探討在蠕蟲與腸道型病原菌共同感染中蠕蟲引起的腸道破損對宿主的影響

Abstract

先前研究顯示，蠕蟲感染可能對宿主對抗後續病原體感染的免疫反應產生正向或負向影響，而這些相互矛盾的結果可能受到多種因素影響。我們實驗室建立了一個蠕蟲－細菌雙重感染的小鼠模型，並發現先前感染蠕蟲（Heligmosomoides polygyrus bakeri，簡稱 H. polygyrus 或 Hpb）的小鼠經過驅蟲治療後，在面對非相關腸道細菌病原（Yersinia pseudotuberculosis，Ypt）感染時具有保護作用；然而，若是感染蠕蟲但沒有驅蟲藥物治療的小鼠則對 Ypt 感染更為易感，造成此相反結果的原因尚不清楚。 我假設蠕蟲感染造成的腸道上皮屏障破壞，可能是導致宿主對後續病原感染易感性增加的因素之一。在我的研究中發現，H. polygyrus 感染會導致近端小腸組織明顯受損，並增加宿主腸道屏障的通透性，而在驅蟲治療後，通透性可恢復正常。這些結果顯示，蠕蟲感染會導致腸道滲漏，而受蠕蟲破壞的腸道在驅蟲治療後可被修復。此外，與有驅蟲治療的小鼠相比，沒有驅蟲治療的小鼠其空腸的連接蛋白claudin-15和myosin light chain kinase (MLCK) 基因表現有顯著上調的現象，而 claudin-3、occludin 的基因表現則下降。這些結果顯示claudin-15和MLCK可能與 H. polygyrus 感染後腸道通透性增加有關。此外，持續受蠕蟲感染的小鼠其空腸TNF-α的基因表現量也有顯著上調，顯示其有可能與腸道通透性的調控有關。另外，在腸道特異性缺失zonula occluden-1 (ZO-1KO.IEC) 小鼠的實驗中，發現此缺失似乎不會導致ZO-1KO.IEC小鼠對Ypt感染更加易感，然而菌落在不同組織的分布有所不同。 最後，我探討Ypt感染是否會使宿主原本已被蠕蟲破壞的腸道破損更嚴重，導致未驅蟲治療的宿主面對Ypt後更易感。雖然結果顯示Ypt並沒有加劇腸道破損，然而在雙重感染但未驅蟲的小鼠腸道中發現claudin-2和MLCK的基因表現有上調現象。而腸道IFN-γ、TNF-α、IL-1β和IL-33基因表現量在不同處理小鼠之間皆無顯著差異，說明在雙重感染的小鼠其腸道緊密連接蛋白可能並非由這四種細胞激素調控。然而後續仍需要透過蛋白質定量才能確定這四種細胞激素在雙重感染的情況下是否有參與腸道緊密連接的變化。

Previous studies have shown that helminth infection can either positively or negatively impact the host's immunity to subsequent pathogen infections, and various factors may contribute to these conflicting outcomes. Our laboratory has established a mouse model of helminth-bacterial dual infection and found that prior helminth (Heligmosomoides polygyrus bakeri, H. polygyrus, or Hpb) infection can protect the host from subsequent unrelated intestinal bacterial pathogen (Yersinia pseudotuberculosis, Ypt) following deworm treatment (i.e., cured). In contrast, helminth-infected mice without deworm treatment (i.e., uncured) were more susceptible to Ypt infection, but the reason for this opposite outcome is unclear. Here, I hypothesized that intestinal epithelial barrier disruption caused by helminth infection is one of the factors that increase host susceptibility to subsequent pathogen infections. In my study, I found significant damage in the proximal small intestine of H. polygyrus-infected mice, and confirmed that helminth infection increased intestinal barrier permeability. Importantly, the permeability was restored in cured mice. These findings suggested that helminth infection can lead to gut leakage, and the intestines disrupted by helminth are likely repaired after deworm treatment. Furthermore, compared to cured mice, gene expressions of jejunal claudin-15 and myosin light chain kinase (MLCK) were significantly upregulated in uncured mice, while the claudin-3 and occludin levels were downregulated, suggesting that claudin-15 and MLCK may contribute to increased gut permeability after H. polygyrus infection. The gene expression of TNF-α was significantly upregulated in uncured mice, suggesting that it may be involved in gut permeability regulation. Furthermore, using intestinal epithelial-specific deletion of zonula occluden-1 (ZO-1KO.IEC) mice, I found the overall susceptibility of ZO-1KO.IEC mice to Ypt infection did not seem to increase, but the bacterial distribution was changed. Lastly, I explored whether Ypt infection would exacerbate helminth-induced intestinal damage and thereby increase susceptibility in the helminth-bacterial-dual infection model. Although the results did not indicate aggravated gut damage following Ypt infection, the gene expressions of intestinal claudin-2 and MLCK were upregulated in Ypt-infected uncured mice. Moreover, the gene expression levels of IFN-γ, TNF-α, IL-1β, and IL-33 were comparable among different treatment groups, suggesting that the expression of tight junctional proteins in dual-infected mice may not be regulated by these four cytokines. Protein-level quantification will be required to clarify whether these cytokines are involved in tight junction modulation during dual-infection.