Polydopamine-mediated immobilization of BMP-2 onto electrospun nanofibers enhances bone regeneration.

Dealing with bone defects is a significant challenge to global health. Electrospinning in bone tissue engineering has emerged as a solution to this problem. In this study, we designed a PVDF-b-PTFE block copolymer by incorporating TFE, which induced a phase shift in PVDF fromαtoß, thereby enhancing the piezoelectric effect. Utilizing the electrospinning process, we not only converted the material into a film with a significant surface area and high porosity but also intensified the piezoelectric effect. Then we used polydopamine to immobilize BMP-2 onto PVDF-b-PTFE electrospun nanofibrous membranes, achieving a controlled release of BMP-2. The scaffold's characters were examined using SEM and XRD. To assess its osteogenic effectsin vitro, we monitored the proliferation of MC3T3-E1 cells on the fibers, conducted ARS staining, and measured the expression of osteogenic genes.In vivo, bone regeneration effects were analyzed through micro-CT scanning and HE staining. ELISA assays confirmed that the sustained release of BMP-2 can be maintained for at least 28 d. SEM images and CCK-8 results demonstrated enhanced cell viability and improved adhesion in the experimental group. Furthermore, the experimental group exhibited more calcium nodules and higher expression levels of osteogenic genes, including COL-I, OCN, and RUNX2. HE staining and micro-CT scans revealed enhanced bone tissue regeneration in the defective area of the PDB group. Through extensive experimentation, we evaluated the scaffold's effectiveness in augmenting osteoblast proliferation and differentiation. This study emphasized the potential of piezoelectric PVDF-b-PTFE nanofibrous membranes with controlled BMP-2 release as a promising approach for bone tissue engineering, providing a viable solution for addressing bone defects.

Acacetin inhibits RANKL-induced osteoclastogenesis and LPS-induced bone loss by modulating NFATc1 transcription.

Osteolytic disorders are characterized by impaired bone volume and trabecular structure that leads to severe fragility fractures. Studies have shown that excessive osteoclast activity causes impaired bone microstructure, a sign of osteolytic diseases such as osteoporosis. Approaches of inhibiting osteoclastogenesis and bone resorption specifically could prevent osteoporosis and other osteolytic disorders. Acacetin is a potent molecule extracted from plants with anti-cancer and anti-inflammatory bioactivities. Here, we demonstrated, for the first time, that acacetin repressed osteoclastogenesis, formation of F-actin rings, bone resorption activity, and osteoclast-related gene expression in vitro through modulating ERK, P38, and NF-κB signaling pathways and preventing expression of NFATc1. Micro-CT and H & E staining results indicated that acacetin alleviated LPS-induced osteolysis in vivo. Overall, our findings suggested that acacetin could help to prevent osteoporosis and other osteoclast-related osteolytic disorders.

Molecular mechanisms of mechanical load-induced osteoarthritis.

INTRODUCTION: Mechanical loading enhances the progression of osteoarthritis. However, its molecular mechanisms have not been established. OBJECTIVE: The aim of this review was to summarize the probable mechanisms of mechanical load-induced osteoarthritis. METHODS: A comprehensive search strategy was used to search PubMed and EMBASE databases (from the 15th of January 2015 to the 20th of October 2020). Search terms included "osteoarthritis", "mechanical load", and "mechanism". RESULTS: Abnormal mechanical loading activates the interleukin-1ß, tumour necrosis factor-α, nuclear factor kappa-B, Wnt, transforming growth factor-ß, microRNAs pathways, and the oxidative stress pathway. These pathways induce the pathological progression of osteoarthritis. Mechanical stress signal receptors such as integrin, ion channel receptors, hydrogen peroxide-inducible clone-5, Gremlin-1, and transient receptor potential channel 4 are present in the articular cartilages. CONCLUSION: This review highlights the molecular mechanisms of mechanical loading in inducing chondrocyte apoptosis and extracellular matrix degradation. These mechanisms provide potential targets for osteoarthritis prevention and treatment.

Improving the Detection of Urine Sediment with a Modified Urinalysis Review Procedure.

