Moso bamboo is one of the most important economic bamboo species in China, but cadmium (Cd) pollution has become a potential threat of its sustainable development. Silicon (Si) reduces Cd accumulation in many plant species, however, the exact mechanisms of this effect in Moso bamboo are still poorly understood. Here, we investigated the effect of Si on Cd accumulation in Moso bamboo in terms of Cd concentration in roots, Cd cellular and subcellular distribution, root cell morphology and genes expression. The effect of Si on alleviation of Cd-induced inhibition of root elongation was not obvious, but Si could significantly reduce Cd accumulation in roots at all tested Cd concentrations. Cd was localized in all cells of roots but Si treatment alter Cd distribution from all cells to distal side of exodermis cells in roots. Semi-quantitative determination of Cd by energy-dispersive X-rays revealed higher Cd concentrations in exodermis but lower concentrations in the stele when Si was present. However, Si increased Cd accumulation in root cell walls but decreased Cd concentrations in cell sap. Moreover, more than 70% of Cd and Si was found in hemicellulose 1 fraction of the cell wall. These results suggested that Si reduces Cd accumulation by sequestering Cd in hemicellulose 1 in the cell wall at the subcellular level and retaining most of the Cd in the root exodermis at the cellular level in Moso bamboo when short-term Si was applied.