Robert M. Lafrenie

Biophoton emissions from cell cultures: biochemical evidence for the plasma membrane as the primary source.

Photon emissions were measured at ambient temperature (21°C) in complete darkness once per min from cultures of 10(6) cells during the 12 h following removal from 37°C. The energy of emission was about 10(-20) J/s/cell. Of 8 different cell lines, B16-BL6 (mouse melanoma cells) demonstrated the most conspicuous emission profile. Acridine orange and ethidium bromide indicated the membranes were intact with no indication of (trypan blue) cell necrosis. Treatments with EGF and ionomycin produced rapid early (first 3 h) increases in energy emission while glutamine-free, sodium azide and wortmanin-treated cells showed a general diminishment 3 to 9 h later. The results suggested the most probable origin of the photon emission was the plasma cell membrane. Measures from cells synchronized at the M- and S-phase supported this inference.

Chloroquine-Mediated Radiosensitization is due to the Destabilization of the Lysosomal Membrane and Subsequent Induction of Cell Death by Necrosis

Abstract Zhao, H., Cai, Y., Santi, S., Lafrenie, R. and Lee, H. Chloroquine-Mediated Radiosensitization is due to the Destabilization of the Lysosomal Membrane and Subsequent Induction of Cell Death by Necrosis. Radiat. Res. 164, 250–257 (2005). The anti-malarial drug chloroquine (CQ) is also thought to be a potential radiation sensitizer. To gain a better understanding of how the lysomotropic CQ can potentiate the effects of ionizing radiation, we investigated the effects of CQ on lysosomal and mitochondrial membrane stability, the subcellular localization of ceramide, plasma membrane permeability, and the mode of cell death in response to irradiation. We found that CQ accumulated in the lysosomes and thus lysosomal volumes increased. As a result, both the lysosomal and plasma membranes were destabilized. After 7 Gy irradiation, most ceramide was associated with the lysosomes in the cells treated with CQ but not in the CQ-untreated control. The elevated levels of ceramide in the lysosomes of the CQ-treated cells appeared to further destabilize the lysosomal and plasma membranes of the cell. Both CQ-treated and -untreated cells had approximately the same rate of cell death by apoptosis after 7 Gy irradiation (P > 0.05, ns). However, in contrast to the CQ-untreated control, the CQ-treated cells underwent massive cell death by necrosis at 24–48 h after irradiation (P < 0.05). Taken together, our data support the idea that the increase in cytotoxic effects by the combination of CQ and radiation is due to radiation-mediated apoptosis and CQ-mediated necrosis.

Photon emissions from human brain and cell culture exposed to distally rotating magnetic fields shared by separate light-stimulated brains and cells

Light flashes delivered to one aggregate of cells evoked increased photon emission in another aggregate of cells maintained in the dark in another room if both aggregates shared the same temporospatial configuration of changing rate, circular magnetic fields. During the presentation of the same shared circumcerebral magnetic fields increases in photon emission occurred beside the heads of human volunteers if others in another room saw light flashes. Both cellular and human photon emissions during the light flashes did not occur when the shared magnetic fields were not present. The summed energy emissions from the dark location during light stimulation to others was about 10(-11) W/m(2) and calculated to be in the order of 10(-20) J per cell which is coupled to membrane function. These results support accumulating data that under specific conditions changes in photon emissions may reflect intercellular and interbrain communications with potential quantum-like properties.

Attenuation of Pseudomonas aeruginosa virulence factors and biofilms by co-encapsulation of bismuth–ethanedithiol with tobramycin in liposomes

