Plasma Rich in Growth Factors Promotes Autophagy in ARPE19 Cells in Response to Oxidative Stress Induced by Blue Light.

Age-related macular degeneration (AMD) causes the degeneration of photoreceptors and retinal cells leading to vision loss in older subjects. Among possible exogenous risk factors, it has been recently proposed that long-term exposure to blue light could aggravate the course of AMD. In the search for therapeutic options, plasma rich in growth factors (PRGF) has been shown to enhance cell antioxidant pathways and protect photoreceptors against the harm produced by blue light, although its mechanism of action remains unknown. One possible mechanism, autophagy, is one of the most conservative cell renewal systems used in eukaryotes to destroy cellular components that have been damaged by some kind of insult. The oxidative stress of exposure to blue light is known to induce cell autophagy. In this study, we examined the combined effects on autophagy of blue light and PRGF in a retinal cell line, ARPE19. In response to treatment with both PRGF and blue light, we detected the modulated expression of autophagy markers such as NF-kB, p62/sqstm1, Atg5, LC3 and Beclin1, and inflammatory markers such as IL1B and IL18. Our findings suggest that PRGF promotes cell autophagy in response to exposure to blue light.

The effect of pathogen inactivation on cryoprecipitate: a functional and quantitative evaluation.

BACKGROUND: As a pooled donor blood product, cryoprecipitate (cryo) carries risks of pathogen transmission. Pathogen inactivation (PI) improves the safety of cryoprecipitate, but its effects on haemostatic properties remain unclear. This study investigated protein expression in samples of pathogen inactivated cryoprecipitate (PI-cryo) using non-targeted quantitative proteomics and in vitro haemostatic capacity of PI-cryo. MATERIALS AND METHODS: Whole blood (WB)- and apheresis (APH)-derived plasma was subject to PI with INTERCEPT® Blood System (Cerus Corporation, Concord, CA, USA) and cryo was prepared from treated plasma. Protein levels in PI-cryo and paired controls were quantified using liquid chromatography-tandem mass spectrometry. Functional haemostatic properties of PI-cryo were assessed using a microparticle (MP) prothrombinase assay, thrombin generation assay, and an in vitro coagulopathy model subjected to thromboelastometry. RESULTS: Over 300 proteins were quantified across paired PI-cryo and controls. PI did not alter the expression of coagulation factors, but levels of platelet-derived proteins and platelet-derived MPs were markedly lower in the WB PI-cryo group. Compared to controls, WB (but not APH) cryo samples demonstrated significantly lower MP prothrombinase activity, prolonged clotting time, and lower clot firmness on thromboelastometry after PI. However, PI did not affect overall thrombin generation variables in either group. DISCUSSION: Data from this study suggest that PI via INTERCEPT® Blood System does not significantly impact the coagulation factor content or function of cryo but reduces the higher MP content in WB-derived cryo. PI-cryo products may confer benefits in reducing pathogen transmission without affecting haemostatic function, but further in vivo assessment is warranted.

Preoperative Radiotherapy Leads to Significant Differences in the Plasma Protein Profile of Rectal Cancer Patients.

INTRODUCTION: Colorectal cancer (CRC) is the third most common cancer worldwide, accounting for 10% of the global cancer burden. Rectal cancer accounts for around 30% of CRC cases, and patients with resectable rectal cancer are often given preoperative radiotherapy (PRT) to reduce the rate of local recurrence. The human plasma proteome is an exceptionally complex proteome and ideal to study due to its ability to reflect the presence of diseases such as cancer and the ease of obtaining blood samples. Previous proteomic studies involving rectal cancer patients have mostly focused on the identification of proteins involved in resistance to radiotherapy. OBJECTIVE: The aim of this study was to investigate the overall effects of PRT on plasma protein expression in rectal cancer patients, as there is a lack of such studies. METHODS: Here, we have used mass spectrometry and subsequent statistical analyses to analyze the plasma samples of 30 rectal cancer patients according to PRT status (positive or negative) and tumor stage (II or III). RESULTS AND CONCLUSIONS: We discovered 42 proteins whose levels differed significantly between stage II and III rectal cancer patients who did or did not receive PRT. This study shows that PRT, although localized to the pelvis, leads to measurable, tumor stage-specific changes in plasma protein expression. Future studies of plasma proteins should, when relevant, take this into account and be aware of the widespread effects that PRT has on the plasma proteome.