BACKGROUND: Currently, numerous review procedures are applied to perform the urine sediment examination. Clinical technologists and nephrologists use different procedures for the determination of specimen concentration and for particle counting. These techniques may underestimate the formed elements such as pathological casts (CASTs) and renal tubular epithelial (RTE) cells and might interfere with clinical diagnosis. The aim of this study was to evaluate a modified review procedure for urinary analysis and to narrow the gap between nephrologists and technologists by increasing the detection positivity rate for pathological formed elements. METHODS: We implemented a modified urinalysis procedure between October 2016 and January 2017 based on strict manual microscopic criteria and the currently available equipment. We confirmed the agreement between methods using a review procedure and Sysmex UF-1000i urinary flow cytometer (Pairwise Agreement > 0.88 for WBCs, RBCs, CASTs, and SRCs). Then we derived the review procedure that was based on the optimal sensitivity and specificity as follows: RBC > 26.1/µL, WBC > 37.0/µL, CAST > 1.0/µL, SRC > 8.2/µL, XTAL > 1.5/µL, YLC > 10.0/µL, BACT > 287.5/µL. RESULTS: Of the 317 specimens investigated, 17.4% (26/149) and 31.5% (39/124) of the specimens for RTEs and Path. CASTs, respectively, were correctly detected using the proposed review procedure. Sensitivity and specificity for this procedure was 96.9% and 46.2%, respectively. In addition, we verified the ability of the procedure to detect the pathological elements with technologists and nephrologists and the agreement was satisfactory. CONCLUSIONS: This modified review procedure can significantly improve the quality of urinalysis and reduce the risk of underestimating the detection of pathological particles.

A magnetically recyclable carboxyl-functionalized chitosan composite for efficiently removing methyl orange from wastewater: Isotherm, kinetics, thermodynamic, and adsorption mechanism.

In this study, a kind of magnetically recyclable adsorbent for dyes was synthesized by grafting diethylenetriamine pentaacetate acid (DTPA) to the composite of Fe3O4 microspheres and crosslinked chitosan (CS). The microstructures, molecular structure, crystal structure, and magnetic hysteresis loops of the chitosan matrix adsorbent before and after grafting was characterized. The results suggested that DTPA was covalent bonded with the composite of Fe3O4 microspheres and chitosan. The modified composite has larger specific surface area and can realize rapid solid-liquid separation. Batch experiments were conducted to optimize the parameters affecting the adsorption of methyl orange (MO). The adsorption process could be better described by pseudo-second-order kinetics model and Langmuir isotherm equation, and its saturated adsorption capacity of the modified adsorbents was 1541.5 mg·g-1 at 25 °C, which was 1.40 times of that the unmodified adsorbent (1104.1 mg·g-1). The obtained values of the thermodynamic parameters indicated that the adsorption was a spontaneous process. The regeneration experiment proved the stability and reproducibility of the adsorbent even after five cycles of adsorption-desorption. The primary adsorption mechanism was electrostatic interaction and hydrogen bonding. The adsorbent could be potentially applied for removing dyes from wastewater in wide pH of range, especially acid wastewater.

Butyrate enhances erastin-induced ferroptosis of osteosarcoma cells via regulating ATF3/SLC7A11 pathway.

Osteosarcoma (OS) is a highly fatal bone tumor characterized by high degree of malignancy and early lung metastasis. Traditional chemotherapy fails in improving the efficacy and survival rate of patients with OS. Butyrate (NaBu) has been reported as a new antitumor drug for inhibiting proliferation and inducing apoptosis in various cancer cells. However, the effect of NaBu on the ferroptosis of OS is still unknown. This study aimed to investigate whether NaBu promotes erastin-induced ferroptosis in OS cells and to uncover the underlying mechanism. Here, we found that NaBu significantly enhanced erastin-induced ferroptosis in vitro and in vivo. Compared with the group that erastin used alonely, pre-treating with NaBu exacerbated erastin-meditated GSH depletion, lipid peroxidation, and mitochondrial morphologic changes in OS cells. In a subcutaneous OS model, NaBu combined with erastin significantly reduced tumor growth and increased the levels of 4-HNE. Mechanistically, NaBu downregulated SLC7A11 transcription via regulating ATF3 expression. Overexpression of ATF3 facilitated erastin to induce ferroptosis, while ATF3 knockdown attenuated NaBu-induced ferroptosis sensitivity. In conclusion, our findings revealed a previously unidentified role of NaBu in erastin-induced ferroptosis by regulating SLC7A11, suggesting that NaBu may be a potential therapeutic agent for OS treatment.

