Colitis Model Research Articles

Sturgeon-derived peptide LLLE alleviated colitis via regulating gut microbiota and its metabolites

Inflammatory bowel disease (IBD), encompassing ulcerative colitis (UC) and Crohn’s disease, entails chronic inflammation of the gastrointestinal tract. The pathogenesis of IBD implicates genetic factors, gut microbiome alterations, and immune dysregulation, contributing to its increasing global prevalence. The sturgeon-derived peptide, which exhibits promising anti-inflammatory effects, provides potential therapeutic insights for managing IBD symptoms. This study aims to elucidate the therapeutic mechanisms of novel sturgeon-derived peptide (LLLE, Leu-Leu-Leu-Glu) by investigating their effects on intestinal inflammation, gut microbiota composition, and fecal metabolites in a mouse model of IBD. LLLE administration alleviated weight loss and disease activity index (DAI) scores in dextran sulfate sodium salt (DSS)-induced colitis in mice. Histopathological examination showed LLLE pretreatment improved colon morphology and histopathological condition and decreased serum interleukin-6 (IL-6) levels. 16S rRNA sequencing indicated LLLE-modulation of gut microbiota, especially alleviated DSS-elevated Bacteroidetes. Fecal metabolomic analysis unveiled that LLLE restores critical metabolites such as indole-3-propionic acid, which is pivotal in anti-inflammatory responses. Altogether, sturgeon peptide exhibits considerable promise as a therapeutic agent for colitis, owing to its anti-inflammatory effects, modulation of gut microbiota, and restoration of essential fecal metabolites.

Huaiyu pill alleviates inflammatory bowel disease in mice blocking toll like receptor 4/ myeloid differentiation primary response gene 88/ nuclear factor kappa B subunit 1 pathway.

To investigate the therapeutic effects of Huaiyu pill (, HYP) on inflammatory bowel disease (IBD) and the underlying mechanisms have not been elucidated. To establish the IBD model, mice were administered with dextran sulfate sodium (DSS). Mice were intragastrically pre-treated with sulfasalazine (SASP) and HYP. Disease activity index (DAI) and colon length were monitored, and the colonic tissues were subjected to hematoxylin-eosin staining. Pro-inflammatory factors and vascular inflammation-related proteins were determined using enzyme-linked immunosorbent assay (ELISA). The potential mechanisms of HYP were examined using network pharmacology analysis.The expressions of zona occludens 1 (ZO-1), occludin, toll like receptor 4 (TLR4), myeloid differentiation primary response gene 88 (MYD88), and nuclear factor kappa B p65 subunit (NF-κB p65) in colon tissues were examined using Western blotting or immunohistochemical analyses. Pre-treatment with HYP enhanced the colon length, decreased DAI scores, and mitigated histopathological alterations in DSS-treated mice. HYP alleviated intestinal inflammation by downregulating the levels of interleukin 1 beta (IL-1β), interleukin 6 (IL-6), tumor necrosis factor alpha (TNF-α) and interleukin 17 (IL-17). Additionally, HYP suppressed the disruption of the gut barrier by upregulating the ZO-1, occludin, and mucin 2 (MUC2) levels and downregulating the endothelin 1 (ET-1) and erythropoietin (EPO) levels. Network pharmacological analysis and experimental results revealed that HYP downregulated the colonic tissue levels of TLR4, MYD88, and NF-κB p65 in DSS-treated mice. This study investigated the in vivotherapeutic effects of HYP on IBD and the underlying molecular mechanisms. These findings provide an experimental foundation for the clinical application of HYP.