Irreversible oxidations of platelet proteins after riboflavin-UVB pathogen inactivation.

Pathogen inactivation technologies are known to alter in vitro phenotype and functional properties of platelets. Because pathogen inactivation generates reactive oxygen species, oxidative stress is considered as one of the plausible cause at the origin of the platelet storage lesion acceleration after treatment. To date proteomics has been used to document the protein variations to picture out the impact. Here, platelet concentrates were prepared from buffy-coats in Intersol additive solution, leukoreduced and pathogen inactivated using a riboflavin/UVB treatment. At day 2 of storage the platelet proteomes of control (untreated) and treated platelet concentrates were investigated against the site specific oxidation by liquid chromatography coupled to tandem mass spectrometry in a shotgun experiment. The shotgun approach detected 9350 peptides (and 2534 proteins) of which 1714 were oxidized. Eighteen peptides were found exclusively oxidized in treated platelets whereas 3 peptides were only found oxidized in control. The present data evidenced an interference with several proteins involved in platelet aggregation and platelet shape change (such as talin and vinculin).

Persistent depletion of plasma gelsolin (pGSN) after exposure of mice to heavy silicon ions.

Little is known about plasma proteins that can be used as biomarkers for early and late responses to radiation. The purpose of this study was to determine a link between depletion of plasma gelsolin (pGSN) and cell-death as well as inflammatory responses in the lung (one of the tissues known to be radiosensitive) of the same exposed CBA/CaJ mice after exposure to heavy silicon (28Si) ions. To prevent the development of multiple organ dysfunctions, pGSN (an important component of the extracellular actin-scavenging system) is responsible for the removal of actin that is released into the circulation during inflammation and from dying cells. We evaluated the levels of pGSN in plasma collected from groups of mice (5 mice in each) at 1 week (wk) and 1 month (1 mo) after exposure whole body to different doses of 28Si ions, i.e. 0, 0.1, 0.25, or 0.5 Gy (2 fractionated exposures, 15 days apart that totaled each selected dose). In the same mouse, the measurements of pGSN levels were coupled with the quantitation of injuries in the lung, determined by (a) the levels of cleaved poly (ADP-ribose) polymerase (cleaved-PARP), a marker of apoptotic cell-death, (b) the levels of activated nuclear factor-kappa B (NF-κB) and selected cytokines, i.e. tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), and IL-6, from tissue-lysates of the lung. Further, the ratio of neutrophils and lymphocytes (N/L) was determined in the same mouse. Our data indicated: (i) the magnitude of pGSN depletion was dependent to radiation dose at both harvest times, (ii) a persistent depletion of pGSN up to 1 mo post-exposure to 0.25 or 0.5 Gy of 28Si ions, (iii) an inverse-correlation between pGSN depletion and increased levels of cleaved-PARP, including activated NF-κB/pro-inflammatory cytokines in the lung, and (iv) at both harvest times, statistically significant increases in the N/L ratio in groups of mice exposed to 0.5 Gy only. Our findings suggested that depletion in pGSN levels reflects not only the responses to 28Si-ion exposure at both harvest times but also early and late-occurring damage.

Plasma temperature during methylene blue/light treatment influences virus inactivation capacity and product quality.

BACKGROUND: Photodynamic treatment using methylene blue (MB) and visible light is in routine use for pathogen inactivation of human plasma in different countries. Ambient and product temperature conditions for human plasma during production may vary between production sites. The influence of different temperature conditions on virus inactivation capacity and plasma quality of the THERAFLEX MB-Plasma procedure was investigated in this study. METHODS: Plasma units equilibrated to 5 ± 2°C, room temperature (22 ± 2°C) or 30 ± 2°C were treated with MB/light and comparatively assessed for the inactivation capacity for three different viruses, concentrations of MB and its photoproducts, activity of various plasma coagulation factors and clotting time. RESULTS: Reduced solubility of the MB pill was observed at 5 ± 2°C. Photocatalytic degradation of MB increased with increasing temperature, and the greatest formation of photoproducts (mainly azure B) occurred at 30 ± 2°C. Inactivation of suid herpesvirus, bovine viral diarrhoea virus and vesicular stomatitis virus was significantly lower at 5 ± 2°C than at higher temperatures. MB/light treatment affected clotting times and the activity of almost all investigated plasma proteins. Factor VIII (-17·7 ± 8·3%, 22 ± 2°C) and fibrinogen (-14·4 ± 16·4%, 22 ± 2°C) showed the highest decreases in activity. Increasing plasma temperatures resulted in greater changes in clotting time and higher losses of plasma coagulation factor activity. CONCLUSIONS: Temperature conditions for THERAFLEX MB-Plasma treatment must be carefully controlled to assure uniform quality of pathogen-reduced plasma in routine production. Inactivation of cooled plasma is not recommended.

