Infliximab is not superior to placebo in preventing clinical recurrence after CD-related resection. However, infliximab does reduce endoscopic recurrence. ClinicalTrials.gov ID NCT01190839.

Protein kinase CK2 has been implicated in the regulation of a wide range of proteins that are important in cell proliferation and differentiation. Here we demonstrate that the stress signaling agents anisomycin, arsenite, and tumor necrosis factor-␣ stimulate the specific enzyme activity of CK2 in the human cervical carcinoma HeLa cells by up to 8-fold, and this could be blocked by the p38 MAP kinase inhibitor SB203580. We show that p38␣ MAP kinase, in a phosphorylation-dependent manner, can directly interact with the ␣ and ␤ subunits of CK2 to activate the holoenzyme through what appears to be an allosteric mechanism. Furthermore, we demonstrate that anisomycin-and tumor necrosis factor-␣-induced phosphorylation of p53 at Ser-392, which is important for the transcriptional activity of this growth suppressor protein, requires p38 MAP kinase and CK2 activities.The signaling pathways that regulated protein kinase CK2 1 (formerly casein kinase II) have been elusive since the discovery of this highly conserved and ubiquitous protein-serine/ threonine kinase over 45 years ago (1-5). Despite the identification of over 160 putative substrates for CK2, the overriding function of this kinase has been evasive. In most cells, the holoenzyme form of CK2 is a constitutively active heterotetramer of two catalytic ␣ and/or ␣Ј) and two regulatory (␤) subunits, but CK2␤-free pools of CK2␣ also exist. Further increases (2-3-fold) in the phosphotransferase activity of CK2 in cell lysates toward synthetic peptide substrates have been reported following treatment of murine and human fibroblastic cell lines, A431 and 3T3-L1 adipocytes with serum (6), insulin (7-9), insulin-like growth factor-18 (10), epidermal growth factor (7, 11), bombesin (12), and tumor necrosis factor (TNF) (13). CK2 phosphotransferase activity and protein levels are also commonly elevated in solid human tumors and transformed cell lines (2-4, 14, 15). Furthermore, CK2␣ overexpression along with c-Myc induces lymphomas in mice (16). While the increased expression of CK2 is linked with neoplastic transformation, little is known concerning acute regulation of this protein kinase. EXPERIMENTAL PROCEDURESCell Culture and Lysates Preparation-HeLa cells were grown at 37°C on 125-cm 2 flasks in M199 medium (Life Technologies, Inc.) supplemented with 10% fetal calf serum. For cell stimulation, HeLa cells were split at 2 ϫ 10 6 cells/10-cm 2 dish 24 h prior to starvation. Cells were starved with 0.5% fetal calf serum for 15-20 h before exposure to stimuli. Starved cells were incubated with 10 M SB203580 (Calbiochem) for 30 min prior to addition of 10 g/ml anisomycin (Sigma) for 30 min, 50 M arsenite (Sigma) for 15 min, or 20 ng/ml TNF␣ for 15 min. Adhering cells were washed twice with ice-cold phosphate-buffered saline, scraped, then lysed in 50 mM Hepes (pH 7.2), 100 mM NaCl, 1 mM EGTA, and 20 mM sodium fluoride in Buffer A (1 mM sodium orthovanadate, 1% aprotinin, 1 mM phenylmethylsulfonyl fluoride, 1 M pepstatin, 0.5 g/ml leupeptin, 10 g/ml soybean trypsin inhibitor, ...

