Snail1 expression in colorectal cancer and its correlation with clinical and pathological parameters.

BACKGROUND: Snail1 is a transcription regulator of E-cadherin. The loss of E-cadherin seems to be a crucial step in the process of Epithelial-mesenchymal transition (EMT). EMT initiates invasion and proliferation in many tumours. Overexpression of Snail1 is known to be associated with poor outcome in several solid tumours. The aim of this study was to analyse its expression profile and prognostic significance in colorectal cancer. METHODS: Tissue microarrays (TMA) containing paraffin-embedded primary colorectal cancer (CRC) tissue samples from 251 patients were used in this study. The expression of Snail1 and E-cadherin was assessed by immunohistochemistry in different tumour compartments, corresponding lymph node metastases and normal colonic mucosa. Intensity of staining was classified according to the Remmele score (standardized scoring system) as well as the semiquantitative score established by Blechschmidt et al. RESULTS: Snail1 expression was observed in 76% of the CRC. Loss of E-cadherin was noted in 87% of the CRC. Snail1 positive tumours were significantly correlated with Snail1 positive lymph node metastases (p=0.03). There was no significant correlation between loss of E-cadherin and Snail1 expression, or between N-stage or grading and Snail1 expression. Kaplan-Meier survival analysis identified no prognostic impact of Snail1 expression on overall survival. CONCLUSION: Snail1 expression was detectable in most of the CRC but showed no significant association with E-cadherin loss, clinical pathological characteristics or overall survival. The observed loss of E-cadherin could be explained by effects of other important EMT pathways, such as the Wnt-signalling cascade.

The evolution of carbapenem resistance determinants and major epidemiological lineages among carbapenem-resistant Acinetobacter baumannii isolates in Germany, 2010-2019.

The aim of this study was to investigate and compare the molecular epidemiology and carbapenem resistance determinants in clinical Acinetobacter baumannii isolates collected during four multicentre surveillance studies conducted by the Paul-Ehrlich-Society for Infection Therapy. Isolates were collected prospectively from hospital in-patients at 17 medical centres in Germany over four periods of three- to six-months starting in October of each of 2010, 2013, 2016 and 2019. Species identification was performed by MALDI-TOF, gyrB multiplex polymerase chain reaction (PCR), and detection of the intrinsic blaOXA-51-like gene. Minimum inhibitory concentrations were determined by broth microdilution. The prevalence of carbapenemase-encoding genes was investigated by OXA-multiplex PCR and whole-genome sequencing. Molecular epidemiology was examined by rep-PCR and core-genome multi-locus sequence typing. A total of 302 A. baumannii isolates were collected. Resistance to imipenem and/or meropenem was detected in 58 isolates (19.2%) from 14 centres. The proportion of carbapenem-resistant isolates increased from 21.3% in 2010 to 33.3% in 2013, and then decreased to 13.8% in 2016 and 12.3% in 2019. Forty-six of these isolates were associated with the international clonal lineage IC2 and five with IC1. The most prevalent carbapenemase gene detected was blaOXA-23-like (n=51). Further carbapenem-resistance determinants were blaOXA-40-like (n=1), blaOXA-58-like (n=3) and blaNDM-1 (n=2). In one isolate, ISAba1 was detected upstream of blaOXA-51-like. In conclusion, IC2 was the most prevalent clonal lineage detected in this study. Interestingly, in Germany, carbapenem resistance seems to have decreased in A. baumannii between 2013 and 2019.

Potential role of vasomotor effects of fibrinogen in bradykinin-induced angioedema.