Reversal of drug-induced gingival overgrowth by UV-mediated apoptosis of gingival fibroblasts - an in vitro study.

Gingival overgrowth (GO) is an undesirable result of certain drugs like Cyclosporine A (CsA). Histopathology of GO shows hyperplasia of gingival epithelium, expansion of connective tissue with increased collagen, or a combination. Factors such as age, gender, oral hygiene, duration, and dosage also influence onset and severity of GO. One of the mechanisms behind uncontrolled cell proliferation in drug-induced GO is inhibition of apoptotic pathways, with a consequent effect on normal cell turnover. Our objective was to determine if UV photo-treatment would activate apoptosis in the gingival fibroblast component. Human gingival fibroblast cells (HGF-1) were exposed to 200ng/ml or 400ng/ml CsA and maintained for 3, 6, and 9 days, followed by UV radiation for 2, 5, or 10min (N=6). Naïve (no CsA or UV), negative (UV, no CsA), and positive controls (CsA, no UV) were designated. Prior to UV treatment, growth media was replaced with 1M PBS to prevent absorption of UV radiation by serum proteins, and cells were incubated in growth media for 24h post-UV before processing for TUNEL assay, cell proliferation assays, or immunofluorescence. Data showed a temporal increase in proliferation of HGF-1 cells under the influence of CsA. The 200ng/ml dose was more effective in causing over-proliferation. UV treatment for 10min resulted in significant reduction in cell numbers, as evidenced by counts and proliferation assays. Our study is a first step to further evaluate UV-mediated apoptosis as a mechanism to control certain forms of GO.

Formation of long-lived reactive species of blood serum proteins induced by low-intensity irradiation of helium-neon laser and their involvement in the generation of reactive oxygen species.

It was demonstrated that low-intensity radiation of helium-neon (He-Ne) laser at 632.8nm, which leads to the transition of oxygen to a singlet state, causes the formation of reactive oxygen species (ROS) - hydrogen peroxide, hydroxyl and superoxide (hydroperoxide) radicals - in aqueous solutions. The oxygen effect - dependence of hydrogen peroxide formation on the concentration of molecular oxygen - was shown, and the participation of singlet oxygen, hydroxyl radicals and superoxide (hydroperoxide) radicals in this process was testified. Laser radiation-induced ROS in solutions of blood serum proteins, bovine serum albumin and bovine gamma-globulin, cause the formation of long-lived reactive protein species (LRPS) with a half-life of about 4h. The generation of LRPS caused by laser irradiation results in prolonged several-hour generation of ROS - hydrogen peroxide, hydroxyl and superoxide radicals. As affected by LRPS, coupled radical reactions lead to conversion of dissolved molecular oxygen to hydrogen peroxide. Irradiation with light sources away from the oxygen absorption band is not attended by formation of ROS and LRPS. A consideration is provided for the possible molecular mechanisms of ROS formation under the influence of He-Ne laser irradiation, the role of proteins in their generation and the biological significance of these processes.

Temperature Influence on Myocardial Cell Cytotoxicity of the Extracellular Photosensitization Reaction with Talaporfin Sodium and Serum Proteins at 17°-37°C.

BACKGROUND: We investigated the binding of talaporfin sodium with albumin and its photocytotoxicity during temperature changes by measuring absorbance spectra. The targeted tissue temperature differs according to the procedure. The photocytotoxicity efficiency should be investigated quantitatively because efficiency changes arising from temperature changes are expected. MATERIALS AND METHODS: The temperature dependence of talaporfin sodium binding with human serum albumin (0-20 mg/mL), high-density lipoprotein (0-0.04 mg/mL), and low-density lipoprotein (0-0.14 mg/mL) was investigated at 17°C, 27°C, and 37°C by measurement of absorbance spectra. Cell lethality was measured using a water-soluble tetrazolium-8 assay at 2 h after the photosensitization reaction at 17°C and 37°C. RESULTS: The binding ratios of talaporfin sodium with high-density lipoprotein decreased by 6.3% and those with low-density lipoprotein decreased by 12.8% when the temperature increased from 17°C to 37°C. Cell lethality increased significantly with a temperature rise from 17°C to 37°C at irradiation exposure of 20 and 40 J/cm2 and talaporfin sodium concentration of 20 µg/mL. CONCLUSIONS: From our in vitro data, we can predict that the change in photocytotoxicity efficiency would be negligible with a temperature decrease of <5°C from the body temperature in the case of photodynamic ablation with a short drug-light interval.

