OXYGEN-DEPENDENCE OF NEAR UV (365 NM) LETHALITY AND THE INTERACTION OF NEAR UV AND X-RAYS IN TWO MAMMALIAN CELL LINES

Abstract— The colony-forming ability of Chinese hamster cells (V-79) and HeLa cells has been measured after near-ultraviolet (UV) irradiation, predominantly at 365 nm. To avoid the production of toxic photoproducts, cells were irradiated in an inorganic buffer rather than in tissue culture medium. Under these circumstances near-UV lethality was strongly oxygen-dependent. Both cell lines were approximately 10⁴ times more sensitive to 254 nm irradiation than to 365 nm radiation when irradiated aerobically. Pretreatment with 6 times 10⁵ Jm^-2 365 nm radiation sensitised the HeLa, but not the V-79 cell line to subsequent X-irradiation. Pretreatment of cells with 17 Jm^-2 254 nm radiation, a dose calculated to produce twenty times more pyrimidine dimers than the 365 nm dose, produced only slight sensitisa-tion to X-rays. It is suggested that the sensitisation to X-rays seen in the HeLa cells after 365 nm treatment is not the result of lesions induced in DNA by the near-UV radiation, but may reflect the disruption of DNA-repair systems.     

DIFFERENT (DIRECT and INDIRECT) MECHANISMS FOR THE INDUCTION OF DNA-PROTEIN CROSSLINKS IN HUMAN CELLS BY FAR- and NEAR-ULTRAVIOLET RADIATIONS (290 and 405 nm)

Abstract— Apparent DNA-protein crosslinking induced by monochromatic 290 and 405 nm Tadiations was measured in cultured human P3 teratocarcinoma cells with DNA alkaline elution techniques. The rates of the induction of crosslinks by 290 nm radiation were the same when the cells were irradiated either aerobically or anaerobically or when the cells were in an H₂O or D₂O aqueous environment. With 405 nm radiation, anaerobic irradiation reduced the induction of the crosslinks (dose modifying factor is about 0.2), and about twice as many crosslinks were observed when the cells were irradiated in an environment of D₂O rather than H₂O. The results are consistent with the hypothesis that far-UV radiation induces DNA-protein crosslinks by a direct mechanism, whereas near-UV radiation induces crosslinks via indirect photodynamic photosensitizations in which unidentified cellular endogenous photosensitizers and reactive species of oxygen are used.     

UNSCHEDULED DNA SYNTHESIS: A QUANTITATIVE INDICATOR OF RESIDUAL IMMUNODETECTABLE PYRIMIDINE DIMERS IN HUMAN FIBROBLASTS AFTER ULTRAVIOLET-B IRRADIATION

We have addressed the question whether the level of UV-B induced DNA damage can be accurately assessed by the measurement of the rate of unscheduled DNA synthesis (UDS). Cultured human fibroblasts were irradiated with UV radiation at 290, 313 or 365 nm. The LD50 was 85 J/m2 at 290 nm, 4500 J/m2 at 313 nm, and 70 kJ/m2 at 365 nm. The analysis of UDS measurements indicate complete arrest of repair processes within 24 h after irradiation, irrespective of the dose (in the range 10-60 J/m2 at 290 nm, and 250-1000 J/m2 at 313 nm). Irradiation at 365 nm failed to yield detectable evidence of UDS. Incubation of irradiated cells with an antiserum directed against both 6-4 type and cyclobutane-type pyrimidine dimers shows a clear parallelism between the disappearance of the antibody-binding determinants and the variation of the rate of UDS vs time after the end of the irradiation. Thus it is concluded that in UV-B irradiated normal cultured human fibroblasts, the lack of UDS reflects the absence of immunodetectable pyrimidine dimers.     

The effect of gamma irradiation on mechanical, and thermal properties of recycling polyethylene terephthalate and low density polyethylene (R-PET/LDPE) blend compatibilized by ethylene vinyl acetate (EVA)

A study has been made on the compatibility of recycled polyethylene terephthalate (R-PET) and low density polyethylene (LDPE) blend in the presence of ethylene vinyl acetate (EVA) as a compatibilizing agent prepared by extrusion hot stretching process. EVA content in the blend as a compatibilizing agent was an enhancement effect on radiation crosslinking of R-PET/EVA/LDPE blends and the highest radiation crosslinking was obtained when the EVA content was reached at 10 % EVA when irradiated by gamma irradiation. Blends containing different (EVA) ratios were irradiated to different doses of gamma irradiation 25, 50 and 100 kGy. The effect of the compatibilizer and radiation on mechanical, thermal properties of R-PET together with LDPE and morphology has been investigated. It was found that gamma irradiation together with the presence of compatibilizing agent (EVA) has positive effect on the mechanical and thermal properties of R-PET/LDPE blend. The structural properties of R-PET/LDPE modified by gamma irradiation and EVA as compatibilizing agent was examined by SEM. Also, it was found that the optimum concentration of EVA and gamma irradiation dose was found to be 10 % EVA and 100 kGy, respectively.     

