Novel nanosensors for rapid analysis of telomerase activity

Elevated telomerase levels are found in many malignancies, offering an attractive target for therapeutic intervention and diagnostic or prognostic purposes. Here we describe the use of a novel nanosensor developed for rapid screens of telomerase activity in biological samples. The technique utilizes magnetic nanoparticles that, on annealing with telomerase synthesized TTAGGG repeats, switch their magnet state, a phenomenon readily detectable by magnetic readers. We tested the efficacy of different telomerase inhibitors in crude human and murine samples and show that phosphorylation of telomerase regulates its activity. High-throughput adaptation of the technique by magnetic resonance imaging allowed processing of hundreds of samples within tens of minutes at ultrahigh sensitivities. Together, these studies establish and validate a novel and powerful tool for rapidly sensing telomerase activity and provide the rationale for developing analogous magnetic nanoparticles for in vivo sensing.     

Progression and distribution of plantar pressure in Type 2 diabetic patients

The aim of this prospective 4-yr study was to analyse changes in mean plantar pressure (PP) over time and local shifts of maximal PP in Type 2 diabetic patients. One-hundred fifty-five Type 2 diabetic patients (age 58.9 +/- 7.5 yr, diabetes duration 11.0 +/- 7.6 yr, baseline HbA1c 9.6 +/- 1.6%) were examined with regard to foot abnormalities, neuropathy and measurement of PP during walking (pedobarography). They were assigned to two subgroups, namely normal PP (n=94) and elevated PP (n=57). Patients with an abnormal mean PP did not significantly differ from subjects with a normal PP with regard to sex, age, duration of diabetes and HbA1c. With the exception of the hallux, the mean PP was significantly increased in both groups at all other plantar sites. Maximum PP was located below the metatarsal heads (MTH) 2-5 and significantly increased from baseline (median, lower/upper quartile: 475, 355/715 kPa) to the end of the study (540, 435/749; p<0.0001). On the other hand, PP was normalized in 17 subjects (29.8%) who had an elevated PP at baseline. Furthermore, we observed a local shift in maximal PP towards the MTH 2-5 region. The percentage of patients who had their highest PP under MTH 2-5 was increased from 54.0% at baseline to 61.1% at the end of the study. In general, we registered an elevation of PP over time and a centralization towards sites which are generally prone to ulceration.     

Inhibition of nuclear factor-kappaB and nitric oxide by curcumin induces G2/M cell cycle arrest and apoptosis in human melanoma cells

Curcumin (diferuloylmethane) inhibits tumour cell growth by inducing apoptosis in many tumour types, including melanoma, via complex and ill-defined pathways. Recent studies have shown that curcumin is both a nitric oxide scavenger and an inhibitor of inducible nitric oxide synthase (iNOS) expression, low levels of which correlate with antiapoptotic function and poor survival and which may be regulated by inhibition of nuclear factor-kappaB (NFkappaB) activation. To elucidate the mechanisms by which curcumin inhibits melanoma proliferation, we tested the in vitro effects of curcumin on specific cell cycle pathways and melanoma cell survival, including NFkappaB activation. Curcumin induced melanoma cell apoptosis and cell cycle arrest, which is associated with the downregulation of NFkappaB activation, iNOS and DNA-dependent protein kinase catalytic subunit expression, and upregulation of p53, p21(Cip1), p27(Kip1) and checkpoint kinase 2. Curcumin also downregulated constitutive iNOS activity in melanoma cells. Our results demonstrate that curcumin arrested cell growth at the G(2)/M phase and induced apoptosis in human melanoma cells by inhibiting NFkappaB activation and thus depletion of endogenous nitric oxide. Therefore, curcumin should be considered further as a potential therapy for patients with melanoma.     

