The relative contribution of calcium, zinc and oxidation-based cross-links to the stiffness of Arion subfuscus glue.

Metal ions are present in many different biological materials, and are capable of forming strong cross-links in aqueous environments. The relative contribution of different metal-based cross-links was measured in the defensive glue produced by the terrestrial slug Arion subfuscus. This glue contains calcium, magnesium, zinc, manganese, iron and copper. These metals are essential to the integrity of the glue and to gel stiffening. Removal of all metals caused at least a 15-fold decrease in the storage modulus of the glue. Selectively disrupting cross-links involving hard Lewis acids such as calcium reduced the stiffness of the glue, while disrupting cross-links involving borderline Lewis acids such as zinc did not. Calcium is the most common cation bound to the glue (40 mmol l(-1)), and its charge is balanced primarily by sulphate at 82-84 mmol l(-1). Thus these ions probably play a primary role in bringing polymers together directly. Imine bonds formed as a result of protein oxidation also contribute substantially to the stiffness of the glue. Disrupting these bonds with hydroxylamine caused a 33% decrease in storage modulus of the glue, while stabilizing them by reduction with sodium borohydride increased the storage modulus by 40%. Thus a combination of metal-based bonds operates in this glue. Most likely, cross-links directly involving calcium play a primary role in bringing together and stabilizing the polymer network, followed by imine bond formation and possible iron coordination.

Segmental haemorrhagic colitis under ciprofloxacin therapy.

The segmental haemorrhagic colitis has been described as a particular form of antibiotics-associated colitis under penicillin therapy. A 34-year-old male presented in reduced general condition. He suffered from haematoschezia and abdominal pain. Due to a sinusitis maxillaris he had taken ciprofloxacin tablets (500 mg b. i. d.) for four days prior to the day of admission. Upon physical examination a slight general abdominal pain on palpation and some fresh blood at rectal digital examination were found. Apart from a slightly elevated CK level, all other routinely checked laboratory parameters were within normal ranges. All stool cultures remained negative for pathogenous germs. On colonoscopy we saw a severe haemorrhagic colitis with map-like mucosal lesions. It spread over the sigmoid and the distal part of the descending colon. The pathohistological assessment of step biopsies showed an acute erosive, ulcerative colitis. After cessation of antibiotic therapy the disorders subsided within three days without specific treatment. To the best of our knowledge a segmental haemorrhagic colitis under ciprofloxacin has not been described previously. A single Japanese paper mentions three cases of patients with a segmental haemorrhagic colitis under quinolones taken for four weeks before the symptoms started. In contrast, our patient developed symptoms within a couple of hours after intake of ciprofloxacin. We assume a toxic or idiosyncratic reaction to ciprofloxacin.

Absence of the interstitial cell of Cajal network in mitochondrial neurogastrointestinal encephalomyopathy.

Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is a rare autosomal-recessive multisystemic disorder with predominant gastrointestinal involvement, presenting with variable degrees of gut dysmotility up to frank chronic intestinal pseudoobstruction. Despite major advances in understanding its basic molecular pathogenesis in recent years, the distinct mechanisms and pathoanatomical substrate underlying MNGIE-associated gastrointestinal dysmotility are still widely unknown. As yet, though their critical role in proper gastrointestinal transit in terms of spontaneous pacemaker activity and enteric neurotransmission is well established, the population of the interstitial cells of Cajal (ICC) has not been investigated in MNGIE. Therefore, we examined small bowel samples of a well-characterized MNGIE patient by using conventional histology and immunohistochemistry techniques. The ICC network was studied by immunohistochemistry for the tyrosine kinase Kit (CD117), known to reliably detect ICCs, while mucosal mast cells served as an internal and normal small bowel specimen as external controls. At a light microscopic level, no gross structural alteration of the bowel wall composition and its neuromuscular elements was noted. However, a complete absence of Kit immunoreactive cells could be demonstrated in regions where ICCs are normally abundant, while internal and external controls retained strong Kit positivity. In conclusion, our preliminary results provide a first evidence for an alteration of the ICC network in MNGIE, and support the notion that ICC loss might be an early pathogenetic event in MNGIE-associated gut motor dysfunction before significant myopathic and/or neuropathic structural changes occur.

