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. 2024 Mar 8;72(11):5710–5724. doi: 10.1021/acs.jafc.3c03432

Exploring the Role of Inulin in Targeting the Gut Microbiota: An Innovative Strategy for Alleviating Colonic Fibrosis Induced By Irradiation

Kaihua Ji 1, Manman Zhang 1, Liqing Du 1, Jinhan Wang 1, Yang Liu 1, Chang Xu 1, Ningning He 1, Qin Wang 1, Yeqing Gu 1, Huijuan Song 1, Yan Wang 1,*, Qiang Liu 1,*
PMCID: PMC10958509  PMID: 38457473

Abstract

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The use of radiation therapy to treat pelvic and abdominal cancers can lead to the development of either acute or chronic radiation enteropathy. Radiation-induced chronic colonic fibrosis is a common gastrointestinal disorder resulting from the above radiation therapy. In this study, we establish the efficacy of inulin supplements in safeguarding against colonic fibrosis caused by irradiation therapy. Studies have demonstrated that inulin supplements enhance the proliferation of bacteria responsible to produce short-chain fatty acids (SCFAs) and elevate the levels of SCFAs in feces. In a mouse model of chronic radiation enteropathy, the transplantation of gut microbiota and its metabolites from feces of inulin-treated mice were found to reduce colonic fibrosis in validation experiments. Administering inulin-derived metabolites from gut microbiota led to a notable decrease in the expression of genes linked to fibrosis and collagen production in mouse embryonic fibroblast cell line NIH/3T3. In the cell line, inulin-derived metabolites also suppressed the expression of genes linked to the extracellular matrix synthesis pathway. The results indicate a novel and practical approach to safeguarding against chronic radiation-induced colonic fibrosis.

Keywords: inulin, chronic radiation-induced colonic fibrosis, gut microbiota-derived metabolites, short-chain fatty acids (SCFAs), extracellular matrix synthesis pathway

Introduction

In the past few years, radiotherapy has become a major supplementary treatment for malignant tumors in the intraperitoneal, retroperitoneal, and pelvic areas.1−4 In a recent study, it was found that over 20% of individuals with abdominal or pelvic tumors underwent radiotherapy.5 Radiation enteropathy refers to an intestinal damage resulting from radiation treatment administered to malignant tumors in the abdomen and pelvis, which can be categorized into acute and chronic two types.6 The main reason for the limitation in the radiation dose of radiotherapy is radiation enteropathy, which directly impacts the effectiveness of cancer therapy. Intestinal fibrosis caused by radiation represents a grave form of long-term enteropathy, marked by an overabundance of extracellular matrix (ECM) elements, potentially leading to intestinal scarring and dysfunction.7 Consequently, it is essential to explore ways to prevent and treat radiation-induced intestinal disease, particularly chronic radiation-induced intestinal fibrosis.

Studies have suggested a potential connection between the gut microbiota in the digestive system and the prognosis, prevention, or management of radiation treatment induced enteropathy. Research have indicated that radiation may disrupt gut microbiota balance, leading to use of probiotics, prebiotics, particular diets, and microbiome restoration to focus on microbiome in managing radiation enteropathy.8−10 The results of a mouse model demonstrate that FMT improves gut health and intestinal integrity postirradiation, indicating that microbiome restoration might be beneficial in managing radiation enteropathy.11 Studies have shown that probiotics are safe and effective matters to regulate the microbial population and reduce the effects of radiation enteropathy in humans.12 A preliminary investigation conducted on individuals suffering from radiation enteropathy revealed that the utilization of low-fermented diets, such as monosaccharides, disaccharides, oligosaccharides, and polyols is believed to reduce radiation-induced enteropathy symptoms, albeit without evaluating of the gut microbiota.13 In addition, Lam and colleagues observed variations in microbial SCFAs metabolic pathway levels between individuals with and without radiation enteropathy.6 Patients exhibiting signs of radiation enteropathy experienced a reduction in the number of microbial SCFA metabolic pathways. These studies provide a new perspective on preventing and treating radiation enteropathy by focusing on gut microbiota, particularly those bacteria that produce SCFAs.

Studies have shown that inulin, a prebiotic, can greatly enhance physiological performance. Inulin was reported to facilitate the proliferation of intestinal epithelial cells, inhibit colonic atrophy, diminish the invasion of microbiota into the mucosa, and nourish the gut microbiota, thereby offering defense against metabolic syndrome in a manner dependent on microbiota and IL-22.14 Schroeder and colleagues discovered that the utilization of the probiotics Bifidobacteria or inulin successfully maintained the normal microbial community and mitigated the harm to the colon mucus layer in mice that were induced by a Western-style diet.15 A double-blinded population research revealed that the combination of soluble dietary fiber including inulin effectively targeted gut microbiota, thereby enhancing the proliferation of those bacteria which can produce SCFAs and significantly reducing the evolution of type 2 diabetes.16 Furthermore, prior research had indicated that SCFAs produced by gut microbiota fermented inulin are associated with the enhancement of the advantageous aspects of host metabolism and alleviation of inflammation.17 To conclude, it appears that inulin has the potential to influence the gut microbiota, particularly bacteria that produce SCFAs, thus impacting the body’s normal function.

The current research explored the possible protective properties of inulin, the gut microbiota in mice treated with inulin, and SCFAs produced by fermentation of inulin by gut microbiota in a mouse model of radiation-induced colonic fibrosis. Additionally, we investigated the potential reverse relationship between microbiota-generated inulin-derived SCFAs and the synthesis of the extracellular matrix in fibroblasts.

Results

Inulin Alleviated Radiation-Induced Intestinal Fibrosis

Prior research had shown that fermentable fiber inulin can promote intestinal epithelial growth and prevent colonic atrophy.14 Additionally, Schroeder et al. discovered that the utilization of inulin effectively alleviated the functional abnormalities in the mucosal layer of colon caused by a Western Style Diet in mice.15 Hence, the primary objective of the current research was to ascertain if the administration of inulin can alleviate intestinal fibrosis caused by radiation. The C57BL/6J mice were divided into four groups and subjected to receive the treatments as illustrated in Figure S1A. On the 120th day postradiation exposure, all mice were euthanized, and an analysis was conducted on their body weights, along with the lengths of both the small intestine and colon and the relative length of each. The IR group and inulin + IR group exhibited a significant decrease in body weight (p < 0.005 for IR; p < 0.01 for inulin + IR) in comparison to the control group, as depicted in Figure 1A. In contrast to the control and inulin groups, the IR group exhibited a reduced length of the small intestine (Figure S1B). Nevertheless, the inulin, IR, and inulin + IR groups exhibited a greater relative length of the small intestine compared to the control group (Figure S1B). The IR group exhibited a notable decrease in colon length in comparison to the control group (Figure 1B). The inulin + IR group exhibited longer colon lengths and relative colon lengths compared to the IR group (Figure 1B,C). The discovery of elongated colon length supports the hypothesis that the inulin, a fermentable diet fiber, prevents colon injury caused by radiation (Figure 1B,C).

Figure 1.

Figure 1

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Inulin supplementation prevented radiation-induced intestine damage and colon fibrosis. (A) Weight of the body. (B) Colon’s length and colon’s relative length. (C) Illustrative colons image captured from a single mouse within each group. (D) Colon tissue staining using Haematoxylin-eosin (HE) stain, Masson trichrome stain, and immunohistochemistry (alpha-SMA and Collagen III) stain techniques on the 120th day postradiation exposure. (E) Protein levels of alpha-SMA and GAPDH in colon tissue homogenate on the 120th day of postradiation exposure. (F) Quantification of the alpha-SMA protein levels. (G) mRNA levels quantification of alpha-Sma. Results were derived from a single experiment and presented as an average ± standard error (n = 6–10). * represents p < 0.05; ** represents p < 0.01; *** represents p < 0.001 (using one-way ANOVA, Tukey test). The areas indicated by the red arrow were the area with more collagen deposition. The areas circled by the red boxes were the areas with more positive staining for alpha-SMA protein. The areas circled by the blue boxes were the areas with more positive staining for Collagen III protein.

