Possible humoral mechanism of 2450-MHz microwave-induced increase in complement receptor positive cells

These studies indicate that the increase in the frequency of complement receptor-positive (CR⁺) spleen cells observed 6 days after a 30-min exposure to 2450-MHz microwaves is not the result of microwave-induced alterations of lymphocyte recirculation patterns, but is mediated by a soluble, humoral factor produced by cells within the spleen.     

Interaction of Microwaves and a Temporally Incoherent Magnetic Field on Single and Double DNA Strand Breaks in Rat Brain Cells

The effect of a temporally incoherent magnetic field noise on microwave-induced DNA single and double strand breaks in rat brain cells was investigated. Four treatment groups of rats were studied: microwave-exposure (continuous-wave 2450-MHz microwaves, power density 1 mW/cm², average whole-body specific absorption rate of 0.6 W/kg), noise-exposure (45 mG), microwave + noise-exposure, and sham-exposure. Animals were exposed to these conditions for 2h. DNA single- and double-strand breaks in brain cells of these animals were assayed 4h later using a microgel electrophoresis assay. Results show that brain cells of microwave-exposed rats had significantly higher levels of DNA single- and double-strand breaks when compared with sham-exposed animals. Exposure to noise alone did not significantly affect the levels (i.e., they were similar to those of the sham-exposed rats). However, simultaneous noise exposure blocked microwave-induced increases in DNA strand breaks. These data indicate that simultaneous exposure to a temporally incoherent magnetic field could block microwave-induced DNA damage in brain cells of the rat.     

Microwave radiation (2450 MHz) alters the endotoxin-induced hypothermic response of rats

The parenteral administration of bacterial endotoxin to rats causes a hypothermia that is maximal after approximately 90 minutes. When endotoxin-injected rats were held in a controlled environment at 22°C and 50% relative humidity and exposed for 90 minutes to microwaves (2450 MHz, CW) at 1 mW/cm², significant increases were observed in body temperature compared with endotoxintreated, sham-irradiated rats. The magnitude of the response was related to power density (10 mW/cm² > 5 mW/cm² > 1 mW/cm²). Saline-injected rats exposed for 90 minutes at 5 mW/cm² (specific absorption rate approximately 1.0 mW/g) showed no significant increase in body temperature compared with saline-injected, sham-irradiated rats. The hypothermia induced by endotoxin in rats was also found to be affected by ambient temperature alone. Increases in ambient temperature above 22°C in the absence of microwaves caused a concomitant increase in body temperature. This study reveals that subtle microwave heating is detectable in endotoxin-treated rats that have an impaired thermoregulatory capability. These results indicate that the interpretation of microwave-induced biological effects observed in animals at comparable rates and levels of energy absorption should include a consideration of the thermogenic potential of microwaves.     

Immunological properties of Fc receptor on lymphocytes. 6. Characterization of suppressive B-cell factor (SBF) released from Fc receptor-bearing B cells.

EA, i.e., antigen-antibody complexes are able to induce an antigen-nonspecific suppressive factor(s) from FcR⁺ B cells by binding on FcR. This factor, termed “suppressive B-cell factor (SBF)” was only effective on H-2 compatible, but not on H-2 incompatible spleen cells in an adoptive cell transfer system. Furthermore, SBF, prepared from B10.A (H-2^a) splenic FcR⁺ B cells, suppressed the adoptive primary response of B10.D2 mice (H-2^d), in addition to A/J mice (H-2^a) against DNP-DE, by the pretreatment of cells with SBF in vitro. Absorption with affinity columns demonstrated that active components) of SBF from C3H/He mice (H-2^k) was eliminated by both B6 anti-CBA (H-2^b anti-H-2^k) and B10.D2 anti-B10.BR (H-2^d anti-H-2^k), but not B10 anti-B10.A (H-2^b anti-H-2^a). In contrast, the suppressive activity of SBF was eliminated neither by anti-mouse Ig nor by a heat-aggregated human γ-globulin column. These results indicate that SBF contains a product coded by the right-hand side of H-2 gene complex, but does not contain Ig determinants nor FcR. Thus, it is conceivable that a compatibility of the right-hand side of H-2 gene complex is required for inducing effective suppression of spleen cells by SBF. SBF was considered to be a trypsin-resistant and heat-labile substance with a molecular weight of 30,000–63,000. The target cells for SBF were FcR^? B precursors, but not helper T cells.     

