MAPK regulation of prostaglandin E2 production by lipopolysaccharide-stimulated macrophages is not dependent on nuclear factor kappaB

BACKGROUND: Prostaglandin E2 (PGE2) is a major contributor to the production and maintenance of immunosuppression in injured and septic patients. Although the synthesis of PGE2 by various enzymes has been elucidated, the regulatory mechanism of these enzymes is not clear. The purpose of this study was to determine the role of MAPK cascades in COX-2 gene activation by lipopolysaccharide (LPS)-stimulated macrophages (Mphi). MATERIALS AND METHODS: RAW 264.7 cells, a mouse Mphi cell line, were exposed to Escherichia coli LPS (10 microg/ml) in the presence of PD98059, a selective inhibitor of MAPKP44/P42, and SB202190, a selective inhibitor of MAPKP38. COX-2 mRNA expression and PGE2 production were measured by Northern Blot assay and ELISA, respectively. Mphi nuclear factor (NF)kappaB and cAMP-response element (CRE) activities were determined by electrophoretic mobility shift assays. RESULTS: LPS stimulation increased Mphi COX-2 mRNA expression and PGE2 production. PD98059 or SB202190 attenuated LPS-induced COX-2 mRNA as well as PGE2 production in a dose-dependent fashion. Inhibition of MAPKP44/P42 or MAPKP38 had no effect on NFkappaB activation but reduced CRE activity induced by LPS stimulation. CONCLUSION: Our data show that MAPK cascades regulate COX-2 gene expression and PGE2 production in LPS-stimulated Mphi through NFkappaB independent pathways. The regulatory mechanism is likely to be mediated through CRE.     

Tissue distribution of a new photosensitizer ATX-S10Na(II) and effect of a diode laser (670 nm) in photodynamic therapy

The aims of the present study were to analyse the quantitative tissue distribution of ATX-S10Na(II) and to investigate the maximal effect of a diode laser and the irradiation conditions required to obtain this effect in photodynamic therapy (PDT) with ATX-S10Na(II). Spectrofluorometry was used to obtain quantitative tissue distribution of ATX-S10Na(II) in Colon 26 carcinoma-bearing mice as a function of time following administration. Next, transplanted tumours of mice with or without ATX-S10Na(II) were treated with the diode laser under conditions in which power density and irradiation time were varied. Tumour tissue concentrations of ATX-S10Na(II) were higher than in all tissues at all intervals following administration. The uptake of ATX-S10Na(II) by most tissues was rapid, with maximal concentrations occurring 1 h after i.v. injection, and ATX-S10Na(II) was almost excreted within 24 h after administration. The maximal depth of necrosis induced by PDT in the treated tumour was 7.9 mm under conditions in which power density was 160 mW/cm² and total dose was above 100 J/cm². PDT with ATX-S10Na(II) and the diode laser is useful for the treatment of superficial cancers.     

Transactivation of ErbB1 and ErbB2 receptors by angiotensin II in normal human prostate stromal cells

BACKGROUND: Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is synthesized primarily in the stromal compartment of the human prostate and may regulate stromal as well as epithelial cell growth and survival. The primary cognate HB-EGF receptor, ErbB1, has been shown recently to be transactivated by G-protein coupled receptors (GPCRs) through regulated proteolytic cleavage of the membrane-bound, precursor form of HB-EGF. Previous studies have demonstrated that human prostate tissue, especially tissue from benign prostatic hyperplasia (BPH), has high angiotensin converting enzyme activity, and a high density of angiotensin (Ang) receptors in periurethral stromal cells. Because the pressor peptide Ang II signals through GPCRs, we tested the possibility that Ang II could transactivate ErbB1/ErbB2 in human prostate stromal (hPS) cells. METHODS: Primary and early passage hPS cells were used as an in vitro model. Cells were stimulated by recombinant HB-EGF or Ang II and total cell lysates were harvested for immunoprecipitation and Western blot. Cell growth was measured by [(3)H]thymidine incorporation assay and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazoliumbromide (MTT) assay. RESULTS: Ang II receptors AT1 and AT2 were expressed in hPS cells. ErbB1 and ErbB2 receptors were activated by HB-EGF. Furthermore, Ang II was able to transactivate both ErbB1 and ErbB2, and this transactivation activity could be abolished by pretreatment with [Glu-52]-diphtheria toxin/CRM197, a specific inhibitor of HB-EGF bioactivity. Consistent with its transactivation activity, Ang II modestly promoted hPS cell growth and this effect was abolished by pretreatment with the ErbB1 antagonist AG1478. CONCLUSION: These experiments suggest a regulatory role for Ang II in the prostate stroma and implicate the endogenous stromal growth factor HB-EGF as a mediator of Ang II signaling in the prostate.     