Paclitaxel and carboplatin vs gemcitabine and vinorelbine in patients with adeno- or undifferentiated carcinoma of unknown primary: a randomised prospective phase II trial.

Platinum/taxane combinations are widely used in patients with carcinoma of unknown primary (CUP), yielding response rates of 30% and median overall survival of 9-11 months in selected patients. Yet these combinations have not been subject to a randomised trial to overcome selection bias, a major problem in CUP. We randomised 92 patients to either paclitaxel/carboplatin (arm A) or the non-platinum non-taxane regimen gemcitabine/vinorelbine (arm B). The primary endpoint was rate of practicability as defined: application of >or=2 cycles of therapy (1) with a maximal delay of 1 week (2) and survival of >or=8 months (3). Practicability was shown in 52.4% (95% CI 36-68%) in arm A and in 42.2% (95% CI 28-58%) in arm B, respectively. The median overall survival, 1-year survival -rate and response rate of patients treated in arm A was 11.0 months, 38, and 23.8%, arm B 7.0 months, 29, and 20%. In conclusion, the paclitaxel/carboplatin regimen showed clinically meaningful activity in this randomised trial (Clinical trial registration number 219, 'Deutsches KrebsStudienRegister', German Cancer Society.)

Para-[(123)I]iodo-L-phenylalanine in patients with pancreatic adenocarcinoma: tumour uptake, whole-body kinetics, dosimetry.

UNLABELLED: Recently, p-[(123)I]iodo-L-phenylalanine (IPA) was clinically validated for brain tumour imaging. Preclinical studies demonstrated uptake of IPA into pancreatic adenocarcinoma suggesting its diagnostic application in patients with pancreatic tumours. The aim was to study the tumour uptake of IPA in patients with pancreatic adenocarcinoma and to analyse its biodistribution and dosimetry to assess the radiation dose resulting from its diagnostic use. PATIENTS, METHODS: Seven patients with pancreatic adenocarcinoma underwent whole-body scintigraphies and SPECT up to 24 h after administration of 250 MBq of IPA. Tumour uptake of IPA was assessed visually. Time activity curves and the corresponding residence times were determined for whole-body, kidneys, liver, spleen, lung, heart content, brain, and testes. Mean absorbed doses for various organs and the effective dose were assessed based on the MIRD formalism using OLINDA/EXM. RESULTS: IPA exhibited no accumulation in proven manifestations of pancreatic adenocarcinomas. IPA was exclusively eliminated by the urine and showed a delayed clearance from blood. Residence times were 0.26 +/- 0.09 h for kidneys, 0.38 +/- 0.19 h for liver, 0.15 +/- 0.07 h for spleen, 0.51 +/- 0.20 h for lungs, 0.22 +/- 0.07 h for heart content, 0.11 +/- 0.05 h for brain, 0.014 +/- 0.005 h for testes and 6.4 +/- 2.2 h for the remainder. The highest absorbed doses were determined in the urinary bladder wall and in the kidneys. According to the ICRP 60 the effective dose resulting from 250 MBq IPA was 3.6 +/- 0.7 mSv. CONCLUSION: Para-[(123)I]iodo-L-phenylalanine can be used in diagnostic nuclear medicine with acceptable radiation doses. Besides its proven validity for brain tumour imaging, IPA does not appear to be suitable as tracer for pancreatic cancer.

Increased cytokine transcripts in pouchitis reflect the degree of inflammation but not the underlying entity.