Effect of nonionizing radiation on the purkinje cells of the rat cerebellum

In one experiment, Sprague Dawley rats (16–21 days of gestation) and their offspring were exposed to 100-MHz (CW) electromagnetic radiation at 46 mW/cm² (SAR 2.77 mW/g) for 4 h/day for 97 days. In another experiment, the pregnant rats were irradiated daily from 17 to 21 days of gestation with 2450-MHz (CW) microwaves at 10 mW/cm² (SAR 2 mW/g) for 21 h/day. In a third experiment, 6-day-old rat pups were irradiated 7 h/day for five days with 2450-MHz radiation at 10 mW/cm². Equal numbers of animals were sham irradiated in each group. Quantitative studies of Purkinje cells showed a significant and irreversible decrease in rats irradiated during fetal or fetal and early postnatal life. In animals exposed postnatally, and euthanized immediately after irradiation, significant decrease in the relative number of Purkinje cells was apparent. However, restoration apparently occurred after forty days of recovery.     

Expression of subregion and haplotype preference for H-2Kk restricted cytotoxic T-cell responses in vivo and at the level of the precursor frequencies

Several strains of mice bearing the H-2K^k allele were found to generate in vivo strong CTL responses against TNP-haptenated syngeneic cells, while several other strains of mice were found to generate comparably weak or no responses. C3H × DBA/2)F1 mice (H-2^k × H-2^d) and A/J mice with the recombinant haplotype $\text{H-2}{}^{\mathrm{<mtext>k</mtext><mtext>d</mtext>}}$ generated CTL responses in vivo that were completely restricted toward the H-2^k haplotype or the K end of the $\text{H-2}{}^{\mathrm{<mtext>k</mtext><mtext>d</mtext>}}$ haplotype, respectively. The CTL activity of C3H × DBA/2)F1 and A/J mice against haptenated H-2^k targets was found to be more than 25-fold higher than the CTL activity on H-2^d targets. The CTL responses in vitro under macroculture conditions showed, on the other hand, only a 3- to 6-fold higher cytotoxic activity against the haptenated H-2^k targets as compared with haptenated allogeneic or H-2^d targets; and limiting dilution experiments in microcultures revealed that the CTL precursor frequencies were only 2- to 3-fold smaller for TNP-haptenated H-2^d or haptenated allogeneic targets than for haptenated H-2^k target cells. This indicated that sufficient numbers of H-2^d-restricted and allorestricted CTL precursors were actually present in these strains, but did not develop detectable cytotoxic activity in vivo. The exceptional property of the H-2^k haplotype is, therefore, only partly determined by a difference in the CTL precursor frequencies, and to the larger extent determined at the level of the activation of the CTL response.     

Syngeneic sensitization of mouse lymphocytes on monolayers of thyroid epithelial cells: IV. Correlation with H-2 haplotypes

In vitro primary syngeneic sensitization on monolayers of thyroid epithelial cells was performed with 21 inbred strains of mice representing 11 original H-2 haplotypes. Significant differences in the proliferative responses, assessed by thymidine uptake, were found to be related to the major histocompatibility complex haplotype. This result was further confirmed using congenic resistant strains of mice. In comparison with the experimental autoimmune thyroiditis induced by syngeneic thyroglobulin and adjuvant, primary syngeneic sensitization on monolayers of thyroid epithelial cells appeared to be under the same genetic control (H-2^k strains being good responders, while H-2^b mice are poor responders).     

In vitro microwave effects on human neutrophil precursor cells (CFU-C)

Human marrow cells were irradiated with 2450-MHz CW microwaves in a fluid-filled waveguide irradiation system. Cell exposure was conducted by placing a marrow cell suspension in 20-μl glass microcapillary tubes that were positioned in the exposure chamber, and irradiated at power densities from 31 to 1,000 mW/cm² (with corresponding specific absorption rates of 62 to 2,000 mW/g) for 15 minutes. The temperature of the sample was maintained at a fixed point. Sham-irradiated (SI) and microwave-irradiated (MWI) cells were cultured in a methylcellulose culture system for neutrophil colony proliferation. There was no reduction in neutrophil colony number on days 6–7 or 12–14 in cells exposed at 31 or 62 mW/cm², but as the power density was increased to 1,000 mW/cm², there was a reduction in colony number of MWI cells compared with SI cells. The microwave interaction with the human neutrophil colony-forming cells was apparently not related to temperature rise, or to the state of cell cycle, and was irreversible.     

