Objective: Pulmonary fibrosis is a common pathological phenomena in lung cancer patients after chemotherapy or radiotherapy, which is a key factor hindering to transport ion of high concentrated drug to the lung tissu...Objective: Pulmonary fibrosis is a common pathological phenomena in lung cancer patients after chemotherapy or radiotherapy, which is a key factor hindering to transport ion of high concentrated drug to the lung tissue, peptide transporter has become targets of the rational design of peptides and peptide drug. The purpose of the study is to investigate the expression of PEPT2 mRNA in the lung of rats with bleomycin(BLM)-induced pulmonary fibrosis. Methods: Fifty healthy adult Spragne-Dawley rats were randomized into five groups, the rats in BLM 7d, 14d and 28d groups were treated with a single instillation of 5 mg/kg of BLM, to induced pulmonary fibrosis models. On days 7, 14 and 28, the animals were killed by exsanguination respectively. Normal saline(NS) group were treated by NS, on days 14, the animals were killed by exsanguinations.Control group were untreated. The lung samples were processed for light microscopy and determined the hydroxyproline(HYP) concentration. The expression of PEPT2 mRNA were measured by RT-PCR. PEPT2 cDNA fragments were tested by dideoxy chain termination. Results: Compared with control and NS group, HYP levels increased on day 7 of BLM group, but there was no statistical significant difference(P > 0.05). HYP levels markedly increased on days 14 and 28 of BLM group,there was statistical significant difference(P < 0.01). The morphological study showed that collagenous fiber proliferated on days 14 and 28 of BLM group, especially on day 28, formed pulmonary fibrosis. There were no significant changes of pulmonary PEPT2 mRNA expression at different groups(P > 0.05). Conclusion: The pulmonary fibrosis models of SD rats can be induced by a single instillation of 5 mg/kg of bleomycin on 28d. There were no significant changes of PEPT2 mRNA expression in the lung of rats with bleomycin-induced pulmonary fibrosis.展开更多
Peptide transporter 2(PepT2)transports short peptides from the blood into bovine mammary epithelial cells(BMEC)to stimulate milk protein synthesis.Despite the fact that the effect of PepT2 is acknowledged in BMEC,litt...Peptide transporter 2(PepT2)transports short peptides from the blood into bovine mammary epithelial cells(BMEC)to stimulate milk protein synthesis.Despite the fact that the effect of PepT2 is acknowledged in BMEC,little is known about its regulation.This study was completed to investigate the role of mammalian target of the rapamycin(mTOR)signaling in regulating the expression and function of PepT2 in BMEC.The regulation of PepT2 by mTOR in BMEC was studied in vitro using peptide transport assay,gene silencing,Western blot.The membrane expression of PepT2 and the uptake of b-Ala-Lys-N-7-amino-4-methylcoumarin-3-acetic acid(b-Ala-Lys-AMCA),a model dipeptide,in BMEC were reduced by rapamycin(a mTOR inhibitor)and silencing of either mTOR complex 1(mTORC1)or mTOR complex 2(mTORC2),stimulated by DEP domain-containing mTOR-interacting protein(DEPTOR,endogenous inhibitor of mTORC1 and mTORC2)silencing.The trafficking of PepT2 to the membrane and the uptake of b-Ala-Lys-AMCA was promoted by neuronal precursor cell-expressed developmentally down-regulated 4 isoform 2(Nedd4-2)silencing.The effects of knockdown of mTORC1,but not mTORC2,on cell membrane expression and transport activity of PepT2 was abolished by Nedd4-2 silencing.With immunofluorescence staining,PepT2 was identified to be interacting with Nedd4-2.The Nedd4-2 expression and the interaction between PepT2 and Nedd4-2 was increased through mTORC1 knockdown,indicating an increased ubiquitination of PepT2.The results revealed that mTORC1 can regulate the expression and function of PepT2 through Nedd4-2 in BMEC.展开更多
基金Supported by a grant from the Natural Science Foundation of Yunnan Province(No.2011FZ129)
文摘Objective: Pulmonary fibrosis is a common pathological phenomena in lung cancer patients after chemotherapy or radiotherapy, which is a key factor hindering to transport ion of high concentrated drug to the lung tissue, peptide transporter has become targets of the rational design of peptides and peptide drug. The purpose of the study is to investigate the expression of PEPT2 mRNA in the lung of rats with bleomycin(BLM)-induced pulmonary fibrosis. Methods: Fifty healthy adult Spragne-Dawley rats were randomized into five groups, the rats in BLM 7d, 14d and 28d groups were treated with a single instillation of 5 mg/kg of BLM, to induced pulmonary fibrosis models. On days 7, 14 and 28, the animals were killed by exsanguination respectively. Normal saline(NS) group were treated by NS, on days 14, the animals were killed by exsanguinations.Control group were untreated. The lung samples were processed for light microscopy and determined the hydroxyproline(HYP) concentration. The expression of PEPT2 mRNA were measured by RT-PCR. PEPT2 cDNA fragments were tested by dideoxy chain termination. Results: Compared with control and NS group, HYP levels increased on day 7 of BLM group, but there was no statistical significant difference(P > 0.05). HYP levels markedly increased on days 14 and 28 of BLM group,there was statistical significant difference(P < 0.01). The morphological study showed that collagenous fiber proliferated on days 14 and 28 of BLM group, especially on day 28, formed pulmonary fibrosis. There were no significant changes of pulmonary PEPT2 mRNA expression at different groups(P > 0.05). Conclusion: The pulmonary fibrosis models of SD rats can be induced by a single instillation of 5 mg/kg of bleomycin on 28d. There were no significant changes of PEPT2 mRNA expression in the lung of rats with bleomycin-induced pulmonary fibrosis.
基金the National Natural Science Foundation of China 32072756 and 31872989.
文摘Peptide transporter 2(PepT2)transports short peptides from the blood into bovine mammary epithelial cells(BMEC)to stimulate milk protein synthesis.Despite the fact that the effect of PepT2 is acknowledged in BMEC,little is known about its regulation.This study was completed to investigate the role of mammalian target of the rapamycin(mTOR)signaling in regulating the expression and function of PepT2 in BMEC.The regulation of PepT2 by mTOR in BMEC was studied in vitro using peptide transport assay,gene silencing,Western blot.The membrane expression of PepT2 and the uptake of b-Ala-Lys-N-7-amino-4-methylcoumarin-3-acetic acid(b-Ala-Lys-AMCA),a model dipeptide,in BMEC were reduced by rapamycin(a mTOR inhibitor)and silencing of either mTOR complex 1(mTORC1)or mTOR complex 2(mTORC2),stimulated by DEP domain-containing mTOR-interacting protein(DEPTOR,endogenous inhibitor of mTORC1 and mTORC2)silencing.The trafficking of PepT2 to the membrane and the uptake of b-Ala-Lys-AMCA was promoted by neuronal precursor cell-expressed developmentally down-regulated 4 isoform 2(Nedd4-2)silencing.The effects of knockdown of mTORC1,but not mTORC2,on cell membrane expression and transport activity of PepT2 was abolished by Nedd4-2 silencing.With immunofluorescence staining,PepT2 was identified to be interacting with Nedd4-2.The Nedd4-2 expression and the interaction between PepT2 and Nedd4-2 was increased through mTORC1 knockdown,indicating an increased ubiquitination of PepT2.The results revealed that mTORC1 can regulate the expression and function of PepT2 through Nedd4-2 in BMEC.