Macrophages play a central role in the development and progression of atherosclerotic lesions. It is well known that oxidized low density lipoprotein (ox-LDL) promotes the recruitment of monocytes (which differentiate to macrophages) into the intima. We reported recently that ox-LDL blocks apoptosis in bone marrow-derived macrophages deprived of macrophage colonystimulating factor (M-CSF) by a mechanism involving protein kinase B (PKB) (Hundal, R., Salh, B., Schrader, J., Gó mez-Muñ oz, A., Duronio, V., and Steinbrecher, U. (2001) J. Lipid Res. 42, 1483-1491). The aims of the present study were 1) to define the apoptotic pathway involved in the pro-survival effect of ox-LDL; 2) to determine which PKB target mediated this effect; and 3) to identify mechanisms responsible for PKB activation by ox-LDL. Apoptosis following M-CSF withdrawal was accompanied by activation of the caspase 9-caspase 3 cascade and cytochrome c release from mitochondria, but the caspase 8 pathway was unaffected. M-CSF withdrawal resulted in a marked and selective reduction in Bcl-X L protein and mRNA levels, and this decrease was prevented by ox-LDL. The ability of ox-LDL to preserve Bcl-X L levels was blocked by NFB antagonists, thereby implicating IB kinase as a key PKB target. M-CSF deprivation resulted in activation of acid sphingomyelinase and an increase in ceramide levels. Desipramine (a sphingomyelinase inhibitor) prevented the increase in ceramide and inhibited apoptosis after M-CSF deprivation. Ox-LDL completely blocked the increase in acid sphingomyelinase activity as well as the increase in ceramide after M-CSF deprivation. Pretreatment of macrophages with C 2 -ceramide reversed the effect of ox-LDL on PKB and macrophage survival. These results indicate that ox-LDL prevents apoptosis in M-CSFdeprived macrophages at least in part by inhibiting acid sphingomyelinase. This in turn prevents ceramide-induced down-regulation of PKB, the activity of which is required to maintain production of Bcl-X L .

Numerous therapies used for inflammatory bowel disease (IBD) target the transcription factor NF-kappaB, which is involved in the production of cytokines and chemokines integral for inflammation. Here we show that curcumin, a component of the spice turmeric, is able to attenuate colitis in the dinitrobenzene sulfonic acid (DNB)-induced murine model of colitis. When given before the induction of colitis it reduced macroscopic damage scores and NF-kappaB activation. This was accompanied by a reduction in myeloperoxidase activity, and using semiquantitative RT-PCR, an attenuation of the DNB-induced message for IL-1beta was detected. Western blotting analysis revealed that there was a reproducible DNB-induced activation of p38 MAPK detected in intestinal lysates by using a phosphospecific antibody. This signal was significantly attenuated by curcumin. Furthermore, we show that the immunohistochemical signal is dramatically attenuated at the level of the mucosa by curcumin. We conclude that the widely used food additive curcumin is able to attenuate experimental colitis through a mechanism correlated with the inhibition of the activation of NF-kappaB and effects a reduction in the activity of p38 MAPK. We propose that this agent may have therapeutic implications for human IBD.

Summary Stress‐activated protein kinases (SAPKs) are activated in human inflammatory bowel disease (IBD). Recently it has been demonstrated that p38MAPK (mitogen‐activated protein kinase) inhibition using SB203580 is effective in reducing disease in both dextran sulphate sodium (DSS)‐induced and 2,4,6‐trinitrobenzenesulphonic acid (TNBS)‐induced murine colitides, underscoring the importance of this pathway in gastrointestinal inflammation. However, the contribution of c‐Jun N‐terminal kinase (JNK) in intestinal inflammation is unknown. Based on the known involvement of JNK in tumour necrosis factor‐α (TNF‐α) expression and in mediating the effects of oxidant stress, we hypothesized that JNK inhibition would also affect colitis. Our studies in mice with DSS‐induced colitis treated with the JNK inhibitor SP600125, indicate that there is a significant reduction in wasting as well as a significant reduction in histological damage scores. Both total colonic and mesenteric lymphocyte CD3/CD28‐stimulated TNF‐α levels were dramatically reduced under the same circumstances. This was associated with a reduction in JNK protein expression and activity, as well as a reduction in AP‐1 DNA binding with SP600125. Interestingly, there were no apparent changes in either p38MAPK or p42/44ERKs. Immunofluorescence of the colon for the active form of JNK revealed a prominent signal arising from the infiltrating inflammatory cells. SP600125 reduced this as well as, specifically, macrophage infiltration. Strikingly, we also demonstrate reduced epithelial cell apoptosis in response to treatment with SP600125. We conclude that specific inhibition of JNK is beneficial in the DSS model of colitis, and may be of value in human IBD.