Prokineticin 2 is a catabolic regulator of osteoarthritic cartilage destruction in mouse.

BACKGROUND: Our preliminary study indicates that the multi-functional protein, prokineticin 2 (Prok2), is upregulated in osteoarthritic (OA) chondrocytes as a target of the hypoxia-inducible factor (HIF)-2α. This study aims to elucidate the potential roles of Prok2 in OA. METHODS: Prok2 expression was assessed through microarray analysis in chondrocytes and confirmed via immunostaining in OA cartilage. Experimental OA was induced through destabilization of the medial meniscus (DMM). Functions of Prok2 were assessed by adenoviral overexpression, intra-articular (IA) injection of recombinant Prok2 (rProk2), and knockdown of Prok2 in joint tissues. We also explored the potential utility of Prok2 as an OA biomarker using enzyme-linked immunosorbent assay (ELISA). RESULTS: HIF-2α upregulated Prok2, one of the prokineticin signaling components, in OA chondrocytes of mice and humans. Adenoviral overexpression of Prok2 in chondrocytes and cartilage explants, as well as the application of rProk2, led to an upregulation of matrix metalloproteinase (MMP)3 and MMP13. Consistently, the overexpression of Prok2 in joint tissues or IA injection of rProk2 exacerbated cartilage destruction and hindpaw mechanical allodynia induced by DMM. However, the knockdown of Prok2 in joint tissues did not significantly affect DMM-induced cartilage destruction. Additionally, despite being a secreted protein, the serum levels of Prok2 in OA mice and human OA patients were found to be below the range detected by ELISA. CONCLUSION: The upregulation of Prok2 exacerbates OA cartilage destruction and hindpaw mechanical allodynia. However, its knockdown is not sufficient to inhibit experimental OA and Prok2 is not a potential candidate serum biomarker of OA.

Suppression of HIF-1α accumulation by betulinic acid through proteasome activation in hypoxic cervical cancer.

Betulinic acid (BA) exhibits various biological activities such as anti-bacterial, anti-inflammatory, anti-human papilloma virus (HPV), and anti-cancer activities. HPV infection is associated with a high risk of cervical cancer, which is the leading cause of deaths among women worldwide. Therefore, BA is an attractive therapeutic agent for treating cervical cancer. In this study, we investigated the role of BA in regulating the hypoxia-mediated response in HeLa cells and clarified the underlying mechanism of action. We found that BA inhibited the hypoxia-induced accumulation of HIF-1α without affecting HIF-1α mRNA levels and suppressed the expression of HIF target genes, including VEGF, GLUT1, and PDK1 in HeLa cells. Additionally, BA enhanced the ß1, ß2, and ß5 activities of the proteasome, which resulted in reduced levels of ubiquitinated proteins and HIF-1α protein in HeLa cells. However, BA treatment did not affect the deubiquitinase enzyme activity in HeLa cells. These results indicate that inhibition of HIF-1α accumulation by BA is mediated by activation of the proteasome, and BA is a potential anticancer agent for the regulation of the HIF signaling pathway.

Apigenin alleviates TGF-ß1-induced nasal mucosa remodeling by inhibiting MAPK / NF-kB signaling pathways in chronic rhinosinusitis.

BACKGROUND: Chronic rhinosinusitis is involved in tissue remodeling of nasal mucosa such as nasal myofibroblast differentiation and extracellular matrix production. Apigenin (4',5,7-trihydroxyflavone) is a bioflavonoid compound and has anti-tissue remodeling characteristics. The aims of this study were to evaluate the effect of apigenin on TGF-ß1-induced myofibroblast differentiation and extracellular matrix accumulation and to determine the underlying mechanism. METHODS: Nasal fibroblasts and ex vivo nasal inferior turbinate tissues were stimulated with TGF-ß1 with or without apigenin. The expression levels of α-SMA, fibronectin and collagen type I were determined by real-time PCR, western blot and immunocytochemical staining. Mitogen-activated protein kinase (MAPK) phosphorylation induced by TGF-ß1 were determined by western blot analysis. The transcriptional activity of NF-κB was measured by luciferase assay. Migration effects of fibroblasts were evaluated by wound scratch and transwell migration assay. Contractile activity was determined by collagen gel contraction assay. RESULTS: The expression levels of α-SMA, fibronectin, and collagen type I significantly increased in TGF-ß1-stimulated nasal fibroblasts. In TGF-ß1-stimulated nasal fibroblasts, apigenin inhibited the expressions of α-SMA, fibronectin, and collagen type I. Inhibitors of MAPK (p-38, JNK) and NF-κB blocked the expression of α-SMA, fibronectin and collagen type I. Apigenin suppressed the activation of MAPK (p-38, JNK) and NF-κB induced by TGF-ß1 treatment. Apigenin also inhibited the functional activity of fibroblasts by reducing the migration and collagen contractile activities. CONCLUSIONS: These results suggests the possible use of apigenin as a chronic rhinosinusitis therapeutic agent which can suppress tissue remodeling in nasal mucosa.

Cigarette smoke extract increases vascular endothelial growth factor production via TLR4/ROS/MAPKs/NF-kappaB pathway in nasal fibroblast.

PURPOSE: Cigarette smoke is a complex mixture of various chemical compounds, including free radicals and highly toxic compounds. Cigarette smoke exposure has been shown to be associated with chronic rhinosinusitis and tissue remodeling in upper airway. Vascular endothelial growth factor (VEGF) is one of the cytokines with a crucial role in tissue remodeling of airway. The aims of this study were to determine the effects of cigarette smoke extract (CSE) on VEGF expression and to investigate the underlying molecular mechanisms of CSE in nasal fibroblasts. METHODS: Nasal fibroblasts were stimulated with CSE. Cytotoxicity was evaluated by 3-(4,5- dimethylthiazol-2yl)-2,5-diphenyl-tetrazolium bromide assay. The expression level of VEGF was measured using reverse transcription-polymerase chain reaction (RT-PCR), and enzyme-linked immunosorbent assay. Messenger RNA (mRNA) expression level of TLR4 were determined by RT-PCR. Small interfering RNA (siRNA) for TLR4 was transfected to suppress TLR4 expression. Activation of reactive oxygen species (ROS) was analyzed by using dichloro-dihydro-fluorescein diacetate assay. Mitogen-activated protein kinase (MAPK) and NF-kappaB activations were determined by using western blot and/or luciferase assay. RESULTS: CSE had no significant cytotoxic effect in nasal fibroblast up to 5%. CSE significantly increased both VEGF mRNA and protein expression dose-dependently. The down-regulation of TLR4 transcription by siRNA treatment suppressed CSE-induced expressions of both TLR4 and VEGF. Pretreatment with ROS scavengers, specific inhibitors of each MAPK, and NF-kappaB inhibitor significantly decreased CSE-induced VEGF expression. CONCLUSIONS: CSE has a stimulatory effect on VEGF expression through the TLR4, ROS, MAPK, and NF-kappaB signaling pathway in nasal fibroblasts.