Mesenchymal stem cells and macrophages and their interactions in tendon-bone healing.

Tendon-bone insertion injuries (TBI), such as anterior cruciate ligament (ACL) and rotator cuff injuries, are common degenerative or traumatic pathologies with a negative impact on the patient's daily life, and they cause huge economic losses every year. The healing process after an injury is complex and is dependent on the surrounding environment. Macrophages accumulate during the entire process of tendon and bone healing and their phenotypes progressively transform as they regenerate. As the "sensor and switch of the immune system", mesenchymal stem cells (MSCs) respond to the inflammatory environment and exert immunomodulatory effects during the tendon-bone healing process. When exposed to appropriate stimuli, they can differentiate into different tissues, including chondrocytes, osteocytes, and epithelial cells, promoting reconstruction of the complex transitional structure of the enthesis. It is well known that MSCs and macrophages communicate with each other during tissue repair. In this review, we discuss the roles of macrophages and MSCs in TBI injury and healing. Reciprocal interactions between MSCs and macrophages and some biological processes utilizing their mutual relations in tendon-bone healing are also described. Additionally, we discuss the limitations in our understanding of tendon-bone healing and propose feasible ways to exploit MSC-macrophage interplay to develop an effective therapeutic strategy for TBI injuries. The Translational potential of this article: This paper reviewed the important functions of macrophages and mesenchymal stem cells in tendon-bone healing and described the reciprocal interactions between them during the healing process. By managing macrophage phenotypes, mesenchymal stem cells and the interactions between them, some possible novel therapies for tendon-bone injury may be proposed to promote tendon-bone healing after restoration surgery.

TiO2nanotubes-MoS2/PDA-LL-37 exhibits efficient anti-bacterial activity and facilitates new bone formation under near-infrared laser irradiation.

Titanium dioxide (TiO2), as one of the titanium (Ti)-based implants, holds a promise for a variety of anti-bacterial application in medical research. In the current study, a functional molybdenum disulfide (MoS2)/polydopamine (PDA)-LL-37 coating on titanium dioxide (TiO2) implant was prepared. Anodic oxidation and hydrothermal treatment was given to prepare TiO2nanotubes-MoS2/PDA-LL-37 (T-M/P-L). Thein vitroosteogenic effect of T-M/P-L was evaluated by measuring mesenchymal stem cell (MSC) adhesion, proliferation, alkaline phosphatase (ALP) activity, extracellular matrix (ECM) mineralization, collagen secretion and osteoblast-specific messenger RNAs (mRNAs) expression. The determination on the anti-bacterial ability of T-M/P-L was followed. Furthermore, the ability of T-M/P-L to promote bone formationin vivowas evaluated. Near-infrared (NIR) laser irradiation exposure enabled the T-M/P-L coating-endowed Ti substrates to hold effective anti-bacterial ability. T-M/P-L promoted the adhesion and proliferation of MSCs. In addition, an increase was witnessed regarding the ALP activity, collagen secretion and ECM mineralization, along with the expression of runt-related transcription factor 2, ALP and osteocalcin in the presence of T-M/P-L. Additionally, T-M/P-L could stimulate endothelial cells to secrete vascular endothelial growth factor (VEGF) and promote capillary-like tubule formation. Upon NIR laser irradiation exposure, T-M/P-L not only exhibited efficientin vivoanti-bacterial activity but also facilitated new bone formation. Collectively, T-M/P-L had enhanced anti-bacterial and osteogenic activity under NIR laser irradiation.

Development and Verification of a Hypoxic Gene Signature for Predicting Prognosis, Immune Microenvironment, and Chemosensitivity for Osteosarcoma.