Changes of protein glycosylation in the course of radiotherapy.

This is the first study of changes in protein glycosylation due to exposure of human subjects to ionizing radiation. Site specific glycosylation patterns of 7 major plasma proteins were analyzed; 171 glycoforms were identified; and the abundance of 99 of these was followed in the course of cancer radiotherapy in 10 individual patients. It was found that glycosylation of plasma proteins does change in response to partial body irradiation (∼ 60 Gy), and the effects last during follow-up; the abundance of some glycoforms changed more than twofold. Both the degree of changes and their time-evolution showed large inter-individual variability.

The effectiveness of riboflavin photochemical-mediated virus inactivation and changes in protein retention in fresh-frozen plasma treated using a flow-based treatment device.

BACKGROUND: A flow-based treatment device using riboflavin and ultraviolet (UV) light was developed to inactivate viruses in fresh-frozen plasma (FFP). The objective of this study was to evaluate the in vitro effectiveness of virus inactivation and changes in protein quality in FFP treated with this device. STUDY DESIGN AND METHODS: FFP-contaminating viruses were treated with riboflavin and UV light using a one-pass linear flow device. The infectivity of viruses was measured using established biologic assays. Real-time polymerase chain reaction (PCR) was performed to detect damage to viral nucleotides after treatment. Treated plasma was analyzed using standard coagulation assays. RESULTS: FFP treated at the UV dose of 3.6 J/cm(2) (J) exhibited a mean reduction of virus titer of more than 4 logs. The effectiveness increased significantly at higher doses. Real-time PCR showed that the cycle threshold values for both complete inactivation and virus recultivation were higher than that of the untreated sample. At doses of 3.6, 5.4, and 7.2 J, the protein recovery rates were 60.2 ± 8.6, 46.6 ± 9.4, and 28.0 ± 1.0% for fibrinogen; 67.0 ± 3.1, 57.3 ± 8.0, and 49.2 ± 3.8% for Factor VIII; 93.6 ± 2.8, 89.6 ± 6.1, and 86.5 ± 5.3% for antithrombin-III; and 72.1 ± 5.6, 59.8 ± 14.2, and 49.2 ± 8.4% for Protein C, respectively. CONCLUSION: The effectiveness of virus inactivation was enhanced, but total activity of plasma factors was reduced, in a UV dose-dependent manner.

Photodynamic inactivation of enveloped virus in protein plasma preparations by solid-phase fullerene-based photosensitizer.

BACKGROUND: The problem of transfusion-transmitted infections still remains serious and actual for health care despite the detailed testing of donors. Human immunodeficiency virus, hepatitis B and C viruses and human cytomegalovirus are among the most dangerous pathogens that can be transmitted with blood. Previously, a composition consisting of fullerene layer applied on silica gel particles was shown to inactivate influenza virus up to complete loss of infectivity. METHODS: In the present study the unit has been developed with source of irradiation whose spectrum is appropriate for solid-phase fullerene. The ability of the unit to inactivate the enveloped influenza virus in protein fraction of donor blood has been studied. RESULTS: It was shown that at optimized conditions complete inactivation of enveloped virus of extremely high initial titer (7.0-9.5 log 10 EID 50/0.2 mL) in the solution of albumin was achieved after as short time as 30 min of irradiation. This process did not affect the oxidative metabolism of neutrophils and membranes of erythrocytes evaluated by NBT reduction test and morphological analysis of erythrocytes, respectively. CONCLUSION: The data obtained suggests that the method described can be recommended for further development and optimization of the procedure of inactivation of viruses in the preparations of the plasma of donor blood.

Profiling of low molecular weight proteins in plasma from locally irradiated individuals.

