Non-irradiated Research Articles

Adipose-derived Stem Cells and Wound Healing Are Progressively Impaired Long-term After Radiotherapy in Mice.

The pathogenesis of deterministic radiation damage is not clearly understood, but it has been reported that fibroinflammatory pathways are up-regulated. We hypothesized that the number of adipose-derived stem/stromal cells (ASCs) decline after radiotherapies, preventing normalization of fibrosis and angiogenesis, resulting in chronic radiation damages that progress over time. Dorsal skin of 8-week-old male BALB/cfC3H mice was irradiated with 10 Gy weekly for 4 weeks. At 1, 3, 6, 9, and 12 months after radiotherapy (n = 5, 5, 5, 5, and 4), tissue hemoglobin oxygen saturation, and time until epithelialization were evaluated. Skin biopsies were measured for thickness and CD34+/isolectin- stem/stromal cell count. Nonirradiated (NRT) controls were evaluated at each time point as well (n = 5 each). Compared with NRT controls, time until epithelialization was significantly longer at 1 month (28 ± 3, P < 0.01); not statistically different at 3 months (16 ± 2, P = 0.32); and lengthened over time at 6 months (20 ± 2, P = 0.21), 9 months (28 ± 2, P < 0.01), and 12 months (26 ± 3, P < 0.01), as did tissue oxygen saturation. The number of CD34+/isolectin- ASCs decreased over time, at 1 month (5.3 ± 1.3, P = 0.01), 3 months (6.0 ± 1.4, P = 0.03), 6 months (4.0 ± 0.8, P < 0.01), 9 months (1.7 ± 0.5, P < 0.01), and 12 months (0.3 ± 0.5, P < 0.01). The subcutaneous fatty layer was significantly thinner at 3 months (116 ± 33, P < 0.01), 6 months (147 ± 22, P = 0.02), 9 months (52 ± 12, P = 0.04), and 12 months (89 ± 19, P = 0.04), but not at 1 month (141 ± 18, P = 0.43). After 6 months postirradiation, the number of ASCs continued to decline over time, accompanied by irreversible progression of fibrosis, atrophy, and ischemia, which resulted in impaired wound healing.

Retention of ZnO nanoparticles onto polypropylene and polystyrene microplastics: Aging-associated interactions and the role of aqueous chemistry

Microplastics (MPs) are pervasive and undergo environmental aging processes, which alters potential interaction with the co-contaminants. Hence, to assess their contaminant-carrying capacity, mimicking the weathering characteristics of secondary MPs is crucial. To this end, the present study investigated the interaction of Zinc oxide (nZnO) nanoparticles with non-irradiated (NI) and UV-irradiated (UI) forms of the most abundant MPs, such as polypropylene (PP) and polystyrene (PS), in aqueous environments. SEM images revealed mechanical abrasions on the surfaces of NI-MPs and their subsequent photoaging caused the formation of close-ended and open-ended cracks in UI-PP and UI-PS, respectively. Batch-sorption experiments elucidated nZnO uptake kinetics by PP and PS MPs, suggesting a sorption-desorption pathway due to weaker and stronger sorption sites until equilibrium was achieved. UI-PP showed higher nZnO (∼3000 mg/kg) uptake compared to NI-PP, while UI-PS showed similar or slightly decreased nZnO (∼2000 mg/kg) uptake compared to NI-PS. FTIR spectra and zeta potential measurements revealed electrostatic interaction as the dominant interaction mechanism. Higher nZnO uptake by MPs was noted between pH 6.5 and 8.5, whereas it decreased beyond this range. Despite DOM, MPs always retained ∼874 mg/kg nZnO irrespective of MPs type and extent of aging. The experimental results in river water showed higher nZnO uptake on MPs compared to DI water, attributed to mutual effect of ionic competition, DOM, and MP hydrophobicity. In the case of humic acids, complex synthetic and natural water matrices, NI-MPs retained more nZnO than UI-MPs, suggesting that photoaged MPs sorb less nZnO under environmental conditions than non-photoaged MPs. These findings enhance our understanding on interaction of the MPs with co-contaminants in natural environments.

