This study investigated how Haemophilus influenzae type b, a major bacterial cause of meningitis, interacts with bovine aortic endothelial cells (BAOEC). In serum-free medium, H. influenzae showed extensive proliferation when cultured with BAOEC, compared to minimal growth in medium alone. Dialysis experiments revealed that BAOEC secrete unidentified low-molecular-mass compounds (less than 2000 kDa) that promote bacterial growth. Outer-membrane protein analysis showed differences between bacteria grown in standard broth versus those grown with BAOEC, particularly in the 45-70 kDa protein range, suggesting phenotypic adaptation to the endothelial cell environment. Both culture conditions produced serum-sensitive bacteria that could be converted to a serum-resistant phenotype resembling in vivo isolates after exposure to serum filtrate. The findings suggest H. influenzae can utilize endothelial cell-derived factors to sustain growth, and that the endothelial cell environment may better mimic in vivo conditions than standard laboratory media, potentially offering insights into bacterial pathogenesis during bloodstream dissemination and meningitis development.

Key findings

• H. influenzae type b proliferates extensively in serum-free medium containing BAOEC, versus minimal growth in medium alone, indicating endothelial cells provide growth-supporting factors
• BAOEC secrete unidentified low-molecular-mass compounds (<2000 kDa) responsible for enhanced bacterial growth, as demonstrated by dialysis experiments
• Bacteria grown with BAOEC show altered outer-membrane protein profiles compared to broth-grown bacteria, with increased minor proteins in the 45-70 kDa range
• Both BAOEC-grown and broth-grown H. influenzae are serum-sensitive in vitro but convert to serum-resistant phenotype after exposure to serum filtrate
• The BAOEC growth model may better approximate in vivo conditions than standard laboratory media, offering a valuable system for studying H. influenzae pathogenesis