Human tumor cell strains defective in the repair of alkylation damage

We have previously identified four human astrocytoma cell strainsas defective in the repair of N-methyl-N'-nitro-N-nitrosoguanidine(MNNG) damaged adenovirus 5. We now show that two of these strains(the only two tested), in comparison to other tumor strainsor normal human skin fibroblasts, are very sensitive to MNNG-producedkilling as measured by colony forming ability, but are normallysensitive to ultraviolet light. Further, such repair deficientcells may be cultured from tumors of the colon, lung, skin,and neck. The phenotype of deficient repair of MNNG-treatedadenovirus 5 has now been found in a subgroup of 9 of the 39human tumor strains tested. We propose to call this phenotypethe Mer^– phenotype. None of the 22 strains of normal humanskin fibroblasts tested showed deficient repair of MNNG damage.MNNG treatment (80 µM) causes a decrease in semi-conservativeDNA synthesis from which Mer^– tumor cells do not recover,but from which cells capable of normal repair of MNNG damage(Mer⁺) do. Somewhat paradoxically, Mer^–cells show moreMNNG-stimulated DNA synthesis (‘repair synthesis’)than do Mer⁺ cells. Besides being deficient in the repair ofMNNG-damaged adeno-viruses Mer– cells also have difficultyin repairing viruses damaged either by other N-alkyl-N'-nitro-N-nitrosoguanidines,or by N-methyl- or N-ethyl-N-nitrosoureas.     

Chronic Oral Anticoagulation and Clinical Outcome in Hospitalized COVID-19 Patients

Cardiovascular Drugs and Therapy - The clinical course of COVID-19 may be complicated by acute respiratory distress syndrome (ARDS) and thromboembolic events, which are associated with high risk of...     

Hypersensitivity to DNA-damaging agents in cultured cells from patients with Usher''s syndrome and Duchenne muscular dystrophy.

Lymphoblastoid lines from nine Usher's syndrome (recessively inherited retinitis pigmentosa and congenital sensorineural deafness) patients (representing eight kindreds) and from ten Duchenne muscular dystrophy patients (representing seven kindreds) showed a small but statistically significant hypersensitivity to the lethal effects of X-rays, as measured by the cellular ability to exclude the vital dye trypan blue, when compared with lines from 26 normal control subjects. Fibroblast lines from the Usher's syndrome patients, treated with X-rays or the radiomimetic, DNA-damaging chemical N-methyl-N'-nitro-N-nitrosoguanidine, also showed a statistically significant hypersensitivity when compared to normal fibroblast lines. These findings are consistent with the possibility that defective DNA repair mechanisms may be involved in the pathogenesis of these degenerative diseases.     

Abdominal aortic aneurysm in patients affected by intermittent claudication: prevalence and clinical predictors

Background

Abdominal aortic aneurysm (AAA) is a frequent cause of death among elderly. Patients affected by lower extremity peripheral arterial disease (LE-PAD) seem to be particularly at high risk for AAA. We aimed this study at assessing the prevalence and the clinical predictors of the presence of AAA in a homogeneous cohort of LE-PAD patients affected by intermittent claudication.

Methods

We performed an abdominal ultrasound in 213 consecutive patients with documented LE-PAD (ankle/brachial index ≤0.90) attending our outpatient clinic for intermittent claudication. For each patient we registered cardiovascular risk factors and comorbidities, and measured neutrophil count.

Results

The ultrasound was inconclusive in 3 patients (1.4%), thus 210 patients (169 males, 41 females, mean age 65.9 ± 9.8 yr) entered the study. Overall, AAA was present in 19 patients (9.0%), with a not significant higher prevalence in men than in women (10.1% vs 4.9%, p = 0.300). Patients with AAA were older (71.2 ± 7.0 vs 65.4 ± 9.9 years, p = 0.015), were more likely to have hypertension (94.7% vs 71.2%, p = 0.027), and greater neutrophil count (5.5 [4.5 – 6.2] vs 4.1 [3.2 – 5.5] x10³/μL, p = 0.010). Importantly, the c-statistic for neutrophil count (0.73, 95% CI 0.60 – 0.86, p =0.010) was higher than that for age (0.67, CI 0.56–0.78, p = 0.017). The prevalence of AAA in claudicant patients with a neutrophil count ≥ 5.1 x10³/μL (cut-off identified at ROC analysis) was as high as 29.0%.

Conclusions

Prevalence of AAA in claudicant patients is much higher than that reported in the general population. Ultrasound screening should be considered in these patients, especially in those with an elevated neutrophil count.

     

Ligand-directed surface profiling of human cancer cells with combinatorial peptide libraries

A collection of 60 cell lines derived from human tumors (NCI-60) has been widely explored as a tool for anticancer drug discovery. Here, we profiled the cell surface of the NCI-60 by high-throughput screening of a phage-displayed random peptide library and classified the cell lines according to the binding selectivity of 26,031 recovered tripeptide motifs. By analyzing selected cell-homing peptide motifs and their NCI-60 recognition patterns, we established that some of these motifs (a) are similar to domains of human proteins known as ligands for tumor cell receptors and (b) segregate among the NCI-60 in a pattern correlating with expression profiles of the corresponding receptors. We biochemically validated some of the motifs as mimic peptides of native ligands for the epidermal growth factor receptor. Our results indicate that ligand-directed profiling of tumor cell lines can select functional peptides from combinatorial libraries based on the expression of tumor cell surface molecules, which in turn could be exploited as "druggable" receptors in specific types of cancer.     

Aequorin chimeras as valuable tool in the measurement of Ca2+ concentration during cadmium injury

The ability of cadmium to disrupt calcium homeostasis has been known since a long time, but the precise cellular targets of its toxic action are still debated. A great problem in the interpretation of data has been associated with the ability of cadmium to strongly bind traditional calcium probes. Aequorin, the well-characterized calcium-sensitive photoprotein, was used as intracellular calcium indicator during cadmium injury in NIH 3T3 murine fibroblasts. NIH 3T3 cells were transfected with a cDNA construct containing aequorin fused to a truncated glutamate receptor, which directs the probe to the outer surface of intracellular membranes. At first, we tested if different cadmium concentrations were able to modify the rate of light emission by aequorin showing that cadmium concentrations <15 microM were ineffective on aequorin luminescence. Hence, aequorin chimeras revealed as a useful tool in the analyses of Cd2+/Ca2+ interference. To directly investigate the role of Cd2+ in Ca2+ homeostasis, we have started to selectively measure the free Ca2+ concentration in different cell compartments. Here, we report that cadmium reduces the transient free calcium signal after stimulation of cells with bradykinin. Further studies are in progress to clarify the role of mitochondria and endoplasmic reticulum in cadmium-induced alterations of Ca2+ homeostasis in order to link signal transduction modifications with the onset of apoptosis induced by cadmium exposure.