BMC Biotechnology最新文献

Background: Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne zoonotic disease that presents with severe hemorrhagic manifestations and is associated with significant fatality rates. The causative agent, Crimean-Congo Hemorrhagic Fever Virus (CCHFV), is a high-priority pathogen identified by the World Health Organization with no approved vaccine or specific treatment available. In addition, there is a critical need for enhanced diagnostic tools to improve public health awareness, prevention measures, and disease control strategies.

Methods: We designed plasmids to enable the purification of soluble CCHFV glycoprotein Gc expressed in mammalian 293 F cells, followed by purification using affinity and size exclusion chromatography. The purified antigen was analyzed by SDS-PAGE and Western blotting to confirm its reactivity to antibodies from CCHF survivors. Additionally, an in-house indirect ELISA was developed using the purified Gc as a coating antigen.

Results: The optimized expression system successfully produced soluble and pure Gc antigen after affinity chromatography. The protein showed specific reactivity with CCHFV-positive serum antibodies in Western blot analysis. The indirect ELISA assay demonstrated high efficacy in distinguishing between CCHFV-positive and -negative serum samples, indicating its potential as a valuable diagnostic tool. Size exclusion chromatography further confirmed the presence of aggregates in our protein preparation.

Conclusions: The purified Gc antigen shows promise for developing direct diagnostic assays for CCHFV. The antigen's suitability for subunit vaccine development and its application as bait for monoclonal antibody isolation from survivors could be investigated further. This work lays the foundation for future research into the development of rapid diagnostic tests for field deployment.

Based on our previous findings that salicylic acid and jasmonic acid increased Nostoc flagelliforme polysaccharide yield by regulating intracellular nitric oxide (NO) levels, the mechanism through which NO affects polysaccharide biosynthesis in Nostoc flagelliforme was explored from the perspective of S-nitrosylation (SNO). The addition of NO donor and scavenger showed that intracellular NO had a significant positive effect on the polysaccharide yield of N. flagelliforme. To explore the mechanism, we investigated the relationship between NO levels and the activity of several key enzymes involved in polysaccharide biosynthesis, including fructose 1,6-bisphosphate aldolase (FBA), glucokinase (GK), glucose 6-phosphate dehydrogenase (G6PDH), mitochondrial isocitrate dehydrogenase (ICDH), and UDP-glucose dehydrogenase (UGDH). The enzymatic activities of G6PDH, ICDH, and UGDH were shown to be significantly correlated with the shifts in intracellular NO levels. For further validation, G6PDH, ICDH, and UGDH were heterologously expressed in Escherichia coli and purified via Ni⁺-NAT affinity chromatography, and subjected to a biotin switch assay and western blot analysis, which revealed that UGDH and G6PDH were susceptible to SNO. Furthermore, mass spectrometry analysis of proteins treated with S-nitrosoglutathione (GSNO) identified the SNO modification sites for UGDH and G6PDH as cysteine 423 and cysteine 249, respectively. These findings suggest that NO modulates polysaccharide biosynthesis in N. flagelliforme through SNO of UGDH and G6PDH. This reveals a potential mechanism through which NO promotes polysaccharide synthesis in N. flagelliforme, while also providing a new strategy for improving the industrial production of polysaccharides.

