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Biomedical subjects

Michael Karin

Publications and source records attributed to Michael Karin.

At least 73 records · Page 4Linked to original sources
PubMed2004

The human CYP1A1 gene is regulated in a developmental and tissue-specific fashion in transgenic mice.

Regulation and expression of human CYP1A1 is demonstrated in transgenic mice. We have developed two transgenic mouse lines. One mouse strain (CYPLucR) carries a functional human CYP1A1 promoter (-1612 to +293)-luciferase reporter gene, and the other strain (CYP1A1N) expresses CYP1A1 under control of the full-length human CYP1A1 gene and 9 kb of flanking regulatory DNA. With CYPLucR(+/-) mice, 2,3,7,8-tetrachlordibenzo-p-dioxin (TCDD) and several other aryl hydrocarbon receptor ligands induced hepatocyte-specific luciferase activity. When other tissues were examined, TCDD induced luciferase activity in brain with limited induction in lung and no detectable luciferase activity in kidney. Treatment of CYP1A1N(+/-) mice with TCDD resulted in induction of human CYP1A1 in liver and lung, while mouse Cyp1a1 was induced in liver, lung, and kidney. Although induced CYP1A1/Cyp1a1 could not be detected by Western blot analysis in brains from CYP1A1N(+/-) mice, induction in brain was verified by detection of CYP1A1/Cyp1a1 RNA. The administration of TCDD to nursing mothers to examine the effect of lactational exposure via milk demonstrated prominent induction of luciferase activity in livers of CYPLucR(+/-) newborn pups with limited induction in brain. However, TCDD treatment of adult CYPLucR(+/-) mice led to a 7-10-fold induction of brain luciferase activity. Combined these results indicate that tissue-specific and developmental factors are controlling aryl hydrocarbon receptor-mediated induction of human CYP1A1.

Animals↗
PubMed2004

The protein kinase PKR is required for macrophage apoptosis after activation of Toll-like receptor 4.

Macrophages are pivotal constituents of the innate immune system, vital for recognition and elimination of microbial pathogens. Macrophages use Toll-like receptors (TLRs) to detect pathogen-associated molecular patterns--including bacterial cell wall components, such as lipopolysaccharide or lipoteichoic acid, and viral nucleic acids, such as double-stranded (ds)RNA--and in turn activate effector functions, including anti-apoptotic signalling pathways. Certain pathogens, however, such as Salmonella spp., Shigellae spp. and Yersiniae spp., use specialized virulence factors to overcome these protective responses and induce macrophage apoptosis. We found that the anthrax bacterium, Bacillus anthracis, selectively induces apoptosis of activated macrophages through its lethal toxin, which prevents activation of the anti-apoptotic p38 mitogen-activated protein kinase. We now demonstrate that macrophage apoptosis by three different bacterial pathogens depends on activation of TLR4. Dissection of anti- and pro-apoptotic signalling events triggered by TLR4 identified the dsRNA responsive protein kinase PKR as a critical mediator of pathogen-induced macrophage apoptosis. The pro-apoptotic actions of PKR are mediated both through inhibition of protein synthesis and activation of interferon response factor 3.

Animals↗
PubMed2004

Targeting of TAK1 by the NF-kappa B protein Relish regulates the JNK-mediated immune response in Drosophila.

The molecular circuitry underlying innate immunity is constructed of multiple, evolutionarily conserved signaling modules with distinct regulatory targets. The MAP kinases and the IKK-NF-kappa B molecules play important roles in the initiation of immune effector responses. We have found that the Drosophila NF-kappa B protein Relish plays a crucial role in limiting the duration of JNK activation and output in response to Gram-negative infections. Relish activation is linked to proteasomal degradation of TAK1, the upstream MAP kinase kinase kinase required for JNK activation. Degradation of TAK1 leads to a rapid termination of JNK signaling, resulting in a transient JNK-dependent response that precedes the sustained induction of Relish-dependent innate immune loci. Because the IKK-NF-kappa B module also negatively regulates JNK activation in mammals, thereby controlling inflammation-induced apoptosis, the regulatory cross-talk between the JNK and NF-kappa B pathways appears to be broadly conserved.