BACKGROUND: Although bradykinin is known to play a major role in the pathophysiology of hereditary and angiotensin-converting enzyme inhibitor (ACEi)-induced angioedema, other factors acting as triggers or enhancers are likely important as well. OBJECTIVE: We hypothesized that fibrinogen might contribute to ACEi-induced angioedema (eg, through direct actions on vascular tone). METHODS: Plasma levels of fibrinogen were determined in 59 patients with acute angioedema. Vascular activity of human and bovine fibrinogen and its effects on bradykinin-induced vasodilation and phosphorylation of vasodilator-stimulated phosphoprotein were investigated in small (0.8-1.4 mm in diameter) porcine coronary artery and human internal thoracic artery (ITA) segments. RESULTS: In patients with ACEi-induced angioedema, fibrinogen levels (481 +/- 22 mg/dL, n = 39) were significantly higher than in patients with idiopathic angioedema (302 +/- 15 mg/dL, P < .001). Fibrinogen (1-15 mumol/L) induced a concentration-dependent vasodilation in preconstricted small porcine coronary arteries (n = 13), reaching a maximum vasodilator effect of 70% +/- 4.7%. Likewise, fibrinogen induced a 52.1% +/- 9.1% (n = 7) vasodilation in ITA rings. Fibrinogen vasorelaxations were completely inhibited by abciximab and diminished by endothelial denudation and treatment with the nitric oxide synthase inhibitor L-nitroargininemethylester and glibenclamide (P < .01). Importantly, fibrinogen increased the vasodilator potency of bradykinin by 10-fold (P < .0001) and increased bradykinin-induced vasodilator-stimulated phosphoprotein phosphorylation (P < .01). CONCLUSION: The increase of plasma fibrinogen levels, its vasodilator activity in human ITAs, and the potentiation of bradykinin-induced vasodilation suggest that fibrinogen might contribute to the pathophysiology of ACEi-induced angioedema. Thus acute-phase proteins, such as fibrinogen, might be viewed as risk factors for bradykinin-induced angioedema.

Reduction of peripheral flow reserve impairs endothelial function in conduit arteries of patients with essential hypertension.

BACKGROUND: A diminished flow reserve in resistance vessels is a hallmark of hypertensive microvascular disease. Hypertension is associated with structural alterations in the microcirculation and a reduced endothelium-dependent dilation in conduit arteries. Both have been demonstrated to predict future cardiovascular events. OBJECTIVE: We hypothesized that a reduced peripheral flow reserve impairs endothelial function in upstream conduit arteries in patients with arterial hypertension. DESIGN: In 43 hypertensive patients (HT) and 38 normotensive controls (NT) endothelial function of the brachial artery was assessed by measurement of flow-mediated dilatation (FMD), using high-resolution ultrasound. Peripheral flow reserve (FR) was determined via measurements of forearm blood flow at rest and during increments of reactive hyperaemia, using venous occlusion plethysmography. RESULTS: FMD was markedly impaired in HT (3.6 +/- 0.3%) as compared with NT (10.2 +/- 0.3%), whereas maximum brachial artery diameter following endothelium-independent dilatation was similar in both groups. In hypertensive patients FR was significantly reduced (HT, 3.2 versus NT, 6.0) during reactive hyperaemia after 5 min of ischaemia. FR was associated with FMD (r = 0.68, P < 0.01). Multiple stepwise regression analysis identified FR as a strong independent variable determining the extent of FMD (r2 = 0.46, P < 0.01). In HT the dose-response curve of FMD upon stepwise increases of FR was shifted significantly to the right. Normalization of FR improved FMD in HT by more than 60%. CONCLUSIONS: In essential hypertension a reduced FR contributes to the endothelial dysfunction of upstream conduit arteries. These findings may have therapeutic and prognostic implications in patients with arterial hypertension.

Prevalence of mupirocin resistance in clinical isolates of Staphylococcus aureus and Staphylococcus epidermidis: results of the Antimicrobial Resistance Surveillance Study of the Paul-Ehrlich-Society for Chemotherapy, 2001.