Identification of SUMOylation-related signature genes associated with immune infiltration in ulcerative colitis through bioinformatics analysis and experimental validation

ObjectiveUlcerative colitis (UC) is a chronic inflammatory disorder challenging to diagnose clinically. We focused on identifying and validating SUMOylation-related signature genes in UC and their association with immune infiltration. MethodsFive eligible gene expression profiles were selected from the Gene Expression Omnibus (GEO) database and merged into a single dataset comprising 260 UC patients and 76 healthy controls (HC). Differentially expressed genes (DEGs) were identified, and these were intersected with SUMOylation-related genes to obtain differentially expressed SUMOylation-related genes (DESRGs). Next, we identify the signature genes and validate them through comprehensive analyses employing GO, KEGG, GSVA, Lasso-cox regression, ROC curves, and clustering analysis. The infiltrating immune cells were analyzed using the CIBERSORT algorithm and Pearson correlation analysis. Finally, in vitro and in vivo experiments validated the identified signature genes. ResultsPALMD, THRB, MAGED1, PARP1, and SLC16A1 were identified. Next, an excellent predictive model for UC was established and distinct subgroups of patients associated with SUMOylation were identified. Moreover, the NF-κB signaling pathway likely plays a pivotal role in the regulation of SUMOylation in UC. Additionally, we validated that the alterations in PALMD, THRB, and MAGED1 expression in LPSinduced Caco-2 cells concurred with our bioinformatics findings, particularly demonstrating statistically significant differences in PALMD and THRB expression. Finally, in a DSS-induced mouse colitis model, we observed a significant upregulation of PALMD expression. Powered by Editorial Manager® and ProduXion Manager® from Aries Systems Corporation. ConclusionThis study comprehensively elucidates the biological roles of SUMOylation-related genes in UC, identifying PALMD, MAGED1, THRB, PARP1, and SLC16A1 as signature genes that represent promising biomarkers for UC diagnosis and prognosis.

Weizmannia coagulans spores alleviates DSS-induced ulcerative colitis model by modulation of gut flora, metabolites and suppressing TLR4/MyD88/NF-κB pathway

<em>Weizmannia coagulans</em> spores alleviates DSS-induced ulcerative colitis model by modulation of gut flora, metabolites and suppressing TLR4/MyD88/NF-κB pathway

Phage cocktail inhibits inflammation and protects the integrity of the intestinal barrier in dextran sulfate sodium-induced colitis mice model

Ulcerative colitis (UC) is an inflammatory bowel disease characterized by abdominal pain, diarrhea, and rectal bleeding. This study aims to explore the protective effects of a phage cocktail (108 PFU/mL of Clostridium perfringens phage, 108 PFU/mL of Escherichia coli phage, and 108 PFU/mL of Salmonella phage) on a mouse colitis model induced by dextran sulfate sodium (DSS) and its potential toxic effects on normal mice. The results demonstrate that the phage cocktail significantly alleviates clinical symptoms in mice, reduces colon shortening, weight loss, and colonic pathological damage. Furthermore, the phage cocktail markedly suppresses the inflammatory response and safeguards intestinal barrier integrity in the colonic tissues of the mouse colitis model. Preliminary investigation of the toxic effects of the phage cocktail in mice indicates that continuous administration for 14 days does not yield statistically significant differences in hematological and blood biochemical parameters, and specific pathological changes are absent in histopathological examination results. The aforementioned findings suggest that the phage cocktail exhibits anti-inflammatory and intestinal barrier protective effects in the mouse colitis model, and it does not exert significant toxic side effects on mice.

Therapeutic Effects of Bacteroides fragilis Vesicles in a Model of Chemically Induced Colitis in Rats.

The anti-inflammatory properties of Bacteroides fragilis vesicles were studied in a rat model of dextran sodium sulfate-induced colitis. According to the histology results, addition of B. fragilis vesicles to the therapy promoted colon repair. Evaluation of the disease activity index confirms the high rate of colon recovery: against the background of vesicle administration, the absence of blood in stool, normal stool consistency, and body weight normalization were observed.