Objective: Hypoxic tumors contribute to local failure and distant metastases. Nevertheless, the molecular hallmarks of hypoxia remain ill-defined in osteosarcoma. Here, we developed a hypoxic gene signature in osteosarcoma prognoses. Methods: With the random survival forest algorithm, a prognostic hypoxia-related gene signature was constructed for osteosarcoma in the TARGET cohort. Overall survival (OS) analysis, receiver operating characteristic (ROC) curve, multivariate cox regression analysis, and subgroup analysis were utilized for assessing the predictive efficacy of this signature. Also, external validation was presented in the GSE21257 cohort. GSEA was applied for signaling pathways involved in the high- and low-risk samples. Correlation analyses between risk score and immune cells, stromal/immune score, immune checkpoints, and sensitivity of chemotherapy drugs were performed in osteosarcoma. Then, a nomogram was built by integrating risk score, age, and gender. Results: A five-hypoxic gene signature was developed for predicting survival outcomes of osteosarcoma patients. ROC curves confirmed that this signature possessed the well predictive performance on osteosarcoma prognosis. Furthermore, it could be independently predictive of prognosis. Metabolism of xenobiotics by cytochrome P450 and nitrogen metabolism were activated in the high-risk samples while cell adhesion molecules cams and intestinal immune network for IgA production were enriched in the low-risk samples. The low-risk samples were characterized by elevated immune cell infiltrations, stromal/immune scores, TNFRSF4 expression, and sensitivity to cisplatin. The nomogram accurately predicted 1-, 3-, and 5-years survival duration. Conclusion: These findings might offer an insight into the optimization of prognosis risk stratification and individualized therapy for osteosarcoma patients.

Hydroxyapatite-Coated Titanium by Micro-Arc Oxidation and Steam-Hydrothermal Treatment Promotes Osseointegration.

Titanium (Ti)-based alloys are widely used in tissue regeneration with advantages of improved biocompatibility, high mechanical strength, corrosion resistance, and cell attachment. To obtain bioactive bone-implant interfaces with enhanced osteogenic capacity, various methods have been developed to modify the surface physicochemical properties of bio-inert Ti and Ti alloys. Nano-structured hydroxyapatite (HA) formed by micro-arc oxidation (MAO) is a synthetic material, which could facilitate osteoconductivity, osteoinductivity, and angiogenesis on the Ti surface. In this paper, we applied MAO and steam-hydrothermal treatment (SHT) to produce HA-coated Ti, hereafter called Ti-M-H. The surface morphology of Ti-M-H1 was observed by scanning electron microscopy (SEM), and the element composition and the roughness of Ti-M-H1 were analyzed by energy-dispersive X-ray analysis, an X-ray diffractometer (XRD), and Bruker stylus profiler, demonstrating the deposition of nano-HA particles on Ti surfaces that were composed of Ca, P, Ti, and O. Then, the role of Ti-M-H in osteogenesis and angiogenesis in vitro was evaluated. The data illustrated that Ti-M-H1 showed a good compatibility with osteoblasts (OBs), which promoted adhesion, spreading, and proliferation. Additionally, the secretion of ALP, Col-1, and extracellular matrix mineralization was increased by OBs treated with Ti-M-H1. Ti-M-H1 could stimulate endothelial cells to secrete vascular endothelial growth factor and promote the formation of capillary-like networks. Next, it was revealed that Ti-M-H1 also suppressed inflammation by activating macrophages, while releasing multiple active factors to mediate osteogenesis and angiogenesis. Finally, in vivo results uncovered that Ti-M-H1 facilitated a higher bone-to-implant interface and was more attractive for the dendrites, which promoted osseointegration. In summary, MAO and SHT-treated Ti-M-H1 not only promotes in vitro osteogenesis and angiogenesis but also induces M2 macrophages to regulate the immune environment, which enhances the crosstalk between osteogenesis and angiogenesis and ultimately accelerates the process of osseointegration in vivo.

Composite Coating of Graphene Oxide/TiO2 Nanotubes/HHC-36 Antibacterial Peptide Construction and an Exploration of Its Bacteriostat and Osteogenesis Effects.

