ATF2 与ATF3 在结直肠癌中表达及临床病理意义

目的:观察结直肠癌组织中活化转录因子2(ATF2)和活化转录因子3(ATF3)的表达并分析其表达的临床病理意义。方法:收集结直肠癌病例,明确其病理诊断并收集临床资料,应用免疫组织化学SP法检测活化转录因子2和活化转录因子3蛋白的表达。结果:ATF2在癌旁肠组织、腺瘤、腺癌组的阳性表达率分别为38%,32%,64%,差异有统计学意义,其中癌旁肠组织、腺瘤分别与腺癌有显著性差异。ATF3在癌旁肠组织、腺瘤、腺癌组的阳性表达率分别为56%,44%,52%,差异无统计学意义。ATF2表达与浸润肠壁深度,淋巴结转移有关,而与肿块大小、部位、分化程度无关。ATF3表达与肿块直径、浸润肠壁深度,而与淋巴结转移、部位、分化程度无关。结论:ATF2与结直肠癌反生和发展有关,而ATF3与结直肠癌的恶性演进有关。     

ATF2与ATF3在结直肠癌中表达及临床病理意义

目的：观察结直肠癌组织中活化转录因子2（ATF2）和活化转录因子3（ATF3）的表达并分析其表达的临床病理意义。方法：收集结直肠癌病例,明确其病理诊断并收集临床资料,应用免疫组织化学SP法检测活化转录因子2和活化转录因子3蛋白的表达。结果：ATF2在癌旁肠组织、腺瘤、腺癌组的阳性表达率分别为38%,32%,64%,差异有统计学意义,其中癌旁肠组织、腺瘤分别与腺癌有显著性差异。ATF3在癌旁肠组织、腺瘤、腺癌组的阳性表达率分别为56%,44%,52%,差异无统计学意义。ATF2表达与浸润肠壁深度,淋巴结转移有关,而与肿块大小、部位、分化程度无关。ATF3表达与肿块直径、浸润肠壁深度,而与淋巴结转移、部位、分化程度无关。结论：ATF2与结直肠癌反生和发展有关,而ATF3与结直肠癌的恶性演进有关。     

Nrf2 activity is lost in the spinal cord and its astrocytes of aged mice

We report herein a study of aging using in vitro and in vivo models. Glial fibrillary acidic protein and ferritin expression levels increased, and the levels of glutamate transporter 1 and transferrin receptor 1 decreased in aging mouse spinal cord and its astrocytes. Mitochondrial transmembrane potential in astrocytes decreased after 60 d of culture. Given the relationship between aging and loss of antioxidant tolerance capacity, we examined the expression of heme oxygenase 1 (HO1) and NAD(P)H/quinone oxidoreductase 1 (NQO1) in the old mouse astrocytes and spinal cord. Indeed, both antioxidant enzymes decreased there. Total nuclear factor E2-related factor 2, which governs basal and inducible expression of HO1 and NQO1, decreased significantly. Significantly, epigallocatechin gallate restored the Nrf2 activity.     

Lipopolysaccharide-induced inhibition of connexin43 gap junction communication in astrocytes is mediated by downregulation of caveolin-3

Astrocytes play a crucial role in maintaining the homeostasis of the brain. Changes to gap junctional intercellular communication (GJIC) in astrocytes and excessive inflammation may trigger brain damage and neurodegenerative diseases. In this study, we investigated the effect of lipopolysaccharide (LPS) on connexin43 (Cx43) gap junctions in rat primary astrocytes. Following LPS treatment, dose- and time-dependent inhibition of Cx43 expression was seen. Moreover, LPS induced a reduction in Cx43 immunoreactivity at cell–cell contacts and significantly inhibited GJIC, as revealed by the fluorescent dye scrape loading assay. Toll-like receptor 4 (TLR4) protein expression was increased 2–3-fold following LPS treatment. To study the pathways underlying these LPS-induced effects, we examined downstream effectors of TLR4 signaling and found that LPS induced a significant increase in phosphorylated extracellular signal-regulated kinase (pERK) levels up to 6 h, followed by signal attenuation and downregulation of caveolin-3 expression. Interestingly, LPS treatment also induced a dramatic increase in inducible nitric oxide synthase (iNOS) levels at 6 h, which were sustained up to 18–24 h. The LPS-induced downregulation of Cx43 and caveolin-3 was prevented by co-treatment of astrocytes with the iNOS cofactor inhibitor 1400W, but not the ERK inhibitor PD98059. Specific knockdown of caveolin-3 using siRNA had a significant inhibitory effect on GJIC and resulted in a downregulation of Cx43. Our results suggest that long-term LPS treatment of astrocytes leads to inhibition of Cx43 gap junction communication by the activation of iNOS and downregulation of caveolin-3 via a TLR4-mediated signaling pathway.     