Application of the biotin-dUTP chromosome labelling technique to study the role of 5-bromo-2'-deoxyuridine in the formation of UV-induced sister chromatid exchanges in CHO cells

The role of 5-bromo-2'-deoxyuridine (BrdU) in the formation of sister chromatid exchanges (SCEs) in cells exposed to UV radiation was studied. Cells were unifilarily labelled (labelling of one strand of chromosomal DNA) with BrdU or biotin-16-2'-deoxyuridine (biotin-dU) and irradiated in G(1) phase of the cell cycle either with 254 nm, which is absorbed by all nucleobases including bromouracil (BrU) or with 313 nm radiation, which is predominantly absorbed by the BrU moiety. Elevated SCE frequencies were observed in cells irradiated at 254 nm (1.2 and 3.0 J m(-2)) which were pre-labelled with BrdU or biotin-dU. Following irradiation at 313 nm (38 and 96 J m(-2)) a statistically elevated SCE frequency was observed in cells pre-labelled with BrdU but not with biotin-dU. In cells pre-labelled with BrdU, UV-radiation at 254 nm was 50-80 times more effective in inducing SCEs than that at 313 nm. This result can be accounted for by the fact that in BrdU-DNA the cross-section for uracilyl radical and bromine atom formation is approximately 100-fold higher at 254 nm than that at 313 nm. Upon irradiation at 254 nm, BrdU had a strong sensitising effect on SCE induction: the SCE frequencies observed in cells pre-labelled with BrdU are approximately 6 times higher than in cells pre-labelled with biotin-dU. From this it follows that BrdU-induced damage is responsible for more than 80% of the SCEs formed in UV irradiated cells unifilarily labelled with BrdU. Based on photochemical considerations and the fact that chemical agents which form DNA interstrand cross-links are among the most potent inducers of SCEs, we propose that an interstrand cross-link may be the major lesion leading to SCEs in BrdU-labelled cells.     

A SURVEY OF PHOTOPRODUCTS OF AN IRRADIATED OLIGODEOXYNUCLEOTIDE BY MONOCHROMATIC PHOTONS WITH THE ENERGY RANGED FROM 6.5 TO 22.5 eV

We have irradiated 2'-deoxyadenylyl-(3'-5')-2'-deoxyadenosine (dephospho(dA)₂) in solid state with a newly developed synchrotron irradiation system over 6.5 to 22.5 eV photon energy range (190 to 55 nm in wavelength). Densitometric analysis of thin-layer chromatogram of photoproducts indicated that adenine and deoxyadenosine monophosphate were commonly produced above 7.3 eV (170 nm) in approximately equal amount. Deoxyribose was not detected by the fluorescence method with scratched materials from the chromatogram at any location except deoxyadenosine monophosphate and parent oligonucleotide positions. The photoproducts above mentioned were not observed at 6.5 eV (190 nm) even after very high radiation fluence. The energy dependence of this degradation indicates that the shorter the wavelength in use the smaller the fluence it requires. Careful examination of irradiated oligonucleotide on the chromatograms shows that the major portion of deoxyadenosine monophosphate so produced was the 5'phospho derivative. This site-selective degradation, independent of photon energy, seemed to be a remarkable characteristic of vacuum-UV induced effects above 7 eV. From these results a working hypothesis was advanced that the deoxypentose moiety was first destroyed in one of the nucleoside residues due to the strong absorption in the sugar-phosphate group, liberating adenine on the one hand and leaving 5'-deoxyadenosine monophosphate on the other hand.     

