Sheng wu gong cheng xue bao = Chinese journal of biotechnology最新文献

Sepsis is a leading life-threatening problem in intensive care medicine. The recent studies have given insights into the transition from inflammation to long-term immunosuppression in sepsis. This condition might cause physiological changes that comprise the lipopolysaccharide (LPS) tolerance. Most studies about the LPS tolerance focus on the reduced ability of macrophages to secrete pro-inflammatory cytokines. Although this method has identified various molecular changes, it remains ambiguous since changes in the whole cell population are measured as an average and markers are required for cell recognition. A fast and label-free method is in demand to detect cell tolerance and screen therapeutic agents that might reverse the process. In this study, direct current insulator-based dielectrophoresis (DC-iDEP) was used to characterize the biophysical properties (EKMr) of inflamed cells, LPS-tolerant cells, and cells treated with therapeutic agents. The results showed that the EKMr of these cells was 4.28×10⁸, 3.13×10⁸, and 4.25×10⁸ V/m², respectively, suggesting that the established method was useful in distinguishing LPS-tolerant cells. The device holds the promise to be applied in medical diagnosis and medicine screening.

Urate oxidase (Uox) plays a pivotal role in uric acid (UA) degradation, and it has been applied in controlling serum UA level in clinical treatment of hyperuricemia (HUA). However, because Uox is a heterogenous protein to the human body, the immune rejections typically occur after intravenous administration, which greatly hampers the application of Uox-based agents. In this study, we used Lactococcus lactis NZ9000, a food-grade bacterium, as a host to express exogenous Uox genes, to generate the Uox-expressing engineered strains to treat HUA. Aspergillus flavus-derived Uox (aUox) and the "resurrected" human-derived Uox (hUox) were cloned into vector and expressed in NZ9000, to generate engineered strains, respectively. The engineered NZ9000 strains were confirmed to express Uox and showed UA-lowering activity in a time-dependent manner in vitro. Next, in an HUA mice model established by oral gavage of yeast paste, the UA levels were increased by 85.4% and 106.2% at day 7 and day 14. By contrast, in mice fed with NZ9000-aUox, the UA levels were increased by 39.5% and 48.3% while in mice fed with NZ9000-hUox were increased by 57.0% and 82.9%, suggesting a UA-lowering activity of both engineered strains. Furthermore, compared with allopurinol, the first-line agent for HUA treatment, mice fed with NZ9000-aUox exhibited comparable liver safety but better kidney safety than allopurinol, indicating that the use of engineered NZ9000 strains not only alleviated kidney injury caused by HUA, but could also avoided the risk of kidney injury elicited by using allopurinol. Collectively, our study offers an effective and safe therapeutic approach for HUA long-term treatment and controlling.

By targeting the key gene csgD involved in the biofilm formation of Escherichia coli, we employed molecular docking and molecular dynamics simulation to screen the active components of Chinese medicine with inhibitory effects on the biofilm formation from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP). After the anti-biofilm properties of the active components were validated in vitro, data-independent acquisition (DIA) proteomics was employed to further identify the differential proteins involved in interfering with the biofilm formation of Escherichia coli. The mechanisms of inhibition were explored with consideration to the phenotype. Through virtual screening, we identified four candidate active components, including tannic acid, narirutin, salvianolic acid B, and rosmarinic acid. Among them, tannic acid demonstrated significant inhibitory effect on the biofilm formation of E. coli. The analysis of differential proteins, combined with relevant phenotype validation, suggested that tannic acid primarily affected E. coli by intervening in pilus assembly, succinic acid metabolism, and the quorum sensing system. This study provided a lead compound for the development of new drugs against biofilm-associated infections in the future.