Animals↗
PubMed2004

IkappaB-kinasebeta-dependent NF-kappaB activation provides radioprotection to the intestinal epithelium.

Acute injury to the intestinal mucosa is a major dose-limiting complication of abdominal radiation therapy. We studied the role of the transcription factor NF-kappaB in protection against radiation-induced apoptosis in the intestinal epithelium in vivo. We use mice in which NF-kappaB signaling through IkappaB-kinase (IKK)-beta is selectively ablated in intestinal epithelial cells to show that failure to activate epithelial cell NF-kappaB in vivo results in a significant increase in radiation-induced epithelial cell apoptosis. Furthermore, bacterial lipopolysaccharide, which is normally a radioprotective agent, is radiosensitizing in IKKbeta-deficient intestinal epithelial cells. Increased apoptosis in IKKbeta-deficient intestinal epithelial cells was accompanied by increased expression and activation of the tumor suppressor p53 and decreased expression of antiapoptotic Bcl-2 family proteins. These results demonstrate the physiological importance of the NF-kappaB system in protection against radiation-induced death in the intestinal epithelium in vivo and identify IKKbeta as a key molecular target for radioprotection in the intestine. Selective preactivation of NF-kappaB through IKKbeta in intestinal epithelial cells could provide a therapeutic modality that allows higher doses of radiation to be tolerated during cancer radiotherapy.

Animals↗
PubMed2004

Inhibition of NF-kappaB in cancer cells converts inflammation- induced tumor growth mediated by TNFalpha to TRAIL-mediated tumor regression.

We used an experimental murine cancer metastasis model in which a colon adenocarcinoma cell line generates lung metastases, whose growth is stimulated in response to injection of bacterial lipopolysaccharide (LPS), to investigate the role of NF-kappaB in inflammation-induced tumor growth. We found that LPS-induced metastatic growth response in this model depends on both TNFalpha production by host hematopoietic cells and NF-kappaB activation in tumor cells. Inhibition of NF-kappaB in both colon and mammary carcinoma cells converts the LPS-induced growth response to LPS-induced tumor regression. The latter response is TNFalpha-independent, but depends on another member of the TNF superfamily, TRAIL, whose receptor is induced in NF-kappaB-deficient cancer cells.

Adenocarcinoma↗
PubMed2004

CpG oligodeoxynucleotides induce expression of proinflammatory cytokines and chemokines in astrocytes: the role of c-Jun N-terminal kinase in CpG ODN-mediated NF-kappaB activation.

Bacterial DNA and synthetic oligodeoxynucleotides (ODN) containing unmethylated CpG motifs stimulate the cells of the innate immune system through a specific receptor called Toll-like receptor-9 (TLR9). It was reported that CpG ODN stimulation induces activation of astrocytes and microglia. However, the precise intracellular signaling pathways that lead to this glial cell activation have not been clearly elucidated. In this study, we found that CpG ODN induce mRNA expression of adhesion molecules and matrix metalloproteinase-9 (MMP-9), as well as proinflammatory cytokines and chemokines, in mouse astrocytes. CpG ODN stimulation in astrocytes induces the activation of IkappaB kinase (IKK) and c-Jun N-terminal kinase (JNK), whereas it inhibits the constitutive ERK1/2 activation. The abrogation of JNK activity using a pharmacological inhibitor showed that JNK activation is essential for the induction of cytokine and chemokine gene expression. This effect of JNK does not require the phosphorylation of c-Jun; rather, it works via the potentiation of NF-kappaB signaling.

Animals↗
PubMed2004

Genetic dissection of antigen receptor induced-NF-kappaB activation.