OBJECTIVES This study examined the activities of tobramycin and bismuth against quorum sensing, virulence factors and biofilms of Pseudomonas aeruginosa by co-encapsulating the agents in liposomes in order to achieve greater delivery of the agents. METHODS The inhibitory effects of the agents, in either their conventional (free) or vesicle-entrapped (liposomal) formulations, were assessed by measuring the changes in the quorum-sensing signal molecule N-acyl homoserine lactone, pyoverdine, pyocyanin, elastase, protease, chitinase, bacterial attachment and biofilms in vitro. RESULTS The effectiveness of tobramycin and bismuth was superior when they were co-administered as a liposomal formulation as measured by their ability to attenuate the production of N-acyl homoserine lactone, elastase (P < 0.01), protease (P < 0.05) and chitinase (P < 0.01). In the presence of non-lethal concentrations of free and liposomal tobramycin and bismuth, bacterial attachment was attenuated. Biofilm formation was also attenuated with free tobramycin and bismuth, yet, in the presence of liposomal tobramycin and bismuth, biofilm complexes could form but contained mostly dead bacteria. When established biofilms were treated with higher concentrations, free tobramycin and bismuth killed and detached bacteria, while the liposomal tobramycin and bismuth penetrated and killed bacteria in the cores of the biofilms. CONCLUSIONS These data suggest that treatment of P. aeruginosa with tobramycin and bismuth, as measured by the changes in quorum sensing, virulence factors and biofilms, is most effective when delivered as a liposomal formulation at a lower concentration compared with the free formulation.

Uncaria tomentosa acts as a potent TNF-α inhibitor through NF-κB

AIM OF THE STUDY Uncaria tomentosa, commonly known as Cats Claw or Uña de gato, is a medicinal plant that has been shown to have effective anti-inflammatory activities. We have previously shown that treatment of monocyte-like THP-1 cells with Uncaria tomentosa inhibits the production of the pro-inflammatory cytokine TNF-alpha while augmenting the production of IL-1beta. Since TNF-alpha and IL-1beta are usually regulated similarly and share a number of common promoter elements, including NF-kappaB and AP-1, the ability of Uncaria tomentosa to differentially regulate these inflammatory cytokines is of particular interest. MATERIALS AND METHODS To determine the mechanism of action of Uncaria tomentosa, we investigated the effects of specific inhibitors of NF-kappaB on cellular responses including transcription factor activation using TransAM assays, the expression of cytokines as measured by ELISA, and cell survival as measured by changes in cell number following treatment. RESULTS Treatment with Uncaria tomentosa inhibited the LPS-dependent activation of specific NF-kappaB and AP-1 components. In addition, treatment with Uncaria tomentosa enhanced cell death when NF-kappaB was inhibited. The ability of Uncaria tomentosa to inhibit TNF-alpha production was diminished when NF-kappaB activation was prevented by drugs that mask NF-kappaB subunit nuclear localization signals, while IL-1beta expression was unchanged. CONCLUSIONS These results demonstrate that Uncaria tomentosa is able to elicit a response via an NF-kappaB-dependent mechanism. Further studies to characterize the mechanism by which Uncaria tomentosa can affect this pathway could provide a means to develop anti-TNF-alpha therapies.

Inhibition of Cancer Cell Growth by Exposure to a Specific Time-Varying Electromagnetic Field Involves T-Type Calcium Channels

Electromagnetic field (EMF) exposures affect many biological systems. The reproducibility of these effects is related to the intensity, duration, frequency, and pattern of the EMF. We have shown that exposure to a specific time-varying EMF can inhibit the growth of malignant cells. Thomas-EMF is a low-intensity, frequency-modulated (25-6 Hz) EMF pattern. Daily, 1 h, exposures to Thomas-EMF inhibited the growth of malignant cell lines including B16-BL6, MDA-MB-231, MCF-7, and HeLa cells but did not affect the growth of non-malignant cells. Thomas-EMF also inhibited B16-BL6 cell proliferation in vivo. B16-BL6 cells implanted in syngeneic C57b mice and exposed daily to Thomas-EMF produced smaller tumours than in sham-treated controls. In vitro studies showed that exposure of malignant cells to Thomas-EMF for > 15 min promoted Ca2+ influx which could be blocked by inhibitors of voltage-gated T-type Ca2+ channels. Blocking Ca2+ uptake also blocked Thomas-EMF-dependent inhibition of cell proliferation. Exposure to Thomas-EMF delayed cell cycle progression and altered cyclin expression consistent with the decrease in cell proliferation. Non-malignant cells did not show any EMF-dependent changes in Ca2+ influx or cell growth. These data confirm that exposure to a specific EMF pattern can affect cellular processes and that exposure to Thomas-EMF may provide a potential anti-cancer therapy.