Structural determinants for the activation mechanism of the angiotensin II type 1 receptor differ for phosphoinositide hydrolysis and mitogen-activated protein kinase pathways

While the mechanism whereby the angiotensin II type 1 receptor (AT(1) receptor) activates its classical effector phospholipase C-beta (PLC-beta) has largely been elucidated, there is little consensus on how this receptor activates a more recently identified effector, the p42/44 mitogen-activated protein kinases (p42/44(MAPK)). Using transfected COS-1 cells, we investigated the activation of this signaling pathway at the receptor level itself. Previous mutational studies that relied on phosphoinositide turnover as an index of receptor activation have indicated that key residues in the second and seventh transmembrane domains participate in AT(1) receptor activation mechanisms. Thus, we introduced a variety of mutations-AT(1)[D74N], AT(1)[Y292F], AT(1)[N295S], and AT(1)[AT(2) TM7], which is composed of a chimeric substitution of the AT(1) seventh transmembrane domain with its AT(2) counterpart. These mutations that strongly diminished the receptor's ability to activate PLC-beta had little to no effect on its ability to activate p42/44(MAPK), which not only suggests that p42/44(MAPK) does not exclusively lie downstream of the G-protein G(q)/PLC-beta pathway but also indicates that more than one activation state may exist for the AT(1) receptor. The failure of a protein kinase C inhibitor to block AT(1) receptor activation of p42/44(MAPK) further corroborated evidence that the receptor's activation of p42/44(MAPK) is largely independent of the G(q)/PLC-beta/PKC pathway. Taken together, the experimental evidence strongly suggests that the mechanism whereby the AT(1) receptor activates p42/44(MAPK) is fundamentally different from that for PLC-beta, even at the level of the receptor itself.     

Elevated neuronal nitric oxide synthase expression in chronic haloperidol-treated rats

Long-term administration of neuroleptic drugs, such as haloperidol, in the management of psychiatric disorders may adversely cause an irreversible neurological syndrome of tardive dyskinesia, which is associated with dopamine (DA(2)) receptor supersensitivity in the basal ganglia. Recent studies also indirectly suggest an involvement of nitric oxide synthase (NOS) in dopaminergic supersensitivity; however, chronic neuroleptic effects on neuronal NOS (nNOS) expression in the basal ganglia have not been reported. In this investigation, we treated rats with saline or haloperidol (1 mg/kg, s.c.) daily for 21 days. Five days later, we detected a significant increase of NOS activity in the striatum of haloperidol-treated rats when compared to saline controls. This effect was associated with elevated levels of nNOS mRNA and protein expression in the striatum, but not in the nucleus accumbens, as evidenced by the use of in situ hybridization, Western blot and immunohistochemical techniques. The involvement of the nNOS system after chronic haloperidol treatment coincides with increases of striatal DA(2) receptor sites, calmodulin kinase II activity, animal locomotor and stereotypy behaviors. This study suggests an integral role between nNOS, DA(2) receptor and calmodulin system in the development of dopaminergic behavioral supersensitivity resulting from chronic neuroleptic drug treatment. Furthermore, the toxic effect of chronic haloperidol on NOS system selectivity takes place in the neostriatum.     