NF-kappaB is a master transcription factor in the immune system. It regulates various aspects of immune cell development, homeostasis, survival and function. It is in these contexts that the signaling pathways leading to NF-kappaB activation are best understood. This review addresses the cell signaling from antigen receptors as well as Toll-like receptors to NF-kappaB in both innate and adaptive immunity as revealed by the analysis of mice lacking specific signaling intermediates.

Animals↗
PubMed2004

The control of cell motility and epithelial morphogenesis by Jun kinases.

Originally identified as stress-activated protein kinases that control cell survival and proliferation through transcription factor c-Jun, the Jun N-terminal kinase (JNK) subgroup of MAP kinases (MAPKs) have recently emerged as crucial regulators of cell migration and the morphogenetic movement of epithelial sheets. In Drosophila, a well-orchestrated JNK signaling pathway controls formation of actin stress fibers and cell shape changes, which are required for the sealing of embryonic epidermis in a process known as dorsal closure. The JNK pathway is also involved in morphogenetic processes in mice including closure of the eyelid, neural tube and optic fissure. This article focuses on recent advances in understanding the role of JNK pathway in the regulation of cell migration, cytoskeleton rearrangement and the morphogenesis of epithelial sheets.

Animals↗
PubMed2004

A dual role for Ikk alpha in tooth development.

IKK alpha is a component of the I kappa B kinase (IKK) complex that plays a key role in the activation of NF-kappa B. In Ikk alpha mutant mice and mice expressing a transdominant negative mutant of I kappa B alpha (cI kappa B alpha Delta N), molars have abnormal cusps, indicating that Ikk alpha is involved in cusp formation through the NF-kappa B pathway. However, Ikk alpha mutant incisors also have an earlier phenotype where epithelium evaginates outward into the developing oral cavity rather than invaginating into the underlying mesenchyme. A similar evagination of epithelium was also observed in whisker development, suggesting that Ikk alpha contributes to the direction of epithelial growth during the early stages of development in many ectodermal appendages. Since cI kappa B alpha Delta N mice have normal incisor epithelial invagination, Ikk alpha's role appears to be NF-kappa B independent. Changes in Notch1, Notch2, Wnt7b, and Shh expression found in incisor epithelium of Ikk alpha mutants suggest that this NF-kappa B-independent function is mediated by Notch/Wnt/Shh signaling pathways.

Animals↗
PubMed2004

Essential roles of receptor-interacting protein and TRAF2 in oxidative stress-induced cell death.

Oxidative stress and reactive oxygen species (ROS) can elicit and modulate various physiological and pathological processes, including cell death. However, the mechanisms controlling ROS-induced cell death are largely unknown. Data from this study suggest that receptor-interacting protein (RIP) and tumor necrosis factor receptor (TNFR)-associated factor 2 (TRAF2), two key effector molecules of TNF signaling, are essential for ROS-induced cell death. We found that RIP(-/-) or TRAF2(-/-) mouse embryonic fibroblasts (MEF) are resistant to ROS-induced cell death when compared to wild-type cells, and reconstitution of RIP and TRAF2 gene expression in their respective deficient MEF cells restored their sensitivity to H(2)O(2)-induced cell death. We also found that RIP and TRAF2 form a complex upon H(2)O(2) exposure, but without the participation of TNFR1. The colocalization of RIP with a membrane lipid raft marker revealed a possible role of lipid rafts in the transduction of cell death signal initiated by H(2)O(2). Finally, our results demonstrate that activation of c-Jun NH(2)-terminal kinase 1 is a critical event downstream of RIP and TRAF2 in mediating ROS-induced cell death. Therefore, our study uncovers a novel signaling pathway regulating oxidative stress-induced cell death.

Animals↗
PubMed2003

Inhibitor of nuclear factor kappaB kinase deficiency enhances oxidative stress and prolongs c-Jun NH2-terminal kinase activation induced by arsenic.

