Objective To explore the effects of down-regulation of the expression of heat shock protein A12B (HSPA12B) of mouse pulmonary microvascular endothelia cells (mPMVECs) on the inflammatory reaction, migration, and ultrastructural changes induced by lipopolysaccharide (LPS). Methods The mPMVECs subcultured in vitro were divided into the LPS group (LPS of 1 μg/mL was added), LPS+siRNA group (siRNA oligonucleotides of relevant gene segments of HSPA12B were transiently transfected into mPMVECs and then LPS of 1 μg/mL was added), and LPS+NC group (siRNA oligonucleotides of negative controls were transiently transfected into mPMVECs and then LPS of 1 μg/mL was added). The migration of cells of the LPS+siRNA group and LPS+NC group was observed by the Transwell assay and wounding assay. The ultrastructural changes of mPMVECs were observed by the transmission electron microscopy. The expressions of TNF-α, IL-6, IL-10, and IL-1β mRNA of cells of each group were detected by the Real-Time PCR. Results Compared to the LPS+NC group, the mitochondrion damage of mPMVECs of the LPS+siRNA group was severer; the edema of endoplasmic reticulum was more significant; the expressions of TNF-α, IL-6, and IL-10 mRNA significantly up-regulated (P<0.05); the expression of IL-1β down-regulated, but the difference was not statistically significant (P>0.05); and the ability of cell migration was significantly decreased (P<0.05). Conclusion The inflammatory reaction of LPS-induced mPMVECs was enhanced and the cell migration was inhibited after HSPA12B was down-regulated. HSPA12B may be involved in the process of protecting mPMVECs from being invaded by LPS.