TALANTA
·
2022
·
IF 6.556
Biyun Fang,Zhenzhen Jia,Cui Liu,Kangsheng Tu,Mingzhen Zhang,Lei Zhang
Pathogen nucleic acid detection is of great significance to control the spread of diseases caused by the viruses. Nevertheless, traditional methods for nucleic acid detection such as polymerase chain reaction (PCR) and oligonucleotide microarrays require bulky instruments, which restrain their point-of-care (POC) testing application. Here, we proposed a POC method enabling sensitive detection of pathogen nucleic acids by combining the clustered regularly interspaced short palindromic repeat (CRISPR) Cas12a-based assay and personal glucometer readout (PGM). The quantification of target pathogen DNA by PGM was achieved based on pathogen DNA activates Cas12a ssDNase to cleave magnetic bead-DNA-invertase reporter probe, and separated free invertase to catalyze hydrolysis of sucrose to glucose. Without using nucleic acid amplification technology, we demonstrated here dual signal amplifications based on Cas12a and invertase-mediated catalytic reactions, making it possible to sensitively detect HIV-related DNA or SARS-CoV-2 pseudovirus with the limits of detection of 11.0 fM and 50 copies/μL, respectively. This strategy also showed excellent selectivity as well as potential applicability for detection of HIV in human serum samples or of SARS-CoV-2 in saliva samples. Therefore, our CRISPR-PGM-based dual signal amplifications detection platform might offer a great promise in POC diagnosis of pathogen nucleic acids.
研究领域:
分子生物学
·
医学诊断
·
生物技术
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DOI
PubMed
高影响因子
LANGMUIR
·
2024
·
IF 3.7
Xianhui Cheng,Yujuan Miao,Juntao Zhou,Feng Lu,Jingji Jin,Lianghai Hu
Plasma membranes not only maintain the intracellular microenvironment through their phospholipid bilayer but also eliminate exogenous compounds outside the cell membranes. Most drugs especially with high polarity are prevented from entering into cells to exert their effects. Therefore, it is of great significance to design effective drug carriers with a penetrating ability toward plasma membranes. In this study, a dual-templated MIP (dt-MIPs) carrier with controllable microstructure and high drug loading capacity was prepared using highly expressed sphingomyelin on the plasma membrane and tenofovir (TFV), a first-line drug for HIV and chronic hepatitis B, as template molecules. The drug release experiments performed in vitro under simulated physiological conditions demonstrated that sustained and stable adsorption of TFV on dt-MIPs was more than 80% over 50 h. By a combination of flow cytometry and confocal microscopy, dt-MIPs were found to have efficient cell permeability. Furthermore, mass-spectrometry-based intracellular pharmacokinetic studies demonstrated that TFV was delivered completely into cells within 30 min with the delivery of dt-MIPs. The study presented above suggested that dt-MIPs are expected to be alternative nanoscale drug carriers for enhanced drug permeability and controlled release.
研究领域:
药学
·
纳米技术
·
药物递送系统
·
生物医学工程
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产品关联
DOI
PubMed
高影响因子
ACS Synthetic Biology
·
2024
·
IF 3.7
Mei Tang,Xianhao Xu,Yanfeng Liu,Jianghua Li,Guocheng Du,Xueqin Lv,Long Liu
Betulinic acid (BA) is a lupane-type triterpenoid with potent anticancer and anti-HIV activities. Its great potential in clinical applications necessitates the development of an efficient strategy for BA synthesis. This study attempted to achieve efficient BA biosynthesis in Saccharomyces cerevisiae using systematic metabolic engineering strategies. First, a de novo BA biosynthesis pathway in S. cerevisiae was constructed, which yielded a titer of 14.01 ± 0.21 mg/L. Then, by enhancing the BA synthesis pathway and dynamic inhibition of the competitive pathway, a greater proportion of the metabolic flow was directed toward BA synthesis, achieving a titer of 88.07 ± 5.83 mg/L. Next, acetyl-CoA and NADPH supply was enhanced, which increased the BA titer to 166.43 ± 1.83 mg/L. Finally, another BA synthesis pathway in the peroxisome was constructed. Dual regulation of the peroxisome and cytoplasmic metabolism increased the BA titer to 210.88 ± 4.76 mg/L. Following fed-batch fermentation process modification, the BA titer reached 682.29 ± 8.16 mg/L. Overall, this work offers a guide for building microbial cell factories that are capable of producing terpenoids with efficiency.