MEMBRANE DAMAGE CAN BE A SIGNIFICANT FACTOR IN THE INACTIVATION OF ESCHERICHIA COLI BY NEAR-ULTRAVIOLET RADIATION

Abstract A DNA repair competent strain of Escherichia coli K-12 showed sensitivity to inorganic salts (at concentrations routinely used in minimal media) after irradiation with broad spectrum near–UV radiation, at fluences that caused little inactivation when plated on complex growth medium. This effect was not observed with cells that had been exposed to 254 nm radiation. This sensitivity to minimal medium was increased by increasing the salt concentration of the medium and by increasing the pH of the medium. This sensitivity was greatly increased by adding to the medium a low concentration of commercial glassware cleaning detergent that had no effect on unirradiated cells or far-UV irradiated cells. These findings may explain the large variability often observed in near-UV radiation survival data, and demonstrate that, at least on minimal medium plates, membrane damage contributes significantly towards cell killing. This phenomenon is largely oxygen dependent.     

AN ACTION SPECTRUM FOR ULTRAVIOLET RADIATION-INDUCED MEMBRANE DAMAGE IN Escherichia coli K-12

A wild-type Escherichia coli K-12 strain was irradiated using monochromatic radiation in the range 254 to 405 nm. A measure of the cell membrane damage induced at each wavelength was investigated by comparing cell viability after irradiation on nutrient agar and on minimal medium containing either a low or high inorganic salt concentration. An action spectrum for lethality and for cell membrane damage was then determined. From 254 to 310 nm lethality closely corresponded to the absorption spectrum of DNA, and there was no indication of membrane damage. However, above a wavelength of 310 nm, the direct absorption of radiation by DNA could not account for the sensitivity observed. Moreover, at wavelengths longer than 310 nm, cell membrane damage was induced and by an increasing factor up to a peak at 334 nm. At the longer wavelengths of 365 and 405 nm, there was a gradual decrease from the peak of damage to cell membranes induced by 334 nm radiation. These results indicate that cell membrane damage may contribute significantly to near-UV radiation-induced cell lethality in wild-type E. coli K-12.     

LEAKAGE OF 86Rb+ AFTER ULTRAVIOLET IRRADIATION OF Escherichia coli K-12

Abstract— Stationary phase cultures of a DNA repair proficient Escherichia coli K-12 strain showed a release of intracellular material as assessed by three different methods (260 nm absorption; [methyl-³H]thymidine leakage and ⁸⁶Rb⁺ leakage) after broad-band (Black-Light Blue) near-UV radiation but not after far-UV (254 nm) radiation. As a control response for membrane damage to cells, this leakage of intracellular material was also determined by each method after mild-heat (52°C) treatment of E. coli K-12. An action spectrum for the release of ⁸⁶Rb⁺ from E. coli K-12 after irradiation with monochromatic wavelengths, from 254 to 405 nm, is also presented. The action spectrum for lethality (F₃₇ values) obtained for this strain, shows that leakage of ⁸⁶Rb⁺ occurs at fluences equivalent to or slightly less than fluences causing inactivation at wavelengths above 305 nm. In contrast, at wavelengths below 305 nm, leakage of ⁸⁶Rb⁺ from irradiated cells can be induced but only at fluences significantly greater than was required to cause cell inactivation. These results indicate, therefore, that near-UV radiation can induce a damaging effect on the cell's permeability barrier which may be significant in causing the death of the cell, whereas the effect is not significant in causing the death of cells by far-UV radiation where DNA damage is known to be the main cause of lethality.     

Absorption spectra of viral components of Sendai virus in the wavelength region from 130 to 320 nm.

Using synchrotron radiation as a light source, the absorption spectra of purified viral components of the Sendai virus, i.e. messenger RNA, lipids, spike (envelope) proteins, reconstructed envelopes, core proteins and whole virions, were obtained in the wavelength region 130-320 nm by measuring the transmission of thin films. Viral (messenger) RNA two peaks at 260 and 190 nm, and a large increase below 160 nm. The absorption spectrum of lipids exhibited a broad peak at 190 nm and a very sharp increase below 160 nm. With spike proteins, a slight peak at 280 nm and a shoulder at 230 nm were observed in addition to a sharper peak at 190 nm and a rather slow increasing absorption below 160 nm. Reconstructed envelopes showed the features of a combination of lipids and proteins. The absorption spectra of core proteins and whole virions exhibited similar characteristics to spike proteins. Conventional UV data were also obtained in the wavelength range 210-320 nm with RNA and lipids. The UV and synchrotron radiation data were in good agreement in terms of the mass absorption coefficients. The molecular splitting of spike proteins was also examined. Proteins gave more diffuse reflection than their subunits, causing a reduction in absorption. This was explained by a loss of transparency with increasing molecular weight.     