In order to assess the potential improvement of radiation-induced colon fibrosis caused by fermentable fiber inulin, we utilized hematoxylin and eosin, and Masson trichrome to stain colonic tissue sections. The colon’s histopathological analysis, conducted at 120 days after irradiation, revealed the presence of collagen deposition, confirmed through Masson trichrome and collagen III immunohistochemistry staining in the colonic tissue sections exposed to radiation (Figure 1D). The Masson trichrome and collagen III immunohistochemistry staining revealed a significant reduction in collagen deposition due to irradiation exposure when pretreated with inulin. The alpha-Sma expression (Figure 1G) and alpha-SMA protein level (Figure 1E,F), which were induced by radiation, were also reduced by administrating of inulin. To summarize, the presence of collagen deposition was linked to irradiation-induced colon injury, and the introduction of fermentable fiber inulin for a duration of 14 days prior to irradiation greatly alleviated the fibrosis of the colon.

Inulin Facilitates the Proliferation of SCFA-Producing Bacteria in the Gut Microbiota

Intestinal damage induced by radiation is associated with alterations in gut microbiota composition in mice.11 Additionally, inulin is a type of fermentable fiber that is recognized for its ability to enhance the proliferation of advantageous bacteria.14 Consequently, the composition of gut microbiota was investigated through high-throughput sequencing of the 16s rRNA gene V4 region. Five fecal specimens from each group were collected on the day before irradiation, as well as on the 30th day and 120th day of postirradiation exposure, and then examined using Illumina sequencing. The Shannon–Wiener index demonstrated that exposure to radiation led to alterations in the community composition’s richness at 120 days after irradiation in comparison to the samples collected before irradiation (Figure 2A). The Simpson index showed that community uniformity within the irradiation group at 120 days post-irradiation was significantly different from that in pre-irradiation samples, and significantly different from the control group and the inulin group at 120 days post-irradiation (Figure 2B). The beta-diversity of the gut microbiota in the IR and inulin + IR groups (II group) was significantly reduced at 30 days post-irradiation according to the weighted-UniFrac measure (Figure 2C,D). The inulin + IR group exhibited a greater beta-diversity at 120 days compared to the IR group, although this difference did not reach statistical significance (Figure 2C,D). The principal coordinates analysis revealed a notable disparity in community diversity between the inulin treatment group and the other groups after 120 days. Nevertheless, the beta diversity of the community was significantly affected by irradiation. The IR group exhibited minimal variation in beta diversity compared to the Inulin+IR group, both at the 30th day and the 120th day postirradiation (Figure 2E). The metastat examination showed a notable reduction in the IR group’s Bacteroidetes, in contrast to a significant rise in the Firmicutes, Proteobacteria, and Actinobacteria on the 120th day of postirradiation exposure (Figure S2A). The findings suggested that specific bacterial species may vary significantly between these groups. Subsequently, we employed an Lefse calculation to ascertain the influence of inulin administration and radiation exposure on the distinctive biomarkers. On the 120th day of postirradiation, Parasutterella, and Burkholderiaceae (Figure 2F and Figure S2B) were identified as the biomarkers for the IR group. Studies have suggested that Parasutterella may be linked to irritable bowel syndrome and persistent intestinal inflammation.18 There have been reports of certain Burkholderiaceae species being pathogenic microorganisms.19 The inulin + IR group was identified by the biomarkers Ruminococcaceae, Bacteroides acidifaciens, and Clostridium papyrosolvens (Figure 2F). It has been reported that all of these three biomarkers can generate SCFAs.20,21 The findings indicated that the bacterial composition of the mouse fecal flora is primarily influenced by irradiation, rather than inulin supplementation. Despite the minimal impact of inulin on the structure of the intestinal flora community, distinct biomarker characteristics were observed in the inulin + IR and IR groups.

Figure 2.

Figure 2

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Gut bacterial composition profile was modified by irradiation. (A) Shannon–Wiener index. (B) Simpson’s index. (C) The beta-diversity in each group was measured prior to irradiation, at 30 days and 120 days after irradiation, utilizing 16s rRNA gene high-throughput sequencing with a sample size of 5. A Student’s t test is used to determine the statistical significance of asterisks. The 75th and 25th quartile values are indicated by the top and bottom boundaries of each box, respectively, while the median values are indicated by the lines within each box. (D): Each group’s weighted UniFrac tree was assessed preirradiation, at the 30th day, and the 120th day after irradiation. (E) Each group’s Principal coordinate analysis were assessed preirradiation, at the 30th day, and the 120th day after irradiation. (F) Each group’s LefSe analysis were assessed at 120 days after irradiation. Species that exhibit a higher LDA score than 4 in each group were considered biomarkers.

Studies have shown that nondigestible carbohydrates, like inulin, aid in the growth of gut bacteria, which in turn produce SCFAs to alleviate inflammation and supply energy to colonocytes, among various other roles.22,23 The Lefse analysis revealed that inulin stimulated the proliferation of SCFAs-producing bacteria. Consequently, our analysis focused on the quantities of SCFAs-producing gut bacteria within the top 10 and 30 operational taxonomic units (OTUs) at the genus level in various treatment groups. Allobaculum,24Bacteroides,23Odoribacter,25 and Alloprevotella(26) have been found to generate SCFAs among the top 10 OTUs. On the 30th and 120th days of postradiation, the inulin + IR group exhibited a 14.5% and 20.6% rate of SCFAs-producing bacteria, respectively, a significant increase from the IR group’s 8.0 and 13.2% (Figure 3A,B). A comprehensive analysis of the pertinent literature affirms that Parasutterella,27Unidentified_Lachnospiraceae,28Bifidobacterium,23Unidentified_Clostridiales,23Blautia,23 and Intestinimonas(29) are capable of generating SCFAs as well. Among the top 30 OTUs, the inulin + IR group exhibited a 32.1% prevalence of SCFA-producers, surpassing the 23.8% observed in the IR group at the 120th day after radiation exposure (Figure 3C,D). The results of the pathway analysis indicated a significant rise in the pyruvate metabolism pathway within the inulin + IR group on the 120th day of postirradiation (Figure S3). Reports indicated that the primary route for producing SCFAs was the pyruvate metabolism pathway. To evaluate how pretreatment with inulin affects SCFAs levels in mouse feces, the SCFAs concentration in fecal specimens from both IR and inulin + IR groups were analyzed on the 120th day postirradiation. The inulin + IR group feces exhibited a significantly greater concentration of acetic acid and butyric acid compared to the IR group, as illustrated in Figure 3E,F. The results indicated that although inulin does not significantly alter the gut microbiota’s community structure and composition in mouse feces, it can stimulate the growth of SCFAs-producing gut bacteria in the intestines, consequently elevating the SCFAs levels in mouse feces.

Figure 3.

Figure 3

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In the intestinal flora, the proliferation of SCFAs-producing bacteria is facilitated by inulin. (A, C) The relative abundances of the top 10 (A) and top 30 (C) bacteria were analyzed at the genus level before irradiation, at the 30th day, and 120th day after irradiation in different groups. (B, D) Quantification of the SCFAs producers in the top 10 (B) and top 30 (D) bacteria OTUs at the genus level. (E, F) The concentrations of six SCFAs in the feces of IR and II groups were measured using gas chromatography–mass spectrometry (GC-MS) at the 120th day after irradiation. Data were obtained from one experiment and presented as mean ± standard error. * p < 0.05; ** p < 0.01; *** p < 0.001 (one-way ANOVA, Tukey test).