研究领域:
生物技术
·
代谢工程
·
药学
·
微生物学
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DOI
PubMed
高影响因子
Frontiers in Immunology
·
2012
·
IF 0
Slyker Jennifer Ann,Lohman-Payne Barbara Louise,John-Stewart Grace,Dong Tao,Mbori-Ngacha Dorothy,Tapia Kenneth,Atzberger Ann,Taylor Stephen,Rowland-Jones Sarah,Blish Catherine A
Natural killer (NK) cells play an important role in the containment of HIV replication during primary infection, though their functions are impaired during chronic HIV infection. Infants experience more rapid HIV disease progression than adults, but contributions of infant NK cells to containing HIV infection are unknown. The aim of this study was to determine the impact of HIV infection on infant NK cell phenotype by evaluating samples and data from a cohort study of women and their infants, conducted in Nairobi, Kenya between 1999 and 2003. The percentage and phenotype of NK cells was evaluated longitudinally by multi-parameter flow cytometry over the first year of life in HIV-infected (HIV+, = 16), HIV-exposed uninfected (HIV-EU, n = 6), and healthy unexposed controls (HIV–, n = 4). At birth, NK subset distributions based on expression of CD56 and CD16 did not differ between HIV+, HIV-EU, or HIV– infants. However, HIV infection was associated with a subsequent decline in NK cells as a percentage of total lymphocytes (p < 0.001), and an expanding proportion of CD56-CD16+ NK cells (p < 0.001). Activated CD38brightCD69+ NK cells were more frequent in the HIV+ infants, followed by HIV-EU and HIV- infants, in both CD56dim (p = 0.005) and CD56bright compartments (p = 0.03). HIV infection and exposure was also associated with a significant decline in the percentage of perforin-expressing NK cells in the CD56dim compartment over the first year of life, with HIV+ infants losing approximately 2.5% (p < 0.001) and HIV-EU infants losing 3.0% (p = 0.01) of perforin+ cells per month. Thus, infant HIV infection is associated with alterations in NK cell subsets, activation, and cytolytic potential that could contribute to their poor control over HIV infection. Furthermore, exposure to HIV infection in infants who escaped infection is also associated with alterations in NK cells that may contribute to the reduced ability to fight infections that is observed in HIV-EU infants.
研究领域:
免疫学
·
传染病学
·
儿科
✓ 已验证
DOI
PubMed
ANALYTICAL CHEMISTRY
·
2017
·
IF 6.32
Shanglin Li,Yin Gu,Yi Lyu,Yan Jiang,Peng Liu
An integrated graphene oxide purification-lateral flow test strip (iGOP-LFTS) was developed for on-strip purifying and visually detecting polymerase chain reaction (PCR) products with an improved sensitivity as well as a more stringent specificity. PCR products amplified with a pair of biotin- and digoxin-labeled primers were directly pipetted onto GO pads, on which graphene oxide selectively adsorbed residual primers and primer-dimers with the aid of a running buffer containing MgCl2 and Tween 20. By stacking up three GO pads to increase the surface area for adsorption, 83.4% of double-stranded DNA with a length of 30 bp and 98.6% of 20-nt primers could be removed from a 10-μL DNA mixture. Since no primers interfered with detection, the increase of the sample loading volume from 5 to 20 μL could improve the signal-to-noise ratio of the test line 1.6 fold using the iGOP-LFTS while no changes were observed using the conventional LFTS. The limit of detection of the iGOP-LFTS was determined to be 30 copies of bacteriophage λ-DNA with naked eyes and this limit could be further decreased to 3 copies by loading 20 μL of the sample, which corresponded to a 1000-fold improvement compared to that of the LFTS detected by naked eyes. When the ImageJ analysis was employed, a 100-fold decrease of the detection limit can be obtained. In addition, due to the removal of the primer-dimers, the dim test line observed in the negative control of the LFTS was eliminated using the iGOP-LFTS. A mock clinical specimen spiked with defective HIV-1 (human immunodeficiency virus) viruses was successfully analyzed using a two-step reverse transcription-PCR with 30 amplification cycles followed by the iGOP-LFTS detection. These significant improvements were achieved without introducing any additional hands-on operations and instrumentations.