QUANTITATION OF PYRIMIDINE DIMERS BY IMMUNOSLOT BLOT FOLLOWING SUBLETHAL UV-IRRADIATION OF HUMAN CELLS

An immunoslot blot assay was developed to detect pyrimidine dimers induced in DNA by sublethal doses of UV (254 nm) radiation. Using this assay, one dimer could be detected in 10 megabase DNA using 200 ng or 0.5 megabase DNA using 20 ng irradiated DNA. The level of detection, as measured by dimer specific antibody binding, was proportional to the dose of UV and amount of irradiated DNA used. The repair of pyrimidine dimers was measured in human skin fibroblastic cells in culture following exposure to 0.5 to 5 J m^-2 of 254 nm UV radiation. The half-life of repair was approximately 24, 7 and 6 h in cells exposed to 0.5, 2 and 5 J m^-2 UV radiation, respectively. This immunological approach utilizing irradiated DNA immobilized to nitrocellulose should allow the direct quantitation of dimers following very low levels of irradiation in small biological samples and isolated gene fragments.     

WAVELENGTH DEPENDENCE FOR THE INACTIVATION OF ATP IN THE VACUUM-ULTRAVIOLET REGION ABOVE 140 nm

Radiation effects were investigated on the activity and the structure of adenosine triphosphate in the wavelength range from 140 nm to 260 nm, using monochromatized synchrotron radiation from the INS-SOR storage ring. The sample was irradiated as a thin film in vacuum. The activity of adenosine triphosphate decreased sharply below 180 nm as judged by the luminescence in the luciferin-luciferase assay. From the exponential decay of function, the cross-section for inactivation was calculated to be of the order of 10^-21 m²/photon in the range from 140 to 170 nm. No decrease was detected at wavelengths of 190 nm and above. The calculated quantum yield increased as the wavelength became shorter and reached to 0.20 at 150 nm. The release of adenine at 160 nm-irradiation was detected by thin layer chromatography; no adenosine diphosphate or adenosine monophosphate occurred. Only a trace of adenine was found after 190 nm-irradiation. These results indicate that the broad absorption peak for higher excitations attributable to the base moiety around 190 nm does not cause both structural and functional changes, while the absorption by the sugar-phosphate group produces the rupture of N -glycosidic bond, and probably leads to the loss of function.     

WAVELENGTH DEPENDENT FORMATION OF THYMINE DIMERS AND (6-4)PHOTOPRODUCTS IN DNA BY MONOCHROMATIC ULTRAVIOLET LIGHT RANGING FROM 150 TO 365 nm

We investigated the wavelength dependence of cyclobutane thymine dimer and (6-4)photoproduct induction by monochromatic UV in the region extending from 150 to 365 nm, using an enzyme-linked immunosorbent assay with two monoclonal antibodies. Calf thymus DNA solution was irradiated with 254-365 nm monochromatic UV from a spectrograph, or with 220-300 nm monochromatic UV from synchrotron radiation. Thymine dimers and (6-4)photoproducts were fluence-dependently induced by every UV below 220 nm extending to 150 nm under dry condition. We detected the efficient formation of both types of damage in the shorter UV region, as well as at 260 nm, which had been believed to be the most efficient wavelength for the formation of UV lesions. The action spectra for the induction of thymine dimers and (6-4)photoproducts were similar from 180 to 300 nm, whereas the action spectrum values for thymine dimer induction were about 9- and 1.4-fold or more higher than the values for (6-4)photoproduct induction below 160 nm and above 313 nm, respectively.     