Pentacyclic Triterpenoid Content in Cranberry Raw Materials and Products

Cranberry fruit extracts have been shown to inhibit expression of pro-inflammatory cytokines in THP-1 cells and reduce colonic tumor burden and tissue inflammation in a mouse model of colitis. These activities are attributed to both the triterpenoid and polyphenol constituents of the fruit. The pentacyclic triterpenoids ursolic acid (UA), oleanolic acid (OA), corosolic acid (CA), maslinic acid (MA), and esters of UA and OA occur in the waxy layer of cranberry peel, and their content in cranberry products is likely to vary with the fruit source and processing methods. UPLC-MS (ultra performance liquid chromatography-mass spectrometry) was applied to determine the four triterpenoid acids and their esters in cranberry products and raw materials. Cranberry pomace, a side stream in juice production, was a rich source at 64,090 µg total triterpenoids/g DW. Cranberry juice beverages ranged from 0.018 to 0.26 µg/g of product, fruit samples ranged from 6542 to 17,070 µg/g DW, and whole berry products contained up to 2665 µg/g DW. Free UA was the most plentiful triterpenoid in all samples. These analyses illustrate the potential value of an underutilized side stream in cranberry juice production and highlight potential benefits of whole fruit products.

Isatidis Folium Represses Dextran Sulfate Sodium-Induced Colitis and Suppresses the Inflammatory Response by Inhibiting Inflammasome Activation

Background/Objectives: Isatidis Folium (IF) has been used in traditional medicine for various ailments, and recent research highlights its anti-inflammatory, antiviral, and detoxifying properties. This study investigated the anti-inflammatory effects of a hydroethanolic extract of IF (EIF) on inflammasomes and colitis. Methods: Dextran sulfate sodium (DSS)-induced colitis model C57BL/6 mice were treated with DSS, mesalamine, or EIF (200 mg/kg). Parameters such as daily disease activity index (DAI), spleen weight, colon length, and histopathology were evaluated. Intestinal fibrosis, mucin, and tight junction proteins were assessed using Masson’s trichrome, periodic acid–Schiff, and immunohistochemistry staining. RAW264.7 and J774a.1 macrophages were treated with EIF and lipopolysaccharide, with cell viability assessed via the cell counting kit-8 assay, nitric oxide (NO) production with Griess reagent, and cytokine levels with the enzyme-linked immunosorbent assay. NF-κB inhibition was analyzed using the luciferase assay, and phytochemical analysis was performed using UPLC-MS/MS. Results: EIF mitigated weight loss, reduced DAI scores, prevented colon shortening, and attenuated mucosal damage, fibrosis, and goblet cell loss while enhancing the tight junction protein occludin. The anti-inflammatory effects of EIF in RAW264.7 cells included reduced NO production, pro-inflammatory cytokines, and NF-κB activity, along with inhibition of NLRP3 inflammasome responses in J774a.1 cells. The key constituents identified were tryptanthrin, indigo, and indirubin. Conclusions: Animal studies demonstrated the efficacy of EIF in alleviating colitis, suggesting its potential for treating inflammatory diseases.

Effectiveness of Propolis on Experimental Colitis Model In Rats

Aim: This study investigated the therapeutic potential of propolis in an experimental colitis model induced by acetic acid in female Wistar albino rats. Materials and Methods: Thirty rats were divided into five groups: a control group and four experimental groups. Colitis was induced in the second, third, and fifth groups by rectal administration of 1 ml of 4% acetic acid. The third group received rectal propolis solution (50 mg/ml), while the fourth group was given only rectal propolis solution. The fifth group received 1 ml of olive oil rectally after the onset of colitis. Stool consistency and weight loss were monitored, and colon tissue samples were collected for microscopic and macroscopic evaluation. The levels of MDA, MPO, and caspase-3 in tissue, as well as TNF-α and IL-10 levels in blood samples, were examined. Results: The group administered propolis showed a significant decrease in microscopic and macroscopic scores compared to the other experimental groups. The levels of MDA, MPO, and caspase-3 in the tissue, as well as TNF-α and IL-10 levels in blood samples, were significantly decreased in the propolis group compared to the other experimental groups. Weight loss and stool consistency also showed improvement in the propolis group compared to the other experimental groups. Discussion: Propolis may have therapeutic effects in experimental colitis induced by acetic acid. The decrease in oxidative damage and inflammation seen in the propolis group indicates that it may be a useful therapeutic agent for colitis treatment.

An Intelligent Intestine-on-a-Chip for Rapid Screening of Probiotics with Relief-Enteritis Function.

