Antonella Lisi

Differentiation of human adult cardiac stem cells exposed to extremely low-frequency electromagnetic fields

AIMS Modulation of cardiac stem cell (CSC) differentiation with minimal manipulation is one of the main goals of clinical applicability of cell therapy for heart failure. CSCs, obtained from human myocardial bioptic specimens and grown as cardiospheres (CSps) and cardiosphere-derived cells (CDCs), can engraft and partially regenerate the infarcted myocardium, as previously described. In this paper we assessed the hypothesis that exposure of CSps and CDCs to extremely low-frequency electromagnetic fields (ELF-EMFs), tuned at Ca2+ ion cyclotron energy resonance (Ca2+-ICR), may drive their differentiation towards a cardiac-specific phenotype. METHODS AND RESULTS A significant increase in the expression of cardiac markers was observed after 5 days of exposure to Ca2+-ICR in both human CSps and CDCs, as evidenced at transcriptional, translational, and phenotypical levels. Ca2+ mobilization among intracellular storages was observed and confirmed by compartmentalized analysis of Ca2+ fluorescent probes. CONCLUSIONS These results suggest that ELF-EMFs tuned at Ca2+-ICR could be used to drive cardiac-specific differentiation in adult cardiac progenitor cells without any pharmacological or genetic manipulation of the cells that will be used for therapeutic purposes.

Three dimensional (3D) analysis of the morphological changes induced by 50 Hz magnetic field exposure on human lymphoblastoid cells (Raji).

Human Raji B lymphoid cells after exposure for 64 h to a 1 mT (rms) 50 Hz sinusoidal magnetic field showed a reorganization of membrane and cytoskeletal components. Atomic force microscopy in air revealed several modifications in 80% of the exposed cells, such as loss of microvilli-like structures followed by progressive appearance of membrane introflections. This change in plasma membrane morphology was also accompanied by a different actin distribution, as detected by phalloidin fluorescence. These observations support our previous hypothesis that electric and magnetic fields may modify the plasma membrane structure.

Action of combined magnetic fields on aqueous solution of glutamic acid: the further development of investigations

In the present work the results of the known investigation of the influence of combined static (40 μT) and alternating (amplitude of 40 nT) parallel magnetic fields on the current through the aqueous solution of glutamic acid, were successfully replicated. Fourteen experiments were carried out by the application of the combined magnetic fields to the solution placed into a Plexiglas reaction vessel at application of static voltage to golden electrodes placed into the solution. Six experiments were carried out by the application of the combined magnetic fields to the solution placed in a Plexiglas reaction vessel, without electrodes, within an electric field, generated by means of a capacitor at the voltage of 27 mV. The frequency of the alternating field was scanned within the bounds of 1.0 Hz including the cyclotron frequency corresponding to a glutamic acid ion and to the applied static magnetic field. In this study the prominent peaks with half-width of ~0.5 Hz and with different heights (till 80 nA) were registered at the alternating magnetic field frequency equal to the cyclotron frequency (4.2 Hz). The general reproducibility of the investigated effects was 70% among the all solutions studied by us and they arose usually after 40–60 min. after preparation of the solution. In some made-up solutions the appearance of instability in the registered current was noted in 30–45 min after the solution preparation. This instability endured for 20–40 min. At the end of such instability period the effects of combined fields action appeared practically every time. The possible mechanisms of revealed effects were discussed on the basis of modern quantum electrodynamics.

Bioelectromagnetic medicine: The role of resonance signaling

Only recently has the critical importance of electromagnetic (EM) field interactions in biology and medicine been recognized. We review the phenomenon of resonance signaling, discussing how specific frequencies modulate cellular function to restore or maintain health. The application of EM-tuned signals represents more than merely a new tool in information medicine. It can also be viewed in the larger context of EM medicine, the all-encompassing view that elevates the EM over the biochemical. The discovery by Zhadin that ultrasmall magnetic intensities are biologically significant suggests that EM signaling is endogenous to cell regulation, and consequently that the remarkable effectiveness of EM resonance treatments reflects a fundamental aspect of biological systems. The concept that organisms contain mechanisms for generating biologically useful electric signals is not new, dating back to the nineteenth century discovery of currents of injury by Matteucci. The corresponding modern-day version is that ion cyclotron resonance magnetic field combinations help regulate biological information. The next advance in medicine will be to discern and apply those EM signaling parameters acting to promote wellness, with decreasing reliance on marginal biochemical remediation and pharmaceuticals.