To preliminarily understand the pathogenic mechanism of Mycoplasma genitalium (Mg) GroEL protein, we used bioinformatics tools to predict the structure and function of Mg GroEL protein and then constructed the recombinant plasmid pET-28a-GroEL. The protein expression was induced by 0.2 mmol/L IPTG, and the expressed protein was purified by Ni-iminodicitic acid (IDA) column affinity. Tohoku Hospital Pediatrics-1 (THP-1) cells were exposed to 2 μg/mL Mg rGroEL. The levels of interleukin (IL)-1β and tumor necrosis factor (TNF)-α in the cell supernatant were measured by ELISA, and that of IL-6 was measured by an automatic chemiluminescence instrument. The activation of the nuclear factor-kappa B (NF-κB) signaling pathway was visualized by immunofluorescence and Western blotting. The results showed that Mg GroEL was a stable hydrophilic protein composed of 543 amino acid residues, with the relative molecular mass of 58.44 kDa, an isoelectric point of 5.68, and a molecular formula of C₂₅₆₈H₄₃₀₀N₇₀₀O₈₂₅S₈. The secondary structure was mainly composed of α-helices and random coils. Mg GroEL contained 12 B-cell dominant epitopes and 10 T-cell dominant epitopes. It exhibited high homology with the GroEL proteins from Mycoplasma pneumoniae, M. agalactiae, M. arthritidis, M. hyopneumoniae, and M. bovis. Mg rGroEL activated the NF-κB signaling pathway and promoted the secretion of IL-1β, IL-6, and TNF-α in THP-1 cells. These results suggest that Mg GroEL exhibits substantial antigenicity and possesses the capability of triggering inflammation in host cells. This study establishes a theoretical basis for future investigations pertaining to the role and pathogenic mechanisms of Mg GroEL.

Peptide-based hydrogel, the polymer materials with a special network structure, are widely used in various fields of biomedicine due to their stable properties and biocompatibility. Environment-responsive self-assembled peptide aqueous solutions can respond to environment changes by the self-assembly of peptides into nanofiber networks. Peptide-based hydrogels well simulate the extracellular matrix and cell growth microenvironment, being suitable for 3D cell culture and organoid culture. To establish a tumor organoid culture system with peptide-based hydrogels, we cultured Panc-1, U87, and H358 cells in a 3D spherical manner using CulX Ⅱ peptide-based hydrogels in 24-well plates for 15 days. The organoids showed a 3D spherical shape, and their sizes increased with the extension of the culture time, with a final diameter ranging from 150 to 300 μm. The organoids had a large number, varying sizes, good cell viability, clear edges, and a good shape, which indicated successful organoid construction. The tumor organoid culture system established in this study with CulX Ⅱ peptide-based hydrogels provides a model for studying tumor pathogenesis, drug development, and tumor suppression.

Gelsemium elegans, a vine plant of Loganiaceae, has both medicinal and forage values. However, it is susceptible to low temperatures during growth. Exploring low temperature response genes is of great significance for cold resistance breeding of G. elegans. Ethylene response factors (ERFs) are the transcription factors of the AP2/ERF superfamily and play a crucial role in plant stress response. In this study, based on the unigene GeERF involved in the response to low temperature stress in the transcriptome of G. elegans, a full-length cDNA sequence of the transcription factor GeERF4B-1 was cloned from the leaves of G. elegans by reverse transcription-polymerase chain reaction (RT-PCR). Bioinformatics analysis showed that GeERF4B-1 had an open reading frame of 759 bp, encoding a protein composed of 252 amino acid residues and with a relative molecular weight of 27 kDa. The deduced protein was predicted to be an unstable, alkaline, and hydrophilic protein. The phylogenetic tree showed that GeERF4B-1 was in the same clade as the B-4 subfamily of the ERF family. The results of the subcellular localization experiment revealed that GeERF4B-1 was located in the nucleus. Real time quantitative PCR (RT-qPCR) analysis indicated that GeERF4B-1 was expressed in the root, stem, and leaf of G. elegans, with the highest expression level in the root. Compared with the control, the treatments with a low temperature (4 ℃), methyl jasmonate (MeJA), and abscisic acid (ABA) up-regulated the expression level of GeERF4B-1, which reached the peak at 24-48 h. This result revealed that GeERF4B-1 actively responded to low temperature, MeJA, and ABA stresses. However, both sodium chloride (NaCl) and drought treatments down-regulated the expression of GeERF4B-1. In addition, a prokaryotic expression vector of GeERF4B-1 was constructed, and a fusion protein of approximately 52 kDa was yielded after induced expression. The results of the plate stress assay showed that compared with the control, the prokaryotic strain expressing GeERF4B-1 demonstrated enhanced tolerance to low temperatures and sensitivity to salt and mannitol stresses. This study provides theoretical references and potential genetic resources for breeding G. elegans varieties with stress resistance.