Growth of injected melanoma cells is suppressed by whole body exposure to specific spatial-temporal configurations of weak intensity magnetic fields

Purpose: To measure the effect of exposure to a specific spatial-temporal, hysiologically-patterned electromagnetic field presented using different geometric configurations on the growth of experimental tumours in mice. Methods: C57b male mice were inoculated subcutaneously with B16-BL6 melanoma cells in two blocks of experiments separated by six months (to control for the effects of geomagnetic field). The mice were exposed to the same time-varying electromagnetic field nightly for 3 h in one of six spatial configurations or two control conditions and tumour growth assessed. Results: Mice exposed to the field that was rotated through the three spatial dimensions and through all three planes every 2 sec did not grow tumours after 38 days. However, the mice in the sham-field and reference controls showed massive tumours after 38 days. Tumour growth was also affected by the intensity of the field, with mice exposed to a weak intensity field (1–5 nT) forming smaller tumours than mice exposed to sham or stronger, high intensity (2–5 μT) fields. Immunochemistry of tumours from those mice exposed to the different intensity fields suggested that alterations in leukocyte infiltration or vascularisation could contribute to the differences in tumour growth. Conclusions: Exposure to specific spatial-temporal regulated electromagnetic field configurations had potent effects on the growth of experimental tumours in mice.

Digitized quantitative electroencephalographic patterns applied as magnetic fields inhibit melanoma cell proliferation in culture

Weak (1 μT) physiologically patterned magnetic fields produce changes in behavioral, physiological, and cellular activity. In the present experiments 12 temporal samples of the electroencephalographic anomaly and normal activity of a person (SLH) whose proximity reliably affected the brain activity of others were extracted from QEEG data, digitized, and presented as equivalent magnetic field patterns to B16 mouse melanoma cells. Only two of the patterns, both originating from the primary source (right temporal lobe) of the EEG anomaly reduced the cell growth by one-third compared to the other patterns extracted from his QEEG or sham field exposures. In previous experiments these EEG transients were also associated with marked increases in photon emissions from the right side of SLHs head. The results suggest that the intrinsic complexity of electroencephalographic patterns of some people, when amplified appropriately and applied as computer-generated magnetic fields in the three spatial planes, could diminish cancer cell growth.

Temporally-Patterned Magnetic Fields Induce Complete Fragmentation in Planaria

A tandem sequence composed of weak temporally-patterned magnetic fields was discovered that produced 100% dissolution of planarian in their home environment. After five consecutive days of 6.5 hr exposure to a frequency-modulated magnetic field (0.1 to 2 µT), immediately followed by an additional 6.5 hr exposure on the fifth day, to another complex field (0.5 to 5 µT) with exponentially increasing spectral power 100% of planarian dissolved within 24 hr. Reversal of the sequence of the fields or presentation of only one pattern for the same duration did not produce this effect. Direct video evidence showed expansion (by visual estimation ∼twice normal volume) of the planarian following the first field pattern followed by size reduction (estimated ∼1/2 of normal volume) and death upon activation of the second pattern. The contortions displayed by the planarian during the last field exposure suggest effects on contractile proteins and alterations in the cell membrane’s permeability to water.

Intermittent exposures to nanoTesla range, 7 Hz, amplitude-modulated magnetic fields increase regeneration rates in planarian

Abstract Purpose: To discern if physiologically and naturally-patterned electromagnetic fields presented with base frequencies of 7 Hz within the 100 nT range could facilitate regeneration in planarian similar to microTesla, 60 Hz fields. Methods: In two separate experiments planarian were decapitated and exposed to either 140 or 400 nT peak amplitude-modulated 7 Hz magnetic fields for 6 min once per hour, 8 h per night for 5 days. Daily regeneration rates and movement velocities (cm/min) were measured. Results: The planarian exposed to either intensity magnetic field exhibited faster regeneration of photoreceptors and auricles compared to sham field and reference groups. The magnetic field exposure accommodated 50% of the variance during the faster growth days. Conclusions: Naturally-patterned, intermittently-presented weaker electromagnetic fields may produce enhanced regeneration rates in flat worms similar to those observed for 60 Hz, higher intensity fields.