Transplantation of Inulin-Derived Fecal Microbiota and Gut Microbial Metabolites into the Mice Alleviate Irradiation-Induced Colonic Fibrosis

In order to prove that inulin-derived gut microbiota and gut microbial metabolites can reduce colonic fibrosis caused by irradiation, we treated mice with ABX, the broad-spectrum antibiotics, to significantly decrease the enteric microbiota by more than 99% and carried out fecal microbiota transplantation (FMT) and fecal microbiota metabolite transplantation (MeT) from inulin-treated and control mice (at 14 days) prior to irradiation (15 Gy ABI) as illustrated in Figure 4A. The mice were euthanized at 120 days after irradiation. Transplanting fecal microbiota metabolites from mice treated with inulin led to a significant increase in the body weight of these mice when compared to MeT from the control group following irradiation (Figure 4B). The irradiation-induced colonic shortening was significantly alleviated by transplanting gut microbial metabolites derived from inulin, in contrast to metabolites derived from the control group (Figure 4C). It is worth mentioning that the relative length of the colon did not yield identical outcomes. It is possible that the mice that were given metabolite transplantation from the control group had low body weights (Figure 4D). Furthermore, despite the lack of significant increases in colon length or its relative length in groups treated with inulin or those receiving inulin-derived gut microbiota transplants compared to the control or control-derived gut microbiota transplantation groups, an upward trend was noted (Figure 4C,D). The lack of significant differences in the above results may be due to limitations in the number of mice in groups (n = 5) (Figure 4C,D). Histological staining was employed to conduct the impact of FMT and MeT on colonic fibrosis in animals exposed to irradiation. Samples collected on day 120 postirradiation exhibited a significant reduction in colonic fibrosis following irradiation, as evidenced by hematoxylin and eosin staining, along with Masson staining, in mice treated with inulin or FMT/MeT from inulin-treated mice (Figure 4E–4G). The expression of alpha-Sma in mice receiving inulin or FMT from inulin-treated mice was significantly decreased in comparison to their counterparts (Figure 4H,I). Despite the mice receiving inulin-derived MeT showing no significant decrease in alpha-Sma expression compared to the control mice, there was a noticeable downward trend (Figure 4J). To sum up, these data suggested that transplanting inulin-derived gut microbiota and metabolites into the body can prevent the formation of colonic fibrosis caused by irradiation in vivo.

Figure 4.

Figure 4

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Inulin-derived FMT and MeT blocked the progression of irradiation-induced colonic fibrosis. (A) Scheme for FMT and MeT before ABI (15 Gy). The mice were sacrificed at the 120th day postirradiation. (B) Weight of the body. (C) Colon length in each group. (D) Colon relative length in each group (n = 5). The colon tissue was stained with hematoxylin-eosin (HE) and Masson trichrome (E–G) at the 120th day after irradiation in each group. (H–J) Quantification of the alpha-Sma mRNA level in the homogenate of colon tissue. Results were derived from a single experiment and presented as an average ± standard error (n = 5). * represents p < 0.05; *** represents p < 0.001 (one-wayANOVA, Tukey test).

Fibroblast Differentiation Was Reduced by Inulin-Derived Gut Microbial Metabolites and Sodium Butyrate (NaB) In Vitro

To establish whether the differentiation of fibroblast cell lines is inhibited by inulin-derived gut microbial metabolites or inulin-derived SCFAs, the NIH/3T3 cell line was subjected to treatment using gut microbial metabolites and monocomponent SCFAs. The metabolites derived from fecal were dissolved in PBS buffer, centrifugated, and filtered to eliminate contaminants and microorganisms. The protein level of alpha-SMA was evaluated using Western blot in the NIH/3T3 cell line. At 72 h after irradiation (10 Gy), there was a notable increase in the protein level of alpha-SMA (Figure 5A,B). The introduction of metabolites from both the control and inulin groups resulted in a substantial decline in the alpha-SMA protein levels. It is worth noting that the metabolites derived from the inulin group resulted in a substantial reduction in the alpha-SMA protein level, surpassing the decrease observed in the control group. The administration of NaB (1/5) resulted in a decrease in the alpha-SMA protein level (Figure 5F). The qPCR data showed that the alpha-Sma mRNA expression decreased significantly when cells were treated with NaB (1/5) (Figure 5G). This corresponded to the protein level. The findings indicated that both the control and inulin group metabolites had the potential to decrease the transcription and expression of alpha-Sma upon irradiation, while the inulin-derived metabolite group exhibited greater efficacy (Figure 5C). The alpha-Sma mRNA level exhibited a decrease upon treatment with NaB (Figure 5G). Furthermore, we utilized qPCR to examine the alterations in transcription levels of genes associated with collagen synthesis and fibroblast differentiation. The NIH/3T3 cell line exhibited a significant rise in the expression of tissue inhibitors of metalloproteinase 1 (Timp1) and tissue inhibitors of metalloproteinase 2 (Timp2) genes following irradiation. Nevertheless, the pretreated with inulin-derived metabolites resulted in a substantial reduction in the expression of Timp1 and Timp2, as opposed to the control-derived metabolites (Figure 5D,E). The findings indicated that fibroblast differentiation in the NIH/3T3 cell line can be diminished by inulin-derived fecal microbiota metabolites and monocomponent NaB.

Figure 5.

Figure 5

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In the NIH/3T3 cell line, the mRNA and protein levels of fibrosis-related genes were significantly diminished by the metabolites derived from inulin in fecal microbiota. (A) The protein levels of alpha-SMA and GAPDH were analyzed using Western blot analysis at 72 h after irradiation in the NIH/3T3 cell line. The cells were exposed to fecal microbiota metabolites 3 h prior to irradiation. (B) Quantification of the alpha-SMA expression levels. (C) Quantification of alpha-Sma transcription levels. (D, E) Quantification of fibrosis-related genes transcription levels in NIH/3T3 cell lines in each group. (F, G) Western blot analysis of alpha-SMA and GAPDH protein levels or quantification of alpha-Sma mRNA levels at 72 h postirradiation when treated with monocomponent NaAc (mother-liquor concentration 0.273g/L) or NaB (mother-liquor concentration 0.125g/L) in NIH/3T3 cell line. The TGF-beta 1-treated group was used as a fibrosis-positive control group. Three experiments yielded the results which were then presented as the mean value accompanied by the standard error. The statistical significance levels were denoted as * p < 0.05; ** p < 0.01; *** p < 0.001 (one-way ANOVA, Tukey test).

Inulin-Derived Fecal Microbiota Metabolites Altered the Gene Expression Profile Related to ECM in Fibroblasts Triggered by Irradiation In Vitro

To explore how inulin-derived fecal microbiota metabolites regulated fibroblast differentiation induced by irradiation, RNA sequencing was performed on the NIH/3T3 fibroblast cell line under exposure to either the control or inulin-derived fecal microbiota metabolites. The negative binomial distribution was employed to ascertain the importance of differential gene expression. Utilizing the formula |log2 (FoldChange)| > 1 and setting a Padj value below 0.05, it was ascertained that 395 genes exhibited significant differential expression due to their exposure to inulin-derived metabolites in the NIH/3T3 cell line (Figure S4A). 213 out of the 395 genes experienced a decrease in expression, while 182 genes showed an increase in the cells when treated with inulin-derived metabolites. Subsequently, the analysis of gene ontology (GO) revealed that, in comparison to cells treated with the control-derived metabolite, the majority of the GO terms in the cells treated with inulin-derived metabolite exhibiting notable variations in expression were associated with the extracellular matrix and collagen binding (Figure 6A). RNA-seq analysis revealed significant differences in the transcription levels of 16 genes involved in extracellular matrix and proteinaceous extracellular terms. The NIH/3T3 cells exposed to the control-derived metabolites treated group exhibited a significant increase in cysteine-rich secretory protein LCCL domain containing 2 (Crispld2), dermatopontin (Dpt), SPARC-related modular calcium binding 2 (Smoc2), lumican (Lum), nidogen 2 (Nid), laminin gamma 3 (Lamc3), thrombospondin domain containing 4 (Thsd4), collagen type XXVIII alpha 1 chain (Col28a1), ADAM metallopeptidase with thrombospondin Type 1 motif 15 (Adamts15), and retinoic acid responder 2 (Rarres2), and these genes were found to have a positive association with fibrosis.30−38 In the control treatment group, there was a notable reduction in the transcript levels of leucine rich repeat protein 3 (Lrrn3), which was believed to have a reverse correlation with fibrosis (Figure 6B).39 In contrast, NIH/3T3 cells treated with inulin-derived metabolites exhibited a significant increase in wingless-type MMTV integration site family 5b (Wnt5b) and Von Willebrand factor (Vwf), both recognized for their positive correlation with fibrosis (Figure 6B).40,41 The quantification of these genes via qPCR corresponded with the results of RNA sequencing (Figure 6C). We performed a statistical evaluation of the FPKM for genes in the RNA-seq database, given the upregulation of genes association with fibrosis in the group treated with inulin-derived metabolites. The FPKM of Wnt5b and Vwf were notably lower than that of other genes, as illustrated in Figure S4B. Subsequently, our examination focused on the expression and FPKM of 24 collagen synthesis-related genes in the sequencing data. Eleven genes out of these exhibited an increase in cells subjected to inulin-derived metabolites (Figure 6D), while 13 genes displayed an increase in cells exposed to control-derived metabolites (Figure 6D). The FPKMs of the 11 genes that exhibited high expression in cells treated with inulin-derived metabolites were generally significantly lower in comparison to the 13 genes that displayed high expression in the control group (Figure 6E). Collectively, these discoveries illustrated that metabolites derived from gut microbiota of inulin-treated mice have the ability to control the differentiation of fibroblasts by diminishing the transcription of genes associated with the extracellular matrix and collagen synthesis.

