Investigating the effect of cell substrate on cancer cell stiffness by optical tweezers

The mechanical properties of cells are influenced by their microenvironment. Here we report cell stiffness alteration by changing the cell substrate stiffness for isolated cells and cells in contact with other cells. Polydimethylsiloxane (PDMS) is used to prepare soft substrates with three different stiffness values (173, 88 and 17 kPa respectively). Breast cancer cells lines, namely HBL-100, MCF-7 and MDA-MB-231 with different level of aggressiveness are cultured on these substrates and their local elasticity is investigated by vertical indentation of the cell membrane. Our preliminary results show an unforeseen behavior of the MDA-MB-231 cells. When cultured on glass substrate as isolated cells, they are less stiff than the other two types of cells, in agreement with the general statement that more aggressive and metastatic cells are softer. However, when connected to other cells the stiffness of MDA-MB-231 cells becomes similar to the other two cell lines. Moreover, the stiffness of MDA-MB-231 cells cultured on soft PDMS substrates is significantly higher than the stiffness of the other cell types, demonstrating thus the strong influence of the environmental conditions on the mechanical properties of the cells.     

Mechanical forces: The missing link between idiopathic pulmonary fibrosis and lung cancer

Patients with idiopathic pulmonary fibrosis (IPF) have a high risk of developing lung cancer compared with the general population. The morbidity of lung cancer in IPF patient ranges from 3% to 22%, and in some cases exceeds 50%, and these patients have a reduced survival time. However, the mechanisms through which IPF increases the morbidity and mortality in lung cancer remain unclear.By carefully analyzing the pathological features of these two diseases, we uncovered that, first, similar to IPF, lung carcinomas are more frequently found in the peripheral area of the lungs and, second, lung cancers tend to develop from the honeycomb areas in IPF. In accordance with the above pathological features, due to the spatial location, the peripheral areas of the lung experience a high stretch force because the average distance between adjacent alveolar cells in this area tends to be larger than that at the central lung when inflated; furthermore, the honeycomb areas, comprised of condensed fibrous tissue, are characterized by increased stiffness. Both of these pathological features of lung cancer and IPF are coincidentally related to abnormal mechanical forces (stretch and tissue stiffness). Therefore, we believe that the aberrant mechanical forces that are generated in the lung with IPF may contribute to the onset and progression of lung cancer.In this review, we discuss the possible effects of mechanical forces that are generated in IPF on the initiation and progression of lung cancer from the perspective of the hallmarks of cancer, including proliferation, metastasis, angiogenesis, cancer stem cells, immunology, epigenetics, and metabolism, so as to advance our understanding of the pathogenesis of IPF-related lung cancer and to harness these concepts for lung cancer mechanotherapies.     

Rare earth thiocyanate complexes with tripiperidinophosphine oxide: synthesis and characterization. Crystal structure of Pr(SCN)3(tpppO)3

The synthesis and characterization of rare-earth (La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu and Y) thiocyanate adducts with tripiperidinophosphine oxide (tpppO) with general formula (RE)(SCN)₃(tpppO)₃ are reported. Conductance measurements in acetonitrile indicate the non-electrolytic nature of the complexes. Infrared absorption spectra evidence that the SCN⁻ ion coordinates through the nitrogen atom (isothiocyanate form) and that tpppO coordinates through the phosphoryl oxygen. X-ray powder patterns suggest the existence of three different crystal forms: (1) La; (2) an isomorphous series including Ce, Nd and Pr; and (3) another isomorphous series, including Sm, Gd, Eu, Ho, Er, Tb, Lu and Y. The visible spectra of the Nd adduct and the calculated parameters β = 0.98, b^1/2 = 0.072 and δ = 1.06 indicate that the metal-ligand bonds are essentially electrostactic. The emission spectra of the Eu compound showed ⁵D₀ → ⁷F_J bands (J = 0, 1, 2), suggesting a C_3v symmetry for the coordination polyhedron. The lifetime of the ⁵D₀ state is 1.28 ms. The emission spectra of the Tb complex presented ⁵D₄ → ⁷F_J bands (J = 4, 5, 6) and the Dy complex showed the ⁴F_9/2 → ⁶H_13/2 band. The structure of the Pr complex showed that the coordination polyhedron is a trigonal antiprism, with the isothiocyanate anions in one base and three tpppO ligands in the other. Thermal analyses (TG-DTG) were carried out for the Ce, Nd and Gd adducts. Mass losses start between 250 and 334 °C. The final residues at 1300 °C are the corresponding phosphates.     

Focus: Addiction: Relapse Prevention and the Five Rules of Recovery

There are four main ideas in relapse prevention. First, relapse is a gradual process with distinct stages. The goal of treatment is to help individuals recognize the early stages, in which the chances of success are greatest. Second, recovery is a process of personal growth with developmental milestones. Each stage of recovery has its own risks of relapse. Third, the main tools of relapse prevention are cognitive therapy and mind-body relaxation, which are used to develop healthy coping skills. Fourth, most relapses can be explained in terms of a few basic rules. Educating clients in these rules can help them focus on what is important: 1) change your life (recovery involves creating a new life where it is easier to not use); 2) be completely honest; 3) ask for help; 4) practice self-care; and 5) don’t bend the rules.     

Familial breast/ovarian cancer and BRCA1/2 genetic screening: the role of immunohistochemistry as an additional method in the selection of patients.

