Lu Jiang

Despite recent advances in bacterial profiling across the three anatomical types of pediatric short bowel syndrome (SBS), the gut multi-kingdom remains unexplored. We characterized the four-kingdom gut microbiota using modified DNA extraction and deep shotgun metagenomic sequencing of fecal samples from 26 healthy controls and 34 pediatric SBS patients comprising three types (8 SBS I, 15 SBS II, and 11 SBS III). Overall, children with SBS exhibited a significant reduction in α-diversity compared with controls, with no difference observed among SBS types. Compared with controls, the proportion of archaea was significantly decreased in all SBS types, while bacteria, fungi, and viruses remained similar across types. Different types of SBS exhibited distinct microbial signatures: SBS I was enriched with pathogens (such as species from the Streptococcus and Klebsiella genera); SBS II was marked by a depletion of beneficial short-chain fatty acid-producing species (such as Faecalibacterium prausnitzii ); and SBS III displayed loss of bile acid-metabolizing species, alongside a significant expansion of Lactobacillus species. Ecological networks were rewired in SBS, with disruption pronounced in SBS I. Functional analysis revealed that core metabolic pathways were markedly suppressed in SBS I compared with controls. Integrated correlation analyses revealed ileocecal valve loss as the primary determinant of SBS I microbial profiles, linking it to impaired secretory functions and hepatic injury. Our findings deciphered type-specific alterations in the multi-kingdom microbiota and their functional profiles, providing the basis for designing precision microbial therapies aimed at improving long-term outcomes for children with SBS. IMPORTANCE: Pediatric short bowel syndrome (SBS) is a primary cause of intestinal failure, yet prior research characterizing the gut microbiota has focused almost exclusively on bacteria. In this study, we characterized the multi-kingdom microbiome (including bacteria, fungi, archaea, and viruses) across the three anatomical types of pediatric SBS. We found that different SBS subtypes showed distinct microbial patterns: SBS I was enriched in pathogens, SBS II exhibited a depletion of beneficial short-chain fatty acid-producing species, and SBS III was characterized by a loss of bile acid-metabolizing microbes with an expansion of Lactobacillus . Functional analysis showed that SBS I had markedly suppressed core metabolic pathways, and integrated analyses identified the ileocecal valve as a key determinant of microbial gene profiles, with its loss linked to impaired secretion and liver injury. These findings provide a comprehensive multi-kingdom view of the pediatric SBS microbiome and highlight anatomical determinants shaping host–microbiome dysfunction.

Long‐term parenteral nutrition (PN) can lead to liver and intestinal injury in neonates, but effective prevention strategies are limited. Our previous research demonstrated that activation of farnesoid X receptor (FXR) signaling with tropifexor (TXR) could mitigate PN‐induced liver damage in neonatal piglets, but its effect on intestinal injury remains to be addressed. In this study, two‐day‐old Bama minipigs were randomly allocated into three groups ( n = 6/group): EN (receiving enteral nutrition, serving as a control), PN, and PT (PN + TXR treatment). Consequently, TXR treatment significantly mitigated PN‐induced intestinal injury, including villus atrophy, hyper‐permeability, and impaired defense responses. Transcriptomic profiling identified 1188 differentially expressed genes (DEGs) in the PN group compared to EN, among which 108 genes could be substantially attenuated by TXR. Notably, bioinformatics analysis suggested that these DEGs could be classified into “positive regulation of defense response” and “cell–cell adhesion”. Moreover, the top 25 hub genes were identified, including EPCAM, CD28, and IFNG. Furthermore, we demonstrated that EPCAM decreased significantly upon PN administration, which could be substantially attenuated by TXR. Finally, in order to further validate the role of TXR in modulating intestinal barrier function, patient‐derived organoids (PDOs) were used as a mini‐gut model in vitro. Consequently, pharmacological activation of FXR by TXR substantially induced EPCAM expression and enhanced epithelial barrier integrity, particularly in the PDOs derived from pediatric patients receiving PN, while no significant changes were found in those derived from patients receiving oral feeding. In conclusion, this study suggested that TXR treatment could alleviate PN‐induced impairment of defense responses and epithelial barrier dysfunction, potentially offering a preventive strategy for neonates receiving long‐term PN therapy.