Figure 6.

Figure 6

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Gene expression profiles in the NIH/3T3 cell line were significantly altered and the mRNA levels of collagen synthesis-related genes were reduced by inulin-derived fecal microbiota metabolites. (A) Top 30 significantly enriched gene ontology (GO) terms induced by inulin-derived gut microbiota metabolites in the NIH/3T3 cell line. (B) Expression alterations of genes related to extracellular matrix and proteinaceous extracellular matrix gene ontology in the IM72h group versus the CM72h group. (C) Expression verification of the above genes via QPCR in the IM72h group versus the CM72h group. (D) Heatmap illustrating the variations in collagen synthesis-related genes expression between the CM72h group and IM72h groups. The symbol “#” indicates a high expression of genes related to collagen synthesis in the IM72h group. The symbol “*” indicates a high expression of genes related to collagen synthesis in the CM72h group. (E) FPKM analysis of genes expression related to collagen synthesis in the IM72h versus CM72h groups.

Discussion

This research demonstrated that the soluble dietary fiber prebiotics inulin can provide considerable defense against radiation-induced chronic colonic fibrosis by acting on the gut microbiome-metabolome network. The consumption of prebiotic inulin was linked to a rise in bacteria that produce SCFAs and the increase production of SCFAs in the feces of mice. Transplanting ABX-treated mice with gut microbiota or its metabolites derived from inulin pretreated mice prior to irradiation can significantly reduce the chronic intestinal fibrosis caused by irradiation. The treatment of inulin-derived metabolites to mouse embryonic fibroblasts resulted in a notable reduction in the gene expression associated with fibrosis and the extracellular matrix. This research demonstrated the potential of soluble dietary fiber inulin, inulin-derived gut microbiota, and its metabolites in alleviating chronic colonic fibrosis caused by radiation.

The lumen of the gastrointestinal tract is home to a variety of microorganisms, the majority of which are bacteria which have developed in a mutually beneficial relationship with the host.42 Recent studies have corroborated the essential part that the gut microbiota plays in the regulation of their host’s health and wellbeing.43 Consequently, alterations in the composition of the gut microbiota and metabolites can have an impact on the function of host organs.23,44 Studies have demonstrated a correlation between ionizing radiation and substantial alterations in the gut microbiota across diverse animal and human hosts.45 Chowers et al. discovered that changes in the intestinal microbial composition of irradiated mice were exclusively limited to the impair mucosa.46 Notably, mice unexposed to radiation and inoculated with the gut microbiota of irradiated mice exhibited more intense pathological symotoms postirradiation, suggesting than an imbalance in gut microbiota could worsen the toxicity caused by radiation. Transplanting irradiated mice with normal mouse fecal microbiota can significantly enhance their digestive system and overall health.11 It appears that targeting the gut microbiota could be a potential solution to the irradiation-induced intestinal syndrome. During radiotherapy, acute radiation enteropathy frequently appears and then slowly diminishes. Reports indicate that chronic radiation enteropathy ranks as a leading cause of gastrointestinal distress, with its occurrence surpassing that of inflammatory bowel diseases. Therefore, we aimed to develop an approach to diminish chronic radiation enteropathy, particularly intestinal fibrosis, by focusing on the gut microbiota.

In the past few years, there has been a surge in research regarding the advantages of prebiotic and probiotic therapies.47 Prebiotic is a food ingredient that helps improve health by stimulating the growth of one bacterium or a limited group of bacteria in the intestines.48,49 Studies have shown that inulin can reduce the harm to the intestines caused by various external factors.14,15,47 Our research on mouse models has also revealed that inulin effectively mitigates chronic radiation-induced colonic fibrosis. Initially, inulin plays a role in preventing glycans degradation in the intestinal mucus layer of the host by controlling a wide selection of carbohydrate-active enzymes produced by the gut microbiome, thus providing a protective effect.50,51 Inulin is recognized for its ability to enhance the proliferation of specific advantageous bacteria.47 As a result, by employing high-throughput sequencing on 16s rDNA amplicons to assess inulin’s effect on the gut microbiota, it was found that inulin did not modify the gut microbiota’s composition but instead stimulated SCFA-producers. Following the administration of inulin, there was a notable rise in SCFAs levels in the feces, particularly acetic acid and butyric acid. Microorganisms in the distal intestine ferment indigestible fibers to produce SCFAs, which can then act as signals in tissues to provide protection.52,53 Given the beneficial impacts of SCFAs, our hypothesis suggested that metabolites from gut microbiota derived from inulin could play a role in reducing chronic intestinal fibrosis induced by radiation. The results from our in vivo mouse model demonstrated that gut microbiota, derived from inulin and its metabolites, were extremely efficient in diminishing chronic colonic fibrosis caused by radiation. Through in vitro cell experiments, it was demonstrated that the presence of gut microbial metabolites derived from inulin or mono component NaB resulted in a decrease in the expression of genes associated with fibrosis, as well as the collagen and extracellular matrix synthesis.

In the future, our attention will be directed toward understanding the mechanisms of SCFAs in mitigating chronic colonic fibrosis induced by radiation. SCFAs play a crucial role in controlling the immune system and the inflammatory response.54,55 SCFAs have the potential to significantly reduce systemic inflammation, either by decreasing pro-inflammatory cytokines or by elevating anti-inflammatory cytokines.56,57 The initial inflammatory reaction has been widely recognized as a pivotal stage in the progression of fibrosis.7 We hold the view that SCFAs derived from inulin could mitigate radiation-induced colonic fibrosis by inhibiting the initial inflammatory response. Observations show that the PPAR-gamma signaling pathway is activated in both in vivo and in vitro models when treated with SCFAs, particularly butyric acid.42,58 Research has shown that the PPAR-gamma signaling pathway effectively diminishes fibrosis in multiple organs, notably the liver and lung.59−61 Subsequent research will concentrate on exploring these mechanisms.

To conclude, the use of inulin has been shown to alleviate colonic fibrosis induced by irradiation. Therapies for the adverse effects of radiation therapy is limited in scope and costly. This study provides evidence for future clinical trials aimed to evaluating the potential benefits of dietary inulin in the prevention and treatment of radiation-induced intestinal injury.

Materials and Methods

Animals

Male C57BL/6J mice (Beijing HFK Bioscience, Beijing, China), aged from 6 to 8 weeks, were shared a living space (housing five mice in each cage) in the specific pathogen-free environment at the Institute of Radiation Medicine (IRM), China Academy of Medical Sciences (CAMS). The mice were provided with a consistent diet and sterile water while being fed under standard conditions.

Cell Culture

The NIH/3T3 cell line, purchased from the American Type Culture Collection (ATCC), was cultured in a high-glucose Dulbecco’s modified Eagle’s complete medium (Gibco, Grand Island, New York, USA), supplemented with 10% heat-inactivated fetal bovine serum (FBS), maintained at 37 °C in an environment with 5% CO2 humidity. Upon reaching 85% cell confluency, the cells underwent passage.