Only 20-25% of families screened for BRCA1/2 mutations are found positive. Because only a positive result is informative, we studied the role of BRCA1/2 immunohistochemistry as an additional method for patient selection. From 53 high-risk-affected probands, 18 (34%) had available paraffin blocks of their tumors and were selected for this study. Mutation screening was done by conformation-sensitive gel electrophoresis and multiplex ligation-dependent probe amplification. For immunohistochemistry, 21 neoplastic specimens (15 breast carcinomas, 5 ovary neoplasms, and 1 rectal adenocarcinoma) were analyzed with BRCA1 (monoclonal antibody, Ab-1, oncogene) and BRCA2 (polyclonal antibody, Ab-2, oncogene) antibodies. Absence of the BRCA1 protein was confirmed in negative tumors by Western blotting. Seven patients were positive for BRCA1/2 mutations: 5 for BRCA1 and 2 for BRCA2. Four out of five positive patients had tumors negative for BRCA1 immunostaining, and the remaining 13 BRCA1-negative patients had positive BRCA1 immunostaining in all tumor samples. Sensitivity to predict for BRCA1 mutation carriers was 80%, and specificity was 100%, with a positive predictive value of 100% and a negative predictive value of 93%. This correlation was statistically significant (p=0.001). No correlation was observed for BRCA2. If larger studies confirm these results, high-risk patients with BRCA1-negative tumors should be screened first for this gene.     

Analysis of DNA adducts by accelerator mass spectrometry in human breast tissue after administration of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and benzo[a]pyrene

Epidemiological evidence has suggested an association between meat consumption and the risk of breast cancer. 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a heterocyclic amine found in cooked meat, has been implicated in the aetiology of breast cancer and has been shown to induce tumour formation in rodent mammary glands. In addition, polycyclic aromatic hydrocarbons, such as benzo[a]pyrene (B[a]P) which has also been shown to induce tumour formation at a number of sites in rodents including the breast, are produced during the cooking of meat through the pyrolysis of fats. The aim of this study was to examine the bioavailability of these compounds to human breast tissue and their ability to bind to DNA to form DNA adducts. Patients undergoing breast surgery at York District Hospital were orally administered prior to surgery a capsule containing 20 μg of ¹⁴C PhIP (182 kBq, specific activity 2.05 GBq/mmol) or 5 μg of ¹⁴C B[a]P (36 kBq, specific activity 1.81 GBq/mmol). At surgery, normal and tumour breast tissue was resected and tissue concentrations of carcinogen measured by liquid scintillation counting and DNA adduct levels by accelerator mass spectrometry (AMS) were subsequently determined. It was found that both ¹⁴C PhIP and ¹⁴C B[a]P were able to reach the target organ where they had the ability to form DNA adducts. The level of adducts ranged from 26.22–477.35 and 6.61–208.38 adducts/10¹² nucleotides following administration of ¹⁴C PhIP and ¹⁴C B[a]P, respectively, with no significant difference observed between levels in normal or tumour tissue. In addition, the data obtained in this study were comparable to adduct levels previously found in colon samples following administration of the same compounds to individuals undergoing colorectal surgery. This is the first report that these two carcinogens bind to human breast DNA after administration of a defined low dose.     

An Ecteola-cellulose chromatography assay for 3′-phosphoadenosine 5′-phosphosulfate: Phenol sulfotransferase

A new assay procedure for phenol sulfotransferase which employs [35S]-3'-phosphoadenosine-5'-phosphosulfate as a sulfate donor and a variety of phenols as sulfate acceptors was developed. The appearance of the 35S-sulfated products or the disappearance of the [35S]-3'-phosphoadenosine-5'-phosphosulfate are determined simultaneously by chromatography of the assay incubation mixtures on Ecteola-cellulose columns, eluting with an NH4HCO3 step gradient. Various acidic, neutral, and basic phenols can be employed as substrates for phenol sulfotransferase using this procedure.     

Providing straw to allow exploratory behaviour in a pig experimental system does not modify putative indicators of positive welfare: peripheral oxytocin and serotonin

Numerous studies have shown that providing straw to pigs can reduce undesirable behaviours such as aggression, tail biting and stereotypy. The measurement of various neuromodulators can be helpful in assessing the development of positive behaviours and overall animal welfare. The oxytocin release is frequently linked to positive emotions and positive welfare. It has been suggested that oxytocin modulates the serotoninergic system. This study aims to investigate the potential effect of straw provision in pigs on peripheral levels of oxytocin and serotonin. In total, 18 mini-pigs were involved in an exploratory study conducted in two parallel groups, Enriched (n=10) and Control (n=8) groups. Pigs were divided by group and housed in pens of two individuals. Straw was provided continuously only in Enriched group and renewed each day for 2 weeks. Two blood samples were drawn from each animal 5 to 10 min before providing the straw, and 15 min after providing straw, during the 1^st week, to analyse peripheral changes in oxytocin and serotonin before and after straw provision, and determine the existence of a putative short-term effect. The same procedure was carried out for Control group, without straw provision. Long-term effects of straw provision were also examined using blood samples drawn at the same hour from each animal in the 2^nd and 3^rd weeks. During this time, animals had the permanent possibility to explore the straw in Enriched group but not in Control group. At the end of each week, one animal-keeper completed two visual analogue scales for each mini-pig regarding the difficulty/ease to work with and handle it and its trust in humans. Results showed peripheral oxytocin increases in both groups after 2 weeks (P=0.02). Results did not demonstrate any effect of providing straw to allow exploratory behaviour on peripheral serotonin. Other results were not significant. This preliminary study explored the relationship between peripheral oxytocin and serotonin and the presence of straw that allow pigs to perform exploratory behaviour, suggesting that there was no relationship between them. Some future studies may include crossing oxytocin and serotonin with other parameters, such as behavioural measures, to obtain more information about the true state of the animal and any possible relationship with pig welfare.