Screening probiotics with specific functions is essential for advancing probiotic research. Current screening methods primarily use animal studies or clinical trials, which are inefficient and costly in terms of time, money, and labor. An intelligent intestine-on-a-chip integrating machine learning (ML) is developed to screen relief-enteritis functional probiotics. A high-throughput microfluidic chip combined with environment control systems provides a standardized and scalable intestinal microenvironment for multiple probiotic cocultures. An unsupervised ML-based score analyzer is constructed to accurately, comprehensively, and efficiently evaluate interactions between 12 Bifidobacterium strains and host cells of the colitis model in the intestine-on-a-chips. The most effective contender, Bifidobacterium longum 3-14, is discovered to relieve intestinal inflammation and enhance epithelial barrier function in vitro and in vivo. A distinct advantage of this strategy is that it can intelligently differentiate small therapeutic variations in probiotic strains and prioritize their efficacies, allowing for economical, efficient, accurate functional probiotics screening.

Design, Synthesis, and Bioevaluation of Novel NLRP3 Inhibitor with IBD Immunotherapy from the Virtual Screen.

NLRP3, a crucial member of the NLRP family, plays a pivotal role in immune regulation and inflammatory modulation. Here, we report a potent and specific NLRP3 inhibitor Z48 obtained though docking-based virtual screening and structure-activity relationship studies with an IC50 of 0.26 μM in THP-1 cells and 0.21 μM in mouse bone marrow-derived macrophages. Mechanistic studies indicated that Z48 could bind directly to the NLRP3 protein (KD = 1.05 μM), effectively blocking the assembly and activation of the NLRP3 inflammasome, consequently manifesting anti-inflammatory properties. Crucially, with acceptable mouse pharmacokinetic profiles, Z48 demonstrated notable therapeutic efficacy in a mouse model of DSS-induced ulcerative colitis, while displaying no significant therapeutic impact on NLRP3KO mice. In conclusion, this study provided a promising NLRP3 inflammasome inhibitor with novel molecular scaffold, poised for further development as a therapeutic candidate in the treatment of inflammatory bowel disease.

Qin-Yu-Qing-Chang decoction reshapes colonic metabolism by activating PPAR-γ signaling to inhibit facultative anaerobes against DSS-induced colitis

BackgroundQin-Yu-Qing-Chang decoction (QYQC), an herbal formula from China, is extensively employed to manage ulcerative colitis (UC) and exhibits potential benefits for colonic function. Nevertheless, the fundamental molecular mechanisms of QYQC remain largely uncharted.MethodsThe primary constituents of QYQC were determined utilizing UHPLC-MS/MS analysis and the effectiveness of QYQC was assessed in a mouse model of colitis induced by dextran sulfate sodium. Evaluations of colon inflammatory responses and mucosal barrier function were thoroughly assessed. RNA sequencing, molecular docking, colonic energy metabolism, and 16S rRNA sequencing analysis were applied to uncover the complex mechanisms of QYQC in treating UC. Detect the signal transduction of the peroxisome proliferator-activated receptor-γ (PPAR-γ) both in the nucleus and cytoplasm. Furthermore, a PPAR-γ antagonist was strategically utilized to confirm the functional targets that QYQC exerts.ResultsUtilizing UHPLC-MS/MS, the principal constituents of the nine traditional Chinese medicinal herbs comprising QYQC were systematically identified. QYQC treatment substantially ameliorated colitis in mice, as evidenced by the improvement in symptoms and the reduction in colonic pathological injuries. Besides, QYQC treatment mitigated the inflammatory response and improved mucosal barrier function. Furthermore, QYQC enhanced the mitochondria citrate cycle (TCA cycle) by triggering PPAR-γ signaling and increasing the proportion of PPAR-γ entering the nucleus. This prevented the unconstrained expansion of facultative anaerobes, particularly pathogenic Escherichia coli (E. coli, family Enterobacteriaceae) and thus improved colitis. Results of molecular docking indicated that the representative chemical components of QYQC including Baicalin, Paeoniflorin, Mollugin, and Imperatorin bound well with PPAR-γ. The impact of QYQC on colitis was diminished in the presence of a PPAR-γ antagonist.ConclusionsIn summary, QYQC ameliorates UC by activating PPAR-γ signaling and increasing the proportion of PPAR-γ entering the nucleus, which enhances the energy metabolism of intestinal epithelial cells and thereby preventing the uncontrolled proliferation of facultative anaerobes.

