Patterns of calcium signaling: A link between chronic emotions and cancer.

Intra and inter-cellular calcium signaling is present in all types of cells and body tissues. In the human brain, calcium currents and waves are related to mental activities, including emotions. We present a theoretical interpretation of these phenomena suggesting their involvement in chronic emotional patterns and in the pathology of cancer. Recent developments on biophysics, translational biology and psychoneuroendocrinoimmunology (PNEI) can support explanatory hypotheses about the link between emotional stresses and the origin and development of different types of tumor cells. Chronic stresses may cause perturbations of rhythms of the PNEI system, excessive activation of HPA axis and abnormal activation of calcium signals in somatic tissues, with deleterious effects on different parts of the body. The increasing of calcium signaling inside cells may lead to a deregulation of different pathways and epigenetic systems that promote the production of genomic mutations in a second phase. In particular, the hyperactivation of the transcription nuclear factor kappaB (NF-κB), if is not counterbalanced by the following activation of the nuclear factor (erythroid-derived 2)-like 2 (NFE2L2 or Nrf2), increases the production of oxidative catabolites, as the advanced glycation end products (AGE), which play a key role in the progression of different types of cancer and other degenerative diseases. Cortisol binding to glucocorticoid receptor (GR) reduces the activity of both NF-κB and Nrf2 inside the cells but inhibits the cellular immunity and the anabolic processes of tissue regeneration. The tissue atrophy and the defective anti-ageing mechanisms promotes the tumoral cells growth and their escape from the immune-surveillance.

Corrections to chance fluctuations: quantum mind in biological evolution?

According to neo-Darwinian theory, biological evolution is produced by natural selection of random hereditary variations. This assumption stems from the idea of a mechanical and deterministic world based on the laws of classic physics. However, the increased knowledge of relationships between metabolism, epigenetic systems, and editing of nucleic acids suggests the existence of self-organized processes of adaptive evolution in response to environmental stresses. Living organisms are open thermodynamic systems which use entropic decay of external source of electromagnetic energy to increase their internal dynamic order and to generate new genetic and epigenetic information with a high degree of coherency and teleonomic creativity. Sensing, information processing, and decision making of biological systems might be mainly quantum phenomena. Amplification of microscopic quantum events using the long-range correlation of fractal structures, at the borderline between deterministic order and unpredictable chaos, may be used to direct a reproducible transition of the biological systems towards a defined macroscopic state. The discoveries of many natural genetic engineering systems, the ability to choose the most effective solutions, and the emergence of complex forms of consciousness at different levels confirm the importance of mind-action directed processes in biological evolution, as suggested by Alfred Russel Wallace. Although the main Darwinian principles will remain a crucial component of our understanding of evolution, a radical rethinking of the conceptual structure of the neo-Darwinian theory is needed.

The Bov-A2 retroelement played a crucial role in the evolution of ruminants.

The Bov-A2 is one of the most common short interspersed nucleotide elements (SINEs) in ruminants. The genomic distribution and evolution of this retroelement were analysed in order to highlight its possible functional role. Several regions containing an entire Bov-A2 were amplified and polymorphisms were identified by direct sequencing of the amplification products. The obtained sequences were used together with entire Bov-A2 sequences of the public database to analyse their evolutionary pathway. A site-specific micro-recombination followed by gene conversion or unequal crossing-over might be responsible for the high amount of genetic variation of Bov-A2 sequences. Short cDNAs copied by the reverse transcriptase might be the donor sequences for the micro-recombination, according to the RT-mutatorsome mechanism of the somatic hyper-mutation (SHM) process in the hypervariable regions of immunoglobulin and major histocompatibility complex (MHC) genes. The Bov-A2 is generally present in the non-coding regions of several genes preferentially expressed during the cell response to environmental stresses or activation signals. In particular, the presence of Bov-A2 sequence in the 3' untranslated regions of mRNAs involved in cell growth and differentiation during the immune response might be related to an important functional role in the post-transcriptional regulation of gene expression. This hypothesis is supported by the similarity between the conserved "core" sequences motives CACTn (n = 3, 4, 3) of Bov-A2 and elements affecting the messengers stability as several microRNAs (miRNAs). Using primers based on the "core" sequence and bovine, ovine and human cDNAs in RT-PCR experiments, we demonstrate that the mRNAs containing the "core" sequence are present at high levels in lymphocytes only after their activation. Our results suggest the existence of a system based on environmental and epigenetic signals that is able to spread and mutate the Bov-A2 sequence in the genes expressed during the response to cellular activation signals. By means of this adaptive system a reverse flow of information from environment to genes might reinforce and diversify the stress response at cellular and individual levels.

