Research Advances in the Impact of Probiotic Supplementation on Ulcerative Colitis Management

Comments 1: [Lack of critical synthesis:
The review lists multiple studies but rarely evaluates their quality, design, or bias. A comparative summary table (RCTs, meta-analyses, models used, sample sizes) is recommended.]

Response 1: Thank you for pointing this out. We agree with this comment. Therefore, we have added a table in the paper.

“[updated text in the manuscript if necessary]” Line 445-446

Comments 2: [Overly descriptive sections:
The pathogenesis and mechanistic sections are too long and could be condensed using schematic figures or summarised text.]

Response 2: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Environmental factors, especially diet, are important external conditions that drive disease. Westernized diet with high sugar, high fat and additives (such as emulsifiers and artificial sweeteners) can significantly change the composition of intestinal flora. It leads to an increase in pro-inflammatory bacteria (such as Proteobacteria), a decrease in beneficial bacteria (such as Bifidobacterium and Lactobacillus), and a decrease in the pro-duction of short-chain fatty acids (SCFAs) [22,23]. This imbalance of intestinal flora is the core link in the pathogenesis of UC.

Healthy intestinal flora provides energy for intestinal epithelial cells, maintains barrier integrity, and inhibits pro-inflammatory pathways such as NF-κB by producing metabolites such as SCFAs (such as butyric acid) and vitamins, thereby shaping the immune tolerance environment [27,31]. UC patients showed decreased bacterial diver-sity and reduced SCFAs, which weakened intestinal barrier function and increased intestinal permeability [24-27].

Ultimately, abnormal activation of the immune system is the direct cause of tissue damage. In the context of genetic susceptibility and dysbacteriosis, the response of the intestinal mucosal immune system to intracavitary antigens is out of control. It is characterized by excessive activation of pro-inflammatory T cell subsets (such as Th1, Th17) and their cytokines (such as IFN-γ, IL-17), while regulatory T cells (Treg) function is impaired and cannot effectively inhibit inflammation [28,29]. At the same time, innate immune cells (such as macrophages, neutrophils) are recruited in large numbers and release inflammatory mediators such as reactive oxygen species and proteases, leading to intestinal epithelial damage, erosion and ulceration, forming chronic inflammation. [28]

All in all, the incidence of UC is an individual in a genetic background. Triggered by adverse environmental factors (especially diet), UC is caused by a chain reaction of ' intestinal flora imbalance → mucosal barrier damage → abnormal immune activation '.]

“[updated text in the manuscript if necessary]” Line 95-119

Comments 3: [Limited translational discussion:
The authors should better differentiate between findings from animal models and those from human trials, and discuss the limitations of clinical applicability.]

Response 3: Thank you for pointing this out. We agree with this comment. T herefore, we have added a table in the paper.

“[updated text in the manuscript if necessary]” Line 445-446

Comments 4: [The manuscript is scientifically sound but lacks depth in its evaluation of the strength of the evidence. Some claims (e.g., that probiotics can “shift treatment from symptom control to addressing the underlying condition”) are too general and lack robust clinical support.

A more balanced tone is recommended, emphasising the variability in strain efficacy, dosing, and study duration.]

Response 4: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [4.Application case of probiotics to alleviate UC.

The following sections summarize the findings from several clinical and preclinical studies investigating the effects of probiotic interventions in UC. It should be noted that the evidence is derived from a limited number of studies, and the results should be interpreted with caution pending further validation in larger cohorts.

4.1. Application of Saccharomyces boulardii in the alleviating of the UC mouse model

In this study, female C57BL/6 UC mice were chosen as the experimental subjects [76]. The mice were gavaged with Saccharomyces boulardii for three weeks before being challenged with dextran sulfate sodium (DSS) to induce UC. Various physiological indexes were monitored throughout the experiment. The results indicated that the Saccharomyces boulardii treatment effectively alleviated UC symptoms, including colon shortening, intestinal mucosal injury, and apoptosis of colon tissue cells. Further analysis revealed that Saccharomyces boulardii significantly reduced the levels of pro-inflammatory factors in the serum and maintained intestinal microbial stability by increasing the abundance of Porphyromonadaceae in the mice [74]. Additionally, fructooligosaccharides (FOS), a commonly used prebiotic, selectively promotes the growth of beneficial probiotics such as Bifidobacterium and Lactobacillus in the intestine while reducing the production of pro-inflammatory cytokines, thereby regulating intestinal immunity [75]. A recent study demonstrated that the combination of FOS and Saccharomyces boulardii produced superior anti-inflammatory effects in a mouse colitis model. This combination not only significantly reduced the disease activity index (DAI) and effectively inhibited colitis but also notably increased levels of butyric acid and isobutyric acid. Furthermore, the combination therapy significantly enhanced the abundance of beneficial bacteria, including lactic acid bacteria and bifidobacteria, effectively regulating the composition of the intestinal microbiome [76]. This research provides a solid scientific foundation for the prevention and alleviating of UC.