研究领域:
分子生物学
·
分析化学
·
生物医学工程
·
病毒学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
TALANTA
·
2020
·
IF 5.339
Jiao Zhou,Lei Han,Yu Ling,Lei Wang,Nian Bing Li,Hong Qun Luo
Relying on the specific coordination of Ag + and mismatched cytosine-cytosine (C–C), the high-efficiency inhibition of urease by Ag + ion, and the rapid and sensitive response of phenol red to pH, a sensitive ratiometric sensor has been designed for visual detection of human immunodeficiency virus gene (HIV DNA). This sensor utilizes the HIV DNA to initiate catalytic hairpin assembly (CHA) process, releasing Ag + to inhibit subsequent urease-catalyzed urea hydrolysis and prevent the pH of the solution from rising. The CHA process and the absorbance ratio of phenol red at different wavelengths (A 559 /A 432 ) amplify the signal, allowing the sensor to detect HIV DNA from 10 to 130 nM in a sensitive and highly selective manner with a low detection limit of 7.8 nM. In addition, this sensor can visually distinguish different concentrations of HIV DNA within a certain range and possesses a good recovery in 1% of serum samples, which will provide new ideas for biosensor design, dipstick test, blood test, and other clinical disease prevention.
研究领域:
生物技术
·
纳米技术
·
分析化学
·
分子生物学
·
医学诊断
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Frontiers in Plant Science
·
2017
·
IF 4.291
Su Ping,Guan Hongyu,Zhang Yifeng,Wang Xing,Gao Linhui,Zhao Yujun,Hu Tianyuan,Zhou Jiawei,Ma Baowei,Tu Lichan,Tong Yuru,Huang Luqi,Gao Wei
Tripterygium wilfordii produces not only ent-kaurene, which is an intermediate of gibberellin (GA) biosynthesis in flowering plants, but also 16α-hydroxy-ent-kaurane, whose physiological role has not been characterized. The two compounds are biosynthesized from the universal diterpenoid precursor (E,E,E)-geranylgeranyl diphosphate (GGPP) by diterpene synthases, which have been discovered and functionally characterized in T. wilfordii. Here, we described the functional characterization of four cytochrome P450 reductases (TwCPR) and one ent-kaurene oxidase (TwKO). Four TwCPRs were found to have relatively ubiquitous expression in T. wilfordii root, stem, leaf, and flower tissues. Co-expression of both a TwCPR and TwKO in yeast showed that TwCPR3 has a slightly better activity for providing the electrons required for these reactions, indicating that TwCPR3 is more suitable for use in the functional analysis of other cytochrome P450 monooxygenases. TwKO catalyzed the three-step oxidation of the C4α methyl of the tetracyclic diterpene intermediate ent-kaurene to form ent-kaurenoic acid as an early step in GA biosynthesis. Notably, TwKO could also convert 16α-hydroxy-ent-kaurane to 16α-hydroxy-ent-kaurenoic acid, indicating an important function of 16α-hydroxy-ent-kaurane in the anti-HIV principle tripterifordin biosynthetic pathway in planta. Homology modeling and molecular docking were used to investigate the unknown crucial active amino acid residue involved in the catalytic reaction of TwKO, and one key residue (Leu387) contributed to the formation of 16α-hydroxy-ent-kaurenoic acid, most likely by forming hydrogen bonds with the hydroxyl group (-OH) of 16α-hydroxy-ent-kaurane, which laid the basis for further investigation of the multifunctional nature of KO catalysis. Also, our findings paved the way for the complete biosynthesis of the anti-HIV principle tripterifordin.