NPSR1 promotes chronic colitis through regulating CD4+ T cell effector function in inflammatory bowel disease

BackgroundNeuropeptide S receptor 1 (NPSR1) has been implicated in the the onset of inflammatory bowel disease (IBD), though its exact mechanism remains unclear. This study investigates the role of NPSR1 in regulating CD4+ T cell effector function in IBD. MethodsPeripheral blood and colonic mucosal biopsies from IBD patients, as well as dextran sodium sulfate (DSS)-induced mouse colitis models, were analyzed to assess the effects of NPSR1 on colitis and CD4+ T cell-mediated immune responses. NPSR1 knockdown was conducted both in vitro and in vivo to elucidate underlying mechanisms. Expression of NPSR1 and CD4+ T cell-related factors was measured using quantitative real-time PCR, immunoblotting, cytometric bead array, immunofluorescence, and immunohistochemistry. CD4 + T cell effector functions were evaluated through flow cytometry, EdU incorporation assay, Annexin V-FITC/PI staining, and transwell assay. ResultsNPSR1 expression was elevated in the intestinal tissues from IBD patients. Its downregulation provided protection in DSS-induced mouse colitis models. NPSR1 correlated positively with CD4 + T cell-mediated inflammation, and its knockdown reduced CD4+ T cell-mediated immune responses and inhibited CD4+ T cell differentiation. Additionally, NPSR1 knockdown decreased CD4+ T cell proliferation, increased apoptosis, and enhanced CCL2-induced migration in vitro, while significantly reducing Th1 cell chemotaxis in vivo. ConclusionsThis study demonstrates that NPSR1 promotes chronic colitis by regulating CD4 + T cell effector functions in IBD, offering potential new therapeutic strategies for IBD treatment.

Establishment of an ulcerative colitis model using colon organoids derived from human induced pluripotent stem cells

The etiology of inflammatory bowel disease (IBD) is complex, with much room for a greater understanding and development of improved therapies. Therefore, establishing a reliable IBD model is crucial for future advancements. In this study, human induced pluripotent stem (iPS) cell-derived colon organoids (hiPSC-COs) were treated with a combination of tumor necrosis factor alpha (TNF-α), interferon-gamma (IFN-γ), and interleukin (IL)-1β (3 cytokines [3CK]), known to be elevated in the serum of IBD patients. Inflammatory responses in stromal cells and damage to intestinal epithelial cells were observed in the 3CK-treated hiPSC-COs. Comparison of molecular signatures of 3CK-treated hiPSC-COs with those of ulcerative colitis (UC) patient's colon revealed that 3CK-treated hiPSC-COs resemble UC patient's colon. Furthermore, the elevated production of inflammatory cytokines observed in 3CK-treated hiPSC-COs was attenuated by treatment with tofacitinib. Our UC model will be an essential tool to understand its pathologic mechanisms and identify effective therapeutic approaches.

A CRISPR mediated point‐of‐care assay for the detection of mucosal calprotectin in an animal model of ulcerative colitis

AbstractInflammatory bowel disease (IBD) is a chronic disorder associated with inflammation in the gastrointestinal tract, leading to a range of debilitating symptoms. Fecal calprotectin is an established biomarker for ulcerative colitis (UC), one of the main IBD diseases, which provides indications of the presence and severity of inflammation in the digestive tract. Enzyme‐Linked Immunosorbent Assay (ELISA) as a gold standard approach for fecal calprotectin detection is time‐consuming and impractical in point‐of‐care settings. Moreover, obtaining fecal samples from patients is challenging and inhibits longitudinal monitoring. To address these specific problems, we have developed a novel approach for detecting calprotectin which leverages clustered regularly interspaced short palindromic repeats (CRISPR)/Cas technology. We successfully developed a portable tube‐based CRISPR/Cas assay for point‐of‐care testing of calprotectin. This assay showed a detection range from 1 to 10,000 ng/ml (over 4 log units), using both fluorescent and colorimetric analytical techniques. The established assay was further validated through measurements in mucosal samples obtained in an anesthetised preclinical rodent model of UC, with 2–3 times higher calprotectin concentration detected in UC rat samples compared to that of healthy control animals. This point‐of‐care test may provide a rapid, precise, and user‐friendly approach for the diagnosis and monitoring of IBD through mucosal sample testing.

