The Influence Of Retinoic Acid To Rat Alveolar Epithelial Cells Type Ⅱ Which Affected By Acrolein

Objective To establish a stable method for isolation, primary culture and identification of rat alveolar epithelial cells type Ⅱ (AECII).Explore the effect of acrolein on the cell cycle of AECII and to observe the antagonism action of retinoic acid (RA) to acrolein. Method The lung was instilled with4.2u/ml elastinase solution via the tracheal cannula to isolate AECII cells from the lung tissue.The cell suspension was panned on a rat IgG-coated Petri dish to purify AECII. The primary culture cells were identified by electron microscopy, improved papanicolaou staining, tannic acid staining, alkaline phosphatasestaining (BCIP/NBT) and immunohistochemical staining, simultaneously calculate the purity of AECII. Using MTT method to observe the cell proliferation of AECII treated by acrolein and cell cycle was detected by flow cytometry. The expression level of mRNA of P21and P53was examined by the reverse transcription quantitative polymerase chain reaction. Results The primary culture cells presented the island-like growth figure under the inverted microscope, the lamellar bodies could be observed by electron microscopy, they were presented as black granules in the cytoplasm by tannic acid staining. The cell proliferation was measured by MTT method has downregulated. Cell cycle was blocked in phase G1after treated by acrolein, and RA has no antagonism action to acrolein observed by flow cytometry. Compared to the control group, mRNA expression of p21was downregulated in all three groups, but that of p53showed no significant alteration in all groups. Conclusion Elastinase functions gently and could keep the cell membrane intact during the separation process, so the gained cells had good viability. Immunological attachment with rat IgG could efficiently increase the purity of AECII from cell suspensions. All of the above used assays were stable and specific to identify isolated AECII. The damage of acrolcin to AECII was obvious but RA had no antagonism action to acrolein. Compared to the control group, mRNA expression of p21has downregulated and RA has no antagonism action to acrolein; p53showed no significant alteration in all groups.