Graphene oxide (GO), a kind of polymer, is often selected as a controlled released agent, whereas titanium dioxide (TiO2) nanotubes are commonly used as a drug-coated carrier. This study was conducted to develop methods for manufacturing the GO/TiO2/HHC-36 composite coating and exploring its bacteriostat and osteogenesis properties. The GO/TiO2 nanotubes were prepared by electrochemical methods and HHC-36 was then adsorbed to GO/TiO2to obtain GO/TiO2/HHC-36. Sustained release of HHC-36 was analyzed and the antibacterial effect was examined by the inhibition zone test. The biocompatibility and osteogenesis in vitro of GO/TiO2/HHC-36 were explored. Finally, the osteogenesic property of the composite coating was investigated in a rat femoral defect model in vivo. GO/TiO2/HHC-36 was successfully prepared and had good controlled released performance in vitro. The inhibit zone size of S. aureus was 2.1 mm and that of E. coli was 3.0 mm. GO/TiO2/HHC-36 showed good biocompatibility with mesenchymal stem cells (MSCs) and promoted their adhesion, migration, and differentiation. In addition, the secretion of alkaline phosphatase, collagen, mineralized matrix and osteoblast-related nutrient factors of MSCs was increased after treatment with GO/TiO2/HHC-36. Furthermore, GO/TiO2/HHC-36 also stimulated endotheliocytes to secrete VEGF, leading to angiogenesis. Finally, implantation of GO/TiO2/HHC-36 in the rat femur defect model resulted in MSC migration and increased expression of osteoblast related proteins. The composite coating with controlled released of HHC-36 showed distinct antibacterial properties and promoted osteogenesis in vitro and in vivo.

The impact of miR-9 in osteosarcoma: A study based on meta-analysis, TCGA data, and bioinformatics analysis.

The function of miR-9 in osteosarcoma is not well-investigated and controversial. Therefore, we conducted meta-analysis to explore the role of miR-9 in osteosarcoma, and collected relevant TCGA data to further testify the result. In addition, bioinformatics analysis was conducted to investigate the mechanism and related pathways of miR-9-3p in osteosarcoma.Literature search was operated on databases up to February 19, 2020, including PubMed, Web of Science, Science Direct, Cochrane Central Register of Controlled Trials, and Wiley Online Library, China National Knowledge Infrastructure, China Biology Medicine disc, Chongqing VIP, and Wan Fang Data. The relation of miR-9 expression with survival outcome was estimated by hazard ratio (HRs) and 95% CIs. Meta-analysis was conducted on the Stata 12.0 (Stata Corporation, TX). To further assess the function of miR-9 in osteosarcoma, relevant data from the TCGA database was collected. Three databases, miRDB, miRPathDB 2.0, and Targetscan 7.2, were used for prediction of target genes. Genes present in these 3 databases were considered as predicted target genes of miR-9-3p. Venny 2.1 were used for intersection analysis. Subsequently, GO, KEGG, and PPI network analysis were conducted based on the overlapping target genes of miR-9-3p to explore the possible molecular mechanism in osteosarcoma.Meta-analysis shown that overexpression of miR-9 was associated with worse overall survival (OS) (HR = 4.180, 95% CI: 2.880-6.066, P < .001, I = 23.5%). Based on TCGA data, osteosarcoma patients with overexpression of miR-9-3p (HR = 1.603, 95% CI: 1.028-2.499, P = .037) and miR-9-5p (HR = 1.698, 95% CI: 1.133-2.545, P = .01) also suffered poor OS. In bioinformatics analysis, 2 significant and important pathways were enriched: Wnt signaling pathway from gene ontology analysis (gene ontology:0016055, P-adjust = .008); hippo signaling pathway from Kyoto Encyclopedia of Genes and Genomes analysis (P-adjust = .007). Moreover, network analysis relevant protein-protein interaction was visualized, revealing 117 nodes and 161 edges.High miR-9 expression was associated with poor prognosis. Based on bioinformatics analysis, this study enhanced the understanding of the mechanism and related pathways of miR-9 in osteosarcoma.

Coating of Epimedii Folium Water Extract onto TiO2 Nanotube Surfaces Promotes Differentiation and Proliferation of Osteoblasts.