4.2. Application of mixed probiotics in UC patients

This study involved a randomized controlled trial with 130 patients diagnosed with UC [77]. The participants were divided into two groups: 65 patients in the experimental group received a treatment regimen of mesalazine, somatostatin, and a bifid triple viable capsule, while 65 patients in the control group were treated with only mesalazine and somatostatin. The bifid triple viable capsule, used in the experimental group, is a compound probiotic formulation that primarily contains three active probiotics: Bifidobacterium longum, Lactobacillus acidophilus, and Enterococcus faecalis.

The total effective rate was 92.31% in the experimental group and 76.92% in the control group. Regarding inflammatory markers, the experimental group showed lower levels of IL-6 (90.01 ± 8.02 ng/L vs 101.27 ± 7.27 ng/L) and TNF-α (61.87 ± 6.38 ng/L vs 68.29 ± 7.11 ng/L). Immune profiling revealed higher proportions of CD 4+ T cells (46.37 ± 4.21% vs 40.02 ± 3.21%) and a higher CD4+/CD8+ ratio (1.79 ± 0.14 vs 1.41 ± 0.09) in the experimental group. An indicator of intestinal barrier function, D-lactic acid level, was also lower in the experimental group (4.01 ± 0.08 mmol/L vs 4.61 ± 0.02 mmol/L). The incidence of adverse reactions was 4.62% in the experimental group and 7.69% in the control group. This study provides robust evidence for the use of probiotics as an ad-junctive treatment for UC, highlighting its significant clinical relevance [77]. However, the independent contribution of the probiotic combination and its synergy with con-ventional drugs require further investigation.

4.3. Application of dietary probiotics in UC patients

In a study involving 93 UC patients, participants were randomized into a probiotics combination group (UC-P, n=44) and a control group (UC-NP, n=49) [78]. The UC-P group received a supplement containing five probiotics (Bifidobacterium infan-tis, Bifidobacterium animalis, Lactobacillus bulgaricus, Lactobacillus helveticus, and Enterococcus faecalis, each at ≥ 5 billion CFU), L-glutamine, biotin, and dietary guidance for 12 weeks.

After the intervention, the UC-P group showed increases in muscle mass (MM) and skeletal muscle index (SMI), and a decrease in the extracellular water/total body water ratio (ECW/TBW). The Short Inflammatory Bowel Disease Questionnaire (SIBDQ) score increased from 45.21 to 60.47. The rate of abnormality in the inflammatory marker C-reactive protein (CRP) decreased from 50% to 9.1% [78].

This study not only confirmed the positive effects of the probiotic combination on intestinal symptoms in UC patients but also highlighted its significant role in regulating systemic inflammatory responses. Furthermore, the combination demonstrated beneficial effects on nutritional metabolism and body composition. These findings offer new insights into the use of probiotics for comprehensive management of inflammatory bowel disease. They suggest that a specific strain combination may serve as an adjuvant management strategy to alleviate extraintestinal symptoms, such as muscle loss and metabolic disorders, in UC patients. The mechanisms of action and clinical application potential of this strategy warrant further investigation [78].

4.4. Application of fecal microbiota transplantation in UC patients

Fecal microbiota transplantation (FMT) is an intervention that aims to restore the balance of intestinal microbiota by transferring fecal microorganisms from healthy donors. This method provides a diverse array of probiotics to the body. In this study, 10 patients with moderate to severe active UC underwent six sessions of FMT.