Microwave irradiation affects radial-arm maze performance in the rat

After 45 min of exposure to pulsed 2450 MHz microwaves (2 μsec pulses, 500 pps, 1 mW/cm², average whole body SAR 0.6 W/kg), rats showed retarded learning while performing in the radial-arm maze to obtain food rewards, indicating a deficit in spatial “working memory” function. This behavioral deficit was reversed by pretreatment before exposure with the cholinergic agonist physostigmine or the opiate antagonist naltrexone, whereas pretreatment with the peripheral opiate antagonist naloxone methiodide showed no reversal of effect. These data indicate that both cholinergic and endogenous opioid neurotransmitter systems in the brain are involved in the microwave-induced spatial memory deficit. © 1994 Wiley-Liss, Inc.     

Control of normal differentiation of myeloid leukemic cells. IV. Induction of differentiation by serum from endotoxin treated mice

Sera from different strains of mice injected with endotoxin induced clones (D⁺) from a cultured line of myeloid leukemic cells to undergo normal differentiation to mature granulocytes and macrophages. Other clones (D^?) derived from the same cell line were not inducible by these sera to undergo normal cell differentiation. Sera from the same strains of mice that had not been injected with endotoxin, increased the cloning efficiency of D⁺ and D ^? clones but did not induce differentiation. Endotoxin serum induced differentiation in D⁺ cells at dilutions up to 1:64, but increased the cloning efficiency of these cells at dilutions up to 1:2048. The end point of the dilution of endotoxin serum that induced differentiation in D⁺ cells, was also the end point that induced the formation of colonies with differentiation from normal bone marrow cells. The results indicate that serum from endotoxin treated animals can serve as a good in vivo source to induce normal differentiation in D⁺ myeloid leukemic cells; that the progeny of a single leukemic cell was induced to undergo differentiation to both macrophages and granulocytes; that endotoxin serum contained two activities, one that increased cloning efficiency and the other that induced cell differentiation; and that the same material in endotoxin serum induced cell differentiation in normal and leukemic cells.     

Effect of microwaves (2450-MHz) on the immune system in mice: studies of nucleic acid and protein synthesis

CBA/J adult male mice were given single or triple exposures to 2450-mHz microwaves in an environmentally controlled wave guide facility. The average absorbed dose rate for a single exposure varied from 12 to 15 mW/g. Sham-exposed mice served as controls. Lymphoid cells were collected and tested for metabolic activity on days 3, 6, and 9 following a single exposure, and on days 9, 12, and 16 following triple exposures on days 0, 3, and 6. Cells were cultured in vitro for four hours to seven days before their metabolic rates were assayed. Under these conditions, microwaves failed to produce any detectable change in deoxyribonucleic acid (DNA), ribonucleic acid (RNA), and protein synthesis, as measured by the incorporation of methyl(3H)-thymidine (3H-TDR) (DNA substrate), 3H-uridine (3H-UR) (RNA substrate), and 3H-leucine (protein substrate) by spleen, bone marrow, and peripheral blood lymphocytes (PBL) in vitro. These data suggest that microwave-induced increases in the frequency of complement-receptor (CR)- or surface-immunoglobulin (sIg)-bearing cells were not associated with a concomitant increase in cell proliferation and/or protein synthesis, and favor the concept that microwaves under these conditions stimulate already existing B-cell precursors for maturation.     

Transbilayer movement of 24Na in sonicated phosphatidylcholine vesicles exposed to frequency-modulated microwave radiation

Sonicated egg phosphatidylcholine vesicles loaded with ²⁴Na⁺ were exposed at 20mW to a frequency-modulated (3 Hz) microwave field in the range of 2350 to 2550 MHz, or at 80 mW to a 2450-MHz CW (continuous wave) field, in a waveguide. The vesicle suspension absorbed microwaves at about 1 mW/ml and 25 mW/ml (CW experiment). The average temperature change of the irradiated suspension was < 0.1 °C from ambient. Leakage of ²⁴Na⁺ from the vesicles for up to 19 hours was measured. No difference was noted in the movement of ²⁴Na⁺ from the vesicles in the irradiated and control dispersions.     

The ability of H-2+ and H-2− cell lines to induce or be lysed by Cytotoxic T cells

We have studied the T cell-mediated lysis of two C58 lymphoma lines: R1(TL⁺), which bears serologically detectable H-2^k and TL1,2,3 antigens, and R1(TL^–), an immunoselected variant which lacks these antigens. Unlike R1(TL⁺) cells, the variant cells are not sensitive to specific lysis by T cells directed against either H-2^k or minor H antigens of C58 mice. An injection of R1(TL⁺) cells into allogeneic H-2-identical or H-2-different mice primes for an excellent secondary cytotoxic response in vitro to R1(TL⁺); but not to R1(TL^–). Immunization in vivo and in vitro with R1(TL)^–) leads to little or no priming or generation of cytotoxic T cells. Both cell lines, however, are sensitive to nonspecific lysis by cytotoxic cells in the presence of PHA or Con A, although even under these conditions, R1(TL⁺) is killed more effectively than is R1(TL^–). We conclude that R1(TL^–) does not express any form of H-2 antigen which can be detected by immunization or by sensitivity to cytotoxic T cells.     

