Lung transplantation is one therapeutic treatment for severe lung dysfunctions. Lung transplant recipients often face a higher mortality rate of approximately 50% of the lung transplantation patients within five years and a more frequent loss of graft function, as compared to other organ transplants. Lung ischemia and reperfusion (I/R) injury is the most common cause of respiratory failure during the first 72 hr after transplantation. In fact, I/R injury is the risk factor for the producing diverse pro-inflammatory cytokines leading to pulmonary inflammation and bronchiolitis obliterans, manifesting a process of inflammatory cascades of the small airways, resulting in activation of the inflammatory process against the airway epithelium, which plays crucial role in innate immunity to destroy the harmonious immunity of airway environment in recipients. Glucosamine is a commonly consumed supplement, recently being regarded as a potential anti-inflammatory molecule. Our previous studies have demonstrated an effective suppression of glucosamine on lipopolysaccharide (LPS)-induced inflammatory response, such as interleukin production by attenuating mitogen-activated protein kinase (MAPK) pathways in human bronchial epithelial cells. This study aimed to disclose the glucosamine impact on cellular proliferation in human alveolar epithelial cells (A549) and bronchial epithelial cells (HBECs). With the trypan blue-exclusion assay, we observed that glucosamine (10, 20, 50 mM) caused a decrease in cell number of both A549 cells and HBECs at 24 hr; a more profound and longer effect on cell number reduction of HBECs was noted at 48 hr. With the flow cytometric analysis, we also noted an elevated cell numbers accumulated within the G0/G1 phase by glucosamine (10, 20, 50 mM) at 24 hr in both cell types. By examining the phosphorylation of the retinoblastoma (Rb) protein, an inhibition of Rb protein phosphorylation by glucosamine (20, 50 mM) was revealed in A549 cells and HBECs. By measuring mRNA and protein expression of cell cycle-related regulators, glucosamine at 50 mM was able to upregulate both mRNA and protein expression of HO-1 and p53 in A549 cells, but only HO-1 in HBECs; glucosamine induced p21 mRNA expression, but reduced p21 protein expression in A549 cells. In addition, we further uncovered that glucosamine significantly downregulated the p21 protein stability by the proteasomal proteolytic pathway. Altogether, our findings suggest that high dose of glucosamine may disturb cell cycle progression at least in part by modulating the expression of HO-1, p53, p21 as well as Rb phosphorylation, leading to a halt at G0/G1 phase and an inhibition in cell proliferation.