ULTRAVIOLET ACTION SPECTRA FOR PHOTOBIOLOGICAL EFFECTS IN CULTURED HUMAN LENS EPITHELIAL CELLS

Abstract— The action spectrum for cell killing by UV radiation in human lens epithelial (HLE) cells is not known. Here we report the action spectrum in the 297–365 nm region in cultured HLE cells with an extended lifespan (HLE B-3 cells) and define their usefulness as a model system for photobiological studies. Cells were irradiated with monochromatic radiation at 297, 302, 313, 325, 334 and 365 nm. Cell survival was determined using a clonogenic assay. Analysis of survival curves showed that radiation at 297 nm was six times more effective in cell killing than 302 nm radiation; 297 nm radiation was more than 260, 590, 1400 and 3000 times as effective in cell killing as 313, 325, 334 and 365 nm radiation, respectively. The action spectrum was similar in shape to that for other human epithelial cell lines and rabbit lens epithelial cells. The effect of UV radiation on crystallin synthesis was also determined at different wavelengths. To determine whether exposure to UV radiation affects the synthesis of β-crystallin, cells were exposed to sublethal fluences of UV radiation at 302 and 313 nm, labeled with [³⁵S]methionine and the newly synthesized βY-crystallin was analyzed by immunoprecipitation and western blotting using an antibody to β-crystallin. The results show a decrease in crystallin synthesis in HLE cells irradiated at 302 and 313 nm at fluences causing low cytotoxicity. The effect of radiation on membrane perturbation was determined by measuring enhancement of synthesis of prostaglandin E₂ (PGE₂). Synthesis of PGE₂ occurs at all UV wavelengths tested in the 297–365 nm region. The slope of the PGE₂ response curves was higher than that of cell killing curves in cultured HLE cells. These data show that cultured HLE cells with extended lifespan are a suitable system for investigating photobiological responses of cells to UV radiation.     

Increased levels of 8-hydroxy-2'-deoxyguanosine in Drosophila larval DNA after irradiation with 364-nm laser light but not with X-rays

Exposure to UVA light causes damage to cellular components such as DNA and membrane lipids. We showed previously that UVA irradiation can induce mutations in Drosophila larvae and that the major lesions responsible for mutations were not thymidine dimers when wavelengths tested became longer. The use of a longer wavelength with UVA laser apparatus (364 nm) has made it possible to test the effects of this powerful light in biological organisms. In the present study, we irradiated third instar larvae of the urate-null Drosophila mutant strain y v ma-l, which is sensitive to oxidative stress, and compared the effects of 364 nm light irradiation with the effects of X-rays. To assay viability, some of the larvae were kept at 25 degrees C until they eclosed in order to obtain a measure of viability. The remaining larvae were used to measure the amount of 8-hydroxydeoxyguanosine (8-OHdG), an indicator of oxidative DNA damage. The amount of 8-OHdG increased and viability decreased in response to increased UV dose in both the y v ma-l and wild-type strains. With irradiation of 600 kJ m(-2), 8-OHdG/10(6)dG was 7.2 +/- 3.2 and 6.2 +/- 2.0 in y v ma-l and wild-type strains, respectively, whereas the respective levels were 2.2 +/- 0.6 and 2.3 +/- 0.8 without irradiation. Our results indicated that irradiation with a 364-nm laser light caused significant oxidative damage in Drosophila larval DNA; however, induction of the damage was not prohibited by urate. To the best of our knowledge, this is the first report of a study in whole animals that shows increased levels of 8-OHdG in response to 364-nm UVA. X-ray ionizing radiation is also thought to generate reactive oxygen species in irradiated cells. We found that the amount of 8-OHdG in DNA following X-ray radiation remained unchanged in both strains, though survival rates were affected. X-ray-generated oxidative damage in Drosophila cells was followed by cell death but not DNA base oxidation, and the damage was suppressed by urate. The overall results suggest significant differences in the major in vivo oxidative damage caused by 364-nm light and X-rays.     

Influence of Gamma Radiation on Electrical Conduction of Polyethylene Terephthalate (PET) at Ambient Temperature

The electrical conduction in polyethylene terephthalate (PET) exposed to a gamma radiation dose of 150 kGy was investigated in the applied field range from 4 to 36 kV/mm. Samples were irradiated in air at room temperature by means of a ⁶⁰Co gamma source at a dose rate of approximately 42 Gy/min. The electrical properties of virgin and irradiated materials were examined by charging and discharging current measurements. The current decays with time can be represented according to an inverse power law. The changes of dielectric behavior after irradiation were attributed to scission effects.     