Glutamate-induced swelling of cultured astrocytes is mediated by metabotropic glutamate receptor

The effects of glutamate and its agonists and antagonists on the swelling of cultured astrocytes were studied. Swelling of astrocytes was measured by [3H]-O-methyl-D-glucose uptake. Glutamate at 0.5, 1 and 10mmol/L and irons-l-aminocyclopentane-1,3-dicarboxylic acid (trans-ACPD), a metabotropic glutamate receptor (mGluR) agonist, at 1 mmol/L caused a significant increase in astrocytic volume, whereas alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA) was not effective. L-2-amino-3-phosphonopropionic acid (L-AP3), an antagonist of mGluR, blocked the astrocytic swelling induced by trans-ACPD or glutamate. In Ca2+-free condition, glutamate was no longer effective. Swelling of astrocytes induced by glutamate was not blocked by CdCl2 at 20 μmol/L, but significantly reduced by CdCl2 at 300 μmol/L and dantrolene at 30 μmol/L. These findings indicate that mGluR activation results in astrocytic swelling and both extracellular calcium and internal calcium stores play important roles in the genes     

Igalan induces detoxifying enzymes mediated by the Nrf2 pathway in HepG2 cells

Igalan is one of the sesquiterpene lactones found in Inula helenium L., which is used as the traditional medicine to treat inflammatory diseases. However, the pharmacological effects of igalan have not been characterized. In this study, we isolated igalan from I. helenium L. and evaluated the effects of igalan on signaling pathways and expression of target genes in HepG2 cells. Igalan activated the nuclear factor erythroid 2‐related factor 2 (Nrf2) pathway by increasing the inactive form of GSK3β, the phosphorylated form of AKT, and the nuclear accumulation of Nrf2. Thus, target genes of Nrf2 such as HO‐1 and NQO1 increased in HepG2 cells. Moreover, igalan inhibited the tumor necrosis factor‐α (TNF‐α)‐induced nuclear factor‐κB activation and suppressed the expression of its target genes, including TNF‐α, interleukin (IL)‐6, and IL‐8 in HepG2 cells. Our results indicate the potential of igalan as an activator of cellular defense mechanisms and a detoxifying agent.     

Protection by pyruvate against glutamate neurotoxicity is mediated by astrocytes through a glutathione-dependent mechanism

Pyruvate, an endogenous metabolite of glycolysis, is an anti-toxicity agent. Recent studies have suggested possible roles for pyruvate in protecting CNS neurons from excitotoxic and metabolic insults. Utilizing cultures derived from embryonic rat cortex, the studies presented in this paper indicate that an astroglia-mediated mechanism is involved in the neuroprotective effects of pyruvate against glutamate toxicity. Glutamate-induced toxicity could be reversed by pyruvate in a mixed culture of cortex cells. Importantly, in pure neuronal cultures from the same tissue, pyruvate failed to protect against glutamate toxicity. Addition of astroglia to the pure neuronal cultures restores the ability of pyruvate to protect neurons from glutamate-induced toxicity. Our results further suggest that pyruvate can induce glia to up-regulate the synthesis of glutathione (GSH), an antioxidant that protects cells from toxins such as free radicals. Taken together, our data suggest that astroglia in mixed cultures are essential for mediating the effects of pyruvate, revealing a novel mechanism by which pyruvate, an important intermediate of tricarboxylic acid cycle in the body, may act to protect neurons from damage during insults such as brain ischemia.     