A multicentre surveillance study comprising 26 laboratories located in Austria, Germany, and Switzerland was carried out in November 2001. A total of 787 isolates of Staphylococcus aureus and 456 isolates of Staphylococcus epidermidis mainly recovered from hospitalised patients, were tested. MICs for mupirocin were determined using the broth microdilution procedure. Breakpoints were < or = 4 mg/l (susceptible), 8-256 mg/l (low-level resistance) and > or = 512 mg/l (high-level resistance). Rates of low- and high-level resistances were 2.9 and 0.9% in S. aureus, and 9.4 and 3.3% in S. epidermidis, respectively. Mupirocin resistance was almost exclusively observed in oxacillin-resistant isolates of S. aureus (MRSA) and S. epidermidis (MRSE). High-level mupirocin resistance was detected in 3.1 and 4.5% of MRSA and MRSE, respectively.

Occurrence of multidrug resistance to oral antibiotics among Escherichia coli urine isolates from outpatient departments in Germany: extended-spectrum ß-lactamases and the role of fosfomycin.

The in vitro activities of fosfomycin and seven other antibiotics commonly used for oral treatment of urinary tract infections (UTIs) were evaluated for 499 Escherichia coli isolated from urine samples during a nationwide laboratory-based surveillance study in 2010. Overall, the highest resistance rates were found for amoxicillin (42.9%), followed by amoxicillin/clavulanic acid (32.7%), trimethoprim/sulfamethoxazole (SXT) (30.9%), ciprofloxacin (19.8%), cefuroxime (10.0%), cefpodoxime (8.6%) and cefixime (8.2%). One-half of the isolates (n=252; 50.5%) were fully susceptible to the eight drugs, whilst only 6 strains (1.2%) were resistant to fosfomycin. Combined resistance to amoxicillin, cefuroxime, ciprofloxacin and SXT was detected in 29 isolates (5.8%). Moreover, 40 isolates (8.0%) produced an extended-spectrum ß-lactamase (ESBL), including CTX-M-type ESBLs detected in 39/40 isolates (97.5%) and a TEM-52 ESBL in 1 strain (2.5%). The predominant CTX-M-type ESBL was CTX-M-15 (27/39; 69.2%). Of the 27 CTX-M-15 producers, 19 (70.4%) belonged to the clonal lineage E. coli O25b-ST131. All but one ESBL-producing strains were fosfomycin-susceptible. In view of the emergence of multidrug resistance to standard oral antibiotics, these data support that oral fosfomycin (trometamol salt) may represent a valuable option in the treatment of uncomplicated UTIs.

Laser-supported CD133+ cell therapy in patients with ischemic cardiomyopathy: initial results from a prospective phase I multicenter trial.

OBJECTIVES: This study evaluates the safety, principal feasibility and restoration potential of laser-supported CD133+ intramyocardial cell transplantation in patients with ischemic cardiomyopathy. METHODS: Forty-two patients with severe ischemic cardiomyopathy (left ventricular ejection fraction (LVEF) >15% and <35%) were included in this prospective multicenter phase I trial. They underwent coronary artery bypass grafting (CABG) with subsequent transepicardial low-energy laser treatment and autologous CD133+ cell transplantation, and were followed up for 12 months. To evaluate segmental myocardial contractility as well as perfusion and to identify the areas of scar tissue, cardiac MRI was performed at 6 months and compared to the preoperative baseline. In addition, clinical assessment comprising of CCS scoring, blood and physical examination was performed at 3, 6 and 12 months, respectively. RESULTS: Intraoperative cell isolation resulted in a mean cell count of 9.7±1.2×106. Laser treatment and subsequent CD133+ cell therapy were successfully and safely carried out in all patients and no procedure-related complications occurred. At 6 months, the LVEF was significantly increased (29.7±1.9% versus 24.6±1.5% with p = 0.004). In addition, freedom from angina was achieved, and quality of life significantly improved after therapy (p<0.0001). Interestingly, an extended area of transmural delayed enhancement (>3 myocardial segments) determined in the preoperative MRI was inversely correlated with a LVEF increase after laser-supported cell therapy (p = 0.024). CONCLUSIONS: This multicenter trial demonstrates that laser-supported CD133+ cell transplantation is safe and feasible in patients with ischemic cardiomyopathy undergoing CABG, and in most cases, it appears to significantly improve the myocardial function. Importantly, our data show that the beneficial effect was significantly related to the extent of transmural delayed enhancement, suggesting that MRI-guided selection of patients is mandatory to ensure the effectiveness of the therapy. TRIAL REGISTRATION: EudraCT 2005-004051-35) Controlled-Trials.com ISRCTN49998633.