FLASH Proton Radiation Therapy Mitigates Inflammatory and Fibrotic Pathways and Preserves Cardiac Function in a Preclinical Mouse Model of Radiation-Induced Heart Disease

PurposeStudies during the past 9 years suggest that delivering radiation at dose rates exceeding 40 Gy/s, known as “FLASH” radiotherapy, enhances the therapeutic index of radiation therapy (RT) by decreasing normal tissue damage while maintaining tumor response compared to conventional (or standard) RT. This study demonstrates the cardioprotective benefits of FLASH proton RT (F-PRT) compared to standard (conventional) proton RT (S-PRT), as evidenced by reduced acute and chronic cardiac toxicities. Methods and MaterialsMice were imaged using cone beam computed tomography to precisely determine the heart's apex as the beam isocenter. Irradiation was conducted utilizing a shoot-through technique with a 5mm diameter circular collimator. Bulk RNA-sequencing was performed on non-irradiated (NR) samples, as well as apexes treated with F-PRT or S-PRT, at 2 weeks following a single 40 Gy dose. Inflammatory responses were assessed through multiplex cytokine/chemokine microbead assay and immunofluorescence analyses. Levels of perivascular fibrosis were quantified using Masson's Trichrome and Picrosirius red staining. Additionally, cardiac tissue functionality was evaluated by 2D echocardiograms at 8 and 30 weeks post-PRT. ResultsRadiation damage was specifically localized to the heart's apex. RNA profiling of cardiac tissues treated with PRT revealed that S-PRT uniquely upregulated pathways associated with DNA damage response, induction of tumor necrosis factor superfamily and inflammatory response, while F-PRT primarily affected cytoplasmic translation, mitochondrion organization and ATP synthesis. Notably, F-PRT led to a milder inflammatory response, accompanied by significantly attenuated changes in TGF-β1 and αSMA levels. Critically, F-PRT decreased collagen deposition, and better preserved cardiac functionality compared to S-PRT. ConclusionsThis study demonstrated that F-PRT reduces the induction of an inflammatory environment with lower expression of inflammatory cytokines and profibrotic factors. Importantly, the results indicate that F-PRT better preserves cardiac functionality, as confirmed by echocardiography analysis, while also mitigating the development of long-term fibrosis.

Effect of Periodontal Instrumentation on Tooth Structure Exposure to Ionizing Radiation: In Vitro Study.

This study aimed to evaluate the effects of different scaling protocols on the morphology and roughness of the root dentin substrate exposure to ionizing radiation. One hundred and thirty extracted bovine incisors were randomly divided into two groups (n=65): non-irradiated (NIR) and irradiated (IR). Each group was initially subdivided into three subgroups according to the type of non-surgical periodontal protocol: NIT: no instrumentation; HS: hand scaling with 15 apical-coronal instrument movements; US: ultrasonic scaling with 15 apical-coronal cycles. Subsequently, all samples were subjected to the prophylaxis protocol, being subdivided into the following groups: NIT/PP: prophylaxis with a fine prophylactic paste using a rubber cup for 15 seconds; HS/PP: Hand scaling followed by the prophylaxis protocol; US/PP: Ultrasonic scaling followed by the prophylaxis protocol. The roughness of the root dentin surface was measured with a profilometer (Ra/Rz - μm), and the morphology of the dentin surfaces was analyzed using scanning electron microscopy (SEM). The analyses were conducted before and after the prophylaxis protocol. In the absence of prophylaxis, the roughest surfaces were observed after ultrasonic instrumentation followed by hand instrumentation for both IR and NIR groups. No difference in Ra and RZ values between HS/PP and US/PP was observed for both substrates. For the IR group, the prophylaxis resulted in similar Ra and RZ values for both instrumentation groups in comparison to no instrumentation. Ordinal logistic regression showed that both HS and US resulted in higher scores than NIT, irrespective of IR presence. In conclusion, the IR showed a rougher root surface for both HS and US in comparison to NIR. However, the prophylaxis procedure significantly reduced the roughness of root surfaces after both instrumentation procedures.

Effect of crosslinking/antioxidant agents as final irrigant on the fracture resistance of endodontically treated root after radiotherapy.

