Analysis of the PAX6-Mediated Suppression Pathways in Gliomas: Downregulation of MMP2 Expression

Glioblastoma multiform (GBM) is the most common and most malignantprimary brain tumor. We have reported that the expression ofPAX6 is decreased in most GBM patient samples when comparedto adjacent normal tissue and anaplastic astrocytomas (AAs).Lower PAX6 expression in tumor tissue is an indication of anunfavorable prognosis for patients with malignant astrocyticgliomas (Zhou et. al. Clin. Can. Res. 9:3369-75). PAX6 encodesa transcription factor critical in the development of the eyeand central nervous system. In contrast to the decreased expressionof PAX6 in GBMs, we observed an increase in the expression ofPAX6 in the GBM cell line U251 after introducing a normal chromosome10. Loss of heterozygosity (LOH) of chromosome 10 occurs inup to 80% of human GBM cases. Therefore, the reduced expressionof PAX6 in GBMs may result from the loss of unidentified tumorsuppression gene(s) in chromosome 10; however, this is not asimple side effect of LOH of chromosome 10. Overexpression ofPAX6 via stable transfection of a PAX6/pRC-CMV construct inthe GBM cell line U251HF suppressed anchorage-independent growthin vivo and tumor formation in vitro. This data is consistentwith the hypothesis that PAX6 may have a tumor-suppressor functionin GBM (Zhou et. al. J. Neuro-Oncology, in print). Microarrayhybridization using Affymetrix chips revealed alterations ofmatrix metalloproteinase-2 (MMP2) gene expression in the suppressedU251HF PAX6-transfected cells. This data was confirmed by real-timequantitative reverse transcription PCR from cells grown in vitroand in the brain of nude mice. Moreover, there is a reversecorrelation between the expression of PAX6 and MMP2 in GBMs,but not in AAs. MMP2 is a well-known protease involved in cellinvasion and the malignant progression of gliomas. MMP2 wasalso shown to be suppressed at a functional level in gelatinzymography assays in PAX6 transfectant cells. This suppressioncorrelates with the near abolition of the ability of all PAX6transfectants to invade in matrigel invasion assays; however,cells expressing a dominant negative mutant form of PAX6 (PAX6-344)displayed a significant enhancement in their ability to invadein this assay. Luciferase MMP2 promoter assays have indicatedthat the lower expression level of MMP2 in PAX6-transfectedcells is due to lower MMP2 promoter activity. In conclusion,our data indicate that PAX6 plays an important role in suppressingthe formation of GBM by suppressing the expression of MMP2.Identification and functional characterization of the cis-elementresponsible for the suppression of the MMP2 promoter activityby PAX6 is being examined

Analysis of the PAX6-Mediated Suppression Pathways in Gliomas: Downregulation of MMP2 Expression

Glioblastoma multiform (GBM) is the most common and most malignantprimary brain tumor. We have reported that the expression ofPAX6 is decreased in most GBM patient samples when comparedto adjacent normal tissue and anaplastic astrocytomas (AAs).Lower PAX6 expression in tumor tissue is an indication of anunfavorable prognosis for patients with malignant astrocyticgliomas (Zhou et. al. Clin. Can. Res. 9:3369-75). PAX6 encodesa transcription factor critical in the development of the eyeand central nervous system. In contrast to the decreased expressionof PAX6 in GBMs, we observed an increase in the expression ofPAX6 in the GBM cell line U251 after introducing a normal chromosome10. Loss of heterozygosity (LOH) of chromosome 10 occurs inup to 80% of human GBM cases. Therefore, the reduced expressionof PAX6 in GBMs may result from the loss of unidentified tumorsuppression gene(s) in chromosome 10; however, this is not asimple side effect of LOH of chromosome 10. Overexpression ofPAX6 via stable transfection of a PAX6/pRC-CMV construct inthe GBM cell line U251HF suppressed anchorage-independent growthin vivo and tumor formation in vitro. This data is consistentwith the hypothesis that PAX6 may have a tumor-suppressor functionin GBM (Zhou et. al. J. Neuro-Oncology, in print). Microarrayhybridization using Affymetrix chips revealed alterations ofmatrix metalloproteinase-2 (MMP2) gene expression in the suppressedU251HF PAX6-transfected cells. This data was confirmed by real-timequantitative reverse transcription PCR from cells grown in vitroand in the brain of nude mice. Moreover, there is a reversecorrelation between the expression of PAX6 and MMP2 in GBMs,but not in AAs. MMP2 is a well-known protease involved in cellinvasion and the malignant progression of gliomas. MMP2 wasalso shown to be suppressed at a functional level in gelatinzymography assays in PAX6 transfectant cells. This suppressioncorrelates with the near abolition of the ability of all PAX6transfectants to invade in matrigel invasion assays; however,cells expressing a dominant negative mutant form of PAX6 (PAX6-344)displayed a significant enhancement in their ability to invadein this assay. Luciferase MMP2 promoter assays have indicatedthat the lower expression level of MMP2 in PAX6-transfectedcells is due to lower MMP2 promoter activity. In conclusion,our data indicate that PAX6 plays an important role in suppressingthe formation of GBM by suppressing the expression of MMP2.Identification and functional characterization of the cis-elementresponsible for the suppression of the MMP2 promoter activityby PAX6 is being examined