Is there a role for the Fas-/Fas-Ligand pathway in chemoresistance of human squamous cell carcinomas of the head and neck (SCCHN)?

The aim of the present investigation was to determine the expression of the Fas-receptor/ligand system in established cell lines of squamous cell carcinomas of the head and neck (SCCHN), and to study it's functional impact on chemotherapy-induced apoptosis in these SCCHN cell lines. We observed constitutive expression of Fas and FasL in 13 SCCHN cell lines by RT-PCR, Southern-blotting and immunocytochemistry, respectively. Administration of the agonistic Fas-antibody CH-11 led to a significant reduction of viable cells in the colorimetric MTT-assay in 5 out of 13 (38%) cell lines tested and preincubation with Interferon-gamma (IFN-gamma) rendered 3 (23%) primarily resistant cell lines sensitive. Cisplatin (cDDP) and bleomycin (BLM) caused dose-dependent cytotoxicity in all cell lines as determined by the 50% inhibitory concentration (IC(50)) and induction of apoptosis. Furthermore, both antineoplastic agents led to an enhanced surface expression of Fas and FasL in all cell lines, and this effect was independent of the respective p53-status. This upregulation of Fas/FasL surface expression increased preexisting Fas-sensitivity only, but failed to make primarily resistant cell lines undergo Fas-mediated growth reduction or apoptosis. Vice versa, blockade of Fas-receptor-ligand-interactions by monoclonal antibodies directed against FasL was able to attenuate the cytotoxic effect of cDDP and BLM in 2 out of 5 (40%) cell lines tested only. In conclusion, in contrast to many other solid tumors, the Fas/FasL-system does not seem to play an exclusive role in anticancer drug mediated apoptosis in SCCHN.

Alterations in the p53 pathway and their association with radio- and chemosensitivity in head and neck squamous cell carcinoma.

Chemotherapy and/or radiotherapy are established measures in treatment protocols of head and neck squamous cell carcinoma (HNSCC). However, we still lack reliable predictive markers for the response to radio- and chemotherapy. The p53 pathway is involved in stress response and thus might influence chemo-/radiosensitivity. Using 29 HNSCC cell lines previously characterized for p53 mutations, we simultaneously analyzed several key players in the p53 pathway by RT-PCR, transcript sequencing and immunohistochemistry, and investigated their association with chemosensitivity and radiosensitivity. Cell lines with p53 mutations were slightly more sensitive to cisplatin than those with wild-type p53. The type of mutation did not influence radio- or chemosensitivity. p14(ARF), an activator of p53, was lost or mutated in all cell lines. Three cell lines showed overexpression of HDM-2, a major negative regulator of p53; however, HDM-2 levels did not correlate with radio- or chemosensitivity. HPV-16 oncoproteins were detected in one highly chemoresistant cell line. Our findings suggest that molecular events resulting in the inactivation of the p53 pathway occur in all HNSCC cell lines. However, single alterations in the p53 pathway are not reliable predictors for the response to radio- or chemotherapy in HNSCC.

Pharmacokinetics of oral betaine in healthy subjects and patients with homocystinuria.