Evaluation of TNF-α and IFN-γ primed conditioned medium of mesenchymal stem cell in acetic acid-induced mouse model of acute colitis

IBD, an autoimmune-inflammatory disorder that affects people who are genetically prone to inflammation. There is a lot of interest in MSC-CM therapy, especially when primed with TNF-α + IFN-γ. Throughout the study, data were collected on the percentage of apoptotic cells, gene expression of ZO-1, Foxp3, GATA3, IDO-1, Muc2, T-bet, Notch1, TNFR2, and ROR-γt, colon weight and length, histopathological analysis, and DAI. TNF-α and IL-10 levels were assessed in addition to the NO level. The results suggest that primed MSC-CM improved DAI, mucosal deterioration, intestinal inflammation and NO concentration. The amount of TNF-α was decreased, but IL-10 and the colon’s percentage of apoptotic cells was increased. The mRNA expression of ZO-1, Foxp3, GATA3, IDO-1, and Muc2 genes increased greatly in the treatment groups, while the expression of T-bet, Notch1, TNFR2, and ROR-γt genes has decreased. These studies suggest that primed MSC-CM may combine with common treatments to improve responsiveness.

Fresh bamboo juice regulates the diversity of intestinal microflora in mice by promoting the function of ILC2 cells to mediate intestinal protection

Background and aimUlcerative colitis (UC) is a growing global health concern, with current treatments facing challenges like drug dependence and side effects. Fresh bamboo juice (FBJ), known for its antimicrobial and potential immune-modulating properties, has shown promise as a natural therapeutic agent. The present study aimed to explore the protective effects of FBJ against colitis and further analyze the changes of gut microbiota composition, metabolite profiles, and underlying immune mechanisms. Materials and MethodsA colitis model in mice was established using DSS to investigate the effectiveness of FBJ. Intestinal tissue and fecal samples were also collected for 16S rRNA gene sequencing and liquid chromatography-mass spectrometry (LC-MS) analysis. Additionally, immunofluorescence and flow cytometry were employed to detect the proliferation and function of group 2 innate lymphoid cells (ILC2). Enzyme-linked immunosorbent assay (ELISA) was used to measure the cytokines secreted by immune cells. ResultsFBJ demonstrated significant therapeutic effects against DSS-induced colitis in mice. At the genus level, the abundance of Bacteroides, Akkermansia and unassigned bacteria in the bamboo juice group increased compared with the DSS group. In contrast, the abundance of Alloprevotella, Lactobacillus, Lachnospiraceae_NK4A136_group and Ruminococcaceae_UCG-014 significantly decreased. FBJ partially restored the balance of gut microbiota, as evidenced by the increased levels of beneficial bacteria. Metabolome analysis revealed significant alterations in fecal metabolites, including 3-Hydroxypyridine, Pyridoxine, SM(d18:1/16:0), and DL-Methionine sulfoxide were remarkably altered. Dysregulation of pathways such as Vitamin B6 metabolism, sphingolipid metabolism, and tyrosine metabolism was observed, which may contribute to protection against colitis. Flow cytometry and immunofluorescence showed a significant reduction in the proportion of ILC2 cells following FBJ treatment in the DSS group (1.82 % v.s. 3.18 %, P < 0.05). ELISA showed that the FBJ group had lower levels of IL-5, IL-6, IL-10, IL-13, IL-33, TNF-α, IFN-γ in intestinal tissue. ConclusionsOur findings demonstrate that FBJ exerts a protective effect against colitis, primarily by modulating the intestinal flora and metabolite profiles in mice with colitis. Furthermore, the observed alterations in bacterial flora and metabolites likely affect ILC2 function and cytokine production, thereby mediating the protective effects against colitis through modulation of the immune system.