AT1 antagonism by eprosartan lowers heart rate variability and baroreflex gain

INTRODUCTION: Blockade of the renin-angiotensin system (RAS) by ACE inhibitors has been demonstrated to reduce total mortality in cardiovascular diseases. This advantage was attributed in part to changes of autonomic cardiovascular control, exemplified by an increase of heart rate variability (HRV) and baroreflex gain (BRG). We sought to assess the effects of the angiotensin type 1 (AT1) receptor blocker eprosartan on HRV and BRG. MATERIALS AND METHODS: In a double-blind randomized cross-over design 25 young males took eprosartan (600 mg/day) and placebo each for a period of 7 days with a wash-out period of at least 4 weeks in between. At the end of the intake phases simultaneous recordings of arterial blood pressure (AP; Finapres) and electrocardiogram (ECG) were taken. Power spectra of HRV and arterial blood pressure variability (APV) were calculated by fast Fourier transform (FFT) and served to calculate BRG. Ang-II levels were measured by radioimmunoassay. RESULTS: Eprosartan tended to lower mean AP, it slightly increased heart rate (HR) (p<0.05), and markedly increased circulating Ang-II levels (p<0.01). Eprosartan diminished the total power of HRV (p<0.05) and the BRG (p<0.01). The low/high frequency (LF/HF) ratio of HRV and the APV were not altered. CONCLUSIONS: AT1 antagonism by eprosartan lowers heart rate variability and baroreflex gain. We speculate that these findings are due to the marked increase in circulating angiotensin II (Ang II). Further studies are needed to clarify whether angiotensin type 1 (AT1) blockers with potential actions inside the blood-brain barrier (BBB) may have different effects on HRV and BRG.     

Enterolactone in breast cyst fluid: correlation with EGF and breast cancer risk

The purpose of our study was to investigate whether enterolactone does accumulate into breast cyst fluid and whether it correlates with breast cancer risk. We included 258 women who had at least one cyst aspiration and known intracystic cation and epidermal growth factor (EGF) concentration values. For 191 of such women serum aliquots were also available. The median value of serum enterolactone was 17 nM/l (range 1–140 nM/l). The median intracystic level of enterolactone was much higher (63 nM/l, range 0–872 nM/l) and was significantly higher in type I cysts (p = 0.000). This cyst type contained also significantly higher levels of EGF (p = 0.000). A direct relationship was found between serum and cyst fluid enterolactone levels (p = 0.000) and between cyst enterolactone and EGF levels (p = 0.03), the latter correlation being evident especially in type II cysts. Twelve patients in the cohort of women were found to have developed a breast cancer. After univariate analysis breast cancer risk was associated with cyst type and especially with EGF concentration. No association was evident for enterolactone concentration. However, enterolactone concentration appeared to significantly decrease the risk of patients with high EGF concentrations. Our results show that enterolactone does accumulate in breast cysts, and that it modulates the risk related to the intracystic level of EGF, which is confirmed to be a strong predictor of breast cancer risk.     

A multicentre phase II trial of bryostatin-1 in patients with advanced renal cancer