To investigate the effects of Epigallocatechin gallate (EGCG) and curcumin, as a final irrigant on the fracture resistance of irradiated root that obturated with an epoxy resin sealer. Eighty mandibular premolars were randomly divided into non-irradiated (NIR) and irradiated (IR) groups. The teeth were irradiated at 2 Gy per fraction, 5 times a week for a total dose of 60 Gy over 6 weeks. All specimens were decoronated, remaining 13±1 mm root length. Two groups were subdivided into four groups (n = 10): 1) non-instrumented; the intact root served as control. The other roots were instrumented with a pro-taper NiTi rotary system. The final irrigation used was 17% EDTA, followed by three irrigation solution groups; 2) 2.5% NaOCl, 3) 0.02% EGCG, and 4) 0.1% curcumin. Root canals were filled with gutta-percha and AH plus. All specimens were embedded in self-curing acrylic resin and loaded vertically at 1 mm/min until fracture occurred. Also, sealer penetration was assessed by confocal laser scanning microscopy (CLSM). The data were evaluated statistically using two-way ANOVA and Tukey test (α = 0.05). In irradiated roots, fracture resistance of EGCG and curcumin groups did not differ from non-instrumented roots, but they were higher than the NaOCl group (P = 0.006). However, NaOCl, EGCG, and curcumin in irradiated roots had comparable strength that was higher than in the non-instrumented group (p<0.001). Difference between irradiated and non-irradiated roots was observed only for NaOCl and non-instrumented groups (P≤0.004). In irradiated roots, a higher sealer penetration was observed in EGCG and curcumin groups compared to NaOCl. EGCG and curcumin could be promising final irrigants to reverse the adverse effect of radiotherapy on the strength of irradiated roots obturated with AH Plus sealer.

Chemical and morphological changes of femtosecond laser-irradiated enamel using subablative parameters.

Chemical composition of dental enamel has a great relationship with the prevention of caries. The objective of the present work was to evaluate the chemical and morphological changes of femtosecond laser-irradiated enamel with subablative parameters using Raman spectroscopy, confocal laser scanning microscopy (CLSM), and scanning electron microscopy (SEM). Bovine incisor teeth were used to obtain 30 enamel specimens (5 × 5 mm2 ). The chemical composition of the control sample was analyzed by Raman spectrometry to acquire the absorption spectrum, delimiting the areas under the carbonate and phosphate bands. This analysis was used to evaluate the change in the chemical composition of the sample after irradiation. The specimens were irradiated (IRR) with a Ti:Sapphire laser system (pulsed and focused modes, femtosecond regime 70 fs, average power of 1W and exposure time of 15 s). After irradiation, the areas under the carbonate and phosphate absorption bands were delimited in each specimen. Raman spectrometry data were analyzed using Student's t-test (α=5%). By comparing the spectra of the IRR and non-irradiated (NI) specimens, the results showed a significant increase in the area value for the phosphate peaks and a significant reduction in the area value for the carbonate peak and the carbonate:phosphate ratio. CLSM and SEM analyses did not reveal structural alterations in the subsurface nor morphological alterations in the IRR enamel surface, respectively. It was concluded that femtosecond laser irradiation using subablative parameters reduced the carbonate content and the carbonate/phosphate ratio without altering the structure and morphology of the dental enamel.

TP53 drives abscopal effect by secretion of senescence-associated molecular signals in non-small cell lung cancer

