Suppression of Myd88 Disturbs Gut Microbiota and Activates the NLR Pathway and Hence Fails to Ameliorate DSS-Induced Colitis

Life Sciences Gastroenterology Medicine Immunology
Myd88 gut microbiota NLR pathway colitis inflammatory bowel disease

Inflammatory Bowel Disease (IBD) is a chronic, relapsing inflammatory disorder of the gastrointestinal tract, primarily including Ulcerative Colitis (UC) and Crohn’s Disease (CD). In recent years, the incidence of IBD has been on the rise globally, placing a significant burden on healthcare systems. Although it is widely accepted that IBD is associated with microbial dysbiosis, exaggerated immune responses, impaired intestinal barrier function, and genetic susceptibility, its exact etiology remains unclear. Current treatment strategies have limited efficacy in achieving long-term remission and are often accompanied by significant side effects.

Myeloid Differentiation Factor 88 (Myd88) is the core adaptor protein of Toll-like Receptors (TLRs) and plays a crucial role in host defense against microbial invasion and the initiation of downstream immune responses. However, the role of Myd88 in the pathogenesis of IBD is controversial. Some studies suggest that Myd88 promotes inflammatory responses by activating the nuclear factor kappa-B (NF-κB) pathway, potentially making it a therapeutic target for IBD; while others have found that the absence of Myd88 increases susceptibility to intestinal inflammation. Therefore, further research into the role of Myd88 in intestinal inflammation and its underlying mechanisms is of great importance.

Source of the Paper

This paper was co-authored by Jun-Hua Li, Yu Chen, Zheng-Hao Ye, and others, from the Department of Nephrology, Department of Gastroenterology, and Institute of Organ Transplantation at Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, with some authors affiliated with Hannover Medical School in Germany. The paper was published on May 6, 2024, in the journal Precision Clinical Medicine, with the DOI 10.1093/pcmedi/pbae013.

Research Process

1. Study Design and Experimental Subjects

This study used Myd88 knockout (Myd88−/−) mice and the Myd88 inhibitor (TJ-M2010-5) to investigate the role of Myd88 in acute dextran sodium sulfate (DSS)-induced colitis. The experimental groups included: - Wild-type (WT) mice group - WT-DSS group - Myd88−/− mice group - Myd88−/−-DSS group - TJ-M2010-5 treatment group

2. Disease Model Establishment and Treatment

The acute colitis model was established by administering 3% DSS in drinking water for 7 days. The Myd88 inhibitor TJ-M2010-5 was administered via intraperitoneal injection at a dose of 50 mg/kg/day starting one day before DSS treatment and continuing until the 7th day of DSS challenge. To eliminate the influence of gut microbiota, some mice were treated with broad-spectrum antibiotics (imipenem and vancomycin) 3 days before and during DSS treatment.

3. Disease Activity Index and Histological Scoring

Daily monitoring of body weight, stool consistency, and rectal bleeding was conducted to calculate the Disease Activity Index (DAI). Colon tissues were fixed and stained with hematoxylin and eosin (H&E), and histological scores (HS) were blindly evaluated by three researchers.

4. RNA Transcriptome and 16S rDNA Sequencing

RNA was extracted from colon tissues for transcriptome sequencing to analyze differentially expressed genes and related pathways. Meanwhile, fecal samples were collected for 16S rDNA sequencing to analyze changes in gut microbiota composition.

5. Protein Expression and Signaling Pathway Analysis

Western blotting was used to detect the expression levels of Myd88, NF-κB, and NLR pathway-related proteins, analyzing the impact of Myd88 inhibition on inflammatory signaling pathways.

Main Results

1. Myd88 Inhibition Failed to Alleviate DSS-Induced Colitis

Although the activation levels of NF-κB were significantly lower in Myd88−/− mice and TJ-M2010-5-treated mice compared to the control group, the severity of colitis (DAI, colon length, and histological score) showed no significant difference. Additionally, Myd88 inhibition did not significantly reduce the expression levels of pro-inflammatory cytokines (such as TNF-α, IFN-γ, and IL-1β) in colon tissues.

2. Myd88 Inhibition Altered Gut Microbiota Composition

16S rDNA sequencing results revealed that Myd88 inhibition led to unfavorable changes in gut microbiota composition, primarily characterized by an increase in Proteobacteria abundance and a decrease in Firmicutes abundance. In the DSS-induced colitis model, the abundance of Proteobacteria was significantly higher in TJ-M2010-5-treated mice compared to the control group.

3. Upregulation of the NLR Signaling Pathway

RNA transcriptome analysis showed that the expression of NLR signaling pathway-related genes was significantly upregulated after Myd88 inhibition. Further experiments demonstrated that blocking the NLR signaling pathway or treatment with broad-spectrum antibiotics improved the severity of colitis that was not alleviated by Myd88 inhibition alone.

4. Antibiotic Treatment Improved Colitis

Broad-spectrum antibiotic treatment significantly reduced the abundance of Proteobacteria in Myd88-inhibited mice and downregulated the activation of the NLR signaling pathway, thereby improving the severity of colitis.

Conclusion

This study indicates that while Myd88 inhibition can partially suppress the activation of the NF-κB pathway, it leads to unfavorable changes in gut microbiota composition, subsequently activating the NLR signaling pathway and ultimately failing to alleviate DSS-induced colitis. This finding reveals the complex role of Myd88 in intestinal inflammation, suggesting that simply inhibiting Myd88 may not be sufficient for treating IBD, and a combined approach targeting gut microbiota or the NLR signaling pathway may be a more effective therapeutic strategy.

Research Highlights

  1. Complex Role of Myd88 Inhibition: The study found that while Myd88 inhibition suppresses the NF-κB pathway, it activates the NLR pathway by altering gut microbiota, leading to unmitigated colitis. This provides new insights into the application of Myd88 in IBD treatment.

  2. Interaction Between Gut Microbiota and Immune Pathways: The research highlights the interaction between gut microbiota and the NLR signaling pathway, emphasizing the important role of gut microbes in inflammatory bowel disease.

  3. Proposal of Combined Therapeutic Strategies: The results suggest that simply inhibiting Myd88 may not be sufficient for treating IBD, and a combined approach targeting gut microbiota or the NLR signaling pathway may be more effective.

Research Significance

This study not only reveals the complex role of Myd88 in intestinal inflammation but also provides new insights for IBD treatment. By jointly regulating gut microbiota and immune signaling pathways, more effective IBD treatment strategies may be developed in the future. Additionally, this research offers important experimental evidence for understanding the interaction between gut microbiota and the immune system.