The surface modification of titanium is effective in promoting osseointegration and is widely used in the treatment of bone diseases. Epimedii Folium (EF) plays an important role in the treatment of metabolic bone diseases. However, few studies have so far been reported on their combined use in such treatments. In the present study, EF water extract was coated onto the surface of TiO2 nanotubes (TNT) by electrochemical anodization to obtain EF-TNT. Through analysis of surface morphology characteristics, it was demonstrated that EF was successfully coated on the surface of TiO2 nanotubes. In vitro drug release data suggested that the quantity of EF water extract released was a significant quantity over 4 days, reaching a total of 80%, the release continuing in total for approximately 2 weeks. By using scanning electron microscopy and immunofluorescent staining, it was found that, EF-TNT more strongly promoted adhesion, proliferation, and differentiation of MC3T3-E1 osteoblasts compared with Ti and TNT. Quantitative reverse transcript polymerase chain reaction (qRT-PCR) analysis indicated that the expression of key genes for proliferation and differentiation of osteoblasts, such as COL1a1, ALP, OPN, and Runx2, were up-regulated by EF-TNT. Network pharmacology analysis suggested that EF water extract not only regulated the proliferation and differentiation of osteoblasts but also caused a regulatory effect on osteoclasts via multiple signaling pathways, such as RANKL-RANK-induced signaling and TGF-ß signaling. These findings indicate that the EF-TNT promotes differentiation and proliferation of osteoblasts, and represents considerable potential for use in clinical applications.

In vitro and in vivo evaluation of antibacterial activity of polyhexamethylene guanidine (PHMG)-loaded TiO2 nanotubes.

Artificial joint replacement is an effective surgical method for treating end-stage degenerative joint diseases, but peripheral bacterial infection of prosthesis can compromise the effect of the surgery. Herein, antibacterial effects of titanium dioxide nanotubes (TNTs) coated with polyhexamethylene guanidine (PHMG) were examined via in vitro and in vivo experiments. TNTs with a pore diameter 46.4 ± 5.9 nm and length of 300-500 nm for the slice and 650-800 nm for the rod were fabricated by anodization. Then, 3.46 ± 0.40 mg and 1.27 ± 0.28 mg of PHMG were coated onto the TNT slice and rod, respectively. In vitro studies of the release of PHMG showed that the antibacterial agent was released in two stages: initial burst release and relatively slow release. In vitro and in vivo antibacterial studies showed that the PHMG-loaded TNTs (PHMG-TNTs) had excellent antibacterial abilities to prevent bacterial infections. Clinical pathological analysis of rabbit femurs indicated that the implanted PHMG-TNTs had no apparent pathological changes. Real-time quantitative reverse transcription polymerase chain reaction analysis of the femur tissues around the implants showed that the expression of osteogenic-related genes, including runt-related transcription factor 2, osteocalcin, alkaline phosphatase, bone sialoprotein, bone morphogenetic protein 2 and vascular endothelial growth factor A, was significantly upregulated in the PHMG-TNT implanted group as compared to the other groups. Overall, these findings provide a promising approach for the fabrication of antibacterial and bone biocompatible titanium-based implants in orthopedics.

Rapidly discriminating culture-negative urine specimens from patients with suspected urinary tract infections by UF-5000.

Aim: A rapid and reliable method of discriminating such specimens would be very useful. Materials & methods: We analyzed 566 urine specimens from patients with suspected urinary tract infections using a fully automated urine particle analyzer (UF-5000) and evaluated its performance for culture-negative urine specimens. Results: Using the algorithm cutoff values of bacteria less than 30/µl and/or white blood cell less than 200/µl, we obtained a sensitivity of 97.8%, a specificity of 74.6%, a positive predictive value of 46.9%, a negative predictive value of 99.3%, an agreement of 78.9% with the culture method and reduced 61% unnecessary urine culture. Regarding the discrimination of bacterial Gram groups, 67.7% (63/93) of cases were correctly analyzed using the UF-5000 bacteria information, with a Cohen's kappa concordance coefficient of 0.775 (χ2 = 31.65, p < 0.001). Conclusion: The performance of UF-5000 for rapidly discriminating culture-negative specimens was quite acceptable for clinical use.

A novel electrochemical biosensor for ultrasensitive detection of serum total bile acids based on enzymatic reaction combined with the double oxidation circular amplification strategy.

