Korea Research Institute of Bioscience and Biotechnology
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Development of a novel nannochloropsis strain with enhanced violaxanthin yield for large-scale production
No full textBackground: Nannochloropsis is a marine microalga that has been extensively studied. The major carotenoid produced by this group of microalgae is violaxanthin, which exhibits anti-inflammatory, anti-photoaging, and antiproliferative activities. Therefore, it has a wide range of potential applications. However, large-scale production of this pigment has not been much studied, thereby limiting its industrial application.
Results: To develop a novel strain producing high amount of violaxanthin, various Nannochloropsis species were isolated from seawater samples and their violaxanthin production potential were compared. Of the strains tested, N. oceanica WS-1 exhibited the highest violaxanthin productivity; to further enhance the violaxanthin yield of WS-1, we performed gamma-ray-mediated random mutagenesis followed by colorimetric screening. As a result, Mutant M1 was selected because of its significant higher violaxanthin content and biomass productivity than WS-1 (5.21 ± 0.33 mg g- 1 and 0.2101 g L- 1 d- 1, respectively). Subsequently, we employed a 10 L-scale bioreactor to confirm the large-scale production potential of M1, and the results indicated a 43.54 % increase in violaxanthin production compared with WS-1. In addition, comparative transcriptomic analysis performed under normal light condition identified possible mechanisms associated with remediating photo-inhibitory damage and other key responses in M1, which seemed to at least partially explain enhanced violaxanthin content and delayed growth.
Conclusions: Nannochloropsis oceanica mutant (M1) with enhanced violaxanthin content was developed and its physiological characteristics were investigated. In addition, enhanced production of violaxanthin was demonstrated in the large-scale cultivation. Key transcriptomic responses that are seemingly associated with different physiological responses of M1 were elucidated under normal light condition, the details of which would guide ongoing efforts to further maximize the industrial potential of violaxanthin producing strains.
Hispidulin alleviates 2,4-dinitrochlorobenzene and house dust mite extract-induced atopic dermatitis-like skin inflammation
No full textAtopic dermatitis (AD) is a chronic inflammatory skin disorder that affects 10？20% of the world’s population. Therefore, the discovery of drugs for the treatment of AD is important for human health. Hispidulin (HPD; also known as scutellarein 6-methyl ether or dinatin) is a natural flavone that exerts anti-inflammatory effects. In the present study, the effectiveness of HPD on AD-like skin inflammation was investigated. We used a mouse AD model through repeated exposure of mice to 2,4-dinitrochlorobenzene and house dust mite extract (Dermatophagoides farinae extract, DFE) to the ears. In addition, tumor necrosis factor-α and interferon-γ-activated keratinocytes (HaCaT cells) were used to investigate the underlying mechanism of HPD action. Oral administration of HPD alleviated AD-like skin inflammations: it reduced ear thickness; serum immunoglobulin (Ig)E, DFE-specific IgE, and IgG2a levels; and inflammatory cell infiltration. HPD reduced the expression of pro-inflammatory cytokines and chemokines through inhibition of signal transducer and activator of transcription 1 nuclear factor-κB in HaCaT cells. Taken together, these results suggest that HPD could be a potential drug candidate for the treatment of AD.
Rapid and highly sensitive pathogen detection by real-time DNA monitoring using a nanogap impedimetric sensor with recombinase polymerase amplification
No full textFast detection of pathogens is important for protecting our health and society. Herein, we present a high-performance nanogap impedimetric sensor for monitoring nucleic acid amplification in real time using isothermal recombinase polymerase amplification (RPA) for rapid pathogen detection. The nanogap electrode chip has two pairs of opposing gold electrodes with a 100？nm gap and was fixed to a PCB. Then, the nanogap impedimetric sensor was immersed in RPA reaction solution for the detection of E. coli O157:H7, and target DNA amplification was evaluated through bulk solution impedance changes using impedance spectroscopy every minute during RPA. In addition, target gene amplification in the sample solution during RPA was confirmed with a 2% DNA agarose gel. Our nanogap impedimetric sensor can detect down to a single copy of the eae A gene in gDNA extracted from E. coli O157:H7 as well as a single cell of pathogenic E. coli O157:H7 strain within 5？min during direct RPA, which was performed with the pathogen itself and without the extraction and purification of target gDNA. The miniaturized nanogap impedimetric sensor has potential as a cost-effective point-of-care device for fast and accurate portable pathogen detection via real-time nucleic acid analysis.