Clinical symptoms improved, as indicated by a decrease in the Truelove-Witts index (p < 0.05). Levels of inflammatory markers, including C-reactive protein (CRP) (p = 0.0004) and fecal calprotectin (p = 0.002), were reduced after FMT. Metagenomic analysis indicated changes in the patients' intestinal microbiota, with an increased abundance of Lactobacillus, Prevotellaceae, and Firmicutes, and a decreased abundance of Staphylococcus and Bacillus [79]. These findings indicate that multi-course FMT can effectively regulate the intestinal microbiota in UC patients, alleviate inflammation, and is safe for use. This approach offers a promising new clinical option for UC treatment with broad application potential [79].

Although the above clinical studies have shown the potential of probiotics in the management of UC, there are obvious limitations: 1. Sample size is generally small (such as only 10 cases in FMT study); 2. Short follow-up time (mostly≤ 12 weeks); 3. Lack of standardized strain combinations and doses. For example, Li et al. (2021) showed that triple probiotic capsules significantly improved inflammatory markers, but did not clarify the independent contribution of each strain, and did not set up a probiotic treatment group alone, making it difficult to distinguish its synergistic effect with con-ventional drugs. In addition, most studies did not conduct stratified analysis based on the baseline characteristics of patients ' intestinal microbiota, ignoring the impact of individual differences on efficacy.

4.5. Summary and Critical Appraisal of Evidence

The studies discussed above, while promising, vary considerably in design, quality, and scale, which complicates the formulation of definitive clinical recommendations. To facilitate a comparative overview, key characteristics of selected studies are summarized in Table 2. The table highlights that while several RCTs report positive outcomes with probiotic adjunct therapy [77,78], limitations such as small sample sizes, short duration, and the frequent use of multi-component interventions (making it difficult to isolate the probiotic effect) are common. The most robust evidence often comes from meta-analyses, which nevertheless frequently note significant heterogeneity among trials [12,13]. Preclinical animal studies, though essential for elucidating mechanisms (as detailed in Section 3.2), cannot directly predict human efficacy. This appraisal underscores the necessity for larger, longer-duration, and more rigorously designed RCTs that employ standardized probiotic preparations to move the field toward reliable, evidence-based applications.]

“[updated text in the manuscript if necessary]” Line 341-444

Comments 5: [Would also like to suggest two recent, thematically relevant articles that may enrich the discussion and provide complementary perspectives on probiotic-based modulation of inflammatory bowel diseases. Inclusion of these references is entirely optional and not required for acceptance:

https://www.mdpi.com/2227-9059/11/2/494

https://www.frontiersin.org/journals/nutrition/articles/10.3389/fnut.2025.1575130/full]

Response 4: Thank you for pointing this out. We agree with this comment. Therefore, we have conducted relevant literature research.

Comments 1: [Figure 1

Please highlight the text sections as they are illegible.]

Response 1: Thank you for pointing this out. We agree with this comment. Therefore, We have modified Table 1. “[updated text in the manuscript if necessary]” Line 88-89

Comments 2: [Section 3.2.2
This section, which plays an important role in the context, should be developed in light of recent studies and probiotic strains that affect the expression of tight junctions.]

Response 2: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [In patients with UC, chronic intestinal inflammation leads to damage and apoptosis of intestinal epithelial cells. This damage results in the downregulation of tight junction (TJ) protein expression, structural damage, thinning of the mucus layer, and an imbalance in intestinal microbiota. The increase in pathogenic bacteria and the decrease in beneficial bacterial metabolites compromise barrier function, ultimately leading to the breakdown of intestinal mechanical barriers and a significant rise in intestinal permeability. This phenomenon, often referred to as intestinal leakage, triggers systemic inflammation and exacerbates local intestinal inflammation, creating a vicious cycle linked to the pathogenesis, severity, and recurrence of UC.

Therefore, maintaining the integrity of intestinal epithelial cells and repairing intestinal barrier function is a key therapeutic strategy for managing UC.

A growing body of evidence suggests that specific probiotic strains can help repair the intestinal barrier through multiple mechanisms. For instance, Lacticaseibacillus paracasei CCFM1222 upregulates the expression of tight junction proteins ZO-1, occludin, and claudin-1 in mice, reversing DSS-induced mucosal and barrier dysfunction [59].