Nucleolar complex associated 4 homolog (NOC4L) is a key factor in ribosome biogenesis, and this study aims to investigate its roles in activated T cells from the perspective of translation regulation. Firstly, flow cytometry was employed to determine the expression levels of NOC4L in the CD4⁺ T cells under different conditions in the transgenic reporter mice expressing Noc4l^mCherry. Subsequently, the expression of NOC4L along with cell proliferation was examined under Th1 and Th17 polarization conditions. Finally, in vitro experiments were conducted to identify the proteins interacting with NOC4L during the activation of Th1 and Th17 cells, on the basis of which the potential mechanisms of NOC4L were explored. The results showed that the expression level of NOC4L increased in activated CD4⁺ T cells, and the expression of NOC4L was closely associated with the proliferation and division of activated T cells. The in vitro experiments revealed interactions between NOC4L and proteins involved in ribosome assembly and cell proliferation during T cell activation. These findings lay a foundation for probing into the post-transcriptional regulation in helper T cells and hold profound significance for understanding the activation and regulatory mechanisms of T cells.

The emerging biomedical industry has an increasing demand for the professional talents and puts forward higher requirements for the quality, especially the innovation ability of the talents. Exploring a new model for fostering the innovation ability of postgraduates majoring in biological and medical sciences based on the principle of integrating production, education and research is of practical significance for improving the quality of professional talents and empowering the bio-economic development. Taking the training of innovation ability of postgraduates majoring in biological and medical sciences at Jiangnan University as an example, this paper introduced a new training system of "six integration and six optimizations". This system included ideological guidance, discipline system, training program, faculty, research innovation platform, and communication and cooperation. Satisfactory cultivation results were achieved with this new system. This paper is expected to provide reference for the training of innovative talents in the biological and medical industry.

Prostate cancer is one of the most common malignant tumors in men and posing a serious threat to men's health. Detection methods such as prostate-specific antigen (PSA), prostate biopsy, and magnetic resonance imaging are widely used for prostate cancer screening, but they have low specificity, high cost, and significant risks. Therefore, there is an urgent need to develop highly specific, low-cost, easily obtained, stable, and reliable biomarkers, and use them as the basis to establish non-invasive screening and diagnostic methods for prostate cancer. This paper reviewed the recent advances in the use of prostate cancer biomarkers and combined detection methods for prostate cancer diagnosis and prognosis assessment and provides an in-depth analysis and comparison of different biomarkers and combined detection methods, as well as points out the directions and challenges for future research. The paper emphasizes the importance of developing efficient, cost-effective and easy-to-implement biomarkers to increase the early diagnosis rate of prostate cancer, improve patient prognosis, and reduce the waste of healthcare resources. This paper provides an important theoretical basis and technical guidance for early diagnosis, precise treatment and prognostic evaluation of prostate cancer, and has important reference value for promoting clinical research and practice of prostate cancer.

The dehydration responsive element binding (DREB) transcription factors play an important role in plant growth and development and are extensively involved in plant responses to abiotic stress. The DREB family contains six subfamilies, and TINY belongs to the DREB-A4 subfamily. The Arabidopsis thaliana TINY gene, AtTINY, plays a role in regulating plant growth and responses to stress. In order to investigate the evolutionary characterization of the DREB-A4 subfamily and the biological function of the MdTINY gene in apple (Malus domestica), in this study, we used the databases GDDH13 and TAIR and online tools (Expasy and WoLF PSORT) to study the biological information of the DREB-A4 subfamily in apple. In addition, the tertiary structures of the proteins were predicted. The apple DREB-A4 subfamily contained 22 genes, all of which had a conserved AP2 domain, and subcellular localization predictions showed that DREB-A4 subfamily proteins were mainly located in the nucleus. The transgenic calli of MdTINY were obtained by the Agrobacterium-mediated transformation method, and the main biological functions of MdTINY were explored by quantitative real-time PCR (qRT-PCR) combined with anthocyanin content determination. MdTINY shared the highest amino acid sequence similarity with AtTINY. The coding region of MdTINY had a full length of 759 bp, encoding 252 amino acid residues. Analysis of the promoter elements and expression patterns indicated that MdTINY was responsive to light and multiple stress conditions. MdTINY was localized in the nucleus and had transcriptional autoactivation activity. The overexpression of MdTINY in calli inhibited normal growth and promoted anthocyanoside accumulation. These results indicated that MdTINY negatively regulated apple plant growth and promoted fruit coloring, providing a candidate gene for the breeding of apple varieties with high quality of fruit color.