The Yin and Yang of anti-Darwinian epigenetics and Darwinian genetics.

Darwinian theory, like classic physics, is an incomplete oversimplification of the most complex aspects of the reality. According to the complementarity principle of Taoism and quantum physics, a Lotka-Volterra oscillation between catabolic-entropic and anabolic-syntropic processes might generate the emergence of collective behaviours and fractal structures near the critical points in many physical and biological systems. The proposed mechanism, named metabolic hypercycle, explains many evolutionary dynamics with two alternate and complementary phases at the levels of molecules, cells, organisms, and populations. In the catabolic-entropic phase, the genetic apparatus and the Darwinian processes allow the reproduction and conservation of biological systems. In the anabolic-syntropic phase, the epigenetic apparatus and the Lamarckian anti-Darwinian processes produce innovative adaptations to variable environment. Many experimental evidences, such as for example the distribution of retroelements and DNA mutations in complex genomes, suggest the existence of mechanisms based on epigenetic signals, reverse transcription, and micro-recombination, that promote DNA changes and horizontal transfer of genetic information between organisms and cells, in particular between somatic and germline cells. In complex animals, the acquired immunity regulates the selective proliferation of the cells with adaptive mutations during the physiological responses to stresses. The same immunotrophic process allows the maternal selection of embryos with adaptive mutations. Moreover, the fetal-maternal incompatibility counteracts the negative aspects of Darwinian natural selection and maintains the metabolic biodiversity of animal populations. In conclusion, the increased knowledge of relationships among metabolism, epigenetic systems, genome plasticity, and acquired immunity, supports the idea that these processes could provide a form of Lamarckian adaptive evolution in response to environmental stresses. A better understanding of the biology will increase our ability to design medical and biotechnological applications. Some of these innovative applications are discussed in the paper.

Pro-inflammatory variants of DRB1 and RAGE genes are associated with susceptibility to pediatric type 1 diabetes: a new hypothesis on the adaptive role of autoimmunity.

Functional polymorphisms of two MHC genes (DRB1 and RAGE) were analysed in Italian pediatric patients with Type 1 diabetes and in a control group. The diabetic condition is related positively to the positive electric charge of the pocket 4 of pro-inflammatory DRB1 alleles (R = 0.5072, P = 0.0001) and negatively to at least one anti-inflammatory RAGE allele (R = -0.2200, P = 0.0106). The association DRB1-disease decreases from high risk positively charged alleles to low risk negatively ones. A multiple regression model including the effect of electric charges at positions 70 and 74 of the DRB1 explains more than 31% of the variability of our data. The addition of the RAGE dependent variables does not increase the significance of the model. Our results confirm that the interaction between a negatively charged amino acid of insulin autoantigenic peptides and a positively charged DRB1 is the key event triggering the autoimmune process. The linkage disequilibrium between RAGE and DRB1 is the main cause of the association between the variants of RAGE and the initial outcome of the disease. However, since RAGE ligands increase during the disease progression, the observed association suggests that the proinflammatory RAGE and DRB1 polymorphisms synergize to activate the immune response which leads to the complications of diabetes. These evidences support a new hypothesis that considers the largely unexplored role of the MHC genes in genetic adaptation to a variable environment and in the maintenance of the metabolic biodiversity. A mechanism based on the maternal immunization against the negatively charged autoantigens, such as the insulin peptide B9-23, and on the fetal-maternal interaction might transform the physiological adaptation into adaptive changes of the genetic population structure. According to the "thrifty-genotype" hypothesis, "thrifty DRB alleles" with a positive charge are responsible for the susceptibility to diabetes and for an efficient storage of caloric intake in arctic climates with scarce food availability while "non-thrifty DRB alleles" with a negative or neutral charge are advantaged in tropical climates with constant food supply.