Similarly, Lactiplantibacillus plantarum SC-5 has been shown to reduce mucosal damage in DSS-induced colitis mice and enhance the expression of occludin and claudin-1 [60]. The mechanisms by which probiotics enhance the TJ barrier are diverse. L.rhamnosus GR10 L.rhamnosus GR10 can prevent the dissociation of TJ protein and E-cadherin by down-regulating the key proteins in the MAPK/MLCK/MLC pathway, thereby reducing intestinal permeability [61]. Another study revealed that Lactiplanti-bacillus plantarum NWAFU-BIO-BS29 increased the expression of mucin (MUC2,3) and TJ proteins (occludin and claudin-1) in a mouse model, indicating a strong ability to strengthen the intestinal barrier [62]. Engineering strategies have also been used to improve the efficacy of probiotics. For example, engineered probiotics with a multi-functional coating, derived from Lactobacillus rhamnosus GG (LGG), can reverse the de-crease in the expression of ZO-1 and occludin and significantly reduce goblet cell damage compared to 5-aminosalicylic acid (5-ASA), showing a protective effect on the integrity of the colonic epithelium [63].

 In addition to Lactobacillus, other probiotics also showed similar benefits. Bacillus paralicheniformis HMPM220325 effectively alleviated the decrease of TJ proteins such as ZO-1, occludin and claudin-1 at the mRNA and protein levels. It has been shown to significantly restore colon length and protect goblet cells and mucous layers [64]. In ad-dition, it has been reported that in vitro experiments have shown that the supernatant of Saccharomyces boulardii CNCM I-745 enhances the delivery of E-cadherin to the cell sur-face by redirecting the endocytic E-cadherin to the cell surface. This process involves Rab11A-dependent recycling endosomes, which not only restores the attachment junction of intestinal cells but also restores and strengthens the intestinal barrier function as a whole [65].

 These findings suggest that probiotics can alleviate UC by increasing the levels of tight junction proteins and restoring the mucus layer, thereby reducing intestinal permeability and enhancing the intestinal barrier.]

“[updated text in the manuscript if necessary]” Line 239-279

Comments 3: [Section 3.2.3
This section also needs to be developed, reporting studies on probiotics and their anti-inflammatory effects.]

Response 3: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Recent studies in mice have shown that probiotic Lactic acid bacteria LB-9 can significantly reduce colitis-induced intestinal epithelial cells (IECs) apoptosis mediated by NF-κB and closely related to the inflammatory factor TNF-α [66]. Similarly, Lacticaseibacillus rhamnosus 1.0320 and its progeny showed good remission effects in DSS-induced colitis models. It can inhibit the key proteins of TLR4/MAPK/NF-κB pathway (TLR4, MyD88, JNK, p38 and p65) and reduce the production of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) [67]. In addition to NF-κB, other inflammatory pathways are also regulated. Leuconostoc mesenteroides DRC 1506 has been shown to reduce the levels of pro-inflammatory cytokines (TNF-α, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS)) in colon tissue. At the same time, it significantly increased the level of anti-inflammatory cytokine IL-10, thereby reducing intestinal tissue inflammation [68]. Another key mechanism involves the regulation of immune cell differentiation. Ursodeoxycholic acid (UDCA), a metabolite of Lactobacillus acidophilus (LA), can activate the RapGap/PI3K-AKT/NF-κB pathway. It promotes the differentiation of regulatory T (Treg) cells and inhibits the polarization of pro-inflammatory M1 macrophages. This process reduces the secretion of pro-inflammatory factors such as IL-6 and TNF-α, re-duces intestinal inflammation, and improves colonic pathological damage in UC mice [69]. The anti-inflammatory effects of probiotics can also be strain-specific and involve unique metabolites. It was found that Bifidobacterium FL-276.1 and Bifidobacterium FL-228.1 significantly alleviated DSS-induced colitis. The aryl hydrocarbon receptor (AHR) and nuclear factor erythroid 2-related factor 2 (NRF2) pathways were activated in the colon of mice, while the NLR family pyridine domain containing 3 (NLRP3) was down-regulated in the whole intervention mode. As a possible effector molecule, in-dole-lactic acid (ILA) can up-regulate ZO-1, Claudin-4 and Occludin through AHR/NRF2/NLRP3 pathway to enhance epithelial barrier function. At the same time, it inhibited the increase of TNF-α, IL-6 and IL-1β [70]. Through the above mechanisms, probiotics can not only reduce systemic inflammation, but also have indirect benefits on the emotional health of UC patients [71]. Chronic inflammation is a key association between IBD and ' inflammatory depression ' (anxiety and depression comorbidity) [71]. Among them, pro-inflammatory cytokines such as TNF-α, IL-6 and IL-1β can penetrate the blood-brain barrier, affect neurological function and induce emotional and behavioral changes. By inhibiting these inflammatory factors, probiotics can not only alleviate local intestinal inflammation, but also alleviate inflammation-driven neuropsychiatric symptoms, which is one of the core mechanisms of the role of mental probiotics [71].

