Troglitazone upregulates PTEN expression and induces the apoptosis of pulmonary artery smooth muscle cells under hypoxic conditions.

The increased proliferation and decreased apoptosis of pulmonary artery smooth muscle cells (PASMCs) are the main causes of hypoxic pulmonary hypertension. In this study, we investigated the role of troglitazone [peroxisome proliferator-activated receptor γ (PPARγ) agonist] in the regulation of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) expression and the apoptosis of PASMCs under hypoxic conditions. Normal human PASMCs were cultured in growth medium (GM) and treated with troglitazone (0.5-80 µM) under hypoxic conditions (5% CO2+94% N2+1% O2) for 72 h. The gene expression of PTEN, AKT-1 and AKT-2 was determined by quantitative reverse transcription PCR (qRT-PCR). The protein expression level of PTEN, AKT and phosphorylated AKT (p-AKT) was determined by western blot analysis. The apoptosis of PASMCs was determined by measuring the activities of caspase-3, -8 and -9 and by TUNEL assay. The proliferation rate of the PASMCs was altered in a concentration-dependent manner by troglitazone. A significantly reduced proliferation rate was observed at troglitazone concentrations starting from 20 µM under hypoxic conditions (72 ± 5.8%). Although the gene expression levels of PTEN were increased, the gene expression levels of AKT-1 and AKT-2 remained unaltered. Consistent with this, PTEN protein expression was also altered in a concentration-dependent manner by troglitazone. Although AKT expression was unaltered in all the cell samples, reduced p-AKT expression was observed in the troglitazone-treated PASMCs. Troglitazone increased the activities of caspase-3, -8 and -9 in the PASMCs. bpV(HOpic) (PTEN inhibitor) and GW9662 (PPARγ inhibitor) inhibited PTEN protein expression and recovered the proliferation rate of the PASMCs. TUNEL assay demonstrated that troglitazone significantly increased the apoptosis of PASMCs under hypoxic conditions. In conclusion, troglitazone increases PTEN expression under hypoxic conditions in a concentration-dependent manner. Troglitazone increases the apoptosis of PASMCs under hypoxic conditions. The increase in PTEN expression is mediated through the PPARγ signaling pathway.

Cigarette smoke-induced skeletal muscle atrophy is associated with up-regulation of USP-19 via p38 and ERK MAPKs.

Ubiquitin-specific proteases (USPs) deubiquitinate ubiquitin-protein conjugates in the ubiquitin-proteasome system. Previous research shows that ubiquitin-specific protease-19 (USP-19) is up-regulated in mammalian skeletal muscle in some degradative conditions, such as including fasting, diabetes, dexamethasone treatment, and cancer, and its function is associated with muscle atrophy. However, it is still unclear whether USP-19 is involved in muscle atrophy induced by chronic obstructive pulmonary disease. Rats exposed to chronic cigarette smoke and L6 myotubes incubated with cigarette smoke extract (CSE) were studied here. Using western blot analysis and quantitative real-time polymerase chain reaction (qPCR), we observed over-expression of USP-19 and down-regulation of myosin heavy chain (MHC) in both models. Moreover, CSE exposure inhibited myogenic differentiation and myotube formation in L6 myotubes. To explore the mechanism underlying these effects, we investigated the levels of phosphorylated mitogen-activated protein kinases (MAPKs) and total MAPKs. Exposing myotubes to CSE resulted in the general activation of MAPKs such as p38, JNK, and ERK1/2. The ERK inhibitor PD98059 and the p38 inhibitor SB203580 significantly blocked the increase in USP-19 gene expression induced by CSE. Our findings suggest that USP-19 is associated with muscle atrophy in response to cigarette smoke and is a potential therapeutic target. CSE promotes myotube wasting in culture partly by inhibiting myogenic differentiation and acts via p38 and ERK MAPK to stimulate expression of USP-19 in vitro.

[Activation related genes of memory CD(4)(+) T cells in asthma patients].

OBJECTIVE: To screen and identify the genes of activated memory CD(4)(+) T cells in asthma. METHODS: Differential display polymerase chain reaction (DDPCR) was utilized to identify genes of memory CD(4)(+) T cells after activation from asthmatic patients and normal individuals, and the mRNA levels of the differentially expressed genes were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Nineteen differentially expressed cDNA fragments were isolated. The homology analysis indicated that two fragments possessed a high degree of homology with interleukin-7 (IL-7) mRNA and MM-1 mRNA. RT-PCR confirmed that IL-7 mRNA and MM-1 mRNA levels were increased in memory CD(4)(+) T cells after activation in asthmatic patients (0.390 +/- 0.029, 0.629 +/- 0.047, F = 983, 1264 respectively, all P < 0.05). CONCLUSION: The up-regulated expression of IL-7 and MM-1 genes in memory CD(4)(+) T cells after activation may be a molecular mechanism that differentiates the response to allergens of asthma patients from normal individuals.