Apigenin is a flavonoid of low toxicity and multiple beneficial bioactivities. Published reviews all focused on the findings using eukaryotic cells, animal models, or epidemiological studies covering the pharmacokinetics, cancer chemoprevention, and drug interactions of apigenin; however, no review is available on the antimicrobial effects of apigenin. Research proves that dietary apigenin passes through the upper gastrointestinal tract and reaches the colon after consumption. For that reason, it is worthwhile to study the potential interactions between apigenin and human gut microbiota. This review summarizes studies on antimicrobial effects of apigenin as well as what has been reported on apigenin and human gut microbiota. Various levels of effectiveness have been reported on apigenin's antibacterial, antifungal, and antiparasitic capability. It has been shown that apigenin or its glycosides are degraded into smaller metabolites by certain gut bacteria which can regulate the human body after absorption. How apigenin contributes to the structural and functional changes in human gut microbiota as well as the bioactivities of apigenin bacterial metabolites are worth further investigation.
The Twin Simulator of the Human Intestinal Microbial Ecosystem (TWINSHIME®) was initially developed to study the luminal gut microbiota of the ascending (AC), transverse (TC), and descending (DC) colon regions. Given the unique composition and potential importance of the mucosal microbiota for human health, the TWINSHIME was recently adapted to simulate the mucosal microbiota as well as the luminal community. It has been previously demonstrated that the luminal community in the TWINSHIME reaches a steady state within two weeks post inoculation, and is able to differentiate into region specific communities. However, less is known regarding the mucosal community structure and dynamics. During the current study, the luminal and mucosal communities in each region of the TWINSHIME were evaluated over the course of six weeks. Based on 16S rRNA gene sequencing and short chain fatty acid analysis, it was determined that both the luminal and mucosal communities reached stability 10–20 days after inoculation, and remained stable until the end of the experiment. Bioinformatics analysis revealed the formation of unique community structures between the mucosal and luminal phases in all three colon regions, yet these communities were similar to the inoculum. Specific colonizers of the mucus mainly belonged to the Firmicutes phylum and included Lachnospiraceae (AC/TC/DC), Ruminococcaceae and Eubacteriaceae (AC), Lactobacillaceae (AC/TC), Clostridiaceae and Erysipelotrichaceae (TC/DC). In contrast, Bacteroidaceae were enriched in the gut lumen of all three colon regions. The unique profile of short chain fatty acid (SCFA) production further demonstrated system stability, but also proved to be an area of marked differences between the in vitro system and in vivo reports. Results of this study demonstrate that it is possible to replicate the community structure and composition of the gut microbiota in vitro. Through implementation of this system, the human gut microbiota can be studied in a dynamic and continuous fashion.
Recombinant adeno-associated viral (rAAV) vectors have gained attention for human gene therapy because of their high safety and clinical efficacy profile. For factor VIII gene delivery, splitting the coding region between two AAV vectors remains a viable strategy to avoid the packaging capacity limitation (*5.0 kb). However, it is time-consuming and labor-intensive to produce two rAAV vectors in separate batches. Here we demonstrated successful production of dual rAAV vectors for hemophilia A gene therapy in a single preparation. When the AAV vector plasmids carrying the human factor VIII heavy chain (hHC) and the light chain (hLC) expression cassettes were cotransfected into 293 cells along with the AAV rep&cap and mini-adenovirus helper plasmids, both rAAV-hHC and rAAV-hLC were produced at the desired ratio and in high titer. Interestingly, the rAAVhHC vectors always yielded higher titers than rAAV-hLC vectors as a result of more efficient replication of rAAV-hHC genomes. The resulting vectors were effective in transducing the tissue culture cells in vitro. When these vectors were administered to hemophilia A mice, factor VIII was detected in the mouse plasma by both the activated partial thromboplastin time assay and enzyme-linked immunosorbent assay. The functional activity as well as the antigen levels of secreted factor VIII were similar to those of vectors produced by the traditional method. The dual-vector production method has been successfully extended to both AAV2 and AAV8 serotypes. In conclusion, cotransfection of vector plasmids presents an efficient method for producing dual or multiple AAV vectors at significantly reduced cost and labor.
Environmental pH is a critical parameter for maintenance of the gut microbiota. Here, the impact of pH on the gut microbiota luminal and mucosal community structure and short chain fatty acid (SCFA) production was evaluated in vitro, and data compiled to reveal a donor-independent response to an increase or decrease in environmental pH. The results found that raising environmental pH significantly increased luminal community richness and decreased mucosal community evenness. This corresponded with an increased abundance of Ruminococcaceae Ruminococcus, and Erysipelotrichaceae Erysipelatoclostridium, and a decreased abundance of Coriobacteriaceae Collinsella, and Enterobacteriaceae Shigella for both the luminal and mucosal communities. Total SCFA levels were significantly higher, primarily due to an increase in acetic and 2-methylbutanoic acids. Lowering pH decreased luminal community evenness and decreased mucosal community evenness and richness. This corresponded with an increased abundance of Lachnospiraceae Enterocloster, Veillonellaceae Megasphaera, Veillonellaceae Sporomusa, Erysipelotrichaceae Eubacterium, and Alcaligenaceae Sutterella, and decreased abundance of Odoribacteraceae Butyricimonas, Fusobacteriaceae Fusobacterium, Veillonellaceae Phascolarctobacterium, and multiple Enterobacteriaceae species for both the luminal and mucosal communities. Total SCFA levels were significantly lower, with an observed drop in acetic and propionic acids, and increased butyric and valeric acids. Taken together, these results indicate that alterations to environmental pH can modulate the gut microbiota community structure and function, and some changes may occur in a donor-independent manner.
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