Inulin Treatment, Sample Collection, and Irradiation Procedure

Four distinct groups were formed by dividing the mice (n = 10 each). (1) The mice in the control group were provided with food in the standard conditions. (2) The drinking water of the inulin group was supplemented with 1% (w/v) inulin for a duration of 14 days, after which it was substituted with regular water once more. (3) The mice in the irradiation group received a 15 Gy abdominal irradiation (ABI) with the Gammacell 40 Exactor (Atomic Energy of Canada Lim, Chalk Rive, Canada) at a rate of 1.0 Gy/min. (4) The Inulin+IR group received a 1% (w/v) supply of inulin in drinking water for a duration of 14 days, after which it was substituted with regular water. The mice were exposed to 15Gy ABI on the 14th day, as outlined in group 3. Stool specimens were gathered, 4 days prior to radiation until the day of irradiation, 30 days post irradiation exposure, and 120 days post irradiation exposure. Each mouse’s stool specimens were stored at a temperature of −80 °C for the purpose of analyzing bacterial composition, or alternatively, they were processed for subsequent experimentation.

Transplantation of Microorganisms and Metabolites from the Gut Microbiome

Prior to irradiation, the stool specimens from both control and inulin groups underwent homogenization using phosphate-buffered saline (PBS) (10% w/v), were mixed with vortex, and then subjected to centrifugation at a speed of 800 rpm for 10 min at 4 °C. The liquid on top was gathered and centrifugated at 5000 rpm for a duration of 10 min at 4 °C. Subsequently, the samples were suspended in an equivalent quantity of PBS for the purpose of FMT. The centrifugation yielded a supernatant that underwent sterilization through the utilization of a 0.22-μm membrane filter, followed by its application in the MeT or in vitro cell experiments.

In the experiment involving the FMT and MeT, six distinct groups were formed for the mice, each containing five members. (1) The mice in the control + IR group were provided with standard feeding for a duration of 30 days, followed by treatment with abdominal irradiation (ABI) at a dose of 15 Gy. (2) The inulin + IR group received a 1% (w/v) addition of Inulin to their drinking water for a duration of 30 days, after which it was substituted with regular water. The mice were exposed to 15Gy ABI on the 30th day, following the aforementioned instructions (3–6): mice received a daily regimen of antibiotics, including 200 μL of ampicillin 1g/L, streptomycin 0.5 g/L, neomycin 1g/L, metronidazole 1g/L, vancomycin 0.5 g/L, and sucrose 10g/L, for a period of 15 straight days. Following that, mice were subjected to a 15 day FMT from either the control group (group 3, FMT (C) + IR) or the inulin group (group 4, FMT (I) + IR), as well as MeT from the control group (group 5, MeT (C) + IR) or the inulin group (group 6, MeT (I) + IR). Ultimately, all mice underwent irradiation (ABI, 15 Gy), as previously mentioned.

Quantitative Genetic Analyses

Trizol (Invitrogen, Carlsbad, California, USA) was used to extract the total RNA (ribonucleic acid) from tissues and cells in accordance with the manufacturer’s instructions. The PrimeScript first strand cDNA Synthesis kit (Takara) was utilized to reverse transcribe the extracted RNA. The FastStart Universal SYBR Green Master Mix (purchased from Roche, Mannheim, Germany) was utilized within a CFX96 system (Biorad, Hercules, California, USA) to carry out real-time quantitative PCR (polymerase chain reaction) using the primers outlined in Table S1. The glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene were used as the reference gene to adjust the relative transcript levels of the target genes.

Extraction of Protein and Western Blot Analysis

Tissues and cells underwent homogenization and lysis on ice with cell lysis buffer for western and IP (Beyotime, Nanjing, China) for 0.5 h. The proteins that had been dissolved in the supernatants were gathered, analyzed using SDS-PAGE, and then transferred onto polyvinylidene difluoride membranes. Detection was conducted using the ECL chemiluminescence detection kit by Proteintech, located in Chicago, CA, USA. The antibodies utilized included anti-α-SMA (1:5000, Cell Signaling Technology, #19245) and anti-GAPDH (1:10000, Abcam, ab8245).

Hematoxylin and Eosin (H&E) Staining

Colon tissue samples from dissected mice were promptly preserved in a 4% formaldehyde solution for overnight at ambient temperature. Samples of colon tissues were first dehydrated and then submerged in paraffin through standard methods. Following that, the samples were partitioned into sections with a thickness of 5-μm and subjected to staining using hematoxylin and eosin (H&E). An experienced gastrointestinal pathologist conducted a blind assessment of the samples.

Immunohistochemistry

The paraffin-stored tissue sections were extracted, submerged in distilled water, and obstructed with 3% H2O2 to eliminate inherent catalase at ambient temperature. Subsequently, the samples were submerged in a 0.01 M citrate buffer solution (pH 6.0) and underwent antigen recovery through microwaving for a duration of 10 min. Once the temperature had dropped to ambient temperature, the subsequent procedures were carried out in a damp chamber. The samples underwent three washes with PBS, followed by blocking with standard goat serum at ambient temperature for 20 min, and then left to incubate overnight at 4 °C with either antialpha-SMA or anticollagen III. The sections were washed in PBS and then left to incubate with secondary antibodies for 20 min at ambient temperature. Subsequently, the samples underwent staining with 3,3-diaminobenzidin and hematoxylin.

Masson’s Trichrome Staining

The colon tissues were cut into sections (each 5-μm thick) and then stained using Masson’s trichrome staining following the established procedure. After a short period of deparaffinization, the sections were submerged in Bouin solution and left to incubate at 37 °C overnight. The sections underwent a 2–3 min staining process using Celestine blue after being rinsed with distilled water. Subsequently, the samples were washed with water and colored using Mayer hematoxylin solution for a duration of 2–3 min. After that, the sections underwent another water wash and were differentiated with an acidic ethanol solution for a brief period. Following a 10 min rinse with running water, the sections were stained using Li Chunhong Acid Fuchsin Staining solution, and 1% phosphomolybdic acid was used to differentiate the sections for 10 min. After extraction from phosphomolybdic acid, the samples underwent staining using aniline blue solution, cleansing with distilled water, and a 2 min treatment with a mild acid solution. Eventually, the slices underwent dehydration using 95% and 100% alcohol, followed by sealing.

Bacterial Diversity Analysis

Prior to usage, the fecal samples were promptly gathered and promptly preserved at a temperature of −80 °C. The Power Fecal DNA Isolation Kit (MoBio, Carlsbad, CA, USA) was utilized to extract genomic DNA from feces. The Illumina Hiseq system (Novogene Bioinformatics Technology Co., Ltd., Tianjin, China) was utilized to analyze the bacterial diversity focusing on sequencing the 16S rRNA (rRNA) gene V4 region.

SCFA Measurement

Gas chromatography–mass spectrometry (GC-MS) was employed to quantify the SCFA content of fecal samples, following the previously described procedure.15 Roughly 50–100 mg from every fecal specimen was freeze-dried. The specimen underwent acidification using H2SO4 (0.01M, 1/10 v/v), followed by extraction of SCFA through diethyl ether extraction. After collecting the organic supernatant, it was derivatized overnight at room temperature using N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide. The gas chromatograph, connected to the mass spectrometer (7890B-7000D, Agilent Technologies, Palo Alto, Calif.), was utilized to measure the concentrations of SCFA.

RNA Sequencing

The cells were subjected to Trizol (Invitrogen, Carlsbad, California, USA) for the extraction of total RNA, following the aforementioned procedure. The NanoPhotometer spectrophotometer (Implen, Munchen Germany) and Agilent 2100 bioanalyzer (Agilent Technologies, Palo Alto, Calif) were utilized to evaluate the purity and integrity of the extracted RNA. The Illumina Hiseq system (Novogene Bioinformatics Technology Co., Ltd., Beijing, China) was utilized for mRNA (mRNA) sequencing. Succinctly put, the NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England Biolabs, Ipswich, Massachusetts, USA) was employed to construct the complementary DNA (cDNA) library, which was subsequently subjected to sequencing. Subsequent to conducting quality assurance on the sequencing data, analyses of gene expression differences and gene enrichment were executed using the appropriate software. Genes with a fold change greater than 1 and a significance level of p ≤ 0.05 were chosen if they showed differential expression.