Extremely Low Frequency 7 Hz 100 µT Electromagnetic Radiation Promotes Differentiation in the Human Epithelial Cell Line HaCaT

Electromagnetic therapy is a treatment method in which an electromagnetic or magnetic stimulus is used to achieve physiological changes in the body. The specific aim of the present work concerns the effectiveness of low frequency electromagnetic fields to modify the biochemical properties of human keratinocytes (HaCaT). Cells exposed to a 7 Hz 100 μT electromagnetic field for one hour (twice daily), indicated modification in shape and morphology. These modifications were also associated with different actin distribution as revealed by phalloidin fluorescence analysis. Indirect immunofluorescence with fluorescent antibodies against involucrin and β-Catenin, both differentiation and adhesion markers, revealed an increase in involucrin and β-Catenin expression, supporting the conclusion that exposure to electromagnetic field carries keratinocytes to an upper differentiation level. This study confirms our previous observation and supports the hypothesis that 7 Hz electromagnetic field, may modify cell biochemistry interfering in the differentiation and cellular adhesion of normal keratinocytes.

Ion Cyclotron Resonance as a Tool in Regenerative Medicine

The identification of suitable stem cell cultures and differentiating conditions that are free of xenogenic growth supplements is an important step in finding the clinical applicability of cell therapy in two important fields of human medicine: heart failure and bone remodeling, growth and repair. We recently demonstrated the possibility of obtaining cardiac stem cells (CSCs) from human endomyocardial biopsy specimens. CSCs self-assemble into multi-cellular clusters known as cardiospheres (CSps) that engraft and partially regenerate infarcted myocardium. CSps and cardiosphere-derived-cells (CDCs) were exposed for five days in an incubator regulated for temperature, humidity, and CO2 inside a solenoid system. This system was placed in a magnetically shielded room. The cells were exposed simultaneously to a static magnetic field (MF) and a parallel low-alternating frequency MF, close to the cyclotron frequency corresponding to the charge/mass ratio of the Ca++ ion. In this exposure condition, CSps and CDCs modulate their differentiation turning on cardiogenesis and turning off vasculogenesis. Cardiac markers such as troponin I (TnI) and myosin heavy chain (MHC) were up-regulated. Conversely, angiogenic markers such as vascular endothelial growth factor (VEGF) and kinase domain receptor (KDR) were down-regulated as evidenced by immunocytochemistry. Exposure to the 7 Hz calcium ion cyclotron resonance (ICR) frequency can modulate the cardiogenic vs. angiogenic differentiation process of ex vivo expanded CSCs. This may pave the way for novel approaches in tissue engineering and cell therapy. With regard to bone remodeling, it has been suggested that bone marrow-derived mesenchymal stem cells (MSC) may be considered as a potential therapeutic tool. Using the Ca++-dependent specific differentiation potential of the ELF-MF 7 Hz ICR, we show here that exposure of human MSC to these same MF conditions enhanced the expression of osteoblast differentiation markers such as alkaline phosphatase, osteocalcin, and osteopontin, as analyzed by real-time quantitative PCR, without affecting cell proliferation. As expected, while the differentiation marker factors were up regulated, the ICR electromagnetic field down regulated osteoprotegerin gene expression, a critical regulator of postnatal skeletal development and homeostasis in humans as well as mice.

Attempts to use liposomes and RBC ghosts as vectors in drug and antisense therapy of virus infection

Selective targeting of drugs or oligonucleotide for the treatment of viral diseases or cancer is the objective of new strategies that pursue therapy optimization and reduction of toxicity. In this work we report two protocols based on encapsulation of anti-human immunodeficiency virus drugs within targeted liposomes or erythrocytes. Both have been shown to be effective for the specific delivery of drugs or oligonucleotide in the treatment of viral infection.