BACKGROUND AND AIMS: After ileopouch anal anastomosis (IPAA), 10-40% of patients with ulcerative colitis (UC) but only 5% of patients with familial adenomatous polyposis (FAP) develop pouchitis. Immunoregulatory abnormalities might be of importance in the pathogenesis of the disease. Therefore, we characterized cytokine and chemokine transcripts in inflamed and non-inflamed pouches in patients with UC compared to those with FAP and Crohn's disease (CD). PATIENTS AND METHODS: Mucosal biopsies were taken from 87 patients with IPAA [UC (n=70), CD (n=8) or FAP (n=9)]. Patients with active ileal CD (n=14), active UC (n=17) and non-inflammatory conditions (n=12) served as controls. The expression of 20 gene transcripts was quantified using real-time polymerase chain reaction. RESULTS AND FINDINGS: Pro-inflammatory cytokines and chemokines are significantly increased in IPAA patients with acute pouchitis. This increase is independent of the underlying disease (UC or CD) and reflects the degree of inflammation. A good correlation between pouchitis activity (using the Pouchitis Disease Activity Index) and the MRP-14, interleukin-8, macrophage inflammatory protein-2alpha and matrix metalloproteinase-1 transcripts was observed. INTERPRETATIONS AND CONCLUSIONS: Our data support the view that pouchitis reflects an inflammatory process that is different from that of underlying inflammatory bowel diseases, as the cytokine and chemokine patterns in pouchitis are neither typical of CD nor of UC, but maybe due to bacterial intestinal microflora overgrowth in the pouch lumen. Quantification of transcript levels allows an estimation of the extent of mucosal inflammation and may become helpful in the evaluation of the disease, especially in clinical trials.

Randomised phase II evaluation of irinotecan plus high-dose 5-fluorouracil and leucovorin (ILF) vs 5-fluorouracil, leucovorin, and etoposide (ELF) in untreated metastatic gastric cancer.

An open-label randomised comparison of efficacy and tolerability of irinotecan plus high-dose 5-fluorouracil (5-FU) and leucovorin (LV) (ILF) with etoposide plus 5-FU/LV (ELF) in patients with untreated metastatic or locally advanced gastric cancer. One cycle of ILF comprised six once-weekly infusions of irinotecan 80 mg m(-2), LV 500 mg m(-2), 24-h 5-FU 2000 mg m(-2), and ELF comprised three once-daily doses of etoposide 120 mg m(-2), LV 300 mg m(-2), 5-FU 500 mg m(-2). In all, 56 patients received ILF and 58 ELF. Median age was 62 years, Karnofsky performance 90%, and disease status was comparable for both arms. The objective clinical response rates after 14 weeks treatment (primary end point) were 30% for ILF and 17% for ELF (risk ratio (RR) 0.57, 95% confidence interval (CI) 0.29-1.13, P = 0.0766). Overall response rates over the entire treatment period for ILF and ELF were 43 and 24%, respectively (RR 0.56, 95% CI 0.33-0.97; P = 0.0467). For ILF and ELF, respectively, median progression-free survival was 4.5 vs 2.3 months, time to treatment failure was 3.6 vs 2.2 months (P = 0.4542), and overall survival was 10.8 vs 8.3 months (P = 0.2818). Both regimens were well tolerated, the main grade 3/4 toxicities being diarrhoea (18%, ILF) and neutropenia (57%, ELF). The data from this randomised phase II study indicate that ILF provides a better response rate than ELF, and that ILF should be investigated further for the treatment of metastatic gastric cancer.

[Diagnosis, staging and therapy of gastric cancer]. / Diagnostik, Staging und Therapie des Magenkarzinoms.

In spite of a continuously decreasing incidence in Western countries, gastric cancer remains a major health problem world-wide and a leading cause of cancer-related deaths. The molecular pathogenesis of the vast majority of sporadic tumours has not yet been clarified in detail, while for some familial cancer types causal germline mutations have been identified. Only radical resection offers a chance of cure and the prognosis is determined by the stage of disease at diagnosis. Unfortunately, about two-thirds of patients are still diagnosed in a late stage. The results of surgery have reached a plateau of effectiveness, in stages III and IV (UICC) an R0 resection will be possible in only about 40 % of cases and even, after successful curative resection, the prognosis is limited due to a high recurrence rate. The benefit of neoadjuvant chemotherapy in locally advanced cancers has been verified in phase II studies. Less clear is the adjuvant situation - the benefit of adjuvant radiochemotherapy after systematic lymphadenectomy awaits to be proven. In the palliative situation platinum-based high-dose 5-FU infusion protocols are currently the standard therapy.