Spatial-temporal variation of dissolved N-2 and denitrification in an agricultural river network, southeast China

National Natural Science Foundation of China [41076042, 41376082]; Fundamental Research Funds for the Central Universities [2012121053]The spatio-temporal pattern of excess N-2 production due to denitrification and gaseous nitrogen (N) removal via N-2 emission were investigated for a large agricultural river (the Jiulong River) in southeast China. During 2010-2011, direct measurement of excess dissolved N-2 (Delta N-2, denitrification product) using the N-2:Ar ratio method was carried out along the river network (North River and West River). The results showed that Delta N-2 ranged from 24 to 71 mu mol N-2 L-1 in the West River, and from 3 to 160 mu mol N-2 L-1 in the North River. Areal mean N-2 water-air flux was 8.66 kg N ha(-1) yr(-1) for the whole river network. "Hot spots" of denitrification were found in the upper North River and lower West River, where animal/human wastes dominated the riverine N source. Dissolved inorganic nitrogen (DIN) and dissolved oxygen are the key factors controlling the spatial variation of Delta N-2. Delta N-2 generally peaked in January and during the warm-wet season from May through October, indicating an interactive effect of water temperature and DIN concentration on denitrification. Gaseous N removal fraction (E-d = 22% of [DIN]; annual N removal = 24% of riverine N export) through net denitrification is comparable with other river-estuaries around the world. Gaseous N removal effectiveness generally decreased with N enrichment and was constrained in those hyper-N rich river reaches. Management of N loss should consider the terrestrial and aquatic systems, in order to reduce N export from watershed to coast and mitigate eutrophication in this region. (C) 2014 Elsevier B.V. All rights reserved

Spatial-temporal variation of dissolved N-2 and denitrification in an agricultural river network, southeast China

National Natural Science Foundation of China [41076042, 41376082]; Fundamental Research Funds for the Central Universities [2012121053]The spatio-temporal pattern of excess N-2 production due to denitrification and gaseous nitrogen (N) removal via N-2 emission were investigated for a large agricultural river (the Jiulong River) in southeast China. During 2010-2011, direct measurement of excess dissolved N-2 (Delta N-2, denitrification product) using the N-2:Ar ratio method was carried out along the river network (North River and West River). The results showed that Delta N-2 ranged from 24 to 71 mu mol N-2 L-1 in the West River, and from 3 to 160 mu mol N-2 L-1 in the North River. Areal mean N-2 water-air flux was 8.66 kg N ha(-1) yr(-1) for the whole river network. "Hot spots" of denitrification were found in the upper North River and lower West River, where animal/human wastes dominated the riverine N source. Dissolved inorganic nitrogen (DIN) and dissolved oxygen are the key factors controlling the spatial variation of Delta N-2. Delta N-2 generally peaked in January and during the warm-wet season from May through October, indicating an interactive effect of water temperature and DIN concentration on denitrification. Gaseous N removal fraction (E-d = 22% of [DIN]; annual N removal = 24% of riverine N export) through net denitrification is comparable with other river-estuaries around the world. Gaseous N removal effectiveness generally decreased with N enrichment and was constrained in those hyper-N rich river reaches. Management of N loss should consider the terrestrial and aquatic systems, in order to reduce N export from watershed to coast and mitigate eutrophication in this region. (C) 2014 Elsevier B.V. All rights reserved

Direct measurement of dissolved N2 and denitrification along a subtropical river-estuary gradient, China

The spatial pattern and seasonal variation of denitrification were investigated during 2010-2011 in the Jiulong River Estuary (JRE) in southeast China. Dissolved N2 was directly measured by changes in the N2:Ar ratio. The results showed that excess dissolved N2 ranged from -9.9 to 76.4μmolL-1. Tidal mixing leads to a seaward decline of dissolved gaseous concentrations and water-air fluxes along the river-estuary gradient. Denitrification at freshwater sites varied between seasons, associated with changes in N input and water temperature. The denitrification process was controlled by the nitrate level at freshwater sites, and the excess dissolved N2 observed at the tidal zone largely originated from upstream water transport. Compared to other estuaries, JRE has a relative low gaseous removal efficiency (Ed=12% of [DIN]; annual N removal=24% of DIN load), a fact ascribed to strong tidal mixing, coarse-textured sediment with shallow depth before bedrock and high riverine DIN input. ? 2012 Elsevier Ltd

