Metabolic Signatures of Acinetobacter baumannii and Klebsiella pneumoniae Infections in Acute-on-chronic Liver Failure

Acute-on-chronic liver failure (ACLF) is a life-threatening syndrome characterized by rapid hepatic decompensation, systemic inflammation, and high short-term mortality. Despite its clinical significance, the mechanistic underpinnings of ACLF remain poorly defined, limiting the development of effective diagnostic and therapeutic strategies. This thesis establishes a comprehensive in vitro–ex vivo experimental framework integrating metabolomics and cytokine profiling to advance potential biomarker discovery and translational applications in ACLF. By combining complementary datasets, minimizing cohort-specific biases, and adhering to rigorous biomarker research standards, this work provides a robust methodological foundation that bridges experimental findings with clinical practice. Using infection models with Acinetobacter baumannii and Klebsiella pneumoniae, combined with metabolomic profiling and cell viability assays, four hepatotoxic metabolites — α-ketoglutarate, N-acetylglutamine, indoleacetic acid, and N8-acetylspermidine — were identified and validated via a two-stage (in vitro-ex vivo) strategy. These metabolites illuminate novel aspects of ACLF pathogenesis, particularly in the context of infections with Acinetobacter baumannii and Klebsiella pneumoniae, two clinically relevant Gram-negative pathogens frequently associated with ACLF patients. The findings highlight the interplay between gut-liver axis dysregulation, mitochondrial dysfunction, and microbial contributions as pivotal drivers of metabolic derangements and hepatocyte injury. Importantly, infections with A. baumannii and K. pneumoniae exacerbate oxidative stress and mitochondrial dehydrogenase dysfunction, directly linking microbial metabolism with hepatocyte cell death and progression of liver cells failure. In parallel, cytokine expression profiling in gut and liver infection models revealed immune dysregulation patterns resembling those observed in sepsis, a frequent and severe complication of ACLF. Infections with A. baumannii and K. pneumoniae elicited robust pro-inflammatory and regulatory cytokine responses, suggesting that epithelial cytokine signatures may serve as early indicators of immune activation and systemic inflammation. These immunologic insights complement the metabolic findings, enhancing the translational relevance of the model and underscoring the dual role of microbial metabolites and cytokine disturbances in ACLF pathogenesis. Collectively, this research advances understanding of the metabolic and immunologic underpinnings of ACLF, underscoring the potential of identified metabolites as diagnostic biomarkers and therapeutic targets. By bridging experimental findings with clinical relevance, this thesis lays the foundation for novel strategies to improve ACLF patient outcomes. The integrated framework developed here not only enhances knowledge of ACLF pathophysiology but also provides a basis for future investigations into precision medicine approaches, enabling earlier detection, personalized interventions, and improved management of this high-burden condition.