Statistical Analysis

Each experiment was conducted at least three times. The SPSS 19.0 software (IBM Corp, Armonk, NY, USA) was used to evaluate the statistical significance of the results, which were expressed as mean ± standard deviations. A threshold of p < 0.05 was taken into account for statistical significance.

Acknowledgments

This study was supported by the National Natural Science Foundation of China (31971168, 31900891, and 32071241), CAMS Innovation Fund for Medical Science(2021-I2M-1-042), the Fundamental Research Funds for the Central Universities (3332021066), Natural Science Foundation of Tianjin (21JCYBJC01510), CIRP Open Fund of Radiation Protection Laboratories (CIRP-RRPE20220201). Medical and Health Science and Technology Innovation Project of Chinese Academy of Medical Sciences, and National Medical Health Science and Technology Strategic Platform and System construction Project (2022-I2M-2-003).

Data Availability Statement

Raw 16S rRNA gene sequences and mRNA sequences for all samples used in our study will been deposited in European Nucleotide Archive under accession code PRJEB55246 and PRJEB55247. All data are available from the corresponding author upon reasonable request.

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acs.jafc.3c03432.

  • Effect of gavage of inulin on radiation-induced gastrointestinal tract injury (Figure S1) and on changes in specific bacterial species (Figure S2); ipathway analysis between IR group and inulin + IR group at 120 days (Figure S3); comparative analysis of the effects of control derived fecal microbiota metabolites and inulin derived microbiota metabolites on NIH/3T3 gene expression profiles (Figure S4); primers used in this study (Table S1) (PDF)

Author Contributions

# K.J. and M.Z. contributed equally to this work.

The authors declare no competing financial interest.

Supplementary Material

jf3c03432_si_001.pdf (734.5KB, pdf)