Stress signals activate both inhibitor of nuclear factor-kappaB kinase (IKKbeta) and c-Jun NH(2)-terminal kinase (JNK). It was shown recently that IKK-dependent nuclear factor kappaB activation results in attenuation of tumor necrosis factor alpha-induced JNK activation. How that negative cross-talk between nuclear factor kappaB and JNK occurs is not well-understood. By using wild-type and Ikkbeta gene knockout (Ikkbeta(-/-)) mouse embryo fibroblasts, we found that IKKbeta deficiency results in prolongation of arsenic-induced JNK activation, which was not due to the decreased expression of GADD45beta or X-linked Inhibitor of Apoptosis (XIAP), as suggested previously for RelA(-/-) cells treated with tumor necrosis factor alpha. This enhanced JNK activation was largely associated with an oxidative stress response as indicated by elevated expression of heme oxygenase-1 and the accumulation of H(2)O(2) in Ikkbeta(-/-) cells. Expression profiling experiments revealed an increased expression of p450 family CYP1B1 mRNA in Ikkbeta(-/-) cells compared with wild-type cells. Inhibition of CYP1B1 reduced both oxidative stress and arsenic-stimulated JNK activation. Thus, increased CYP1B1 expression is central to and seems to be responsible for sensitizing Ikkbeta(-/-) cells to stress-induced JNK activation.

Animals↗
PubMed2003

A role for MEK kinase 1 in TGF-beta/activin-induced epithelium movement and embryonic eyelid closure.

MEKK1-deficient mice show an eye open at birth phenotype caused by impairment in embryonic eyelid closure. MEK kinase 1 (MEKK1) is highly expressed in the growing tip of the eyelid epithelium, which displays loose cell-cell contacts and prominent F-actin fibers in wild-type mice, but compact cell contacts, lack of polymerized actin and a concomitant impairment in c-Jun N-terminal phosphorylation in MEKK1-deficient mice. In cultured keratinocytes, MEKK1 is essential for JNK activation by TGF-beta and activin, but not by TGF-alpha. MEKK1-driven JNK activation is required for actin stress fiber formation, c-Jun phosphorylation and cell migration. However, MEKK1 ablation does not impair other TGF-beta/activin functions, such as nuclear translocation of Smad4. These results establish a specific role for the MEKK1-JNK cascade in transmission of TGF-beta and activin signals that control epithelial cell movement, providing the mechanistic basis for the regulation of eyelid closure by MEKK1. This study also suggests that the signaling mechanisms that control eyelid closure in mammals and dorsal closure in Drosophila are evolutionarily conserved.

Actins↗
PubMed2003

Evidence that reactive oxygen species do not mediate NF-kappaB activation.

It has been postulated that reactive oxygen species (ROS) may act as second messengers leading to nuclear factor (NF)-kappaB activation. This hypothesis is mainly based on the findings that N-acetyl-L-cysteine (NAC) and pyrrolidine dithiocarbamate (PDTC), compounds recognized as potential antioxidants, can inhibit NF-kappaB activation in a wide variety of cell types. Here we reveal that both NAC and PDTC inhibit NF-kappaB activation independently of antioxidative function. NAC selectively blocks tumor necrosis factor (TNF)-induced signaling by lowering the affinity of receptor to TNF. PDTC inhibits the IkappaB-ubiquitin ligase activity in the cell-free system where extracellular stimuli-regulated ROS production does not occur. Furthermore, we present evidence that endogenous ROS produced through Rac/NADPH oxidase do not mediate NF-kappaB signaling, but instead lower the magnitude of its activation.

Acetylcysteine↗
PubMed2003

IKK beta is required for peripheral B cell survival and proliferation.