The development and severity of metabolic dysfunction-associated steatotic liver disease (MASLD) in children are closely related to alterations of gut microbiota. This study aims to investigate changes in the gut microbiota signature and microbial metabolites in children with MASLD. We collected fecal samples from children and adolescents aged 6–16 years, and the presence of MASLD was diagnosed by ultrasound. We performed 16S ribosomal DNA sequencing and targeted metabolomics in 36 and 25 subjects, consisting of healthy controls, children with obesity, and children with MASLD. The α-diversity was significantly lower in children with obesity and MASLD compared with healthy controls. Linear discriminant analysis of effect size analysis identified Anaerostipes and A. hadrus as the top biomarkers differentiating the obesity group from the MASLD group. In MASLD patients with high alanine aminotransferase values (≥50 U/L for boys and 44 U/L for girls), we observed a decrease in the gut microbiota health index. MASLD patients with high shear wave elastography ( E ) values (≥6.2 kPa) showed an increased abundance of Ruminococcus torques , which was positively correlated with the levels of deoxycholic acid (DCA) and E values. Importantly, the mediation analysis identified positive associations between R. torques and clinical indicators of MASLD that were mediated by DCA. Overall, our study suggests that gut microbiota and metabolites are significantly altered in children with MASLD, and targeting R. torques may offer potential benefits for disease management. IMPORTANCE: This study investigated alterations in the gut microbiota signature and microbial metabolites in children with metabolic dysfunction-associated steatotic liver disease (MASLD). We found that an increased abundance of Ruminococcus torques was associated with increased levels of deoxycholic acid and the progression of MASLD, suggesting that R. torques may serve as a novel clinical target in pediatric MASLD.

Intestinal failure‐associated liver disease (IFALD) is a serious complication of long‐term parenteral nutrition in patients with short bowel syndrome (SBS), and is the main cause of death in SBS patients. Prevention of IFALD is one of the major challenges in the treatment of SBS. Impairment of intestinal barrier function is a key factor in triggering IFALD, therefore promoting intestinal repair is particularly important. Intestinal repair mainly relies on the function of intestinal stem cells (ISC), which require robust mitochondrial fatty acid oxidation (FAO) for self‐renewal. Herein, we report that aberrant LGR5+ ISC function in IFALD may be attributed to impaired farnesoid X receptor (FXR) signaling, a transcriptional factor activated by steroids and bile acids. In both surgical biopsies and patient‐derived organoids (PDOs), SBS patients with IFALD represented lower population of LGR5+ cells and decreased FXR expression. Moreover, treatment with T‐βMCA in PDOs (an antagonist for FXR) dose‐dependently reduced the population of LGR5+ cells and the proliferation rate of enterocytes, concomitant with decreased key genes involved in FAO including CPT1a. Interestingly, however, treatment with Tropifexor in PDOs (an agonist for FXR) only enhanced FAO capacity, without improvement in ISC function and enterocyte proliferation. In conclusion, these findings suggested that impaired FXR may accelerate the depletion of LGR5 + ISC population through disrupted FAO processes, which may serve as a new potential target of preventive interventions against IFALD for SBS patients.