References

  1. Tam S. Y.; Wu V. W. C. A Review on the Special Radiotherapy Techniques of Colorectal Cancer. Front. Oncol. 2019, 9, 208. 10.3389/fonc.2019.00208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Sassa N. Retroperitoneal tumors: Review of diagnosis and management. Int. J. Urol 2020, 27 (12), 1058–1070. 10.1111/iju.14361. [DOI] [PubMed] [Google Scholar]
  3. Wang J.; Yao M.; Wang Y.; Ho C. T.; Li S.; Shi Y.; Liu Q.; Zhao H. 6-Shogaol ameliorates injury to the intestinal mucosa and increases survival after high-dose abdominal irradiation. Journal of Functional Foods 2017, 36, 63–71. 10.1016/j.jff.2017.06.054. [DOI] [Google Scholar]
  4. Ji K.; Fang L.; Zhao H.; Li Q.; Shi Y.; Xu C.; Wang Y.; Du L.; Wang J.; Liu Q. Ginger Oleoresin Alleviated γ-Ray Irradiation-Induced Reactive Oxygen Species via the Nrf2 Protective Response in Human Mesenchymal Stem Cells. Oxidative Medicine and Cellular Longevity 2017, 2017, 1–12. 10.1155/2017/1480294. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Andreyev H. J. N.; Vlavianos P.; Blake P.; Dearnaley D.; Norman A. R.; Tait D. Gastrointestinal symptoms after pelvic radiotherapy: Role for the gastroenterologist?. Int. J. Radiat. Oncol., Biol., Phys.s 2005, 62 (5), 1464–1471. 10.1016/j.ijrobp.2004.12.087. [DOI] [PubMed] [Google Scholar]
  6. Lam S. Y.; Peppelenbosch M. P.; Fuhler G. M. Prediction and Treatment of Radiation Enteropathy: Can Intestinal Bugs Lead the Way?. Clin. Cancer Res. 2019, 25 (21), 6280–6282. 10.1158/1078-0432.CCR-19-2422. [DOI] [PubMed] [Google Scholar]
  7. Speca S.; Giusti I.; Rieder F.; Latella G. Cellular and molecular mechanisms of intestinal fibrosis. World J. Gastroenterol 2012, 18 (28), 3635–3661. 10.3748/wjg.v18.i28.3635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Nam Y. D.; Kim H. J.; Seo J. G.; Kang S. W.; Bae J. W. Impact of pelvic radiotherapy on gut microbiota of gynecological cancer patients revealed by massive pyrosequencing. PLoS One 2013, 8 (12), e82659 10.1371/journal.pone.0082659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Wang A.; Ling Z.; Yang Z.; Kiela P. R.; Wang T.; Wang C.; Cao L.; Geng F.; Shen M.; Ran X.; Su Y.; Cheng T.; Wang J. Gut microbial dysbiosis may predict diarrhea and fatigue in patients undergoing pelvic cancer radiotherapy: a pilot study. PLoS One 2015, 10 (5), e0126312 10.1371/journal.pone.0126312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Wang Z.; Wang Q.; Wang X.; Zhu L.; Chen J.; Zhang B.; Chen Y.; Yuan Z. Gut microbial dysbiosis is associated with development and progression of radiation enteritis during pelvic radiotherapy. J. Cell Mol. Med. 2019, 23 (5), 3747–3756. 10.1111/jcmm.14289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cui M.; Xiao H.; Li Y.; Zhou L.; Zhao S.; Luo D.; Zheng Q.; Dong J.; Zhao Y.; Zhang X.; Zhang J.; Lu L.; Wang H.; Fan S. Faecal microbiota transplantation protects against radiation-induced toxicity. EMBO Mol. Med. 2017, 9 (4), 448–461. 10.15252/emmm.201606932. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Wang Y. H.; Yao N.; Wei K. K.; Jiang L.; Hanif S.; Wang Z. X.; Pei C. X. The efficacy and safety of probiotics for prevention of chemoradiotherapy-induced diarrhea in people with abdominal and pelvic cancer: a systematic review and meta-analysis. Eur. J. Clin Nutr 2016, 70 (11), 1246–1253. 10.1038/ejcn.2016.102. [DOI] [PubMed] [Google Scholar]
  13. Larsen T.; Hausken T.; Otteraaen Ystad S.; Hovdenak N.; Mueller B.; Lied G. A. Does the low FODMAP diet improve symptoms of radiation-induced enteropathy? A pilot study. Scandinavian Journal of Gastroenterology 2018, 53 (5), 541–548. 10.1080/00365521.2017.1397186. [DOI] [PubMed] [Google Scholar]
  14. Zou J.; Chassaing B.; Singh V.; Pellizzon M.; Ricci M.; Fythe M. D.; Kumar M. V.; Gewirtz A. T. Fiber-Mediated Nourishment of Gut Microbiota Protects against Diet-Induced Obesity by Restoring IL-22-Mediated Colonic Health. Cell Host & Microbe 2018, 23 (1), 41–53. 10.1016/j.chom.2017.11.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Schroeder B. O.; Birchenough G. M. H.; Stahlman M.; Arike L.; Johansson M. E. V.; Hansson G. C.; Backhed F. Bifidobacteria or Fiber Protects against Diet-Induced Microbiota-Mediated Colonic Mucus Deterioration. Cell Host Microbe 2018, 23 (1), 27–40. 10.1016/j.chom.2017.11.004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Zhao L. P.; Zhang F.; Ding X. Y.; Wu G. J.; Lam Y. Y.; Wang X. J.; Fu H. Q.; Xue X. H.; Lu C. H.; Ma J. L.; et al. Gut bacteria selectively promoted by dietary fibers alleviate type 2 diabetes. Science 2018, 359, 1151–1156. 10.1126/science.aao5774. [DOI] [PubMed] [Google Scholar]
  17. Brooks L.; Viardot A.; Tsakmaki A.; Stolarczyk E.; Howard J. K.; Cani P. D.; Everard A.; Sleeth M. L.; Psichas A.; Anastasovskaj J.; et al. Fermentable carbohydrate stimulates FFAR2-dependent colonic PYY cell expansion to increase satiety. Molecular Metabolism 2017, 6 (1), 48–60. 10.1016/j.molmet.2016.10.011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Chen Y. J.; Wu H.; Wu S. D.; Lu N.; Wang Y. T.; Liu H. N.; Dong L.; Liu T. T.; Shen X. Z. Parasutterella, in association with irritable bowel syndrome and intestinal chronic inflammation. J. Gastroenterol Hepatol 2018, 33 (11), 1844–1852. 10.1111/jgh.14281. [DOI] [PubMed] [Google Scholar]
  19. diCenzo G. C.; Mengoni A.; Perrin E. Chromids aid genome expansion and functional diversification in the family Burkholderiaceae. Mol. Biol. Evol. 2019, 36 (3), 562–574. 10.1093/molbev/msy248. [DOI] [PubMed] [Google Scholar]
  20. Biragyn A.; Ferrucci L. Gut dysbiosis: a potential link between increased cancer risk in ageing and inflammaging. Lancet Oncology 2018, 19 (6), e295–e304. 10.1016/S1470-2045(18)30095-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Then C. K.; Paillas S.; Wang X.; Hampson A.; Kiltie A. E. Association of Bacteroides acidifaciens relative abundance with high-fibre diet-associated radiosensitisation. BMC Biol. 2020, 18 (1), 102. 10.1186/s12915-020-00836-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sawicki C. M.; Livingston K. A.; Obin M.; Roberts S. B.; Chung M.; McKeown N. M. Dietary fiber and the human gut microbiota: application of evidence mapping methodology. Nutrients 2017, 9 (2), 125. 10.3390/nu9020125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Koh A.; De Vadder F.; Kovatcheva-Datchary P.; Bäckhed F. From dietary fiber to host physiology: short-chain fatty acids as key bacterial metabolites. Cell 2016, 165 (6), 1332–1345. 10.1016/j.cell.2016.05.041. [DOI] [PubMed] [Google Scholar]
  24. Zhang X.; Zhao Y.; Xu J.; Xue Z.; Zhang M.; Pang X.; Zhang X.; Zhao L. Modulation of gut microbiota by berberine and metformin during the treatment of high-fat diet-induced obesity in rats. Sci. Rep. 2015, 5, 14405. 10.1038/srep14405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hiippala K.; Barreto G.; Burrello C.; Diaz-Basabe A.; Suutarinen M.; Kainulainen V.; Bowers J. R.; Lemmer D.; Engelthaler D. M.; Eklund K. K.; Facciotti F.; Satokari R. Novel Odoribacter Splanchnicus strain and its outer membrane vesicles exert immunoregulatory effects in vitro. Frontiers in Microbiology 2020, 11, 575455 10.3389/fmicb.2020.575455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Yue S. J.; Wang W. X.; Yu J. G.; Chen Y. Y.; Shi X. Q.; Yan D.; Zhou G. S.; Zhang L.; Wang C. Y.; Duan J. A.; et al. Gut microbiota modulation with traditional Chinese medicine: A system biology-driven approach. Pharmacol. Res. 2019, 148, 104453 10.1016/j.phrs.2019.104453. [DOI] [PubMed] [Google Scholar]
  27. Xiao S.; Liu C.; Chen M.; Zou J.; Zhang Z.; Cui X.; Jiang S.; Shang E.; Qian D.; Duan J. Scutellariae radix and coptidis rhizoma ameliorate glycolipid metabolism of type 2 diabetic rats by modulating gut microbiota and its metabolites. Appl. Microbiol. Biotechnol. 2020, 104 (1), 303–317. 10.1007/s00253-019-10174-w. [DOI] [PubMed] [Google Scholar]
  28. Zhang J.; Song L.; Wang Y.; Liu C.; Zhang L.; Zhu S.; Liu S.; Duan L. Beneficial effect of butyrate-producing Lachnospiraceae on stress-induced visceral hypersensitivity in rats. J. Gastroenterol Hepatol 2019, 34 (8), 1368–1376. 10.1111/jgh.14536. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Bui T. P.; Shetty S. A.; Lagkouvardos I.; Ritari J.; Chamlagain B.; Douillard F. P.; Paulin L.; Piironen V.; Clavel T.; Plugge C. M.; et al. Comparative genomics and physiology of the butyrate-producing bacterium Intestinimonas butyriciproducens. Environ. Microbiol Rep 2016, 8 (6), 1024–1037. 10.1111/1758-2229.12483. [DOI] [PubMed] [Google Scholar]
  30. Onursal C.; Dick E.; Angelidis I.; Schiller H. B.; Staab-Weijnitz C. A. Collagen biosynthesis, processing, and maturation in lung ageing. Front Med. (Lausanne) 2021, 8, 593874 10.3389/fmed.2021.593874. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Levavasseur F.; Liétard J.; Ogawa K.; Théret N.; Burbelo P. D.; Yamada Y.; Guillouzo A.; Clément B. Expression of laminin γ1 in cultured hepatocytes involves repeated CTC and GC elements in the LAMC1 promoter. Biochem. J. 1996, 313, 745–752. 10.1042/bj3130745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Zhang H.; Liu Z.; Liu S. HMGB1 induced inflammatory effect is blocked by CRISPLD2 via MiR155 in hepatic fibrogenesis. Mol. Immunol 2016, 69, 1–6. 10.1016/j.molimm.2015.10.018. [DOI] [PubMed] [Google Scholar]