Protein kinase C (PKC) has a critical role in several signal transduction pathways, and is involved in renal cancer pathogenesis. Bryostatin-1 modulates PKC activity and has antitumour effects in preclinical studies. We conducted a multicentre phase II clinical trial in patients with advanced renal cancer to determine the response rate, immunomodulatory activity and toxicity of bryostatin-1 given as a continuous 24 h infusion weekly for 3 out of 4 weeks at a dose of 25 mug m(-2). In all, 16 patients were recruited (11 males and five females). The median age was 59 years (range 44-68). Patients had been treated previously with nephrectomy (8) and/or interferon therapy (9) and/or hormone therapy (4) and/or radiotherapy (6). Eight, five and three patients had performance statuses of 0, 1 and 2, respectively. A total of 181 infusions were administered with a median of 12 infusions per patient (range 1-29). Disease response was evaluable in 13 patients. Three patients achieved stable disease lasting for 10.5, 8 and 5.5 months, respectively. No complete responses or partial responses were seen. Myalgia, fatigue, nausea, headache, vomiting, anorexia, anaemia and lymphopenia were the commonly reported side effects. Assessment of biological activity of bryostatin-1 was carried out using the whole-blood cytokine release assay in six patients, two of whom had a rise in IL-6 levels 24 h after initiating bryostatin-1 therapy compared to pretreatment values. However, the IL-6 level was found to be significantly lower at day 28 compared to the pretreatment level in all six patients analysed.     

抗早2号方治疗女童真性性早熟的临床研究

目的：观察抗早2号方治疗女童真性性早熟的疗效。并探讨其作用机制。方法：68例辨证为痰热互结型本病患儿随机分为两组。分别用抗早2号方（治疗组43例）和知柏地黄汤（对照组25例）治疗,并观察患儿临床体征,疗效,测定促卵泡生成素（FSH）,促黄体生成素（LH）,雌二醇（E2）及子宫卵巢容积,骨龄的变化。结果：临床治疗3个月后,治疗组和对照组总有效率分别为86.0%和76.0%。两组比较差异无显著性（P>0.05)。两组FSH,E2均较用药前降低（P<0.05)。而治疗组FSH,LH降低程度较对照组更显著（P<0.01)。B超显示：治疗组治疗后子宫容积和卵巢容积小于对照组（P<0.05或P<0.01)。用药后6个月骨龄显示;治疗组骨龄增长明显落后于年龄增长,其减缓骨龄增长速度优于对照组（P<0.05)。结论：提早2号方治疗痰热互结型真性性早熟的疗效确切,是治疗女童真性性早熟的一种有效药物。     

Phase I/II trial of weekly cisplatin,etoposide, and irinotecan chemotherapy for metastatic lung cancer: JCOG 9507

Combinations of cisplatin-irinotecan and cisplatin-etoposide are active and well tolerated in patients with both small-cell lung cancer (SCLC) and nonsmall-cell lung cancer (NSCLC). To define the recommended dose for phase II trials of irinotecan combined with cisplatin and etoposide in chemonaive patients with stage IV disease, 56 patients (11 having SCLC and 45 NSCLC) received cisplatin 25 mg m(-2) weekly for 9 weeks, etoposide 60 mg m(-2) for 3 days on weeks 1, 3, 5, 7 and 9, and irinotecan 20-100 mg m(-2) (levels 1-8) on weeks 2, 4, 6 and 8, together with a prophylactical granulocyte colony-stimulating factor support (50 microg m(-2) on days 4-7 on weeks 1, 3, 5, 7 and 9, and on days 2-7 on weeks 2, 4, 6 and 8). Grade 3-4 leukocytopenia, neutropenia and thrombocytopenia were noted in 20 (36%), 28 (50%) and nine (16%) patients, respectively. Grade 3 diarrhoea, grade 3 cardiac toxicity, and grade 4 transaminase elevation developed in one (1.8%) patient each. Totally, four of 56 patients were removed from the study because of toxicity and recovered, and two other patients died in situations where drug toxicity might contribute to their death. Dose-limiting toxicity was noted in less than one-third of patients at dose levels 1-7, but in all patients at dose level 8. Thus, the recommended dose was determined to be level 7 (irinotecan 90 mg m(-2)). The response rates for SCLC and NSCLC were 91% (10/11) and 38% (17/45), respectively. The median survival time and 1-year survival rate were 11.9 months and 46% for SCLC and 10.1 months and 40% for NSCLC, respectively. This regimen was considered to be feasible and promising for the treatment of stage IV SCLC and NSCLC.