Propionate-engineered probiotics reduce radiation-induced intestinal damage
Research Background
Radiation enteritis is a common complication of abdominal radiotherapy patients, and it is also an important pain point in the clinical application of tumor radiotherapy. As the core treatment method of malignant tumors, radiotherapy will inevitably damage the intestinal mucosa and other healthy tissues, often leading to the serious complication of radiation enteritis, and can also cause chronic lesions such as intestinal stenosis, bleeding, and fistula, which seriously affects the quality of life of patients and even affects the process of radiotherapy. At present, clinical treatment methods for radiation enteritis are limited, mainly limited to symptom relief, and lack of internationally recognized specific preventive treatment drugs. Existing drugs such as octreotide and amifostine have limited clinical application due to high cost, large side effects, and insufficient targeting.
Propionic acid in short URL fatty acids has a good radiation protection effect. It can protect the intestinal mucosal barrier and promote repair by upregulating intestinal tight junction protein, promoting intestinal epithelial renewal and repair, and regulating immune inflammation. However, the direct oral supplementation of propionic acid has the disadvantages of low bioavailability and short-term effect, and cannot simulate the physiological state of continuous and local propionic acid production by intestinal flora.
In response to the above issues, this study designed an engineered probiotic that can continuously produce propionic acid in situ in the gut. The strain targets intestinal microbes and host signaling pathways by releasing propionic acid, achieving a synergistic effect of continuous propionic acid delivery and probiotic intestinal repair function. Related research results are published in Bioresources and Bioprocessing
Research ideas
This study used EcN as the chassis. By introducing the propionic acid synthesis pathway (acrylic acid pathway) and knocking out the acetic acid synthesis bypass genes (pflB, poxB), an engineered probiotic PG was constructed, which can achieve sustained and in-situ production of propionic acid. The engineered bacteria overcome the problems of low bioavailability and short-lived effect of direct propionic acid supplementation, and can achieve targeted delivery through probiotic colonization characteristics.
Subsequently, the researchers verified the function and efficacy of PG-engineered probiotics in in vitro fermentation experiments and abdominal irradiation models of C57BL/6J mice. The optimal propionic acid yield and fermentation conditions were clarified, and it was proved that it could repair intestinal villi/crypt structures, inhibit inflammation and oxidative stress, and ultimately achieve efficient protection against radiation-induced intestinal damage.
Key experimental results
First, using E.coli Nissle 1917 (EcN) as the chassis, the complete acrylic pathway (including pct, lcd, acr genes) and the acetic acid synthesis bypass genes (pflB, poxB) were successfully introduced to construct PG engineering probiotics, and it was verified that knocking out the bypass metabolic pathway was an effective strategy to increase the yield of target products. The growth curve analysis of the engineered strain PG showed no significant difference from the original EcN strain and the no-load control strain, indicating that the introduced foreign genes and metabolic modifications did not adversely affect the normal growth of the probiotic host itself, providing preliminary evidence for subsequent in vivo application.
In the mouse model of radiation enteritis, the PG strain exhibited significant therapeutic effect. By comparing the three radiation doses of 8 Gy, 12 Gy and 16 Gy, it was found that PG had the most significant protective effect at the 12 Gy dose, which could effectively increase the level of propionic acid in the intestine, reverse the metabolite loss caused by radiation, reduce the atrophy of the digestive tract and promote the recovery of intestinal length. At the same time, the pro-inflammatory factor IL-22 and the anti-inflammatory factor IL-10 were up-regulated, which effectively reduced the intestinal inflammatory response. It was shown that the engineered probiotic PG could achieve multi-dimensional protection against radiation enteritis by delivering propionic acid continuously and in situ.
Histopathological analysis further confirmed the significant protective effect of the engineered probiotic PG on radiation-induced intestinal injury. The results of HE staining showed that radiation caused severe damage such as typical villus atrophy, crypt reduction and inflammatory cell infiltration in the intestines of mice, while the intestinal tissue structure was significantly improved after PG intervention. The villi were more complete, the crypt morphology was normal, and the inflammatory infiltration was significantly reduced. Quantitative analysis showed that PG could effectively improve the villus height/crypt depth ratio of the ileum and jejunum under three radiation doses of 8 Gy, 12 Gy and 16 Gy, among which the repair effect was the most significant at the dose of 12 Gy.
Through 16S rRNA gene sequencing technology, this study found that the engineered probiotic PG can effectively regulate the imbalance of intestinal flora in mice with radiation enteritis. Compared with the radiation group, PG intervention significantly improved the richness and diversity of intestinal flora, restored the normal abundance of Proteobacteria at the phyla level, and significantly increased the proportion of Verrucella. At the genus level, PG selectively promoted the proliferation of a variety of beneficial bacteria, while effectively inhibiting the excessive proliferation of radiation-induced harmful bacteria and rebuilding the homeostasis of flora.
Further experimental analysis revealed the key molecular mechanism by which the engineered probiotic PG alleviates radiation-induced intestinal injury. Through KEGG pathway enrichment analysis, it was found that the differentially expressed genes after PG intervention were mainly enriched in the JAK-STAT signaling pathway, which plays a central role in maintaining inflammatory homeostasis. The analysis showed that PG significantly down-regulated key genes in the JAK-STAT pathway, and significantly upregulated the expression of negative feedback regulator Socs1, of which Jak2, Stat3 and Socs1 were the most significant changes. This result shows that the engineered probiotic PG effectively alleviated radiation-induced intestinal inflammation and tissue damage by regulating the SOCS1/JAK2/STAT3 signaling axis and inhibiting the abnormal activation of the JAK-STAT signaling pathway. This discovery elucidates the mechanism of PG's protection against radiation-induced intestinal injury at the molecular level, providing an important theoretical basis for the clinical application of engineered probiotics.
Research Significance and Prospects
This study expands the application of synthetic biology in the treatment of radiation intestinal injury, and constructs an engineered probiotic PG that can continuously and in situ produce propionate. It realizes multi-dimensional synergistic protection against radiation intestinal injury by regulating intestinal flora homeostasis, remodeling intestinal metabolic spectrum, and regulating SOCS1/JAK2/STAT3 inflammatory signaling pathway. This strategy not only overcomes the limitations of low bioavailability and short-term effect of direct propionate supplementation, but also makes up for the shortcomings of traditional probiotics in targeting, providing a new biological therapy idea for the clinical prevention and treatment of radiation enteritis. In addition, the design of the engineered probiotic has good safety and stability, and it is expected to be further expanded to the precision treatment of other intestinal diseases in the future.