Revascularization of myocardial scar tissue following prostaglandin E1-therapy in patients with ischemic heart disease

Prostaglandin E1 (PGE-1) treatment has proved to stimulate angiogenesis in vital non-infarcted myocardium of patients with ischemic cardiomyopathy (ICMP). We investigated infarcted myocardial tissue for a possible angiogenic response to PGE-1. Neovascularization was investigated in infarcted areas of 12 hearts explanted from patients with ICMP who had been treated with PGE-1 before heart transplantation (HTX). In transmural sections containing myocardial scar tissue, CD34 and VEGF were immunohistochemically quantified to estimate capillary density and the extent of angiogenesis. To investigate a possible effect of PGE-1 on collagen turnover, the collagen content was determined in myocardial scar tissue by assessing the intensity of the area positively stained with sirius red. PGE-1-treated patients had significantly more CD34- and VEGF-positive cells in infarcted areas, and showed a significant reduction in collagen content as compared with the non-PGE-1 group (CD34: 120.3 +/- 6.1 vs. 47.7 +/- 6.1 capillary profiles/mm2; VEGF: 52.8 +/- 5.6 vs. 24.0 +/- 4.8 capillary profiles/mm2, and collagen content: 2.18 +/- 0.4 eU vs. 3.59 +/- 0.38 eU). Our data demonstrate that PGE-1 stimulates angiogenesis by upregulating VEGF expression, and reduces fibrosis in cardiac scar tissue of ischemic origin. The induction of therapeutic angiogenesis in vital and at sites of putative dead myocardial scar tissue, along with the hemodynamic improvement in patients with severe ICMP, might explain the favorable clinical outcome in PGE-1-treated patients before HTX.     

Association of right ventricular dysfunction and Cheyne-Stokes respiration in patients with chronic heart failure

Cheyne-Stokes respiration (CSR) is present in up to 40% of patients with congestive heart failure (CHF) and is an independent risk factor for increased overall mortality. We examined whether CSR is associated with right ventricular (RV) dysfunction in CHF patients. Parameters of RV function were assessed by two-dimensional echocardiography and tissue velocity imaging in 42 patients (aged 23-75 years) with a left ventricular (LV) ejection fraction below 40%. Respiratory polygraphy revealed CSR with an central apnea-hypopnea index (CAHI) >10 h-1 in 13 of the 42 patients (31%). Demographic characteristics did not differ among the patient groups. The velocity of the tricuspid annular systolic motion (TASM), a parameter reflecting systolic RV function, was significantly reduced in CHF patients with CSR (10.5 +/- 2.3 cm s-1) compared with those without CSR (15.0 +/- 5.1 cm s-1, P = 0.004), and was inversely associated with the CAHI (y = 15.2-0.2x; r = 0.46, P = 0.003). The RV dimensions were significantly increased and the fractional RV area changes significantly reduced in CHF patients with CSR (33 +/- 17 versus 48 +/- 20%; P = 0.04). Doppler parameters of pulmonary artery flow indicate higher pulmonary artery pressures in CSR patients compared with patients without CSR, which is also reflected by an increased RV free-wall thickness in CSR patients (6.5 +/- 1.1 vs. 5.3 +/- 1.3 mm; P = 0.05). Parameters of systolic LV function, forced expiratory volume in 1 s (FEV1), and PaO2 and PaCO2 were not different among patients with or without CSR. In conclusion, CSR is associated with depressed systolic RV function and increased RV dimensions in CHF patients. Future studies will show whether optimized treatment of CSR will improve RV function.     

Rezidivierende Synkopen bei Linksschenkelblock mit normaler PQ-Zeit

Zusammenfassung Wir berichten den Fall einer 53-j?hrigen Patientin, die sich wegen rezidivierender Synkopen zur station?ren Aufnahme vorstellte. Im Oberfl?chen-EKG fand sich ein Sinusrhythmus mit normaler PQ-Zeit und komplettem Linksschenkelblock. Echokardiographisch wurde eine auf ca. 45% reduzierte Pumpfunktion ermittelt. Unter dem Verdacht einer entzündlichen Herzmuskelkrankung wurde eine Herzkatheteruntersuchung mit Endomyokardbiopsie durchgeführt, deren histologische und immunhis-tologische Untersuchung keinen Anhalt für eine Myokarditis oder Speichererkrankung ergab. Die elektrophysiologische Untersuchung zeigte ein stark verl?ngertes HV-Intervall von 101 ms (normal 35–55 ms). Daraufhin wurde ein Zweikammerschrittmachersystem (DDD) implantiert. Eine Untersuchung nach 3 Monaten zeigte die bereits eingetretene Schrittmacherabh?ngigkeit der Patientin bei komplettem AV-Block ohne ausreichenden Ersatzrhythmus.     