In studies reported in the 1960s and since, blood plasma from radiation-exposed individuals has been shown to induce chromosome damage when transferred into lymphocyte cultures of non-irradiated persons. This effect has been described to occur via clastogenic factors, whose nature is still mostly unknown. We have previously examined clastogenic factors from irradiated individuals by looking at plasma-induced DNA damage in reporter cells. Plasma was tested from ca. 30 locally exposed clinical patients receiving fractionated radiation treatment, as well as from three radiological accident victims exposed in 1994, albeit sampled 14 years post-accident. In the current work, proteome changes in the plasma from all subjects were examined with 2D gel electrophoresis-based proteomics techniques, in order to evaluate the level of protein expression with respect to the findings of a clastogenic factor effect. No differences were observed in protein expression due to local radiation exposure (pre- vs post-exposure). In contrast, plasma from the radiation accident victims showed alterations in the expression of 18 protein spots (in comparison with plasma from the control group). Among these, proteins such as haptoglobin, serotransferrin/transferrin, fibrinogen and ubiquitin-60S ribosomal protein L40 were observed, none of them likely to be clastogenic factors. In conclusion, the proteomics techniques applied were unable to identify changes in the proteome of the locally irradiated patients, whereas such differences were observed for the accident victims. However, association with the clastogenic effect or any specific clastogenic factor remains unresolved and thus further studies with more sensitive techniques are warranted.

Human whole-blood (1)H2O longitudinal relaxation with normal and high-relaxivity contrast reagents: influence of trans-cell-membrane water exchange.

PURPOSE: Accurate characterization of contrast reagent (CR) longitudinal relaxivity in whole blood is required to predict arterial signal intensity in contrast-enhanced MR angiography (CE-MRA). This study measured the longitudinal relaxation rate constants (R1 ) over a concentration range for non-protein-binding and protein-binding CRs in ex vivo whole blood and plasma at 1.5 and 3.0 Tesla (T) under physiologic arterial conditions. METHODS: Relaxivities of gadoteridol, gadobutrol, gadobenate, and gadofosveset were measured for [CR] from 0 to 18 mM [mmol(CR)/L(blood)]: the latter being the upper limit of what may be expected in CE-MRA. RESULTS: In plasma, the (1) H2 O R1 [CR]-dependence was nonlinear for gadobenate and gadofosveset secondary to CR interactions with the serum macromolecule albumin, and was well described by an analytical expression for effective 1:1 binding stoichiometry. In whole blood, the (1) H2 O R1 [CR]-dependence was markedly non-linear for all CRs, and was well-predicted by an expression for equilibrium exchange of water molecules between plasma and intracellular spaces using a priori parameter values only. CONCLUSION: In whole blood, (1) H2 O R1 exhibits a nonlinear relationship with [CR] over 0 to 18 mM CR. The nonlinearity is well described by exchange of water between erythrocyte and plasma compartments, and is particularly evident for high relaxivity CRs.

Serum lipids, proteins and electrolyte profiles in rats following total body irradiation.

OBJECTIVE: Serum lipid and electrolyte imbalances are common in critically ill patients undergoing radiation therapy. Although multiple disease states and medication may be responsible for the development of these disorders, the aim of this research is to sequentially document the effect of total body radiation on body function utilizing the sequential changes in the serum lipids, electrolytes and protein in rats. METHODS: Serum protein and lipids contents were assessed using kits while electrolytes were assessed with flame photometry in rats exposed to total body irradiations of 1.27 Gy/min in cumulative doses to the fourth irradiation at five-day intervals. RESULTS: Total cholesterol and triacylglycerols serum levels were significantly reduced by irradiation (p < 0.05). No significant differences between experimental and control groups for HDL-C serum levels were detected. Serum electrolyte concentration remained within the normal range after each total body irradiation. Sodium, bicarbonate and chloride were significantly (p < 0.05) higher than control while potassium and creatinine were significantly reduced after the first irradiation only. Sodium/potassium ratio was significantly (p < 0.05) elevated. Serum protein was significantly (p < 0.05) elevated with increasing radiation. CONCLUSION: There are subtle but significant changes in serum lipids, electrolytes and protein after total body irradiation of normal rats. These variations could be due to non-specific stress reactions; as such, they are important markers in radiation induced injury diagnosis.