In addition, studies have also shown that probiotics have anti-oxidative stress effects, can scavenge reactive oxygen species (ROS) and reduce oxidative stress damage to intestinal tissues. For example, mice treated with Limosilactobacillus fermentum E7 had significantly lower myeloperoxidase (MPO) activity and malondialdehyde (MDA) levels in the colon than DSS)-treated mice, while superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities were significantly increased [72]. Another study on Lactobacillus casei ATCC 393 showed that its synthetic Biogenic Se nanoparticles (SeNPs) can alleviate ROS-mediated mitochondrial dysfunction through the Nrf2 signaling pathway and protect intestinal epithelial barrier function from oxidative damage [73].]

“[updated text in the manuscript if necessary]” Line 287-330

Comments 4: [Sections 4.2, 4.3, and 4.4 report the results of only four studies, also indicating statistical data that I consider inappropriate.
They should limit themselves to reporting and discussing the data.]

Response 4: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [4.2. Application of mixed probiotics in UC patients

This study involved a randomized controlled trial with 130 patients diagnosed with UC [77]. The participants were divided into two groups: 65 patients in the experimental group received a treatment regimen of mesalazine, somatostatin, and a bifid triple viable capsule, while 65 patients in the control group were treated with only mesalazine and somatostatin. The bifid triple viable capsule, used in the experimental group, is a compound probiotic formulation that primarily contains three active probiotics: Bifidobacterium longum, Lactobacillus acidophilus, and Enterococcus faecalis.

The total effective rate was 92.31% in the experimental group and 76.92% in the control group. Regarding inflammatory markers, the experimental group showed lower levels of IL-6 (90.01 ± 8.02 ng/L vs 101.27 ± 7.27 ng/L) and TNF-α (61.87 ± 6.38 ng/L vs 68.29 ± 7.11 ng/L). Immune profiling revealed higher proportions of CD 4+ T cells (46.37 ± 4.21% vs 40.02 ± 3.21%) and a higher CD4+/CD8+ ratio (1.79 ± 0.14 vs 1.41 ± 0.09) in the experimental group. An indicator of intestinal barrier function, D-lactic acid level, was also lower in the experimental group (4.01 ± 0.08 mmol/L vs 4.61 ± 0.02 mmol/L). The incidence of adverse reactions was 4.62% in the experimental group and 7.69% in the control group. This study provides robust evidence for the use of probiotics as an ad-junctive treatment for UC, highlighting its significant clinical relevance [77]. However, the independent contribution of the probiotic combination and its synergy with conventional drugs require further investigation.

4.3. Application of dietary probiotics in UC patients

In a study involving 93 UC patients, participants were randomized into a probiotics combination group (UC-P, n=44) and a control group (UC-NP, n=49) [78]. The UC-P group received a supplement containing five probiotics (Bifidobacterium infan-tis, Bifidobacterium animalis, Lactobacillus bulgaricus, Lactobacillus helveticus, and Enterococcus faecalis, each at ≥ 5 billion CFU), L-glutamine, biotin, and dietary guidance for 12 weeks.

After the intervention, the UC-P group showed increases in muscle mass (MM) and skeletal muscle index (SMI), and a decrease in the extracellular water/total body water ratio (ECW/TBW). The Short Inflammatory Bowel Disease Questionnaire (SIBDQ) score increased from 45.21 to 60.47. The rate of abnormality in the inflammatory marker C-reactive protein (CRP) decreased from 50% to 9.1% [78].

This study not only confirmed the positive effects of the probiotic combination on intestinal symptoms in UC patients but also highlighted its significant role in regulating systemic inflammatory responses. Furthermore, the combination demonstrated beneficial effects on nutritional metabolism and body composition. These findings offer new insights into the use of probiotics for comprehensive management of inflammatory bowel disease. They suggest that a specific strain combination may serve as an adjuvant management strategy to alleviate extraintestinal symptoms, such as muscle loss and metabolic disorders, in UC patients. The mechanisms of action and clinical application potential of this strategy warrant further investigation [78].