Natural resistance of irradiated 129-strain mice to bone marrow allografts: Genetic control by theH-2K region

A major genetic determinant of natural resistance to bone marrow allografts, designated asHh-3, was mapped to theH-2K region. This gene may code for or regulate the expression of cell surface structures selectively expressed on donor hemopoietic cells and recognized by naturally occurring cytotoxic effectors. Resistance was observed as failure of donor cell growth in the spleen of irradiated 129-strain (H-2 ^bc ) recipients of H-2^k bone marrow cells. The mapping was accomplished by substituting donor cells bearingk alleles throughout theH-2 complex with cells of recombinant mouse lines bearingk alleles at definedH-2 regions. The host antigraft reaction underlying resistance was abrogated by pretreating 129-strain mice with either rabbit antimouse lymphocyte serum or the antimacrophage agent silica. Grafting of H-2K^k cells into mice ancestrally unrelated to 129 but sharing theH-2 ^bc or the similarH-2 ^b haplotype, and intoH-2 ^b/k ,H-2 ^k/bc , andH-2 ^k/d F₁ hybrids revealed that resistance was unique to 129 mice, since mice of the other strains, including F₁ hybrids, were susceptible to the grafts. Thus,Hh-3 incompatibility was a necessary but insufficient condition for the manifestation of allogeneic resistance; other genetic factors not associated withH-2 conferred responder status to 129-strain mice and nonresponder status to D1.LP, B10.129(6M), B10, B6, and possibly to F₁ hybrid mice. The possible relationships between allogeneic resistance to H-2^k marrow grafts, hybrid resistance to H-2^k lymphomas, and F₁ hybrid antiparental H-2^k cytotoxicity induced in vitro are discussed.     

Mechanisms of Allografted Tumor Rejection: The Roles of T Cells in Allograft Rejection Mediated by a Type of Bone Marrow-Derived Macrophage

Allografted tumor rejection does not occur in the absence of T cells, but the main effector cells responsible for the rejection are allograft-induced macrophages (AIM). We examined the roles of T cells in the AIM-mediated rejection of Meth A (H-2) tumor cells from C57BL/6 (H-2^b) mice. Irradiation of C57BL/6 mice abrogated both the induction of AIM and the allograft rejection. Reconstitution of the irradiated mice with F1 (C57BL/6 X C3H/He: H-2^b/k) bone marrow cells led to the appearance of H-2^b/k haplo-type of AIM exclusively in the rejection site and to allograft rejection, indicating that radiosensitive cells prerequisite for both the induction of AIM and allograft rejection were bone marrow-derived cells, and that the progenitors of AIM existed in the bone marrow cells to be activated into AIM in the rejection site. To understand the role of T cells in the induction of AIM, we used adult-thymectomized, X-irradiated C57BL/6 mice reconstituted with F1 bone marrow (ATXBM). The ATXBM mice could neither induce AIM nor reject allogeneic Meth A cells, whereas adoptive transfer of F1 lymph node T cells to the ATXBM mice restored not only the induction of AIM but also rejection of the allograft. Among the lymph node T cells, CD4⁺, but not CD8⁺, cells were found to be essential for the activation of AIM progenitors to AIM; and CD8⁺ T cells were further required for rejection, at least in part, to enhance the number of AIM in the rejection site.     

B-Cell responses to male-specific antigen(s) in mice

It has been found that B-cell responses to male-specific antigen(s) can be clearly demonstrated by reversed plaque assays. Female mice injected with syngeneic male spleen cells showed significant increases (greater than 100 × in some strains) in the number of immunoglobulin-secreting cells in lymph nodes draining the injection site. There was a variation in B-cell responsiveness between strains and this correlated only partially with previously reported T-cell responsiveness to the H-Y antigen. C57BL (H-2 ^b ) mice were among the most responsive, while CBA (H-2 ^k ), (CBA × C57BL)F₁, and BALB/c (H-2 ^d ) were all much less responsive. These results apparently open up a new approach to the investigation of B-cell responses to male-specific antigen(s).     