The value of a noninvasive diagnostic approach to mediastinal masses

BACKGROUND: Mediastinal tumors show a wide variability, and therefore, a standardized diagnostic and therapeutic workup is instrumental. We subdivided mediastinal tumors into nonlymphatic mediastinal tumors (NLMTs), most of which require surgical resection without need of preoperative histology, and mediastinal lymphadenopathy (MLA), requiring surgical biopsy for exact histologic classification. We investigated the accuracy of noninvasive diagnostic studies distinguishing between the two groups of MLA and NLMT. METHODS: A retrospective analysis was performed on patients who had previously undergone surgery on mediastinal tumors. Their data were statistically analyzed (chi2 test, logistic regression analysis), and the values of medical history, physical examination, laboratory tests, and computerized tomography scan discriminating between MLA and NLMT were assessed. RESULTS: Out of 299 patients included in the study, 242 (80.9%) had MLA and 57 (19.1%) had NLMT. Sensitivity and specificity of noninvasive investigations for differentiation of MLA and NLMT were 98.2% and 86.0%, respectively. Whereas the prevalence of thoracic symptoms such as shortness of breath, cough, or chest pain was similar in both groups (MLA, 165 [69.3%]; NLMT, 41 [69.5%]; p = 0.98), systemic symptoms, including fever, night sweats, or weight loss (MLA, 110 [49.8%]; NLMT, 17 [29.3%]; p < 0.01), and signs of inflammation, such as c-reactive protein, erythrocyte sedimentation rate, and leukocytosis (MLA, 202 [85.6%]; NLMT, 34 [57.6%]; p < 0.001), were significantly more common in MLA. CONCLUSIONS: Noninvasive diagnostic procedures, including medical history, physical examination, laboratory tests, and computerized tomographic scan, are highly sensitive in detecting MLAs that should undergo surgical biopsy. Our data suggest confirming all suspected NLMTs by fine needle aspiration (FNA) biopsy before surgery.     

Increased light damage susceptibility at night does not correlate with RPE65 levels and rhodopsin regeneration in rats

The susceptibility of rats to light-induced retinal degeneration is increased at night. In mice, an important determinant of light damage susceptibility is the efficacy of rhodopsin regeneration after bleaching. The rate of rhodopsin regeneration is at least partly controlled by RPE65, a protein expressed in the retinal pigment epithelium. We therefore tested a potential involvement of RPE65 and rhodopsin regeneration in the increased light damage susceptibility of rats at night. For this purpose, rats were exposed to visible light at noon or at midnight and extent of light damage was determined by retinal morphology and TUNEL staining. Rpe65 gene expression was analyzed by semiquantitative RT-PCR and levels of RPE65 protein were determined by Western blotting. Rhodopsin regeneration kinetics was determined by measuring rhodopsin content immediately after a strong bleach and after different times of recovery in darkness.Rats were more susceptible to light damage at night as described by Organisciak and collegues [Invest. Ophthalmol. Vis. Sci. 41 (2000) 3694]. Rpe65 gene expression followed a day-night rhythm with highest steady-state mRNA levels at the beginning and lowest levels at the end of the day period. However, RPE65 protein levels remained constant. Rhodopsin regeneration kinetics did not differ during day and night. We conclude that levels of RPE65 protein and rhodopsin regeneration kinetics do not correlate with the increased light damage susceptibility observed in rats at night. Additional genetic or physiologic modifiers may exist in rats that regulate the retinal responsiveness to acute light exposure.