Overexpression of VlPRX21 and VlPRX35 genes in Arabidopsis plants leads to bioconversion of trans-resveratrol to δ-viniferin
No full textStilbenes, including resveratrol and viniferins, a small family of polyphenols, are considered the most important phytoalexin group in Vitis species. In a previous study, we found that co-treatment of methyl jasmonate (MJ) and stevioside (STE) resulted in enhanced extracellular production of viniferins in grapevine cell suspension cultures. Thus, to further understand the mechanisms of viniferin production in grapevine cell cultures, we performed transcriptome analysis and isolated seven candidates of grapevine peroxidase genes (VlAPX6, VlGPX5, VlPRX13, VlPRX21, VlPRX35, VlPRX40, and VlPRX50). Bioconversion of trans-resveratrol to δ-viniferin was examined using crude protein extracts isolated from agroinfiltration-based transient expression of VlPRXs in Nicotiana benthamiana. In addition, we found that crude protein extracts from VlPRX21-, VlPRX35-, and VlPRX40-overexpressing (OX) transgenic Arabidopsis plants led to the conversion of trans-resveratrol to δ-viniferin. We found that in vitro experiments with crude protein extracts from VlPRX21-OX and VlPRX35-OX Arabidopsis plants catalyzed the dimerization of trans-resveratrol to δ-viniferin. Our results suggest that VlPRX21 and VlPRX35 encode functional grapevine class III peroxidases and catalyze the oxidative dimerization of trans-resveratrol to form δ-viniferin in grapevine.
V-set and immunoglobulin domain-containing 1 (Vsig1), predominantly expressed in testicular germ cells, is dispensable for spermatogenesis and male fertility in mice
No full textTo elucidate the functional role of V-set and immunoglobulin domain-containing 1 (VSIG1) in spermatogenesis and fertilization, we knocked out (KO) VSIG1 in a mouse embryo using CRISPR/Cas9 (Clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9) -mediated genome editing. Reverse transcription PCR was performed using cDNA synthesized from VSIG1 KO testis RNA. Although Western blot analysis using a specific antibody to VSIG1 confirmed VSIG1 protein defects in the KO mice, hematoxylin-eosin staining analysis was similar in the KO and wild-type mice. Additionally, computer-assisted sperm analysis and in vitro fertilization experiments were conducted to confirm the activity and fertilization ability of sperm derived from the KO mouse. Mice lacking VSIG1 were viable and had no serious developmental defects. As they got older, the KO mice showed slightly higher weight loss, male mice lacking VSIG1 had functional testes, including normal sperm number and motility, and both male and female mice lacking VSIG1 were fertile. Our results from VSIG1 KO mice suggest that VSIG1 may not play essential roles in spermatogenesis and normal testis development, function, and maintenance. VSIG1 in sperm is dispensable for spermatogenesis and male fertility in mice. As several genes are known to possess slightly different functions depending on the species, the importance and molecular mechanism of VSIG1 in tissues of other species needs further investigation.