Innovative drugs and strategies in the treatment of upper gastrointestinal tract carcinomas.

The results of the standard therapy of upper gastrointestinal tract cancer remain unsatisfactory, mainly because still the majority of cases are diagnosed at a late stage of disease, which implicates a poor prognosis. Innovative strategies focus on neoadjuvant chemotherapy in locally advanced gastric cancer and chemo- or radiochemotherapy in advanced oesophageal cancer. Results of phase II studies published so far have been promising. There is rising evidence that adjuvant radiochemotherapy in gastric cancer may be of benefit for curatively resected patients, whereas the results of trials including a systematic lymphadenectomy have to be awaited. New substances like the taxanes or irinotecan in the palliative setting did not prove to be superior to platinum-based standard chemotherapy protocols so far.

[Oesophageal cancer--diagnosis]. / Osophaguskarzinom--Diagnostik.

The incidence of oesophageal cancer has been rising for the past decades in the western world due to the rapid increase of oesophageal adenocarcinoma. Strategy and extent of the diagnostic work up in case of suspected oesophageal cancer should be focussed on the therapeutic consequences which have to be drawn in the individual patient. Upper endoscopy including biopsy will provide the diagnosis and the location of the tumour, computed tomography and transcutaneous sonography are used to rule out distant metastases. Endoscopic ultrasound is the tool of choice to obtain an exact locoregional staging - an experienced investigator may correctly classify the tumour according to the T-N-categories in about 80% of the cases. To define early cancers restricted to the mucosa, which may be suitable for local ablative therapy, high-resolution endosonography is essential. Intravital staining methods may help to delineate flat or multifocal cancers, they are also used to better recognise dysplastic foci in a Barrett-segment. To rule out osseous metastases a bone scintigraphy is indicated. In tumours located proximal to the bifurcation a bronchoscopy completes the diagnostic procedures, in case of advanced tumours of the oesophagogastric junction laparoscopy may be performed to rule out peritoneal carcinosis.

Downregulation of CD44v6 in colorectal carcinomas is associated with hypermethylation of the CD44 promoter region.

Overexpression of the cell adhesion protein CD44v6 has been demonstrated in colorectal cancer and other gastrointestinal tumors. While CD44v6 is upregulated in benign colorectal adenomas and well-differentiated colorectal cancer tissues, downregulation frequently occurs during disease progression. The mechanism of downregulation, however, is unknown. Therefore, we evaluated the methylation status of the CD44 promoter as a mechanism for decreased CD44v6 expression in advanced colorectal carcinomas. We demonstrated by methylation-sensitive restriction enzyme digestion that the CpG islands of the CD44 promoter were methylated in 6/21 (28%) of benign colorectal adenomas. Interestingly, in colorectal carcinomas the frequency of promoter methylation was significantly increased (10/19; 53%) compared to 7/21 (33%) in the corresponding normal mucosa. Methylation seems to be associated with a more advanced cancer stage, but the trend did not reach statistical significance. In colorectal carcinomas with CD44 promoter methylation CD44v6 mRNA was detected by reverse transcription-polymerase chain reaction in 3/10 carcinomas, whereas in tumors without CD44 promoter methylation CD44v6 expression was observed in 8/9 (P

Lichen planus with oesophageal involvement. A case report and review of the literature.

BACKGROUND: Lichen planus is a common mucocutaneous disease of unknown aetiology. Oral disease affecting the mouth and the pharynx occurs in 30-70% of the cases. Oesophageal disease is considered to be extremely rare. The diagnosis of oesophageal involvement is often not made until complications occur. CASE REPORT: A 56-year-old woman with oral and genital erosive lichen planus for more than 4 years complained of odynophagia and dysphagia. Episodes of oesophageal bolus obstruction started 2 months earlier. Upper endoscopy revealed a high-grade concentric stenosis at 21-24 cm from the incisors. Biopsy specimens taken after bougienage showed a squamous epithelium with dense leukocyte infiltration and Civatte bodies. The bougienage led to complete relief, but due to recurrent symptomatic stenosis, endoscopic dilatation had to be performed another four times within 5 years of follow-up. Immunosuppressive therapy with systemic and local steroid application did not prevent recurrent stenosis. CONCLUSIONS: Patients with lichen planus should be evaluated for gastrointestinal symptoms because oesophageal involvement is a rare but severe complication leading to inflammatory stenosis. The benefit of immunosuppressive therapy in the prevention of recurrent stenosis is not established. A review of all reported cases is done with particular regard to therapy.