Effect of nonionizing radiation on the purkinje cells of the uvula in squirrel monkey cerebellum

Pregnant squirrel monkeys were exposed to 2450-MHz (CW) microwaves at an equivalent power density of 10 mW/cm² (SAR 3.4 mW/g) for three hours daily in a cavity-cage module. The exposure began when pregnancy was determined by a hormonal method, and continued through the offspring's first 9.5 months. After irradiation, the brains of the offspring were fixed with formaldehyde, and the inferior vermis of each cerebella was removed and processed for histologic observations. Purkinje cell density in the uvula was determined in sagittal serial sections. There was no significant difference between control and experimental animals in the number of Purkinje cells per mm of Purkinje cell line (linear density), as well as in the density of Purkinje cells in the Purkinje cell layer.     

Opioid receptor subtypes that mediate a microwave-induced decrease in central cholinergic activity in the rat.

We performed experiments to investigate subtypes of opioid receptors in the brain involved in the effect of acute (45 min) pulsed microwave exposure (2,450-MHz, 2-microseconds pulses, 500 pps, average power density 1 mW/cm2, peak-power density, 1 W/cm2, average whole body SAR 0.6 W/kg) on cholinergic activity in the rat brain. Rats were pretreated by microinjection of specific antagonists of mu, delta, and kappa opioid-receptors into the lateral cerebroventricle before exposure to microwaves. The data showed that all three subtypes of opioid receptors are involved in the microwave-induced decrease in cholinergic activity in the hippocampus. However, the microwave-induced decrease in cholinergic activity in the frontal cortex was not significantly affected by any of the drug treatments, confirming our previous conclusion that the effect of microwaves on the frontal cortex is not mediated by endogenous opioids.     

Quantitative changes in potassium,sodium, and calcium in the submaxillary salivary gland and blood serum of rats exposed to 2880-MHz microwave radiation

Na⁺, K⁺, and Ca²⁺ concentrations in the blood serum and submaxillary salivary gland (SSG) were investigated in adult, male rats exposed to 2880-MHz microwaves modulated with 1.5-μs pulses at a pulse repetition rate of 1000 Hz or in a hyperthermal environment. Rats were exposed, one at a time, for 30 min to microwaves producing a specific absorption rate (SAR) of: 4.2, 6.3,6.8,8.4, 10.8, or 12.6 W/kg, or were sham exposed under similar environmental conditions. In a second series, one group of rats was exposed singly for 15, 30, or 60 min to microwaves producing an SAR of 9.5 W/kg and other rats were exposed for similar periods at 40 °C; and 10 rats were sham exposed. Flame photometric analysis indicated that the thresholds of microwave radiation required to induce a change in Na⁺, K⁺, and Ca²⁺ concentrations in the salivary glands are 6.8, 6.8, and 6.3 W/kg, respectively. The directions of Na⁺, K⁺, and Ca²⁺ ion shifts in exposed rats' salivary glands are similar, whether affected by microwaves or hyperthermia. Greater changes in Na⁺ and K⁺ concentrations in SSG of rats exposed to microwaves for 15 and 30 min were found than in those exposed at 40 °C. On the other hand, exposure to hyperthermia at 40 °C or to microwaves for 1 h caused Na⁺ concentration to be increased by 68.7 and 59.5% and K⁺ concentration to be decreased by 29.6 and 21.7%, respectively.     

Regulation of natural killer cell activity by anti-I-region monoclonal antibodies

The effects of a monoclonal antibody directed against immune response gene products on mouse NK activity were examined. In vivo administration of an anti-I-A^k antibody to C3H/He (H-2^k) mice modulated their peritoneal cell (PC) and spleen cell (SC) natural killer (NK) activity against YAC-1 lymphoma target cells in vitro. No such effect was observed when BALB/c (H-2^d) mice were treated with this antibody. Administration of anti-I-A^k antibody to mice before and after infection with Toxoplasma or treatment with poly(I:C) leads to suppression of NK activity in comparison to NK activity of mice infected with Toxoplasma or injected with poly(I:C) alone. A similar treatment regimen with M5/114 antibody which reacts with I-A^b, I-A^d, I-E^d, and I-E^k molecules resulted in decreased NK activity in B10.D2 (H-2^d) but not in B10.BR (H-2^k) mice. Serum and cell culture supernatant interferon (IFN) concentrations were not altered as a result of anti-I-A^k treatment. Removal of adherent cells did not restore NK activity of anti-I-A^k-treated Toxoplasma-infected mice to levels obtained with mice infected with Toxoplasma. In contrast, depletion of Ly 2.1⁺ cells from nylon-wool nonadherent SC of mice treated with anti-I-A^k antibody, before and after infection with Toxoplasma, resulted in restoration of NK activity to the same level as that observed in Toxoptasma-infected mice.