NF-kappaB activity in mammalian cells is regulated through the IkappaB kinase (IKK) complex, consisting of two catalytic subunits (IKKalpha and IKKbeta) and a regulatory subunit (IKKgamma). Targeted deletion of Ikkbeta results in early embryonic lethality, thus complicating the examination of IKKbeta function in adult tissues. Here we describe the role of IKKbeta in B lymphocytes made possible by generation of a mouse strain that expresses a conditional Ikkbeta allele. We find that the loss of IKKbeta results in a dramatic reduction in all peripheral B cell subsets due to associated defects in cell survival. IKKbeta-deficient B cells are also impaired in mitogenic responses to LPS, anti-CD40, and anti-IgM, indicating a general defect in the ability to activate the canonical NF-kappaB signaling pathway. These findings are consistent with a failure to mount effective Ab responses to T cell-dependent and independent Ags. Thus, IKKbeta provides a requisite role in B cell activation and maintenance and thus is a key determinant of humoral immunity.

Animals↗
PubMed2003

The two faces of IKK and NF-kappaB inhibition: prevention of systemic inflammation but increased local injury following intestinal ischemia-reperfusion.

We studied the role of NF-kappaB in acute inflammation caused by gut ischemia-reperfusion through selective ablation of IkappaB kinase (IKK)-beta, the catalytic subunit of IKK that is essential for NF-kappaB activation. Ablation of IKK-beta in enterocytes prevented the systemic inflammatory response, which culminates in multiple organ dysfunction syndrome (MODS) that is normally triggered by gut ischemia-reperfusion. IKK-beta removal from enterocytes, however, also resulted in severe apoptotic damage to the reperfused intestinal mucosa. These results show the dual function of the NF-kappaB system, which is responsible for both tissue protection and systemic inflammation, and underscore the caution that should be exerted in using NF-kappaB and IKK inhibitors.

Animals↗
PubMed2003

The role of the Ah receptor and p38 in benzo[a]pyrene-7,8-dihydrodiol and benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide-induced apoptosis.

Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous contaminants in the environment. Benzo[a]pyrene (B[a]P), a prototypical member of this class of chemicals, affects cellular signal transduction pathways and induces apoptosis. In this study, the proximate carcinogen of B[a]P metabolism, trans-7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-7,8-dihydrodiol) and the ultimate carcinogen, B[a]P-r-7,t-8-dihydrodiol-t-9,10-epoxide(+/-) (BPDE-2) were found to induce apoptosis in human HepG2 cells. Apoptosis initiated by B[a]P-7,8-dihydrodiol was linked to activation of the Ah receptor and induction of CYP1A1, an event that can lead to the formation of BPDE-2. With both B[a]P-7,8-dihydrodiol and BPDE-2 treatment, changes in anti- and pro-apoptotic events in the Bcl-2 family of proteins correlated with the release of mitochondrial cytochrome c and caspase activation. The onset of apoptosis as monitored by caspase activation was linked to mitogen-activated protein (MAP) kinases. Utilizing mouse hepa1c1c7 cells and the Arnt-deficient BPRc1 cells, activation of MAP kinase p38 by B[a]P-7,8-dihydrodiol was shown to be Ah receptor-dependent, indicating that metabolic activation by CYP1A1 was required. This was in contrast to p38 activation by BPDE-2, an event that was independent of Ah receptor function. Confirmation that MAP kinases play a critical role in BPDE-2-induced apoptosis was shown by inhibiting caspase activation of poly(ADP-ribose)polymerase 1 (PARP-1) by chemical inhibitors of p38 and ERK1/2. Furthermore, mouse embryo p38-/- fibroblasts were shown to be resistant to the actions of BPDE-2-induced apoptosis as determined by annexin V analysis, cytochrome c release, and cleavage of PARP-1. These results confirm that the Ah receptor plays a critical role in B[a]P-7,8-dihydrodiol-induced apoptosis while p38 MAP kinase links the actions of an electrophilic metabolite like BPDE-2 to the regulation of programmed cell death.

7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide↗