Isoform Ratio Rather than Total Content Determines Nonlinear Bioactivity of Red Kidney Bean Phytohemagglutinin in Mice

Abstract Red kidney bean phytohemagglutinin (PHA) is an isolectin system of erythroagglutinating (PHA-E) and leucoagglutinating (PHA-L) subunits, but the biological significance of the natural ratio remains unclear. Here, natural PHA fractions with stepwise PHA-E/PHA-L ratios (N1 = 0.85, N2 = 1.24, N3 = 1.80) were tested in a BALB/c mouse sensitization model. The fractions induced nonlinear, qualitatively distinct responses. N1 and N2 both increased ileal epithelial length by approximately 10.2% relative to PBS; N2 additionally showed the highest IgG2a/IgG1 ratio, a prominent IL-17A response, and restored ZO-1 and occludin expression to PBS levels. N3 evoked the strongest IL-4-dominant Th2-like response and the highest sIgA level among PHA fractions, yet shortened the epithelium by 14.3% with a weaker barrier-gene expression. These phenotypes coincided with fraction-specific gut microbiota remodeling. Thus, the PHA isoform ratio critically determines immune, intestinal, and microbial outcomes and warrants consideration in bean lectin safety evaluation.