Parenteral nutrition (PN), a vital therapy for patients with intestinal failure, can lead to the development of parenteral nutrition-associated liver disease (PNALD). In this study, we aimed to investigate the role of Lactobacillus johnsonii (L. johnsonii) in a rat model of PNALD. Total parenteral nutrition (TPN)-fed rats were used to assess the role of L. johnsonii in liver steatosis, bile acid metabolism, gut microbiota, and hepatocyte apoptosis. We observed a depletion of L. johnsonii that was negatively correlated with the accumulation of glycochenodeoxycholic acid (GCDCA), a known apoptosis inducer, in rats subjected to TPN. L. johnsonii attenuated TPN-induced liver steatosis by inhibiting fatty acid synthesis and promoting fatty acid oxidation. TPN resulted in a decrease in bile acid synthesis and biliary bile secretion, which were partially restored by L. johnsonii treatment. The gut microbial profile revealed depletion of pathogenic bacteria in L. johnsonii-treated rats. L. johnsonii treatment reduced both hepatic GCDCA levels and hepatocyte apoptosis compared with the TPN group. In vitro, L. johnsonii treatment inhibited GCDCA-induced hepatocyte apoptosis via its bile salt hydrolase (BSH) activity. Our findings suggest that L. johnsonii protects against liver steatosis, bile acid dysregulation, and hepatocyte apoptosis in TPN-fed rats.

Pediatric intestinal failure (IF) is the reduction in gut function to below the minimum necessary for the absorption of macronutrients and/or water and electrolytes, such that intravenous supplementation is required to maintain health and/or growth. The overall goal in treating IF is to achieve intestinal adaptation; however, the underlying mechanisms have not been fully understood. In this study, by performing single-cell RNA sequencing in pediatric IF patients, we found that decreased Kruppel-Like Factor 4 (KLF4) may serve as the hub gene responsible for the functional deficit in mature enterocytes in IF patients, leading to the downregulation of solute carrier (SLC) family transporters (e.g., SLC7A9) and, consequently, nutrient malabsorption. We also found that inducible KLF4 was highly sensitive to the loss of certain enteral nutrients: in a rodent model of total parenteral nutrition mimicking the deprivation of enteral nutrition, the expression of KLF4 dramatically decreased only at the tip of the villus and not at the bottom of crypts. By using IF patient-derived intestinal organoids and Caco-2 cells as in vitro models, we demonstrated that the supplementation of decanoic acid (DA) could significantly induce the expression of KLF4 along with SLC6A4 and SLC7A9, suggesting that DA may function as a potential therapeutic strategy to promote cell maturation and functional improvement. In summary, this study provides new insights into the mechanism of intestinal adaptation depending on KLF4, and proposed potential strategies for nutritional management using DA.

Total parenteral nutrition (TPN) is life-saving therapy for the pediatric patients with intestinal failure (IF) who cannot tolerate enteral nutrition (EN). However, TPN-induced metabolic alterations are also a critical issue for the maintenance of intestinal homeostasis, and thus the global metabolomic signatures need to be addressed. In this study, ileal mucosal biopsies were collected from 12 neonatal Bama piglets receiving either EN or TPN for 14 days, and changes in the intestinal metabolism were examined by multi-omics (HM350 Metabolomics + Tandem Mass Tag (TMT)-based proteomics). As a result, a total of 240 compounds were identified by metabolomics, including 56 down-regulated and 9 up-regulated metabolites. Notably, tissue levels of fatty acyl-carnitines (decreased by 35–85%) and succinate (decreased by 89%) dramatically decreased in the TPN group, suggestive of disrupted processes of fatty acid oxidation (FAO) and the citrate cycle, respectively. Interestingly, however, no differences were found in the production of adenosine 5′-triphosphate (ATP) between groups, suggesting that these dysregulated metabolites may have mainly led to the loss of bioactive compounds rather than energy deficit. Additionally, 4813 proteins were identified by proteomics in total, including 179 down-regulated and 329 up-regulated proteins. The analysis of protein–protein interactions (PPI) indicated that most of the differentially expressed proteins were clustered into “lipid metabolism” and “innate immune responses”. In summary, this work provided new findings in TPN-induced intestinal metabolic alterations, which would be useful to the improvement of nutritional management for IF patients.