研究领域:
植物生物化学
·
酶学
·
分子生物学
·
生药学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOMACROMOLECULES
·
2016
·
IF 5.583
Nan Wang,Xin Jin,Dongbo Guo,Gangsheng Tong,Xinyuan Zhu
Iron accumulation in substantia nigra pars compacta (SNpc) has been proved to be a prominent pathophysiological feature of Parkinson’s diseases (PD), which can induce the death of dopaminergic (DA) neurons, up-regulation of reactive oxygen species (ROS), and further loss of motor control. In recent years, iron chelation therapy has been demonstrated to be an effective treatment for PD, which has shown significant improvements in clinical trials. However, the current iron chelators are suboptimal due to their short circulation time, side effects, and lack of proper protection from chelation with ions in blood circulation. In this work, we designed and constructed iron chelation therapeutic nanoparticles protected by a zwitterionic poly(2-methacryloyloxyethyl phosphorylcholine) (PMPC) to delay the saturation of iron chelators in blood circulation and prolong the in vivo lifetime, with HIV-1 trans-activating transcriptor (TAT) served as a shuttle to enhance the blood-brain barrier (BBB) permeability. We explored and investigated whether the Parkinsonian neurodegeneration and the corresponding symptoms in behaviors and physiologies could be prevented or reversed both in vitro and in vivo. The results demonstrated that iron chelator loaded therapeutic nanoparticles could reverse functional deficits in Parkinsonian mice not only physiologically but also behaviorally. On the contrary, both untreated PD mice and non-TAT anchored nanoparticle treated PD mice showed similar loss in DA neurons and difficulties in behaviors. Therefore, with protection of zwitterionic polymer and prolonged in vivo lifetime, iron chelator loaded nanoparticles with delayed saturation provide a PD phenotype reversion therapy and significantly improve the living quality of the Parkinsonian mice.
研究领域:
神经科学
·
药学
·
纳米技术
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
BIOCHIMIE
·
2016
·
IF 3.017
Xiaoqiu He,Huayan Zhang,Yuhua Shi,Xin Gong,Shanshan Guan,He Yin,Lan Yang,Yongjiao Yu,Ziyu Kuai,Dongni Liu,Rui Hua,Song Wang,Yaming Shan
With increasing microbial drug resistance worldwide, antimicrobial peptides (AMPs) are considered promising alternatives to addressing this problem. In this study, a series of synthetic peptides were designed based on the membrane-disrupting properties of the membrane-proximal external region (MPER) of human immunodeficiency virus type 1 (HIV-1) envelope protein. The peptide AP16-A was found to exhibit the most effective antimicrobial activities against both Gram-negative and Gram-positive bacteria. The minimal bactericidal concentration (MBC) of AP16-A ranged from 2 μg/ml to 16 μg/ml. AP16-A had no detectable cytotoxicity in various tissue cultures and a mouse model. Furthermore, results of confocal fluorescence microscopy and the SYTOX Green uptake assay indicated that AP16-A killed Gram-negative bacteria by the combined effects of relatively slow membrane permeabilization and interaction with an intracellular target, while it killed Gram-positive bacteria by a fast membrane permeabilization process, which achieved relatively more rapid bacterial killing kinetics. The results of this study support the potential use of AP16-A as an AMP.
研究领域:
微生物学
·
病毒学
·
药学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
MICROCHEMICAL JOURNAL
·
2024
·
IF 4.9
Yan Liu,Yuetao Zhang,Zijie Li,Zhiqi Li,Shaoke Zhou,Yong Xia,Lei Mou
Rapid, accurate, and cost-effective strategies for the multiplexed and quantitative detection of multiple infectious disease biomarkers are still lack in hospital. Here, we introduce a multiplexed screen-printed electrode for the simultaneous detection of three infectious disease biomarkers: HBsAg, anti-HIV, and anti-TP. The electrode modified with in-situ deposition of gold nanoparticles (AuNPs) and MXene multilayered nanosheets. Comprehensive characterization confirms the morphology and electrochemical response of the AuNPs&MXene modified screen-printed electrode (AuNPs&MXene-SPE). AuNPs&MXene composite contributes to enhance sensitivity by maximizing surface area and improving electron transfer efficiency. The AuNPs&MXene-SPE exhibits outstanding detection performance for HBsAg, anti-HIV, and anti-TP. The AuNPs&MXene-SPE sensors achieve broad detection ranges (0.05–1000 ng/mL for HBsAg, 0.35–140 ng/mL for anti-HIV, and 0.25–100 ng/mL for anti-TP) accompanied by low limits of detection (0.01, 0.11, and 0.10 ng/mL, respectively). Applied to standard serum samples, the sensors showcase precision with relative standard deviations consistently below 5.0 %, affirming their suitability for complex solution environments. These sensors offer notable advantages, including cost-effectiveness, rapid response, high sensitivity, and specificity, eliminating the necessity for expensive equipment, which open avenues for the realization of point-of-care testing (POCT).