4.4. Application of fecal microbiota transplantation in UC patients

Fecal microbiota transplantation (FMT) is an intervention that aims to restore the balance of intestinal microbiota by transferring fecal microorganisms from healthy do-nors. This method provides a diverse array of probiotics to the body. In this study, 10 patients with moderate to severe active UC underwent six sessions of FMT.

Clinical symptoms improved, as indicated by a decrease in the Truelove-Witts in-dex (p < 0.05). Levels of inflammatory markers, including C-reactive protein (CRP) (p = 0.0004) and fecal calprotectin (p = 0.002), were reduced after FMT. Metagenomic analysis indicated changes in the patients' intestinal microbiota, with an increased abun-dance of Lactobacillus, Prevotellaceae, and Firmicutes, and a decreased abundance of Staphylococcus and Bacillus [79]. These findings indicate that multi-course FMT can effectively regulate the intestinal microbiota in UC patients, alleviate inflammation, and is safe for use. This approach offers a promising new clinical option for UC treatment with broad application potential [79].

Although the above clinical studies have shown the potential of probiotics in the management of UC, there are obvious limitations: 1. Sample size is generally small (such as only 10 cases in FMT study); 2. Short follow-up time (mostly≤ 12 weeks); 3. Lack of standardized strain combinations and doses. For example, Li et al. (2021) showed that triple probiotic capsules significantly improved inflammatory markers, but did not clarify the independent contribution of each strain, and did not set up a probiotic treatment group alone, making it difficult to distinguish its synergistic effect with conventional drugs. In addition, most studies did not conduct stratified analysis based on the baseline characteristics of patients ' intestinal microbiota, ignoring the impact of individual differences on efficacy.]

“[updated text in the manuscript if necessary]” Line 366-430

Comments 5: [Line 509, if possible, convert "yuan" to another currency (dollar or pound).]

Response 5: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [A single course of treatment can cost tens of thousands of US dollars, placing an economic strain on ordinary families that far exceeds their financial capacity.]

“[updated text in the manuscript if necessary]” Line 473

Comments 6: [Discuss the role of probiotics on the mood of patients with ulcerative colitis, discussing possible psychobiotics.]

Response 6: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Emerging evidence also suggests that certain probiotics, often termed "psychobiotics," may improve mood and anxiety in UC patients via the gut–brain axis, though clinical data remain limited.

Mental probiotics are defined as living microorganisms that can have a beneficial effect on the host 's mental health after ingestion [43]. They play a regulatory role through the ' gut-brain axis ' - a two-way communication network connecting the intestine and the brain. In UC patients, chronic intestinal inflammation and dysbacteriosis can affect the function of the central nervous system through the gut-brain axis, leading to a high incidence of emotional disorders such as anxiety and depression [44]. Psychiatric probiotics, such as certain strains of Bifidobacterium longum and Lacticaseibacillus rhamnosus, have been shown to regulate the synthesis and metabolism of intestinal neurotransmitters such as 5-hydroxytryptamine and γ-aminobutyric acid (GABA). To improve emotional state and stress response [43, 45]. This provides a new perspective for the comprehensive management of physical and mental symptoms of UC patients from the microecological level.

Through the above mechanisms, probiotics can not only reduce systemic inflammation, but also have indirect benefits on the emotional health of UC patients [71]. Chronic inflammation is a key association between IBD and ' inflammatory depression ' (anxiety and depression comorbidity) [71]. Among them, pro-inflammatory cytokines such as TNF-α, IL-6 and IL-1β can penetrate the blood-brain barrier, affect neurological function and induce emotional and behavioral changes. By inhibiting these inflammatory factors, probiotics can not only alleviate local intestinal inflammation, but also alleviate inflammation-driven neuropsychiatric symptoms, which is one of the core mechanisms of the role of mental probiotics [71].