Direct measurement of dissolved N-2 and denitrification along a subtropical river-estuary gradient, China (vol 66, pg 125, 2013)

The spatial pattern and seasonal variation of denitrification were investigated during 2010-2011 in the Jiulong River Estuary (JRE) in southeast China. Dissolved N₂ was directly measured by changes in the N₂:Ar ratio. The results showed that excess dissolved N₂ ranged from -9.9 to 76.4 μmol L⁻¹. Tidal mixing leads to a seaward decline of dissolved gaseous concentrations and water-air fluxes along the river-estuary gradient. Denitrification at freshwater sites varied between seasons, associated with changes in N input and water temperature. The denitrification process was controlled by the nitrate level at freshwater sites, and the excess dissolved N₂ observed at the tidal zone largely originated from upstream water transport. Compared to other estuaries, JRE has a relative low gaseous removal efficiency (E(d)=12% of [DIN]; annual N removal=24% of DIN load), a fact ascribed to strong tidal mixing, coarse-textured sediment with shallow depth before bedrock and high riverine DIN input

Direct measurement of dissolved N-2 and denitrification along a subtropical river-estuary gradient, China

National Natural Science Foundation of China [41076042, 40810069004]The spatial pattern and seasonal variation of denitrification were investigated during 2010-2011 in the Jiulong River Estuary (JRE) in southeast China. Dissolved N-2 was directly measured by changes in the N-2:Ar ratio. The results showed that excess dissolved N-2 ranged from -9.9 to 76.4 mu mol L-1. Tidal mixing leads to a seaward decline of dissolved gaseous concentrations and water-air fluxes along the river-estuary gradient. Denitrification at freshwater sites varied between seasons, associated with changes in N input and water temperature. The denitrification process was controlled by the nitrate level at freshwater sites, and the excess dissolved N-2 observed at the tidal zone largely originated from upstream water transport. Compared to other estuaries, JRE has a relative low gaseous removal efficiency (E-d = 12% of [DIN]; annual N removal = 24% of DIN load), a fact ascribed to strong tidal mixing, coarse-textured sediment with shallow depth before bedrock and high riverine DIN input. (C) 2012 Elsevier Ltd. All rights reserved

Molecular mechanism of bovine Gasdermin D-mediated pyroptosis

Abstract Pyroptosis is a form of programmed cell death characterized by cell swelling, pore formation in the plasma membrane, lysis, and releases of cytoplasmic contents. To date, the molecular mechanism of human and murine Gasdermin D-mediated pyroptosis have been fully investigated. However, studies focusing on molecular mechanism of bovine Gasdermin D (bGSDMD)-mediated pyroptosis and its function against pathogenic infection were unclear. In the present study, we demonstrate that bovine caspase-1 (bCaspase-1) cleaves bGSDMD at amino acid residue D277 to produce an N-terminal fragment (bGSDMD-p30) which leads to pyroptosis. The amino acid residues T238 and F239 are critical for bGSDMD-p30-mediated pyroptosis. The loop aa 278-299, L293 and A380 are the key sites for autoinhibitory structure of the full length of bGSDMD. In addition, bCaspase-3 also cleaves bGSDMD at residue Asp86 without inducing cell death. Therefore, our study provides the first detailed elucidation of the mechanism of bovine GSDMD-mediated pyroptosis. The results will establish a significant foundation for future research on the role of pyroptosis in bovine infectious diseases

Carbon Materials Containing Single-Atom Co–N₄ Sites Enable Near-Infrared Photooxidation

Near-infrared light occupies 54.3% of the solar spectrum and has greater penetration depth, and its effective utilization is of great significance in the practical application of photocatalysis on a larger scale. However, the development of catalysts that can directly utilize near-infrared light is still a huge challenge. This paper proposes a strategy to directly utilize near-infrared light (excitation wavelength extending to 850 nm) by creating carbon material doped with a high-spin-state Co(II)-Nx single-atom site. In the near-infrared-light-irradiated photooxidation of 1,5-dihydroxynaphthalene, the yield of juglone can reach 45% without a significant decrease, even when the catalytic volume is increased by 20 times, which was much higher than that irradiated by 460 nm wavelength (reduced by about 23%). Our study sets the stage for fabricating stable NIR photocatalysts and provides a solution to directly enhance NIR photooxidation in a large-scale manner