研究领域:
电化学
·
生物传感器
·
纳米材料
·
传染病
·
即时检测
✓ 已验证
产品关联
DOI
高影响因子
EUROPEAN JOURNAL OF PHARMACEUTICAL SCIENCES
·
2017
·
IF 3.756
Yixian Liu,Bingjie Liu,Yan Zhang,Youmei Peng,Chenzheng Huang,Ning Wang,Jinhua Jiang,Qingduan Wang,Junbiao Chang
2′-Deoxy-2′-β-fluoro-4′-azidocytidine (FNC), a cytidine analog, has attracted great interest because of its potent activity against wild-type and multidrug-resistant HIV. The purpose of current study was to investigate the absorption mechanisms of FNC in the small intestine, as well as the interactions between FNC and P-glycoprotein (P-gp), multidrug resistance-associated protein 2 (MRP2) and breast cancer resistance protein (BCRP). The experiments were performed using Caco-2 cells and the rat small intestine. The uptake experiment indicated that FNC concentration, extracellular pH and the incubated temperature could influence the uptake of FNC in Caco-2 cells. NaN 3 , verapamil , probenecid , MK571 and GF120918 could significantly increase the FNC uptake in Caco-2 cells. The transport experiment showed that both the absorption and secretion of FNC were concentration dependent. The secretion of FNC was approximately 2-fold greater than the absorption. In the presence of verapamil, probenecid, MK571 or GF120918, the efflux ratio decreased by > 50%. In everted rat intestine, the absorption of FNC also depended on its concentration and was not significantly different in the different segments of the small intestine. Real-time RT-PCR results indicated that the gene expressions of P-gp, MRP2 and BCRP were up-regulated after exposure to FNC. The reduction in accumulation of rhodamine 123 after treatment with FNC revealed its ability to up-regulate P-gp activity. In conclusion, FNC was completely absorbed by passive diffusion and active transport mechanisms. P-gp, MRP2 and BCRP could influence the absorption of FNC in the small intestine. FNC could modulate the gene expressions of P-gp, MRP2 and BCRP, and increase the activity of P-gp.
研究领域:
药理学
·
艾滋病治疗
·
药物转运
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
·
2018
·Vol. 411
·期 19
·4569-4576
·
IF 3.307
Wang Kun,Zhai Fu-Heng,He Meng-Qi,Wang Jin,Yu Yong-Liang,He Rong-Huan
A simple fluorescence biosensor is developed based on the enzyme-assisted cascade amplification strategy. The amplification system consists of a hairpin-structure DNA (H-DNA) and exonuclease III. The target DNA can hybridize with the H-DNA and initiate exonuclease III-assisted target recycling amplification to generate abundant G-rich DNA (G-DNA). One region of G-DNA is designed to possess the same sequence as target DNA. Thus, the G-DNA can also hybridize with H-DNA and initiate the digestion of H-DNA. The cascade strategy in this amplification system causes the concentration of G-DNA to grow exponentially. The fluorescence intensity of N -methylmesoporphyrin IX (NMM) is highly enhanced due to the formation of G-quadruplex configuration. Under optimal conditions, the cascade system could achieve an admirable sensitivity with a detection limit of 52 fM for HIV DNA, and guarantees a satisfactory specificity. Moreover, the cascade system could be implemented for other target DNA detections by substituting the recognition region of the H-DNA. In this way, a detection limit of 65 fM for HBV DNA could be achieved by the cascade system. The target DNA analysis in a real serum sample further indicates that this biosensor has potential for future application in clinical diagnosis.
A simple and label-free cascade amplification strategy is developed by exploiting hairpin DNA and EXO III for sensitive DNA detection.
研究领域:
生物化学
·
分子生物学
·
分析化学
·
临床诊断
✓ 已验证
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）