Systems-Level Proteomic and Biochemical Profiling of Plasma from Captive Indian Star Tortoise with Reactome Pathway Enrichment Analysis

Background/Objectives: The Indian star tortoise (Geochelone elegans) is a protected species for which physiological and molecular health indicators remain poorly characterized. This study aimed to monitor and analyze plasma proteome profiles and biochemical parameters in captive adult Indian star tortoises and to identify potential diagnostic biomarkers. Methods: Plasma samples from nine clinically healthy adult Indian star tortoises (four males and five females) maintained in captivity were subjected to biochemical profiling and proteomic analysis. Sex-related differences in biochemical parameters were evaluated, and differentially expressed proteins were mapped to Homo sapiens Reactome pathways to identify significantly enriched biological processes. Results: Plasma biochemical profiling established baseline reference values, indicating stable hepatic and metabolic function in captive tortoises. Creatinine and urea concentrations were significantly higher in females than in males (p < 0.05), suggesting sex-related differences in protein metabolism or renal function. No significant sex-related differences were observed in hepatic enzymes (ALP, ALT, AST, and GGT), muscle-associated enzymes (CK and LDH), glucose, cholesterol, triglycerides, total proteins, albumin, or electrolyte concentrations (Na, K, Ca, Mg, Cl, P, and Fe). Proteomic analysis identified 12 differentially expressed proteins, including nine upregulated and three downregulated proteins. Functional pathway analysis revealed 90 significantly enriched Reactome pathways (FDR < 0.05). Upregulated proteins were primarily associated with cytoskeletal organization (KRT75, KRT5, and KRT17), lipid transport and remodeling (APOB), coagulation (F10), extracellular transport (TTR), immune response (WFDC3), transmembrane signaling (KCP), and gamete interaction (ZAN). Downregulated proteins (C7, SERPING1, and PZP) were linked to complement activation and acute-phase response pathways. Conclusions: Captive Indian star tortoises exhibited increased cytoskeletal remodeling and coagulation activity together with reduced complement activation. These findings provide novel insights into the plasma proteome of this species and identify candidate biomarkers that may support future health assessment, physiological monitoring, and diagnostic applications in Indian star tortoises.