The exact molecular mechanism that mediates hypoxia-induced pulmonary fibrosis needs to be further clarified. Ang II increased NF-B-p50 protein manifestation in HLF-1 cells, and the special effect could be inhibited by telmisartan (TST), an AT1R inhibitor, and partially inhibited by PD123319, an AT2R inhibitor. Importantly, hypoxia-induced NF-B nuclear translocation could be nearly completely inhibited by an AT1R or AT2R inhibitor. Furthermore pyrrolidine dithiocarbamate (PDTC), a NF-B blocker, abolished the manifestation of hypoxia-induced AT1R and Col-I in HLF-1 Rabbit polyclonal to NOTCH4 cells. Our results indicate that Ang II-mediated NF-B signalling via ATR is usually involved in hypoxia-induced collagen synthesis in human lung fibroblasts. = 3). * 0.05 0 or 6 h following hypoxia … 2.3. Hypoxia Induced both AT1R and AT2R mRNA Manifestation Both 129101-54-8 AT1R and AT2R are involved in promoting lung fibrosis via different mechanisms of action [7,12,18]; therefore, the effects of hypoxia on the manifestation of AT1R and AT2R were also discovered in this 129101-54-8 study. These results showed that AT1R and AT2R exhibited comparable responses to hypoxia in HLF-1 cells. The mRNA manifestation of both AT1R and AT2R increased 6 h after the hypoxic treatment, and the levels were further increased at 12 and 24 h. Furthermore, AT1R mRNA manifestation was increased to a greater extent than AT2R manifestation in HLF-1 cells during the same study period (Physique 3A,W). Physique 3. The effects of hypoxia on the AT1R and AT2R mRNA manifestation levels in HLF-1 cells. HLF-1 cells 129101-54-8 were treated with hypoxic conditions for up to 24 h. (A) AT1R mRNA manifestation was assessed using RT-PCR. The data are presented as means SD (= 3). … 2.4. Hypoxia-Induced Col-I mRNA and Protein Manifestation via Ang II/ATR Signalling Ang II has been identified as a profibrotic factor in vascular fibrosis ; therefore, we investigated whether Ang II was involved in hypoxia-induced Col-I manifestation in lung fibroblasts. Firstly, the Col-I mRNA and protein manifestation levels were assessed in HLF-1 cells after Ang II treatment under normoxic conditions. We observed that Col-I manifestation increased in a time-dependent manner, and the values were significantly higher 24 h after the Ang II treatment (Physique 4ACC). Subsequently, selective Ang II receptor antagonists (telmisartan [TST] for AT1R and PD123319 for AT2R) were used to elucidate the functions of AT1R and AT2R in Ang II-induced 129101-54-8 collagen production under normoxic conditions. We found that both TST and PD123319 inhibited the Ang II-mediated protein manifestation of Col-I in HLF-1 cells under normoxic conditions (Physique 4DCF). As shown in Figures 1 and ?and2,2, hypoxia not only increased the Col-I manifestation, but it also induced Ang II synthesis in HLF-1 cells. Further studies were performed to evaluate whether exogenous Ang II promotes Col-I synthesis during hypoxia. TST and PD123319 were also used to elucidate the functions of AT1R and AT2R in exogenous Ang II-induced collagen production under hypoxic conditions. We found that both TST and PD123319 obviously inhibited the protein manifestation of Col-I in HLF-1 cells incubated with Ang II under hypoxic conditions (Physique 4GCI), and PD123319 inhibited 129101-54-8 collagen manifestation to a greater extent than TST (Physique 4I). Taken together, these results suggest that the ATR could play an important role in hypoxia-induced Col-I protein manifestation. Physique 4. The role of the ATR in Ang II-induced Col-I manifestation under normoxia and hypoxia. (A) A Masson staining assay was used to observe the changes in the total collagen content in HLF-1 cells after treatment with Ang II (1.0 M) for 0, 6, 12 or 24 … 2.5. Hypoxia-Induced NF-B Manifestation Involved in the Angiotensin System The NF-B transcription factor family is usually involved in controlling multiple aspects of homeostasis, including the functional inflammatory system, immune responses, the cell cycle and cell death in response to various cellular tensions, such as Ang II . Therefore, we investigated whether Ang II is usually involved in.