Up-regulation of aldose reductase expression mediated by phosphatidylinositol 3-kinase/Akt and Nrf2 is involved in the protective effect of curcumin against oxidative damage

Up-regulation of aldose reductase (AR) by reactive oxygen species (ROS) and aldehyde derivatives has been observed in vascular smooth muscle cells. However, the pathophysiological consequences of the induction of AR in vascular tissues are not fully elucidated. Herein we report that an herb-derived polyphenolic compound, curcumin, elicited a dose- and time-dependent increase in AR expression. Inhibition of phosphatidylinositol 3-kinase (PI3K) and p38 mitogen-activated protein kinase (MAPK) significantly suppressed the curcumin-augmented mRNA levels and promoter activity of the AR gene. Luciferase reporter assays indicated that an osmotic response element in the promoter was essential for the responsiveness to curcumin. Curcumin accelerated the nuclear translocation of nuclear factor-erythroid 2-related factor 2 (Nrf2), and overexpression of Nrf2, but not the dominant negative Nrf2, enhanced the promoter activity of the AR gene. Cells preincubated with curcumin demonstrated resistance to ROS-induced apoptotic death. These effects were significantly attenuated in the presence of AR inhibitors or small interfering RNAs, indicating a protective role for AR against ROS-induced cell damage. Taken together, the activation of PI3K and p38 MAPK by curcumin augmented the expression of the AR gene via Nrf2, and increased AR activity may be an important cellular response against oxidative stress.     

Genotoxic stress-induced activation of Plk3 is partly mediated by Chk2

Polo-like kinase 3 (Plk3, alternatively termed Prk) is involved in the regulation of DNA damage checkpoint as well as in M-phase function. Plk3 physically interacts with p53 and phosphorylates this tumor suppressor protein on serine-20, suggesting that the role of Plk3 in cell cycle progression is mediated, at least in part, through direct regulation of p53. Here we show that Plk3 is rapidly activated by reactive oxygen species in normal diploid fibroblast cells (WI-38), correlating with a subsequent increase in p53 protein level. Plk3 physically interacts with Chk2 and the interaction is enhanced upon DNA damage. In addition, Chk2 immunoprecipitated from cell lysates of Daudi (which expressed little Plk3) is capable of stimulating the kinase activity of purified recombinant Plk3 in vitro, and this stimulation is more pronounced when Plk3 is supplemented with Chk2 immunoprecipitated from Daudi after DNA damage. Furthermore, ectopic expression Chk2 activates cellular Plk3. Together, our studies suggest Chk2 may mediate direct activation of Plk3 in response to genotoxic stresses.     

Expression of the 2',3'-cAMP-adenosine pathway in astrocytes and microglia

Many organs express the extracellular 3',5'-cAMP-adenosine pathway (conversion of extracellular 3',5'-cAMP to 5'-AMP and 5'-AMP to adenosine). Some organs release 2',3'-cAMP (isomer of 3',5'-cAMP) and convert extracellular 2',3'-cAMP to 2'- and 3'-AMP and convert these AMPs to adenosine (extracellular 2',3'-cAMP-adenosine pathway). As astrocytes and microglia are important participants in the response to brain injury and adenosine is an endogenous neuroprotectant, we investigated whether these extracellular cAMP-adenosine pathways exist in these cell types. 2',3'-, 3',5'-cAMP, 5'-, 3'-, and 2'-AMP were incubated with mouse primary astrocytes or primary microglia for 1 h and purine metabolites were measured in the medium by mass spectrometry. There was little evidence of a 3',5'-cAMP-adenosine pathway in either astrocytes or microglia. In contrast, both cell types converted 2',3'-cAMP to 2'- and 3'-AMP (with 2'-AMP being the predominant product). Although both cell types converted 2'- and 3'-AMP to adenosine, microglia were five- and sevenfold, respectively, more efficient than astrocytes in this regard. Inhibitor studies indicated that the conversion of 2',3'-cAMP to 2'-AMP was mediated by a different ecto-enzyme than that involved in the metabolism of 2',3'-cAMP to 3'-AMP and that although CD73 mediates the conversion of 5'-AMP to adenosine, an alternative ecto-enzyme metabolizes 2'- or 3'-AMP to adenosine.