In the management of mental health comorbidity, future research should clearly focus on the intervention effect of probiotics on emotional disorders in UC patients. It is recommended that the patient 's emotional score (such as HADS, PHQ-9) be used as a secondary endpoint in clinical trials. The efficacy of specific strains (such as Bifidobacterium longum CCFM1077, Lacticaseibacillus rhamnosus JB-1, etc.) with potential for mental probiotics in alleviating anxiety and depression symptoms in UC patients was system-atically evaluated. By combining microbiome, metabolomics and neuroimaging tech-niques, the specific molecular and neural pathways of probiotics regulating the gut-brain axis and improving mental comorbidities were further revealed. Finally, a personalized microecological treatment plan integrating physical and mental health is provided for UC patients.

]” “[updated text in the manuscript if necessary]” Line 139-141,169-180,312-320，548-557

Comments 7: [Once an abbreviation has been indicated, do not repeat it in full (i.e., Line 514 ulcerative colitis, Line 539 short-chain fatty acid, etc.).]

Response 7: Thank you for pointing this out. We agree with this comment. Therefore, we have revised it in the full text.

Comments 8: [Line 563, indicate the four key areas numerically.]

Response 8: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. 

“[updated text in the manuscript if necessary]” Line 527

Comments 9: [Lines 587, 594, 595, 596, instead of NO, write "not applicable."]

Response 9: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Supplementary Materials: Not Applicable.

Institutional Review Board Statement: Not Applicable.

Informed Consent Statement: Not Applicable.

Data Availability Statement: Not Applicable.]

“[updated text in the manuscript if necessary]” Line 563-572

Comments 1: [Some technical editing is needed in the affiliations section and line 16, 22, 392, 416]

Response 1: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Probiotic supplementation has been shown to alleviate UC symptoms through various mechanisms.

Additionally, some probiotics enhance intestinal barrier function, prevent pathogenic microorganisms from invading, and improve intestinal permeability.

4. The Application of Probiotics in the Alleviation of UC

This study involved a randomized controlled trial with 130 patients diagnosed with UC.]

“[updated text in the manuscript if necessary]” Line 16,22, 341,368

Comments 2: [I do not like how the paragraf from line 44 to 56 is written. It indicates that the existing therapies should be replaced with probiotics and this is a very dangerous insinuations. Please rewrite the part where you mention exsiting treatments or delete it. Also they are not traditional treatments but conventional. There is traditional medicine it is something else. Accordingly the whole Prospects section should be rewritten.

I would not say probiotic are treatment options as they are not indicated in any disease so rephrase this wording in the entire manuscript.]

Response 2: Thank you for pointing this out. We agree with this comment. Therefore, we have revised it in the full text.

 [Currently, the clinical management of UC primarily involves aminosalicylates, glucocorticoids, immunosuppressants, and biologics. While these agents can induce remission, each has limitations. Aminosalicylates are effective for mild to moderate UC but often insufficient for severe cases [5]. Glucocorticoids provide rapid an-ti-inflammatory effects but carry risks of adverse events with long-term use, including osteoporosis, hyperglycemia, and increased infection susceptibility [6]. Immunosuppressants require weeks to months to take effect and may cause hepatorenal toxicity or bone marrow suppression [7]. Biologics, though targeted, are costly, and some patients experience secondary failure or hypersensitivity reactions [88–90]. Moreover, these treatments generally focus on symptom control rather than cure, and many patients experience relapse, necessitating long-term therapy. This situation underscores the need for complementary strategies that address underlying factors such as gut micro-biota dysbiosis.]

“[updated text in the manuscript if necessary]” Line 44-55

Comments 3: [Figure 1 is very hard to read and illegible in some parts. What is written above mental psy...?, and immunological. consider not spinning the letters. It should be completely changed. ]

Response 3: Thank you for pointing this out. We agree with this comment. Therefore, we have modified Table 1. “[updated text in the manuscript if necessary]” Line 88-89

Comments 4: [Line 145 and line 161 need references at the end of paragraph]

Response 4: Thank you for pointing this out. We agree with this comment. Therefore, we have revised it in the text.

Comments 5: [Names of pathogens should be in cursive letters, line 174]

Response 5: Thank you for pointing this out. We agree with this comment. Therefore, we have revised it in the text.

Comments 6: [line 56 the society]

Response 6: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. Therefore, we have revised it in the text.

Comments 7: [line 60 host's health]

Response 7: Thank you for pointing this out. We agree with this comment. Therefore, we have changed. [Probiotics are living microorganisms that can confer beneficial effects on host's health when consumed in adequate amounts.]

“[updated text in the manuscript if necessary]” Line 59