Serum lipids, proteins and electrolyte profiles in rats following total body irradiation / Perfiles de lípidos, proteínas y electrolitos plasmáticos en ratas tras irradiación corporal total

OBJECTIVE: Serum lipid and electrolyte imbalances are common in critically ill patients undergoing radiation therapy. Although multiple disease states and medication may be responsible for the development of these disorders, the aim of this research is to sequentially document the effect of total body radiation on body function utilizing the sequential changes in the serum lipids, electrolytes and protein in rats. METHODS: Serum protein and lipids contents were assessed using kits while electrolytes were assessed with flame photometry in rats exposed to total body irradiations of 1.27 Gy/min in cumulative doses to the fourth irradiation at five-day intervals. RESULTS: Total cholesterol and triacylglycerols serum levels were significantly reduced by irradiation (p < 0.05). No significant differences between experimental and control groups for HDL-C serum levels were detected. Serum electrolyte concentration remained within the normal range after each total body irradiation. Sodium, bicarbonate and chloride were significantly (p < 0.05) higher than control while potassium and creatinine were significantly reduced after the first irradiation only. Sodium/potassium ratio was significantly (p < 0.05) elevated. Serum protein was significantly (p < 0.05) elevated with increasing radiation. CONCLUSION: There are subtle but significant changes in serum lipids, electrolytes and protein after total body irradiation of normal rats. These variations could be due to non-specific stress reactions; as such, they are important markers in radiation induced injury diagnosis.

OBJETIVO: Los desequilibrios de lípido y electrolito plasmáticos son comunes en los pacientes críticos sometidos a terapia radioactiva. Aunque los múltiples estados de la enfermedad y la medicación pueden ser responsables del surgimiento de estos trastornos, el objetivo de esta investigación es documentar de manera secuencial el efecto de la radiación corporal total sobre la función corporal, utilizando los cambios secuenciales en los lípidos, electrolitos y proteínas plasmáticos en las ratas. MÉTODOS: Los contenidos de lípidos y proteínas plasmáticos fueron evaluados utilizando kits, en tanto que los electrolitos fueron evaluados mediante fotometría de llama en ratas expuestas a irradiaciones corporales totales de rayos X de 1.27 Gy/min, en dosis cumulativas hasta la cuarta irradiación en intervalos de cinco días. RESULTADOS: El colesterol total y los niveles plasmáticos de triacilgliceroles fueron reducidos significativamente por la irradiación (p < 0.05). No se detectaron diferencias significativas entre; os grupos experimentales y de control en relación con los niveles plasmáticos de colesterol HDL. La concen-tración de electrolito plasmático se mantuvo dentro de los límites normales luego de cada irradiación corporal total de rayos X. La relación sodio/potasio fue significativamente elevada (p < 0.05). La proteína plasmática se elevaba significativamente (p < 0.05) al aumentar la radiación. CONCLUSIÓN: Tras la irradiación corporal total de las ratas normales, seproducen cambios sutiles pero significativos en los lípidos, electrolitos y proteínas del plasma. Estas variaciones podrían ser debidas a reacciones de estrés no específicas, y como tal, son marcadores importantes en el diagnóstico de las lesiones inducidas por la radiación.

Low-intensity infrared laser increases plasma proteins and induces oxidative stress in vitro.

Low-intensity laser therapy is based on the excitation of endogenous chromophores in biotissues and free-radical generation could be involved in its biological effects. In this work, the effects of the low-intensity infrared laser on plasma protein content and oxidative stress in blood from Wistar rats were studied. Blood samples from Wistar rats were exposed to low-intensity infrared laser in continuous wave and pulsed-emission modes at different fluencies. Plasma protein content and two oxidative stress markers (thiobarbituric acid-reactive species formation and myeloperoxidase activity) were carried out to assess the effects of laser irradiation on blood samples. Low-intensity infrared laser exposure increases plasma protein content, induces lipid peroxidation, and increases myeloperoxidase activity in a dose- and frequency-dependent way in blood samples. The low-intensity infrared laser increases plasma protein content and oxidative stress in blood samples, suggesting that laser therapy protocols should take into account fluencies, frequencies, and wavelengths of the laser before beginning treatment.

[Assessment of bronchopulmonary immunity status for Mayak PA workers at the late period after prolonged exposure].

