MicroRNA­155­5p inhibits trophoblast cell proliferation and invasion by disrupting centrosomal function.

Recurrent miscarriage is used to refer to more than three pregnancy failures before 20 weeks of gestation. Defective trophoblast cell growth and invasion are frequently observed in recurrent miscarriage. Several microRNAs (miRs), including miR­155­5p, are aberrantly upregulated in recurrent miscarriage; however, the underlying molecular mechanisms remain unclear. The centrosome orchestrates microtubule networks and coordinates cell cycle progression. In addition, it is a base for primary cilia, which are antenna­like organelles that coordinate signaling during development and growth. Thus, deficiencies in centrosomal functions can lead to several disease, such as breast cancer and microcephaly. In the present study, the signaling cascades were analyzed by western blotting, and the centrosome and primary cilia were observed and analyzed by immunofluorescence staining. The results showed that overexpression of miR­155­5p induced centrosome amplification and blocked primary cilia formation in trophoblast cells. Notably, centrosome amplification inhibited trophoblast cell growth by upregulating apoptotic cleaved­caspase 3 and cleaved­poly (ADP­ribose) polymerase in miR­155­5p­overexpressing trophoblast cells. In addition, overexpression of miR­155­5p inhibited primary cilia formation, thereby inhibiting epithelial­mesenchymal transition and trophoblast cell invasion. All phenotypes could be rescued when cells were co­transfected with the miR­155­5p inhibitor, thus supporting the role of miR­155­5p in centrosomal functions. It was also found that miR­155­5p activated autophagy, whereas disruption of autophagy via the depletion of autophagy­related 16­like 1 alleviated miR­155­5p­induced apoptosis and restored trophoblast cell invasion. In conclusion, the present study indicated a novel role of miR­55­5p in mediating centrosomal function in recurrent miscarriage.

Manipulation of osteogenic and adipogenic differentiation of human degenerative disc and ligamentum flavum derived progenitor cells using IL-1ß, IL-19, and IL-20.

PURPOSE: To elucidate whether pro-inflammatory cytokines might influence the commitment of intervertebral disc (IVD)- and ligamentum flavum (LF)-derived progenitor cells toward either osteogenesis or adipogenesis, specifically Interleukin-1ß (IL-1ß), IL-19, and IL-20. METHODS: Sixty patients with degenerative spondylolisthesis and lumbar or lumbosacral spinal stenosis were included in the study. Injuries to the spine, infections, and benign or malignant tumors were excluded. From nine patient samples, IVD- and LF-derived cells were isolated after primary culture, and two clinical samples were excluded due to mycoplasma infection. The effects of IL-1ß, IL-19, as well as IL-20 in regulating osteogenic and adipogenic differentiation in vitro were investigated. RESULTS: Primary IVD- and LF-derived cells were found to have a similar cell morphology and profile of surface markers (CD44, CD90, and CD105) as placenta-derived mesenchymal stem cells (MSCs). Primary IVD/LF cells have a high capacity to differentiate into osteocytes and adipocytes. IL-19 had a tendency to promote adipogenesis. IL-20 inhibited osteogenesis and promoted adipogenesis; IL-1ß promoted osteogenesis but inhibited adipogenesis. CONCLUSION: IL-1ß, IL-19, and IL-20 impact the adipogenic and osteogenic differentiation of IVD-derived and LF-derived cells. Modulating the expression of IL-1ß, IL-19, and IL-20 provides a potential avenue for controlling cell differentiation of IVD- and LF-derived cells, which might have beneficial effect for degenerative spondylolisthesis and spinal stenosis.

Aspirin enhances trophoblast invasion and represses soluble fms-like tyrosine kinase 1 production: a putative mechanism for preventing preeclampsia.

OBJECTIVE: Recent studies suggested that prophylactic aspirin prior to 16 weeks of gestation in high-risk patients may reduce the risk of developing preeclampsia; however, the exact mechanism of aspirin's effect on the pathophysiology of preeclampsia is not clear. This study was designed to investigate the effect of aspirin on trophoblast cell function and its effect on soluble fms-like tyrosine kinase 1 (sFlt-1) production to elucidate the preventive mechanisms for preeclampsia. METHODS AND RESULTS: We used two human trophoblastic cell lines (HTR-8/SVneo and JAR) and freshly isolated cytotrophoblasts from normal and preeclamptic placenta at term to determine the effect of aspirin on trophoblast cell function. Trophoblasts were pretreated with aspirin, and then cell functions and sFlt-1 expression were assessed. Our results showed that aspirin promoted trophoblast invasion not only in HTR-8/SVneo and JAR cells, but also in isolated cytotrophoblasts. sFlt-1 production was repressed by aspirin in a dose-dependent manner. By adding Flt-1 recombinant protein, the trophoblast invasion ability was inhibited in HTR-8/SVneo cells, which was reversed by Flt-1 small interfering ribonucleic acid knockdown. In addition, metalloproteinase 2/9 expression and activity were activated by aspirin but inhibited by sFlt-1. Aspirin also downregulated Akt phosphorylation, and trophoblast invasiveness was facilitated under Akt inhibitor treatment. CONCLUSION: Aspirin enhances cell invasiveness and inhibits sFlt-1 production in trophoblasts. Moreover, sFlt-1 itself also inhibits trophoblast invasion. Our novel findings suggest that the preeclampsia prevention effect of aspirin may be exerted through these two mechanisms.

Carbon monoxide-induced early thrombolysis contributes to heme oxygenase-1-mediated inhibition of neointimal growth after vascular injury in hypercholesterolemic mice.