Expression of CD44v6 has no prognostic value in patients with colorectal cancer.

BACKGROUND AND AIMS: In colorectal cancer patients' mortality is largely influenced by spreading of tumour cells from the primary tumour site and subsequent metastasis formation. CD44 is an adhesion molecule and represents a highly variable family of isoforms. The isoform CD44v6 has been associated with metastatic spread and poor prognosis in animal models and several human cancers. Results of immunohistological studies in primary colorectal cancer are mostly retrospective and contradictory. The aim of our prospective study was to assess the controversial role of CD44v6 as a prognostic factor in colorectal cancer. METHODS: In 93 patients we analysed tumour CD44v6 expression in prospectively sampled stage I-IV colorectal adenocarcinomas using RT-PCR and Southern blotting. The prognostic value of the CD44v6 expression was assessed using univariate and multivariate analysis. RESULTS: CD44v6 expression was found in 47 % of the cases. CD44v6 expression failed to show any association with the clinical or histological variables examined. CD44v6 expression did not correlate with survival in long-term follow-up. The most important prognostic factor in this cohort was tumour stage. CONCLUSIONS: Changes in CD446 expression level do not predict tumour spread or patient survival in colorectal cancer.

Arabidopsis E2F1 binds a sequence present in the promoter of S-phase-regulated gene AtCDC6 and is a member of a multigene family with differential activities.

Mammalian E2F transcription factors are composed of E2F and DP subunits, and with their negative regulators, Rb-related proteins, govern expression of cell-division-related genes. Six E2Fs and two DPs are present in mammals, but only single E2F genes are known from wheat, tobacco and carrot. Here we show that E2Fs are a multigene family in Arabidopsis, and report isolation of three E2F-like clones AtE2F1-3, with 45-62% identity to wheat, tobacco and carrot E2Fs. Sequence analysis reveals that AtE2F1 and AtE2F3 are closely related to previously identified plant E2Fs, whereas AtE2F2 is related to human E2F6 and Drosophila dE2F2 which are unusual in lacking transcriptional activation potential. Consistent with this, we show that AtE2FI and AtE2F3 activate transcription in yeast cells and bind a plant Rb protein, but AtE2F2 cannot activate transcription or bind Rb. Consensus E2F-binding sites were identified in promoters of several cell cycle related genes, including the D-type cyclin CycD3 and the Arabidopsis homologue of the replication origin protein CDC6. Accumulation of AtE2F1-3 was observed in partially synchronised Arabidopsis cells re-entering the cell cycle, before induction of CycD3 and CDC6 expression in late G1. AtE2F1 complexes bound to consensus E2F sequences and to the AtCDC6 promoter in vitro. We conclude that Arabidopsis contains a family of functionally distinct E2F genes, most probably involved in the G1-to-S phase progression.

Phased chest and abdominal compression-decompression versus conventional cardiopulmonary resuscitation in out-of-hospital cardiac arrest.