  33. Hardin M.; Silverman E. K. Chronic obstructive pulmonary disease genetics: a review of the past and a look into the future. Chronic Obstr Pulm Dis 2014, 1 (1), 33–46. 10.15326/jcopdf.1.1.2014.0120. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Krishnan A.; Li X.; Kao W. Y.; Viker K.; Butters K.; Masuoka H.; Knudsen B.; Gores G.; Charlton M. Lumican, an extracellular matrix proteoglycan, is a novel requisite for hepatic fibrosis. Lab Invest 2012, 92 (12), 1712–1725. 10.1038/labinvest.2012.121. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Gallego-Muñoz P.; Lorenzo-Martín E.; Fernández I.; Herrero-Pérez C.; Martínez-García M. C. Nidogen-2: location and expression during corneal wound healing. Exp. Eye Res. 2019, 178, 1–9. 10.1016/j.exer.2018.09.004. [DOI] [PubMed] [Google Scholar]
  36. Randles M. J.; Humphries M. J.; Lennon R. Proteomic definitions of basement membrane composition in health and disease. Matrix Biol. 2017, 57–58, 12–28. 10.1016/j.matbio.2016.08.006. [DOI] [PubMed] [Google Scholar]
  37. Gerarduzzi C.; Kumar R. K.; Trivedi P.; Ajay A. K.; Iyer A.; Boswell S.; Hutchinson J. N.; Waikar S. S.; Vaidya V. S. Silencing SMOC2 ameliorates kidney fibrosis by inhibiting fibroblast to myofibroblast transformation. JCI Insight 2017, 2 (8), e90299 10.1172/jci.insight.90299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. de Fonseka D.; Edey A.; Stadon L.; Viner J.; Darby M.; Maskell N. A. The physiological consequences of different distributions of diffuse pleural thickening on CT imaging. Br J. Radiol 2017, 90 (1077), 20170218. 10.1259/bjr.20170218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Bauer Y.; Tedrow J.; de Bernard S.; Birker-Robaczewska M.; Gibson K. F.; Guardela B. J.; Hess P.; Klenk A.; Lindell K. O.; Poirey S.; et al. A novel genomic signature with translational significance for human idiopathic pulmonary fibrosis. Am. J. Respir. Cell Mol. Biol. 2015, 52 (2), 217–231. 10.1165/rcmb.2013-0310OC. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Edeling M.; Ragi G.; Huang S.; Pavenstadt H.; Susztak K. Developmental signalling pathways in renal fibrosis: the roles of Notch, Wnt and Hedgehog. Nat. Rev. Nephrol 2016, 12 (7), 426–439. 10.1038/nrneph.2016.54. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Joshi N.; Kopec A. K.; Ray J. L.; Cline-Fedewa H.; Groeneveld D. J.; Lisman T.; Luyendyk J. P. Von Willebrand factor deficiency reduces liver fibrosis in mice. Toxicol. Appl. Pharmacol. 2017, 328, 54–59. 10.1016/j.taap.2017.05.018. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Postler T. S.; Ghosh S. Understanding the holobiont: how microbial metabolites affect human health and shape the immune system. Cell Metab 2017, 26 (1), 110–130. 10.1016/j.cmet.2017.05.008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Ostaff M. J.; Stange E. F.; Wehkamp J. Antimicrobial peptides and gut microbiota in homeostasis and pathology. EMBO Molecular Medicine 2013, 5 (10), 1465–1483. 10.1002/emmm.201201773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Schroeder B. O.; Backhed F. Signals from the gut microbiota to distant organs in physiology and disease. Nat. Med. 2016, 22 (10), 1079–1089. 10.1038/nm.4185. [DOI] [PubMed] [Google Scholar]
  45. Zhang A.; Steen T. Y. Gut microbiomics—a solution to unloose the gordian knot of biological effects of ionizing radiation. Journal of Heredity 2018, 109 (2), 212–221. 10.1093/jhered/esx059. [DOI] [PubMed] [Google Scholar]
  46. Gerassy-Vainberg S.; Blatt A.; Danin-Poleg Y.; Gershovich K.; Sabo E.; Nevelsky A.; Daniel S.; Dahan A.; Ziv O.; Dheer R.; Abreu M. T.; Koren O.; Kashi Y.; Chowers Y.; et al. Radiation induces proinflammatory dysbiosis: transmission of inflammatory susceptibility by host cytokine induction. Gut 2018, 67 (1), 97–107. 10.1136/gutjnl-2017-313789. [DOI] [PubMed] [Google Scholar]
  47. Vandeputte D.; Falony G.; Vieira-Silva S.; Wang J.; Sailer M.; Theis S.; Verbeke K.; Raes J. Prebiotic inulin-type fructans induce specific changes in the human gut microbiota. Gut 2017, 66 (11), 1968–1974. 10.1136/gutjnl-2016-313271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Hutkins R. W.; Krumbeck J. A.; Bindels L. B.; Cani P. D.; Fahey G.; Goh Y. J.; Hamaker B.; Martens E. C.; Mills D. A.; Rastal R. A.; et al. Prebiotics: why definitions matter. Curr. Opin. Biotechnol. 2016, 37, 1–7. 10.1016/j.copbio.2015.09.001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Bindels L. B.; Delzenne N. M.; Cani P. D.; Walter J. Towards a more comprehensive concept for prebiotics. Nat. Rev. Gastroenterol Hepatol 2015, 12 (5), 303–310. 10.1038/nrgastro.2015.47. [DOI] [PubMed] [Google Scholar]
  50. Desai M. S.; Seekatz A. M.; Koropatkin N. M.; Kamada N.; Hickey C. A.; Wolter M.; Pudlo N. A.; Kitamoto S.; Terrapon N.; Muller A.; et al. A dietary fiber-deprived gut microbiota degrades the colonic mucus barrier and enhances pathogen susceptibility. Cell 2016, 167 (5), 1339–1353. 10.1016/j.cell.2016.10.043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Earle K. A.; Billings G.; Sigal M.; Lichtman J. S.; Hansson G. C.; Elias J. E.; Amieva M. R.; Huang K. C.; Sonnenburg J. L. Quantitative imaging of gut microbiota spatial organization. Cell Host Microbe 2015, 18 (4), 478–488. 10.1016/j.chom.2015.09.002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Macfarlane G. T.; Macfarlane S. Bacteria, colonic fermentation, and gastrointestinal health. Journal of Aoac International 2012, 95 (1), 50–60. 10.5740/jaoacint.SGE_Macfarlane. [DOI] [PubMed] [Google Scholar]
  53. Canfora E. E.; Jocken J. W.; Blaak E. E. Short-chain fatty acids in control of body weight and insulin sensitivity. Nat. Rev. Endocrinol 2015, 11 (10), 577–591. 10.1038/nrendo.2015.128. [DOI] [PubMed] [Google Scholar]
  54. Smith P. M.; Howitt M. R.; Panikov N.; Michaud M.; Gallini C. A.; Bohlooly-Y M.; Glickman J. N.; Garrett W. S. The microbial metabolites, short-chain fatty acids, regulate colonic T cell homeostasis. Science 2013, 341, 569–573. 10.1126/science.1241165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Arpaia N.; Campbell C.; Fan X.; Dikiy S.; van der Veeken J.; deRoos P.; Liu H.; Cross J. R.; Pfeffer K.; Coffer P. J.; et al. Metabolites produced by commensal bacteria promote peripheral regulatory T-cell generation. Nature 2013, 504 (7480), 451–455. 10.1038/nature12726. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Chang P. V.; Hao L.; Offermanns S.; Medzhitov R. The microbial metabolite butyrate regulates intestinal macrophage function via histone deacetylase inhibition. Proc. Natl. Acad. Sci. 2014, 111 (6), 2247–2252. 10.1073/pnas.1322269111. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Trompette A.; Gollwitzer E. S.; Yadava K.; Sichelstiel A. K.; Sprenger N.; Ngom-Bru C.; Blanchard C.; Junt T.; Nicod L. P.; Harris N. L.; et al. Gut microbiota metabolism of dietary fiber influences allergic airway disease and hematopoiesis. Nat. Med. 2014, 20 (2), 159–166. 10.1038/nm.3444. [DOI] [PubMed] [Google Scholar]
  58. Byndloss M. X.; Olsan E. E.; Rivera-Chavez F.; Tiffany C. R.; Cevallos S. A.; Lokken K. L.; Torres T. P.; Byndloss A. J.; Faber F.; Gao Y.; et al. Microbiota-activated PPAR-gamma signaling inhibits dysbiotic Enterobacteriaceae expansion. Science 2017, 357 (6351), 570–575. 10.1126/science.aam9949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Ruzehaji N.; Frantz C.; Ponsoye M.; Avouac J.; Pezet S.; Guilbert T.; Luccarini J. M.; Broqua P.; Junien J. L.; Allanore Y. Pan PPAR agonist IVA337 is effective in prevention and treatment of experimental skin fibrosis. Ann. Rheum Dis 2016, 75 (12), 2175–2183. 10.1136/annrheumdis-2015-208029. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Avouac J.; Konstantinova I.; Guignabert C.; Pezet S.; Sadoine J.; Guilbert T.; Cauvet A.; Tu L.; Luccarini J. M.; Junien J. L.; et al. Pan-PPAR agonist IVA337 is effective in experimental lung fibrosis and pulmonary hypertension. Annals of the Rheumatic Diseases 2017, 76 (11), 1931–1940. 10.1136/annrheumdis-2016-210821. [DOI] [PubMed] [Google Scholar]
  61. Boyer-Diaz Z.; Aristu-Zabalza P.; Andrés-Rozas M.; Robert C.; Ortega-Ribera M.; Fernández-Iglesias A.; Broqua P.; Junien J. L.; Wettstein G.; Bosch J.; Gracia-Sancho J.; et al. Pan-PPAR agonist lanifibranor improves portal hypertension and hepatic fibrosis in experimental advanced chronic liver disease. J. Hepatol 2021, 74 (5), 1188–1199. 10.1016/j.jhep.2020.11.045. [DOI] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

jf3c03432_si_001.pdf (734.5KB, pdf)

Data Availability Statement

Raw 16S rRNA gene sequences and mRNA sequences for all samples used in our study will been deposited in European Nucleotide Archive under accession code PRJEB55246 and PRJEB55247. All data are available from the corresponding author upon reasonable request.

Articles from Journal of Agricultural and Food Chemistry are provided here courtesy of American Chemical Society

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