AIMS: Large oral doses of betaine have proved effective in lowering plasma homocysteine in severe hyperhomocysteinaemia. The pharmacokinetic characteristics and metabolism of betaine in humans have not been assessed and drug monitoring for betaine therapy is not available. We studied the pharmacokinetics of betaine and its metabolite dimethylglycine (DMG) in healthy subjects and in three patients with homocystinuria. METHODS: Twelve male volunteers underwent an open-label study. After one single administration of 50 mg betaine kg-1 body weight and during continuous intake of twice daily 50 mg kg-1 body weight, serial blood samples and 24 h urines were collected to determine betaine and DMG plasma concentrations and urinary excretion, respectively. Patients were evaluated after one single dose of betaine. RESULTS: We found rapid absorption (t(1/2),abs 00.28 h, s.d. 0.17) and distribution (t(1/2), lambda1 00.59 h, s.d. 0.22) of betaine. A Cmax of 0.94 mmol l-1 (s.d. 0.19) was reached after tmax 00.90 h (s.d. 0.33). The elimination half life t(1/2), z was 14.38 h (s.d. 7.17). After repeated dosage, t(1/2), lambda1 (01.77 h, s.d. 0.75) and t(1/2), z (41.17 h, s.d. 13.50) increased significantly (95% CI 0.73, 01.64 h and 19.90, 33.70 h, respectively), whereas absorption remained unchanged. DMG concentrations increased significantly after betaine administration and accumulation occurred to the same extent as with betaine. Renal clearance was low and urinary excretion of betaine was equivalent to 4% of the ingested dose. Distribution and elimination kinetics in homocystinuric patients appeared to be accelerated. CONCLUSIONS: Betaine plasma concentrations change rapidly after ingestion. Elimination half-life increased during continuous dosing over 5 days. Betaine is mainly eliminated by metabolism. More pharmacokinetic and pharmacodynamic studies in hyperhomocysteinaemic patients are needed to refine the current treatment with betaine.

Antitumor activity of protein kinase C inhibitors and cisplatin in human head and neck squamous cell carcinoma lines.

Protein kinase C (PKC) plays a pivotal role in signal transduction involved in the control of cell proliferation, differentiation and apoptosis. Interference with such signaling pathways may result in altered tumor cell response to antineoplastic drugs. We investigated the effects of two selective PKC inhibitors as single agents and in combination with cisplatin in cell lines derived from squamous cell carcinomas of the head and neck (SCCHN). Safingol (Saf) is directed against the regulatory domain, whereas chelerythrine (Che) interacts with the catalytic domain of PKC. In six SCCHN cell lines (UM-SCC 11B, 14A, 14C and 22B, 8029NA, and a 5-fold cisplatin-resistant subline 8029DDP). PKC activities ranged between 1 and 158 IU/1 x 10(7) cells, and they were inversely proportional to the amount of cellular epidermal growth factor receptor. Using the colorimetric MTT assay, PKC inhibitors Saf and Che showed comparable dose-dependent growth inhibition. The 50% inhibitory concentrations (IC50) were between 3.8-8.6 microM for Saf and 8.5-13.6 microM for Che with no relationship to PKC activity or cisplatin sensitivity of the respective cell lines. Combinations of cisplatin (IC50 = 0.4-5.8 microg/ml) and either PKC inhibitor (5 microM Saf, 10 microM Che) led to a significant decrease of cisplatin IC50 values in most cell lines. However, comparison with theoretical additive dose-response curves showed additive rather than synergistic effects for both PKC inhibitors.

Toxin-gene profile heterogeneity among endemic invasive European group A streptococcal isolates.

We determined the toxin-gene profiles of 239 endemic, invasive group A streptococcal (GAS) isolates that circulated, within a 5-year period, in European university hospitals. Profiling was performed by use of multiplex polymerase chain reaction that screened for 9 streptococcal pyrogenic exotoxins (speA, speB, speC, speF, speG, speH, speJ, ssa, and smeZ). Analysis revealed that invasive GAS isolates do not share a common toxin-gene profile. Although all emm types were characterized by several different toxin-gene profiles, a predominance of 1 or 2 toxin-gene profiles could be observed, reflecting that a few invasive clones have spread successfully throughout the world. Remarkably, statistical pair-wise analysis of individual toxin genes revealed that strains that did not share the predominant profile still showed a nonrandom distribution of key toxin genes characteristic of the specific emm type. This could indicate that M proteins function, directly or indirectly, as barriers for horizontal gene exchange.