Samples of induced sputum from 187 individuals were used to estimate the bronchopulmonary immunity status after a prolonged radiation exposure (120 individuals who are the Mayak main plant workers were exposed to combined internal a-radiation due to incorporated plutonium (239Pu) and external y-radiation during their career). The control group included 67 Ozersk residents of a corresponding age and gender, without any occupational exposure, who were examined at the same period. The immune system is the most important component in homeostasis. In occupational workers, the status of the systemic and bronchopulmonary immunity after 30-55 years of career was estimated by the level of regulatory proteins in the samples of blood and the induced sputum supernatant stored at low temperatures in freezers of the Radiobiological human tissue Repository, SUBI (interleukins IL-1beta, IFN-gamma, IL-12+p70, IL-15, IL-17A and growth factors EGF, TGF-beta1, FGF and PDGF-AA), as well as by the content of effector and regulatory lymphocytes in blood. After the examination period, each registrant was provided with dosimetry data: 239Pu body burden was from 0.03 kBq to 11.89 kBq, the absorbed dose to the lung from incorporated radionuclide - from 0.01 Gy to 1.38 Gy, and accumulated doses of external y-radiation during the occupational career - from 0.02 Gy to 7.91 Gy. In the case of the prolonged combined radiation, the tensioned mode of immunity cell link functioning and the increase of activated lymphocyte forms were detected. Comparison of correlation coefficients between the content oflymphocytes, the level ofinterleukins and growth factors and the dose load allowed us to find relation of these ratios to the type of exposure, Pu body burden, the absorbed 239Pu dose to the lung and the accumulated external dose. The reduced concentration of the main growth factors in the group of Mayak workers is the evidence for the declined control function of proteins and immunodeficiency. The analysis of the regulatory proteins content in blood and sputum following occupational exposure allowed us to find specific features of the protein expression in support of the local and systemic immune homeostasis.

Radiation-related changes in serum proteome profiles detected by mass spectrometry in blood of patients treated with radiotherapy due to larynx cancer.

The study aimed to detect features of human serum proteome that were associated with exposure to ionizing radiation. The analyzed group consisted of 46 patients treated with radical radiotherapy for larynx cancer; patients were irradiated with total doses in a range from 51 to 72 Gy. Three consecutive blood samples were collected from each patient: before the start, 2 weeks after the start, and 4-6 weeks after the end of radiotherapy. The low-molecular-weight fraction of the serum proteome (2,000-13,000 Da) was analyzed by the MALDI-ToF mass spectrometry. Proteome profiles of serum samples collected before the start of radiotherapy and during the early stage of the treatment were similar. In marked contrast, mass profiles of serum samples collected several weeks after the end of the treatment revealed clear changes. We found that 41 out of 312 registered peptide ions changed their abundance significantly when serum samples collected after the final irradiation were compared with samples collected at the two earlier time points. We also found that abundances of certain serum peptides were associated with total doses of radiation received by patients. The results of this pilot study indicate that features of serum proteome analyzed by mass spectrometry have potential applicability as a retrospective marker of exposure to ionizing radiation.

Protein stability of previously frozen plasma, riboflavin and UV light-treated, refrozen and stored for up to 2 years at -30 °C.

BACKGROUND: The Mirasol pathogen reduction technology system for plasma is based on a riboflavin and UV light treatment process resulting in pathogen inactivation due to irreversible, photochemically induced damage of nucleic acids. The objective of this study was to evaluate the in vitro protein quality of previously frozen FFP, thawed, Mirasol-treated and refrozen for a final storage period of up to 2 years at -30 °C. MATERIALS AND METHODS: All plasma products were held at 4 ± 2 °C, and frozen within 8h after the start of collection. FFP was stored frozen at -30°C for eight time intervals before riboflavin and UV light treatment and refrozen to -30 °C and stored for varying time intervals for a total storage period of up to 2 years at -30 °C. Results were compared to paired, untreated units thawed and stored for the same time intervals. RESULTS: The overall mean values for all time points in riboflavin and UV light treated plasma samples indicates that all proteins were well preserved following a period of frozen storage for 2 years at -30 °C. Factors V, VII and XI, retained 70%, 65% and 53% activity, respectively. All other protein factors, anticoagulant and inhibitor proteins demonstrated retention between 74% and 100%. CONCLUSION: Riboflavin and UV light-treated FFP maintained both coagulant and anticoagulant in vitro protein quality after double freeze/thaw storage at -30 °C for up to 2 years, a finding that may offer processing flexibility to blood centers.