PAK4-relevant proliferation reduced by cell autophagy via p53/mTOR/p-AKT signaling.

Background: P21-activated kinase 4 (PAK4) involves in cell proliferation in cancer and mutually regulates with p53, a molecule is demonstrated to control cell autophagy by mammalian target of rapamycin (mTOR)/protein kinase B (AKT) signaling. Since the signaling exhibits an association with PAK family members in cell autophagy, it implies that PAK4-relevant proliferation may be impacted by autophagy via p53 with a lack of evidence in cancer cells. Methods: In this research, transient and stable PAK4-knockdown human hepatocarcinoma cell lines (HepG2) were constructed by transfection of PAK4-RNA interference (RNAi) plasmid and lentivirus containing PAK4-RNAi plasmid, respectively. We investigated cell proliferation using methyl thiazolyl tetrazolium (MTT) and Cell Counting Kit 8 (CCK8) assays, cell cycle by flow cytometry (FCM) and cell autophagy by monodansylcadaverine (MDC) staining and autophagic biomarker's expression, and detected the expressions of p53, mTOR, phosphorylated-AKT (p-AKT) and AKT by immunofluorescence and western blot to explore the mechanism. Results: We successfully constructed transient and stable PAK4-knockdown HepG2 cell lines, and detected dysfunction of the cells' proliferation. An increased expression of p53, as a molecule of cell-cycle-surveillance on G1/S phase, was demonstrated in the cells although the cell cycle blocked at G2/M. And then, we detected increased autophagosome and autophagic biomarker LC3-II, and decreased expressions in p-AKT and mTOR. Conclusions: The proliferation is reduced in PAK4-knockdown HepG2 cells, which is relative to not only cell cycle arrest but also cell autophagy, and p53/mTOR/p-AKT signaling involves in the cell progress. The findings provide a new mechanism on PAK4 block in cancer therapy.

Expression of E-prostanoid receptors in nasal polyp tissues of smoking and nonsmoking patients with chronic rhinosinusitis.

BACKGROUND: Different inflammatory reactions have been observed in the polyp tissues of nonsmokers and smokers with chronic rhinosinusitis (CRS). E-prostanoid (EP) receptors play a role in the inflammatory processes. Cigarette smoke (CS) exposure regulates EP-receptor expression levels promoting inflammatory mediator release from various inflammatory cells. In this study, we characterize the EP-receptor expression profiles in the polyps of nonsmoking and smoking CRS patients to explore the possible role of CS in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). METHODS: Polyp biopsies were obtained from 28 non-smoking and 21 smoking CRSwNP patients. Histopathological characteristics were observed under a light microscope. The prostaglandin E2 (PGE2), TNF-α, and IL-8 contents in polyp tissues were detected using enzyme-linked immunosorbent assay. Immunostaining was used to locate EP receptors in polyps. Messenger RNA and protein expression of EP receptors were examined using quantitative real-time polymerase chain reaction and Western blot, respectively. RESULTS: More severe inflammatory reactions occurred in polyp tissues of smoking CRSwNP patients. The PGE2, TNF-α, and IL-8 in tissue homogenate levels were significantly higher in smoking CRSwNP patients than those in nonsmoking CRSwNP patients. Moreover, the distribution of each EP receptor subtype was similar in both groups. Compared with the EP-receptor expression in nonsmokers, messenger RNA and protein of EP2 and EP4 receptor were significantly down-expressed in smoking patients, but EP1 and EP3 receptors did not show significant differences. CONCLUSION: CS exposure downregulates the expression levels of EP2 and EP4 receptors and stimulates the production of PGE2 and the proinflammatory cytokine IL-8 and TNF-α in polyp tissues of CRS patients. The down-expressed EP2 and EP4 receptors might be associated with severe inflammatory reactions in smoking CRSwNP patients.

[Changes of heme oxygenase-1 expression in the nigrostriatal system of MPTP-treated SAMP8 mouse].

AIM: To explore the relationship between the expression of hemeoxygenase-1 (HO-1) and the dopaminergic system impairment in MPTP-treated SAMP8 mice. METHODS: 6-month-old male SAMP8 mice received MPTP (20 mg/kg) subcutaneous injection at 2-h intervals for 4 times, and the control group was treated with an equal volume of normal saline. Mice were sacrificed at 6 h, 24 h, 3 d and 8 d after the first injection for the detection of the changes of tyrosine hydroxylase (TH) and HO-1 in the nigrostriatal system by immunohistochemistry and Western blotting. RESULTS: TH-positive neuronal loss was visible at 6 h (14.23%, P<0.05), 24 h (23.85%, P<0.01), 3 d (36.77%, P<0.001), and 8 d (45.90%, P<0.001), and the significant progression of dopaminergic neuronal loss occurred most prominently in the MPTP group from 24 h to 3 d (24 h vs 3 d, P<0.05). There was a significant decrease of striatal TH immunoreactive cells in the MPTP group (P<0.05). Additionally, HO-1 positive cells were detected in striatum just only at 3 d, with the increase of HO-1 protein expression in MPTP groups. Western blot analysis showed no change of HO-1 protein levels in the midbrain after MPTP treatment compared to those of the normal saline group. CONCLUSION: MPTP caused the loss of dopaminergic neuron number and the decrease of TH protein levels in SAMP8 mice. The up-regulation of HO-1 was ephemeral, and its effects related with Parkinson's disease was limited in this study.

Biomechanical characteristics investigation on long-term free graft with expanded porcine skin.

BACKGROUND: Free skin graft has to be used when the large area skin are burned. The objective of this study is to quantify the influence of free graft and expansion on the skin biomechanical remodeling. METHOD: Four white pigs (weighing from 17 kg to 23 kg) were used. Two 180 ml rectangle expanders were aseptically placed beneath the skin on the back of each pig. Four normal skin flaps and four expanded skin flaps were incised from the foreside back of each pig. Two normal skin flaps and two expanded skin flaps were then grafted to rearward back of the pig. Stress-strain relationship, stress relaxation and creep characters in normal skin: N(n=8), expanded skin without graft: E(n=8), normal skin with free graft after three months: NG(n=8) and expanded skin with free graft after three months: EG(n=8) were measured by using Instron material testing machine. FINDINGS: The strains at 3.5 MPa stress (mean (SD)) are 0.4436(0.1760), 0.4851(0.1401), 0.7750(0.1984) and 0.5854(0.0655) respectively in N, E, NG and EG groups. The maximum relaxations (mean (SD)) are 0.6324(0.0169), 0.6279(0.0401), 0.5630(0.0170) and 0.6057(0.0883) in N, E, NG and EG groups, respectively. The maximum creeps (mean (SD)) are 1.0876(0.0086), 1.1037(0.0116), 1.1948(0.0394) and 1.1328(0.0223) in N, E, NG and EG groups. INTERPRETATION: The biomechanical characteristics of free graft with expanded skin after three months have no significant difference with the normal skin, expansion skin and free graft with normal skin after three months (P>0.05). The free graft and expansion have no significant influence on the skin biomechanical remodeling.