Cellular ELF Signals as a Possible Tool in Informative Medicine

According to Quantum Electro-Dynamical Theory by G. Preparata, liquid water can be viewed as an equilibrium between of two components: coherent and incoherent ones. The coherent component is contained within spherical so called “coherence domains” (CDs) where all molecules synchronously oscillate with the same phase. CDs are surrounded by the incoherent component where molecules oscillate with casual phases regarding each other. The existence of coherent domain in water has been demonstrated in a set of experiments on pure water exposed to high voltage, under this condition the electric field concentrates inside the water, arranging the water molecules to form high ordered structure. Recently has been studied the influence of combined static and alternating parallel magnetic fields on the current through the aqueous solution of glutamic acid; outlining the relevance of low frequency electro-magnetic field in interacting with biological target. Additional results demonstrate that at combined static and alternating parallel, magnetic fields matching the ion cyclotron energy resonance of a particular charged molecule into biological tissue an intrinsic weak magnetic field is generated by ion currents in the cell. These results should increase the reliability and the clinical feasibility of the use of electromagnetic field, tuned at ion cyclotron resonance of charged molecules, as a biophysical approach to interfere with biological mechanisms. We demonstrate that Exposure of human epithelial cell to ion cyclotron energy resonance generated by a commercial electromedical device (Vega select 719) tuned to calcium ion at 7 Hz act as a differentiation factor, thus opening up the possibility to use particular extremely low frequency electro magnetic field protocols, in informative medicine.

Engineered liposomes and virosomes for delivery of macromolecules

In order to utilize virosomes or proteoliposomes for the delivery of drugs or macromolecules to specific pathologic target cells we elaborated a system to shuttle drugs to solid tissue (liver) as well as to the macrophages, a crucial cellular compartment of the immune system. Using virosomes prepared from the P3HR1 strain of Epstein-Barr virus, we demonstrated that these particles fused with human hepatocarcinoma cell line Li7A and therefore might be used as drug vectors. Furthermore, we report that proteoliposomes prepared by reconstituting in a cocktail of phosphatidylserine-phosphatidylcholine the anion transporter band 3 protein markedly increased the phagocytic activity of macrophages in culture. This could represent a new device to be used as a drug delivery system to enhance specific macrophagic functions.

Experimental Finding on the Electromagnetic Information Transfer of Specific Molecular Signals Mediated Through the Aqueous System on Two Human Cellular Models

BACKGROUND Recently the authors reported the experimental evidence of the developing concept of Electro Magnetic Information Transfer (EMIT) of specific molecular signals directly and continuously on target cell picking up the molecular signals from the source chemical effector. This was in agreement with the pioneering work of Jaques Benveniste suggesting that the electronic transmission of the 4-phorbol-12-myristate-13-acetate (PMA) signals could be transferred to target neutrophils by an oscillator when coupled to two electromagnetic coils demonstrating the same biologic activity and so mimicking the biologic function of the original chemical active molecule. The present work is the further development of recent research designed to verify the hypotheses that water could record and replay the EMIT from biologic active chemical molecules. METHODS Retinoic acid, a well-known chemical differentiating agent, was placed at room temperature in the input coil connected to an oscillator (VEGA select 719), while culture medium for human neuroblastoma cell (LAN-5) and NT2/D1 stem teratocarcinoma human cells was placed into the output coil and exposed to signals for 1 hour. At the end the oscillator was switched off and LAN-5 neuroblastoma and NT2/D1 stem teratocarcinoma cells were seeded, respectively, into the medium conditioned as reported into an incubator under controlled conditions. After 5 days of incubations, cells were examined by different strategies such as morphological and biochemical parameters. RESULTS It was demonstrated that the electromagnetic signals coming from the retinoic acid molecule could be recorded and stored by the aqueous system of the cell culture medium. Cells seeded in the electronically conditioned medium received physical information generating a statistically significant decrease in metabolic activity and changes in phenotypical structure with protrusion typical of differentiated neuronal cells. CONCLUSIONS These experimental results provide some evidence that water could be tuned in a resonant manner by the EMIT procedure appropriately carried through a carrier frequency provided by the oscillator in a manner that seems related to the chemical structure of the source molecule as, in this case, retinoic acid.