Serum total bile acids (TBA) level is used as a sensitive and reliable index for hepatobiliary diseases in clinics. Herein, a novel electrochemical biosensor was fabricated using enzymatic reaction coupling with the double oxidation circular amplification strategy for the detection of human serum TBA. With the catalysis of 3α-hydroxysteroid dehydrogenase (3α-HSD), 3α-bile acids reacted specifically with nicotinamide adenine dinucleotide (NAD+). And then, the reduced nicotinamide adenine dinucleotide (NADH) was produced. After that, the NADH reacted with the electron mediator of tris(2,2'-bipyridine) ruthenium(Ⅲ) (Ru(bpy)33+), which was then transformed to Ru(bpy)32+. Ultimately, Ru(bpy)32+ was further oxidized to Ru(bpy)33+ under a certain voltage, which was detected by the chronoamperometry assay. The detection was performed using a disposable unmodified screen-printed carbon electrode (SPCE) without sample preparation. The proposed biosensor showed high sensitivity and accuracy with the linear range from 5.0 to 150.0 pmol/L in 106-fold dilution serum. The established method had a good correlation with the enzymatic cycling method (r = 0.9372, P < 0.001, n = 72) commonly used in clinic. The electrochemical biosensor is simple, ultrasensitive and without sample pretreatment, showing great potential for point-of-care testing (POCT) of serum TBA in clinical samples. In addition, the biosensor is cost-effective with a small volume of samples, especially suitable for those who have difficulties in blood collection, such as infants, children and some small animals.

[Simultaneous determination of plasma indole and skatole in pregnant women with hepatitis B virus infection by high performance liquid chromatography].

A high performance liquid chromatographic (HPLC) method with fluorescence detection (FD) was established to simultaneously determine plasma indole and skatole. Plasma samples were pretreated by liquid-liquid extraction. Chromatographic separation was accomplished on a Shim-Pack VP-ODS column (150 mm×4. 6 mm, 4.6 µ m) using an isocratic mixture of 15 mmol/L sodium dihydrogen phosphate solution and methanol (40:60, v/v). The fluorescence excitation and emission wavelengths were 274 nm and 340 nm, respectively. The linear ranges were 2.22-88.89 µ g/L for indole and 1.11-44.44 µ g/L for skatole. The detection limits were 0.11 µ g/L and 0.06 µ g/L for indole and skatole, respectively. The recoveries were in the range of 95.5%-112.3% with the relative standard deviations less than 6.8%. The method was successfully applied to the analysis of plasma from healthy pregnant women (n=46) and pregnant women with hepatitis B virus (HBV) infection (n=29). The results showed that plasma indole and skatole levels were significantly different between two groups. In pregnant women with HBV infection, the concentrations of indolic compounds were positively associated with transaminase levels.

Double-loop hairpin probe and doxorubicin-loaded gold nanoparticles for the ultrasensitive electrochemical sensing of microRNA.

An electrochemical microRNA (miRNA) analysis platform by combining double-loop hairpin probe (DHP) and doxorubicin-loaded gold nanoparticles (AuNPs@Dox) for ultrasensitive miRNA detection is proposed. Firstly, we here report a DHP that is simultaneously engineered to incorporate a miRNA recognition sequence, an output segment and output's complementary fragment. The important aspect of this hairpin probe is that it would not be degraded by duplex specific nuclease (DSN) and circumvents elaborately chemical modification disadvantages encountered by classic molecular beacon. For the DHP-based DSN signal amplification system, DHP hybridizes with target miRNA to form DNA-miRNA heteroduplexes, and the DSN can hydrolyze the DNA in the heteroduplexes structure selectively, while released target miRNA strand can initiate another cycle resulting in a significant signal amplification and the accumulated output segments could be responsible for strand displacement on the electrode directly. Furthermore, a great deal of doxorubicin (Dox) are loaded on the gold nanoparticles (AuNPs) to fabricate the AuNPs@Dox biocomposites that could magnify the electrochemical signal and enable the ultrasensitive analysis of miRNA. As a result, the miRNA was capable of being detected in a limit of 0.17pM and other kinds of miRNA were discriminated facilely by this method. The described DHP as a toolbox and the nano-biocomposites as a novel signal material would not only promote the design of electrochemical biosensors but also open a good way to promote the establishment of test method in malignant tumors.