Towards the analysis of the genomes of single cells: further characterisation of the multiple displacement amplification.

The development of methods for the analysis and comparison of the nucleic acids contained in single cells is an ambitious and challenging goal that may provide useful insights in many physiopathological processes. We review here some of the published protocols for the amplification of whole genomes (WGA). We focus on the reaction known as Multiple Displacement Amplification (MDA), which probably represents the most reliable and efficient WGA protocol developed to date. We discuss some recent advances and applications, as well as some modifications to the reaction, which should improve its use and enlarge its range of applicability possibly to degraded genomes, and also to RNA via complementary DNA.

Rearranged genomes of bovine blood cells can allow the development of clones till late fetal stages; but rare unrearranged genomes have greater potential and lead to adulthood.

Cloning via nuclear transfer is promising, but rather inefficient. Moreover, to date, relatively few data are available for a satisfactory phenotypic and genotypic characterization of the clones. Here, we analyze the genomes of clones derived from bovine peripheral blood mononucleated cells (PBMC), known to be composed mainly by lymphocytes. Their genomes are rearranged at either the immunoglobulin (Ig) or the T-cell-receptor (TCR) loci. The DNA of the single survivor and of four aborted fetuses were amplified by semi-quantitative PCR and sequenced. We found the expected rearrangements in DNA from lymphocytes, but neither in DNA from chondrocytes of the survivor, nor in DNA from brain cells of three of the aborted fetuses. This indicates that these four clones derived from somatic cells bearing unrearranged genomes and suggests that in a population of variably differentiated cells those harbouring unrearranged genomes are better donors. Brain cells of the fourth fetus present rearrangements at both loci. The sequences of these rearrangements differ from those obtained from PBMC because they appear unique, thus confirming the clonal origin of the fetus from a cell bearing a rearranged genome. To our knowledge, this is the first example in which both the placenta and the soma of a late fetus are coded for by the rearranged genome of a terminally differentiated cell, unambiguously identified through a specific genetic marker.

Evaluation of gene expression in human lymphocytes activated in the presence of melatonin.

The effect of melatonin on the expression of genes previously correlated to T lymphocyte activation (HLA-DRB, thymosin beta 10 (beta-Tim)) and to Lymphokine Activated Killer (LAK) activity (beta-Tim, Tumour Rejection Antigen (TRA 1), nRap 2) was investigated in phytohemagglutinin (PHA)-stimulated human lymphocyte cultures. The aim was to find an enhancing effect of this substance on anti-tumoral immune defences as suggested by studies on tumour progression in mice and clinical immunotherapy trials in humans. mRNA obtained from melatonin-treated and -untreated PHA-stimulated lymphocytes was retrotranscribed and amplified by RT-PCR using primers based on the sequences of the selected genes. The results suggest that melatonin does not increase T and LAK cell responses: in fact, a reduction in the transcription of all the considered genes was observed. These data are correlated with the antiproliferative effect of melatonin observed in in vitro treated lymphocytes.

The hairless (hr) gene is involved in the congenital hypotrichosis of Valle del Belice sheep.

Congenital hypotrichosis in mammalian species consists of partial or complete absence of hair at birth. The hairless gene is often responsible for this disorder in men, mice and rats. Recent experimental data on Valle del Belice sheep reared in Sicily for milk production, support the genetic control of the ovine hypotrichosis as a Mendelian recessive trait. The ovine hairless gene was chosen as the candidate gene involved in this disorder. Blood samples were collected from Valle del Belice sheep with the normal and hypotrichotic phenotypes. Almost the entire hairless gene was successfully amplified using the long PCR technique. Unrelated sheep with differing phenotypes were randomly chosen for sequencing the amplified products. Different mutations related to the hypotrichotic phenotype were found in exon 3. In fact, sequencing revealed an A/T transversion at position 739, a G/A transition at position 823, and a C/T transition at position 1312. From these nucleotide exchanges, three substitutions of the processed mature protein were deduced at the amino acid positions 247 (Thr/Ser), 275 (Ala/Thr), and 438 (Gln/Stop). A PCR-SSCP based test was developed in order to detect the last mutation, which is responsible for the hypotrichotic phenotype.