Gene expression in the striatum of cynomolgus monkeys after chronic administration of cocaine and heroin
No full textCocaine and heroin cause impairment of neural plasticity in the brain including striatum. This study aimed to identify genes differentially expressed in the striatum of cynomolgus monkeys in response to cocaine and heroin. After chronic administration of cocaine and heroin in the monkeys, we performed large？scale transcriptome profiling in the striatum using RNA？Seq technology and analysed functional annotation. We found that 547 and 1238 transcripts were more than 1.5？fold up？ or down？regulated in cocaine？ and heroin？treated groups, respectively, compared to the control group, and 3432 transcripts exhibited differential expression between cocaine？ and heroin？treated groups. Functional annotation analysis indicated that genes associated with nervous system development (NAGLU, MOBP and TTL7) and stress granule disassembly (KIF5B and KLC1) were differentially expressed in the cocaine？treated group compared to the control group, whereas gene associated with neuron apoptotic process (ERBB3) was differentially expressed in the heroin？treated group. In addition, IPA network analysis indicated that genes (TRAF6 and TRAF3IP2) associated with inflammation were increased by the chronic administration of cocaine and heroin. These results provide insight into the correlated molecular mechanisms as well as the upregulation and down？regulation of genes in the striatum after chronic exposure to cocaine and heroin.
Pancreatic cancer induces muscle wasting by promoting the release of pancreatic adenocarcinoma upregulated factor
No full textCancer cachexia is a highly debilitating condition characterized by weight loss and muscle wasting that contributes significantly to the morbidity and mortality of pancreatic cancer. The factors that induce cachexia in pancreatic cancer are largely unknown. We previously showed that pancreatic adenocarcinoma upregulated factor (PAUF) secreted by pancreatic cancer cells is responsible for tumor growth and metastasis. Here, we analyzed the relation between pancreatic cancer-derived PAUF and cancer cachexia in mice and its clinical significance. Body weight loss and muscle weight loss were significantly higher in mice with Panc-1/PAUF tumors than in those with Panc-1/Mock tumors. Direct administration of rPAUF to muscle recapitulated tumor-induced atrophy, and a PAUF-neutralizing antibody abrogated tumor-induced muscle wasting in Panc-1/PAUF tumor-bearing mice. C2C12 myotubes treated with rPAUF exhibited rapid inactivation of Akt-Foxo3a signaling, resulting in Atrogin1/MAFbx upregulation, myosin heavy chain loss, and muscle atrophy. The neutrophil-to-lymphocyte ratio and body weight loss were significantly higher in pancreatic cancer patients with high PAUF expression than in those with low PAUF expression. Analysis of different pancreatic cancer datasets showed that PAUF expression was significantly higher in the pancreatic cancer group than in the nontumor group. Analysis of The Cancer Genome Atlas data found associations between high PAUF expression or a high DNA copy number and poor overall survival. Our data identified tumor-secreted circulating PAUF as a key factor of cachexia, causing muscle wasting in mice. Neutralizing PAUF may be a useful therapeutic strategy for the treatment of pancreatic cancer-induced cachexia.
The role of macrophages in the development of acute and chronic inflammatory lung diseases
No full textMacrophages play an important role in the innate and adaptive immune responses of organ systems, including the lungs, to particles and pathogens. Cumulative results show that macrophages contribute to the development and progression of acute or chronic inflammatory responses through the secretion of inflammatory cytokines/chemokines and the activation of transcription factors in the pathogenesis of inflammatory lung diseases, such as acute lung injury (ALI), acute respiratory distress syndrome (ARDS), ARDS related to COVID-19 (coronavirus disease 2019, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)), allergic asthma, chronic obstructive pulmonary disease (COPD), and idiopathic pulmonary fibrosis (IPF). This review summarizes the functions of macrophages and their associated underlying mechanisms in the development of ALI, ARDS, COVID-19-related ARDS, allergic asthma, COPD, and IPF and briefly introduces the acute and chronic experimental animal models. Thus, this review suggests an effective therapeutic approach that focuses on the regulation of macrophage function in the context of inflammatory lung diseases.
Development of human parabiosis models from stem cell-derived lung organoids
No full text줄기세포 폐오가노이드 기반 인간 파라바이오시스 모델 개발KGM113201