TGF-β1-Induced Airway Smooth Muscle Cell Proliferation Involves TRPM7-Dependent Calcium Influx via TGFβR/SMAD3.

TRPM7 in mast cells plays a key role in asthma. However, the effect of TRPM7 on TGF-β1-induced airway smooth muscle cell (ASMC) proliferation remains unclear. Therefore, we designed this study to explore whether TRPM7 is involved in TGF-β1-induced ASMC proliferation. An asthmatic mouse model was established, and the expressions of TGF-β1 and TRPM7 in mice lungs were detected using enzyme-linked immunosorbent assay (ELISA) and western blotting. In addition, murine ASMCs were cultured and stimulated with TGF-β1. Possible TGF-β1 and TRPM7 interactions were examined using RT-PCR and western blotting analyses with ASMCs. The effect of TRPM7 knockdown on ASMC calcium influx was assessed by the fluorescent indicator indo-1. MTT and flow cytometry were applied to evaluate the effects of TRPM7 knockdown on ASMC proliferation and apoptosis. TGF-β1 elevated the expression of TRPM7 via TGFβR/SMAD3. Knocking down TRPM7 decreased the intracellular Ca2+ concentration and cellular proliferation of ASMCs, although apoptosis remained unaffected. Our initial findings suggest that TRPM7 inhibition may prevent asthma-induced airway remodeling.

Chen M ，Zhang W ，Shi J ，Jiang S ... -  《-》

MiR-23b controls TGF-β1 induced airway smooth muscle cell proliferation via direct targeting of Smad3.

MicroRNAs are small yet versatile gene tuners that regulate a variety of cellular processes, including cell growth and proliferation. Here we report that miR-23b inhibited airway smooth muscle cells (ASMCs) proliferation through directly targeting of Smad3. We obtained ASMCs by laser capture microdissection of normal and asthmatic mice lung tissues. Mice ASMCs were cultured and induced by TGF-β1. The implication between TGF-β1 and miR-23b in ASMCs were detected by RT-PCR. The effects of miR-23b on ASMCs proliferation and apoptosis were assessed by transient transfection of miR-23b mimics and inhibitor. The expression of Smad3 in ASMCs were detected by RT-PCR and Western blotting analysis. Dual-Luciferase Reporter Assay System will be applied to identify whether Smad3 is a target gene of miR-23b. TGF-β1 and miR-23b mRNA expression of in-situ bronchial ASMCs collected by laser capture microdissection were increased in asthmatic mice compared to non-asthma controls. This is accompanied by an increase in miR-23b mRNA expression in TGF-β1 induced ASMCs. miR-23b up-regulation significantly inhibited TGF-β1-induced ASMCs proliferation and promoted apoptosis. MiR-23b negatively regulates the expression of Smad3 in ASMCs. Dual-Luciferase Reporter Assay System demonstrated that Smad3 was a direct target of miR-23b. MiR-23b may function as an inhibitor of asthma airway remodeling by suppressing ASMCs proliferation via direct targeting of Smad3.

Chen M ，Shi J ，Zhang W ，Huang L ，Lin X ，Lv Z ，Zhang W ，Liang R ，Jiang S ... -  《-》

MiR-23b controls TGF-β1 induced airway smooth muscle cell proliferation via TGFβR2/p-Smad3 signals.

Abnormal proliferation of ASM (airway smooth muscle) directly contributes to the airway remodeling during development of lung diseases such as asthma. Here we report that a specific microRNA (miR-23b) controls ASMCs proliferation through directly inhibiting TGFβR2/p-Smad3 pathway. The expression of miR-23b in ASMCs was detected by quantitative real-time polymerase chain reaction (RT-PCR). The effects of miR-23b on cell proliferation and apoptosis of ASMCs were assessed by transient transfection of miR-23b mimics and inhibitor. The target gene of miR-23b and the downstream pathway were further investigated. Overexpression of miR-23b significantly inhibited TGF-β1-induced ASMCs proliferation and promoted apoptosis. RT-PCR and Western blotting analysis showed miR-23b negatively regulates the expression of TGFβR2 and p-Smad3 in ASMCs. Subsequent analyses demonstrated that TGFβR2 was a direct and functional target of miR-23b, which was validated by the dual luciferase reporter assay. MiR-23b may function as an inhibitor of airway smooth muscle cells proliferation through inactivation of TGFβR2/p-Smad3 pathway.

Chen M ，Huang L ，Zhang W ，Shi J ，Lin X ，Lv Z ，Zhang W ，Liang R ，Jiang S ... -  《-》

Triptolide inhibits TGF-β1-induced cell proliferation in rat airway smooth muscle cells by suppressing Smad signaling.

We have reported that triptolide can inhibit airway remodeling in a murine model of asthma via TGF-β1/Smad signaling. In the present study, we aimed to investigate the effect of triptolide on airway smooth muscle cells (ASMCs) proliferation and the possible mechanism. Rat airway smooth muscle cells were cultured and made synchronized, then pretreated with different concentration of triptolide before stimulated by TGF-β1. Cell proliferation was evaluated by MTT assay. Flow cytometry was used to study the influence of triptolide on cell cycle and apoptosis. Signal proteins (Smad2, Smad3 and Smad7) were detected by western blotting analysis. Triptolide significantly inhibited TGF-β1-induced ASMC proliferation (P<0.05). The cell cycle was blocked at G1/S-interphase by triptolide dose dependently. No pro-apoptotic effects were detected under the concentration of triptolide we used. Western blotting analysis showed TGF-β1 induced Smad2 and Smad3 phosphorylation was inhibited by triptolide pretreatment, and the level of Smad7 was increased by triptolide pretreatment. Triptolide may function as an inhibitor of asthma airway remodeling by suppressing ASMCs proliferation via negative regulation of Smad signaling pathway.

Chen M ，Lv Z ，Huang L ，Zhang W ，Lin X ，Shi J ，Zhang W ，Liang R ，Jiang S ... -  《-》

MiR-149 attenuates the proliferation and migration of TGF-β1-induced airway smooth muscle cells by targeting TRPM7 and affecting downstream MAPK signal pathway.

Zhu Z ，Zhang L ，Jiang T ，Qian Y ，Sun Y ，Zhang Q ... -  《-》