Settimio Grimaldi

Effect of extremely low frequency ELF magnetic field exposure on morphological and biophysical properties of human lymphoid cell line / Raji

Human B lymphoid cells (Raji) were exposed for 72 h to a 50 Hz sinusoidal magnetic field at a density of 2 milliTesla (rms). The results of exposure showed a decrease in membrane fluidity as detected by Laurdan emission spectroscopy and DPH fluorescence polarization. Field exposure also resulted in a reorganization of cytoskeletal components. Scanning electron microscopy (SEM) revealed a loss of microvilli in the exposed cells. This change in plasma membrane morphology was accompanied by a different actin distribution, as detected by phalloidin fluorescence. We also present evidence that EMF exposure of Raji cells can interfere with protein phosphorylation. Our observations confirm the hypothesis that electric and magnetic fields may modify the plasma membrane structure and interfere with the initiation of the signal cascade pathways.

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.

Newly Engineered Magnetic Erythrocytes for Sustained and Targeted Delivery of Anti-Cancer Therapeutic Compounds

Cytotoxic chemotherapy of cancer is limited by serious, sometimes life-threatening, side effects that arise from toxicities to sensitive normal cells because the therapies are not selective for malignant cells. So how can they be selectively improved? Alternative pharmaceutical formulations of anti-cancer agents have been investigated in order to improve conventional chemotherapy treatment. These formulations are associated with problems like severe toxic side effects on healthy organs, drug resistance and limited access of the drug to the tumor sites suggested the need to focus on site-specific controlled drug delivery systems. In response to these concerns, we have developed a new drug delivery system based on magnetic erythrocytes engineered with a viral spike fusion protein. This new erythrocyte-based drug delivery system has the potential for magnetic-controlled site-specific localization and highly efficient fusion capability with the targeted cells. Here we show that the erythro-magneto-HA virosomes drug delivery system is able to attach and fuse with the target cells and to efficiently release therapeutic compounds inside the cells. The efficacy of the anti-cancer drug employed is increased and the dose required is 10 time less than that needed with conventional therapy.

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.

A combined synthetic-fibrin scaffold supports growth and cardiomyogenic commitment of human placental derived stem cells.

Aims A potential therapy for myocardial infarction is to deliver isolated stem cells to the infarcted site. A key issue with this therapy is to have at ones disposal a suitable cell delivery system which, besides being able to support cell proliferation and differentiation, may also provide handling and elastic properties which do not affect cardiac contractile function. In this study an elastic scaffold, obtained combining a poly(ether)urethane-polydimethylsiloxane (PEtU-PDMS) semi-interpenetrating polymeric network (s-IPN) with fibrin, was used as a substrate for in vitro studies of human amniotic mesenchymal stromal cells (hAMSC) growth and differentiation. Methodology/Principal Findings After hAMSC seeding on the fibrin side of the scaffold, cell metabolic activity and proliferation were evaluated by WST-1 and bromodeoxyuridine assays. Morphological changes and mRNAs expression for cardiac differentiation markers in the hAMSCs were examined using immunofluorescence and RT-PCR analysis. The beginning of cardiomyogenic commitment of hAMSCs grown on the scaffold was induced, for the first time in this cell population, by a nitric oxide (NO) treatment. Following NO treatment hAMSCs show morphological changes, an increase of the messenger cardiac differentiation markers [troponin I (TnI) and NK2 transcription factor related locus 5 (Nkx2.5)] and a modulation of the endothelial markers [vascular endothelial growth factor (VEGF) and kinase insert domain receptor (KDR)]. Conclusions/Significance The results of this study suggest that the s-IPN PEtU-PDMS/fibrin combined scaffold allows a better proliferation and metabolic activity of hAMSCs cultured up to 14 days, compared to the ones grown on plastic dishes. In addition, the combined scaffold sustains the beginning of hAMSCs differentiation process towards a cardiomyogenic lineage.