Arterial thrombosis is a critical event in the pathogenesis of lesion development. In this study, we evaluated the effect of heme oxygenase-1 (HO-1), a stress-inducible enzyme with vasoprotective functions, on arterial thrombosis following vascular mechanical injury. The carotid arteries of apoE-deficient mice were subjected to angioplasty with a modified beaded-needle. Arterial thrombosis occurred at 12 h after injury. Treatment of the injured vessels with an adenovirus bearing HO-1 gene (Adv-HO-1) (1 x 10(8) pfu), but not saline or empty adenovirus (Adv), immediately after angioplasty resulted in earlier thrombolysis and restoration of blood flow detected at 24 h. Immunohistochemistry revealed that the arterial plasminogen activator inhibitor-1 (PAI-1) expression was markedly reduced in Adv-HO-1-treated injured arteries as compared to control counterparts. The thrombolytic effect was also observed by exposing animals with existing arterial thrombosis to carbon monoxide (CO) (250 ppm, 2 h), a byproduct derived from heme degradation by HO-1. In parallel with less fibrin(ogen) deposition, the macrophage infiltration, monocyte chemoattractant protein-1 expression and neointimal formation assessed at 2 weeks after angioplasty were substantially reduced in injured arteries treated with Adv-HO-1. These results support a role of early thrombolysis induced by CO in HO-1-mediated protection against intimal hyperplasia after vascular injury.

Suppressing tumor progression of in vitro prostate cancer cells by emitted psychosomatic power through Zen meditation.

Human prostate cancer PC3 cells were treated in vitro with psychosomatic power emitted by a Buddhist-Zen Master. A significant decrease of growth rate was observed as determined by MTT assay after 48 hours. These cells also had two- to three-fold higher levels of prostatic acid phosphatase (PAcP) activity, a prostate tissue-specific differentiation antigen. In addition, the treated cells formed fewer and smaller colonies in soft agar as compared with control cells, which displayed anchorage-independent growth. These observations provide insight into the suppressive effects of healing power through the practice of Buddhist-Zen meditation on tumor progression. The emitted bioenergy may be suggested as an alternative and feasible approach for cancer research and patient treatment.

Dietary iron restriction increases plaque stability in apolipoprotein-e-deficient mice.

Accumulative evidence has supported the role of iron in the development of atherosclerosis. To test whether iron-mediated oxidative stress influences plaque stability, apoliporotein-E (ApoE)-deficient mice (3 months old) were placed on a chow diet or a low-iron diet for 3 months, and the abundance of interstitial collagen and the expression of the matrix degradation-associated enzyme, matrix metalloproteinase-9 (MMP-9), in vascular lesions were assessed. A low-iron diet appeared to reduce iron deposition while substantially increasing collagen content of lesions in mice. Immunostaining demonstrated lower expression of MMP-9 in lesions of iron-restricted animals. Likewise, SDS-PAGE zymography revealed lower gelatinolytic activities in aortic tissues and sera of the same group of animals. When older ApoE-deficient mice (5 months old) received a low-iron diet for 2 months, development of the lesion area was not significantly affected. However, the lesional collagen content was much higher in the iron-restricted group of animals, and MMP-9 expression in aortic tissues from the same group of mice was significantly lower. Treatment of murine J774 macrophages with increasing concentrations of ferric ammonium citrate significantly enhanced the amount of MMP-9 secreted. Together, these data indicate that decreased vascular iron content following dietary iron restriction in ApoE-deficient mice leads to lower matrix degradation capacity and increased plaque stability.

Induction of heme oxygenase-1 expression in murine macrophages is essential for the anti-inflammatory effect of low dose 15-deoxy-Delta 12,14-prostaglandin J2.

15-Deoxy-Delta 12,14-prostaglandin J2 (15d-PGJ2), a cyclopentenone prostaglandin, displays a potent anti-inflammatory effect at micromolar concentrations (>2 microM) through direct inhibition of nuclear factor (NF)-kappa B activation. Here we show that at submicromolar concentrations (0.1-0.5 microM) 15d-PGJ2 retains the ability to suppress the production of tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) in lipopolysaccharide (LPS)-activated murine J774 macrophages under the conditions of a prolonged incubation (>12 h). Western blot analysis revealed that the expression of the cytoprotective enzyme, heme oxygenase-1 (HO-1), was induced and coincident with the anti-inflammatory action of 15d-PGJ2. Inhibition of HO-1 activity or scavenging carbon monoxide (CO), a byproduct derived from heme degradation, significantly attenuated the suppressive activity of 15d-PGJ2. Furthermore, LPS-induced NF-kappa B activation assessed by the inhibitory protein of NF-kappa B(I kappa B) degradation and p50 nuclear translocation was diminished in cells subjected to prolonged treatment with the low concentration of 15d-PGJ2. Treatment of cells with the protein synthesis inhibitor, cycloheximide, or the specific p38 MAP kinase inhibitor, SB203580, blocked the induction of HO-1 and suppression of LPS-induced I kappa B degradation mediated by 15d-PGJ2. Likewise, HO inhibitor and CO scavenger were effective in abolishing the inhibitory effects of 15d-PGJ2 on NF-kappa B activation induced by LPS. The functional role of CO was further demonstrated by the use of a CO releasing molecule, tricarbonyldichlororuthenium(II) dimer, which significantly suppressed LPS-induced nuclear translocation of p50 as assessed by confocal immunofluorescence. Collectively, these data suggest that even at submicromolar concentrations 15d-PGJ2 can exert an anti-inflammatory effect in macrophages through a mechanism that involves the action of HO/CO.