Chemotherapy as a cornerstone of cancer treatment is slowly being edged aside owing to its severe side effects and systemic toxicity. In this case, nanomedicine has emerged as an effective tool to address these drawbacks. Herein, a biocompatible carrier based on bovine serum albumin (BSA) coated gadolinium oxide nanoparticles (Gd₂O₃@BSA) was fabricated for curcumin (CUR) delivery and its physicochemical features along with its potential anticancer activity against nasal squamous cell carcinoma were also investigated. It was found that the fabricated Gd₂O₃@BSA containing CUR (Gd₂O₃@BSA-CUR) had spherical morphology with hydrodynamic size of nearly 26 nm, zeta-potential of -36 mV and high drug (CUR) loading capacity. Drug release profile disclosed that the release of CUR from the prepared Gd₂O₃@BSA-CUR nanoparticles occurred in a sustained- and pH-dependent manner. Also, in vitro cytotoxicity analysis revealed that the fabricated Gd₂O₃@BSA nanoparticles possessed excellent biosafety toward HFF2 normal cells, while Gd₂O₃@BSA-CUR appeared to display the greatest anticancer potential against RPMI 2650 and CNE-1 cancer cell lines. The results also show that the Gd₂O₃@BSA nanoparticles were compatible with the blood cells with minor hemolytic effect (< 3%). The manufactured NPs were found to be completely safe for biological applications in an in vivo subacute toxicity study. Taken together, these finding substantiate the potential anticancer activity of Gd₂O₃@BSA-CUR nanoparticles against nasal squamous cell carcinoma, but the results obtained demand further studies to assess their full potential.

Background: Colorectal cancer is a common disease worldwide with non-specific symptoms such as blood in the stool, bowel movements, weight loss and fatigue. Chemotherapy drugs can cause side effects such as nausea, vomiting and a weakened immune system. The use of antioxidants such as hesperidin could reduce the side effects, but its low bioavailability is a major problem. In this research, we aimed to explore the drug delivery and efficiency of this antioxidant on the HCT116 colorectal cancer cell line by loading hesperidin into PLGA nanoparticles.

Materials and methods: Hesperidin loaded PLGA nanoparticles were produced by single emulsion evaporation method. The physicochemical properties of the synthesized hesperidin-loaded nanoparticles were determined using SEM, AFM, FT-IR, DLS and UV-Vis. Subsequently, the effect of the PLGA loaded hesperidin nanoparticles on the HCT116 cell line after 48 h was investigated by MTT assay at three different concentrations of the nanoparticles.

Result: The study showed that 90% of hesperidin were loaded in PLGA nanoparticles by UV-Vis spectrophotometry and FT-IR spectrum. The nanoparticles were found to be spherical and uniform with a hydrodynamic diameter of 76.2 nm in water. The release rate of the drug was about 93% after 144 h. The lowest percentage of cell viability of cancer cells was observed at a concentration of 10 µg/ml of PLGA nanoparticles loaded with hesperidin.

Conclusion: The results indicate that PLGA nanoparticles loaded with hesperidin effectively reduce the survival rate of HCT116 colorectal cancer cells. However, further studies are needed to determine the appropriate therapeutic dosage and to conduct animal and clinical studies.

Background: Measuring collagenase activity is crucial in the field of joint health and disease management. Collagenases, enzymes responsible for collagen degradation, play a vital role in maintaining the balance between collagen synthesis and breakdown in joints. Dysregulation of collagenase activity leads to joint tissue degradation and diseases such as rheumatoid arthritis and osteoarthritis. The development of methods to measure collagenase activity is essential for diagnosis, disease severity assessment, treatment monitoring, and identification of therapeutic targets.

Results: This study aimed to validate a rapid collagenase activity detection technique using synovial fluid samples. Antibody microarray analysis was initially performed to quantify the levels of matrix metalloproteinase-9 (MMP-9), a major collagenase in joints. Subsequently, the developed gelatin-based test utilizing fluorescence measurement was used to determine collagenase activity. There was a significant correlation between the presence of MMP-9 and collagenase activity. In addition, Lower Limit of Detection and Upper Limit of Detection can be preliminary estimated as 8 ng/mL and 48 ng/mL respectively.

Conclusions: The developed technique offers a potential point-of-care assessment of collagenase activity, providing real-time information for clinicians and researchers. By accurately quantifying collagenase activity, healthcare professionals can optimize patient care, improve treatment outcomes, and contribute to the understanding and management of joint-related disorders. Further research and validation are necessary to establish the full potential of this rapid collagenase activity detection method in clinical practice.