Neutrophil elastase-activated carbon dots for non-invasive early diagnosis of inflammatory bowel disease by near-infrared-II fluorescence imaging

Most near-infrared-II (NIR-II) fluorescent probes produce an “always-on” output with poor signal specificity for biomarkers. Neutrophil elastase (NE) is a critical factor in chronic inflammation or in the acute response to infection and injury, and NE activatable NIR-II fluorescent probes can aid in the early clinical diagnosis of inflammatory bowel disease (IBD). In this paper, we synthesized OPDG-PFA@PEI carbon dots (CDs) through a two-step reaction of solvothermal/surface covalent bond modification, which can “turn on” specific NIR-II fluorescence signals related to the level of NE produced by neutrophils and target sites of inflammation effectively, thus realizing the monitoring of in-situ NE in cells and in vivo. OPDG-PFA@PEI CDs fluorescent nanoprobe show high-performance real-time non-invasive NIR-II fluorescence imaging for early diagnostic evaluation of IBD in mouse models of DSS-induced IBD, providing a promising approach for the development of clinical non-invasive and more convenient early diagnosis of IBD.

Enhanced Extracellular Vesicle Cargo Loading via microRNA Biogenesis Pathway Modulation.

Extracellular vesicles (EVs) are physiological vectors for the intercellular transport of a variety of molecules. Among these, small RNAs, and especially microRNAs (miRNAs), have been identified as prevalent components, and there has thus been a robust investigation of EVs for therapeutic miRNAs delivery. However, intrinsic levels of EV-associated miRNAs are generally too low to enable efficient and effective therapeutic outcomes. We hypothesized that miRNA localization to EVs could be improved by limiting competing interactions that occur throughout the miRNA biogenesis process. Using miR-146a-5p as a model, modulation of transcription, nuclear export, and enzymatic cleavage steps of miRNA biogenesis were tested for impact on EV miRNA loading. Working in HEK293T cells, various alterations in the EV biogenesis pathway were shown to impact miRNA localization to EVs. The system was then applied in induced pluripotent stem cells (iPSCs), a more promising substrate for therapeutic EV production, and EVs were separated and assessed for anti-inflammatory efficacy in vitro and in a murine colitis model, where the preservation of function was validated. Overall, the results highlight necessary considerations when designing a cell culture system for the devoted production of miRNA-loaded EVs.

PH-Sensitive Tacrolimus loaded nanostructured lipid carriers for the treatment of inflammatory bowel disease

Inflammatory Bowel Disease is the chronic tissue inflammation of the lower part of the Gastrointestinal tract (GIT). Conventional therapeutic approaches face numerous challenges, often making the delivery system inadequate for treating the disease. This study aimed to integrate a pH-sensitive polymer and nanostructured lipid carriers (NLCs) to develop a hybrid nanocarrier system. Tacrolimus-loaded NLCs coated with Eudragit® FS100 (TAC-NLCs/E FS100) nanoparticles were prepared via double emulsion technique followed by an aqueous enteric coating technique. Various parameters, such as particle size, entrapment efficiency, and zeta potential were optimized using Design Expert software®. Cetyltrimethyl ammonium bromide (CTAB) was used as a cationic surfactant which induces a positive charge on the nanoparticles. These cationic NLCs can adhere to the mucosal surface, thereby enabling prolonged retention. In vitro drug release was assessed, and the results demonstrated that drug release was retarded at pH 1.2 corresponding to upper GIT pH and maximum drug was released at pH 7.4 (colonic pH). Moreover, we evaluated TAC-NLCs/E FS100 nanoparticles in murine colitis models to gauge the efficacy of both coated and uncoated NLCs formulation. The TAC-NLCs/E FS100 showed a pronounced reduction in induced colitis, as evident from the restoration of morphological features, improved histopathological scores, antioxidant levels, and decreased the levels of proinflammatory cytokines. Thus, pH-sensitive TAC-NLCs/EFS 100 are attributed to the enhanced localization and targeted delivery at the specific site.