BACKGROUND: Several methods have been developed to improve the efficacy of mechanical resuscitation, because organ perfusion achieved with conventional manual resuscitation is often insufficient. In animal studies, phased chest and abdominal compression-decompression resuscitation by use of the Lifestick device has resulted in a better outcome compared with that of conventional resuscitation. In end-of-life patients, an increased coronary perfusion pressure was achieved. The aim of the present study was to determine the feasibility, safety, and efficacy of the Lifestick compared with conventional resuscitation in patients with sudden nontraumatic out-of-hospital cardiac arrest. METHODS AND RESULTS: The crews of 4 mobile intensive care units, staffed by an emergency physician and a paramedic, were trained to use the device. Fifty patients were randomized by sealed envelopes to either Lifestick (n=24) or conventional (n=26) resuscitation. No differences were found regarding demographic and logistical conditions between the groups. Nineteen of the patients (73%) with conventional resuscitation had ventricular fibrillation, 13 of whom survived to hospital admission (no survivals with other arrhythmias) and 7 were discharged. In contrast, in the Lifestick-CPR group, only 9 patients had ventricular fibrillation (38%; P=<0.02; OR, 2.5; 95% CI, 0.6 to 10.6). Four of these 9 patients and 5 of 15 patients with other arrhythmias survived to hospital admission, but none survived to hospital discharge. Autopsy in a subgroup of patients who died at the scene revealed less injuries with Lifestick than with conventional resuscitation. CONCLUSION: Lifestick resuscitation is feasible and safe and may be advantageous in patients with asystole or pulseless electric activity.

Cell cycle regulation of cyclin-dependent kinases in tobacco cultivar Bright Yellow-2 cells.

Plants possess two major classes of cyclin-dependent kinases (CDK) with cyclin-binding motifs PSTAIRE (CDK-a) and PPTA/TLRE (CDK-b). Tobacco (Nicotiana tabacum L. cv Bright Yellow-2) cells are the most highly synchronizable plant culture, but no detailed analysis of CDK activities has been reported in this system. Here we describe isolation of new PPTALRE CDKs (Nicta;CdkB1) from Bright Yellow-2 cells and present detailed analysis of the mRNA, protein and kinase activity levels of CdkB1, and the PSTAIRE CDKA during the growth and cell cycles. CdkA and CdkB1 transcripts are more abundant in exponential than in stationary phase cells, but the two genes show strikingly different regulation during the cell cycle. CdkA mRNA and protein accumulate during G1 in cells re-entering the cell cycle, and immunoprecipitated histone H1 kinase activity increases at the G1/S boundary. Aphidicolin synchronized cells show the highest CDKA-associated histone H1 kinase activity during S-G2 phases, although CdkA mRNA and protein levels are not significantly regulated. In contrast, CdkB1 transcripts are present at very low levels until S phase and CDKB1 protein and kinase activity is almost undetectable in G1. CdkB1 mRNA accumulates through S until M phase and its associated kinase activity peaks at the G2/M boundary, confirming that transcription of PPTALRE CDKs is cell cycle regulated. We suggest that CDKA kinase activity likely plays roles at the G1/S phase boundary, during S phase, and at the G2/M phase transition, and that CDKB1 kinase activity is present only at G2/M.

Increased matrix metalloproteinase 2 concentration and transcript expression in advanced colorectal carcinomas.

Colorectal cancer is one of the most common malignant tumors and entails a relatively poor prognosis. Clinical outcome depends on the extent of local and metastatic tumor spread. Results of in vivo and in vitro studies suggest that the balance between matrix metalloproteinases (MMPs) and their inhibitors (tissue inhibitors of metalloproteinases TIMPs) is altered in neoplasia, contributing to the invasive and metastatic properties of malignant tumors. We quantified tissue concentrations of MMP-2 and TIMP-2 in 65 malignant colorectal lesions and corresponding normal mucosa by enzyme-linked immunosorbent assay, western blotting, and in situ hybridization. In situ hybridization and western blot analyses demonstrated a clear increase in both stromal expression of MMP-2 transcripts and protein in primary carcinomas. The protein concentration of MMP-2 was higher in all tumor stages, except stage I tumors, than in normal mucosa and adenomas. MMP-2 concentrations were not related to tumor differentiation or to colonic versus rectal location. Surprisingly, the MMP-2 concentration was not increased in metastases. Interestingly, tissue concentrations and epithelial mRNA expression of TIMP-2 decreased significantly in primary colorectal cancer (UICC stages III and IV) but increased in metastases. Therefore an increased ratio of MMP-2 to TIMP-2 is strongly associated with advanced tumor stages, but a decreased ratio was observed in metastases. These findings suggest that the MMP-2:TIMP-2 ratio may prove useful as a marker of local invasion but not of metastasis in colorectal cancer.