Effects of (60)Co gamma radiation on thylakoid membrane functions in Anacystis nidulans

In photosynthetic organisms oxidative stress is known to result in photoinactivation of photosynthetic machinery. We investigated effects of ⁶⁰Co γ radiation, which generates oxidative stress, on thylakoid structure and function in cyanobacteria. Cells of unicellular, non-nitrogen fixing cyanobacterium Anacystis nidulans (Synechococcus sp.) showed D₁₀ value of 257 Gy of ⁶⁰Co γ radiation. When measured immediately after exposure, cells irradiated with 1500 Gy (lethal dose) of ⁶⁰Co γ radiation did not show any differences in photosynthetic functions such as CO₂ fixation, O₂ evolution and partial reactions of photosynthetic electron transport in comparison to unirradiated cells. Incubation of irradiated cells for 24 h in light or dark resulted in decline in photosynthesis. The decline in photosynthesis was higher in the cells incubated in light as compared to the cells incubated in dark. Among the partial reactions of electron transport, only PSII activity declined drastically after incubation of irradiated samples. This was also supported by the analysis of membrane functions using thermoluminescence. Exposure of cyanobacteria to high doses of ⁶⁰Co γ radiation did not affect the thylakoid membrane ultrastructure immediately after exposure as shown by electron microscopy. The level of reactive oxygen species (ROS) in irradiated cells was 20 times higher as compared to control. In irradiated cells de novo protein synthesis was reduced considerably immediately after irradiation. Treatment of cells with tetracycline also affected photosynthesis as in irradiated cells. The results showed that photoinhibition of photosynthetic apparatus after incubation of irradiated cells was probably augmented due to reduced protein synthesis. Active photosynthesis is known to require uninterrupted replenishment of some of the proteins involved in electron transport chain. The defective thylakoid membrane biogenesis may be leading to photosynthetic decline post-irradiation.     

Small-angle scattering of synchrotron radiation investigations of nanostructured alumina membranes synthesized by sol–gel method

Alumina asymmetric ceramic membranes were prepared by sol–gel method. Research on structure of alumina membrane active layer is conducted by methods of physical nitrogen adsorption and small-angle scattering of synchrotron radiation, the obtained data confirms and supplements each other. Surface area for the studied material varies from 255 to 264 m²/g, average pore diameter varies from 5.3 nm to 5.4 nm.     

NON-NUCLEAR DAMAGE AND CELL LYSIS ARE INDUCED BY UVA, BUT NOT UVB OR UVC, RADIATION IN THREE STRAINS OF L5178Y CELLS

The potential to induce non-nuclear changes in mammalian cells has been examined for (1) UVA1 radiation (340–400 nm, UVASUN 2000 lamp), (2) UVA + UVB (peak at 313 nm) radiation (FS20 lamp), and (3) UVC (254 nm) radiation (GI5T8 lamp). The effects of irradiation were monitored in vitro using three strains of L5178Y (LY) mouse lymphoma cells that markedly differ in sensitivity to UV radiation. Comparisons were made for the effects of approximately equitoxic fluences that reduced cell survival to 1–15%. Depending on the cell strain, the fluences ranged from 830 to 1600 kJ/m² for the UVASUN lamp, 75 to 390 J/m² for the FS20 lamp and 3.8 to 17.2 J/m² for the G15T8 lamp. At the exposure level used in this study, irradiation with the UVASUN, but not the FS20 or G15T8, lamp induced a variety of non-nuclear changes including damage to cytoplasmic organelles and increased plasma membrane permeability and cell lysis. Cell lysis and membrane permeabilization were induced by the UVA1 emission of the UVASUN lamp, but not by its visible + IR components (>400 nm). The results show that the plasma membrane and other organelles of LY cells are highly sensitive to UVA1 but not to UVB or UVC radiation. Also UVA1, but not UVB or UVC radiation, causes rapid and extensive lysis of LY cells. In conclusion, non-nuclear damage contributes substantially to UVA cytotoxicity in all three strains of LY cells.     

Erythrocyte response to near-infrared radiation

The effects of NIR (near-infrared radiation 700-2,000 nm) on bovine erythrocytes in plasma was studied as a continuation of earlier studies. Cell shape was observed and the changes of ratio of hemolysis and electrokinetic potential measured as a function of irradiation time. After 10 min of irradiation, the shape of erythrocyte cells was mainly echinocytic. When these cells were incubated at 311 K for 24 h they regained their initial shape, but fresh erythrocytes that were irradiated for 30 min and aged in vitro did not. These phenomena are due to: (1) the absorption of NIR excitation by hemoglobin; the primary photochemical process being the photo-dissociation of oxyhemoglobin to deoxyhemoglobin. Resulting shape and ratio of hemolysis, structural changes and oxidative stress follow higher deoxyhemoglobin concentration. (2) The absorption of the NIR excitation by proteins, water and lipids. After NIR absorption the membrane surface dehydrates, leading to enhanced protonation and dissociation of hydrogen-bonded complexes. This in turn leads to a change in electrokinetic potential.