for this decrease is still unclear. Therefore, we assume that the implanted MSCs might be influenced by signals delivered from pathological bone tissue, where the exosomes might play a critical role in this delivery. This study showed that exosomes from ONFH bone tissues (ONFH-exos) were able to induce GC-induced ONFH-like damage, in vivo and impair osteogenic differentiation and migration of MSCs, in vitro. Then, we analyzed the differentially expressed proteins (DEPs) in ONFH-exos using proteomic technology and identified 842 differentially expressed proteins (DEPs). On the basis of gene ontology (GO) enrichment analysis of DEPs, fold-changes and previous report, cell adhesion-related CD41 (integrin α2b) was selected for further investigation. Our study showed that the CD41 (integrin α2b) was distinctly decreased in ONFH-exos, compared to NOR-exos, and downregulation of CD41 could impair osteogenic differentiation and migration of the MSCs, where CD41-integrin ß3-FAK-Akt-Runx2 pathway was involved. Finally, our study further suggested that CD41-affluent NOR-exos could restore the glucocorticoid-induced decline of osteogenic differentiation and migration in MSCs, and prevent GC-induced ONFH-like damage in rat models. Taken together, our study results revealed that in the progress of ONFH, exosomes from the pathological bone brought about the failure of MSCs repairing the necrotic bone for lack of some critical proteins, like integrin CD41, and prompted the progression of experimentally induced ONFH-like status in the rat. CD41 could be considered as the target of early diagnosis and therapy in ONFH.