BackgroundRecent developments in abscopal effect strongly support the use of radiotherapy for the treatment of metastatic disease. However, deeper understanding of the molecular mechanisms underlying the abscopal effect are required to best benefit a larger proportion of patients with metastasis. Several groups including ours, reported the involvement of wild-type (wt) p53 in radiation-induced abscopal effects, however very little is known on the role of wtp53 dependent molecular mechanisms.MethodsWe investigated through in vivo and in vitro approaches how wtp53 orchestrates radiation-induced abscopal effects. Wtp53 bearing (A549) and p53-null (H1299) NSCLC lines were xenotransplanted in nude mice, and cultured in 2D monolayers and 3D tumor spheroids. Extracellular vesicles (EVs) were isolated from medium cell culture by ultracentrifugation protocol followed by Nanoparticle Tracking Analysis. Gene expression was evaluated by RT-Real Time, digital qRT-PCR, and dot blot technique. Protein levels were determined by immunohistochemistry, confocal anlysis, western blot techniques, and immunoassay.ResultsWe demonstrated that single high-dose irradiation (20 Gy) induces significant tumor growth inhibition in contralateral non-irradiated (NIR) A549 xenograft tumors but not in NIR p53-null H1299 or p53-silenced A549 (A549sh/p53) xenografts. We further demonstrates that irradiation of A549 cells in vitro induces a senescence-associated secretory phenotype (SASP) producing extracellular vesicles (EVs) expressing CD63 and carrying DNA:RNA hybrids and LINE-1 retrotransposon. IR-A549 EVs also hamper the colony-forming capability of recipient NIR A549 cells, induce senescent phenotype, nuclear expression of DNA:RNA hybrids, and M1 macrophage polarization.ConclusionsIn our models, we demonstrate that high radiation dose in wtp53 tumors induce the onset of SASP and secretion of CD63+ EVs loaded with DNA:RNA hybrids and LINE-1 retrotransposons that convey senescence messages out of the irradiation field triggering abscopal effect in NIR tumors.

Effects of fractionation and ionizing radiation dose on the chemical composition and microhardness of enamel

ObjectiveTo evaluate the chemical and mechanical properties of enamel submitted to different in vitro radiation protocols. DesignThird molars were divided into seven groups (n = 8): non-irradiated (NI); a single dose of 30 Gy (SD30), 50 Gy (SD50), or 70 Gy (SD70) of radiation; or fractional radiation doses of up to 30 Gy (FD30), 50 Gy (FD50), or 70 Gy (FD70). Hemisections were analysed by Fourier transform infrared spectroscopy (FTIR), energy dispersive X-ray spectroscopy (EDS) and Knoop microhardness (KHN) test. One-way ANOVA followed by Bonferroni's post-hoc test compared the test groups with the NI. Two-way ANOVA was performed for the fractionation and radiation dose, followed by Bonferroni’s test (α = 0.05). ResultsFTIR revealed differences for the amide I band between the NI and FD50 and NI and FD70 groups (p < 0.001). For the organic matrix/mineral ratio, the FD70 group presented a lower ratio compared to NI (p = 0.009). Excluding the NI group, there were differences between the FD30 and FD50 (p = 0.045) and the FD30 and FD70 groups (p < 0.001). For EDS, there were differences for Ca (p = 0.011) and Ca/P (p < 0.001), with the FD70 group presenting lower values compared to NI (p = 0.015; p < 0.001). For KHN, the FD70 group presented lower values than the NI (p = 0.002). Two-way ANOVA showed difference for the dose (p < 0.001), with the 70 Gy group presenting a lower KHN value within the fractionated groups. ConclusionFractional doses 70 Gy irradiation caused chemical and mechanical changes to enamel. Radiation applied in single or fractional doses produced different effects to enamel.

The secretome of non-tumorigenic mammary cells MCF-10A elicits DNA damage in MCF-7 and MDA-MB-231 breast cancer cells

Radiotherapy is used in breast cancer to destroy tumor cells lingering after surgery. It is accepted that lethal effects of ionizing radiation occur as a result of damage to DNA in irradiated (IR) cells. However, response mechanisms may promote cell survival with efficient DNA repair or genomic alterations. Chromosomal aberrations are frequent in surviving cells and may enhance chromosomal instability (CIN) which is associated with increased risk of recurrence and metastasis. Intercellular communication can affect the response in IR cells and cause damage in non-irradiated (N-IR) cells. We evaluated the effect of the secretome of non-tumorigenic mammary cells (MCF-10A) on proliferation and DNA damage in breast cancer cells (MCF-7 and MDA-MB-231). Results showed that conditioned media from IR and N-IR MCF-10A cells produced cycles of DNA double-strand breaks in N-IR and IR tumor cells leaving them with residual damage. CIN markers (micronuclei, nucleoplasmic bridges, nuclear buds) were also increased in IR and N-IR tumor cells, being the effect of conditioned media from IR MCF-10A greater in many cases. The inhibition of phosphorylation/activation of Src kinase in cancer cells hindered CIN markers' increment. Besides, clonogenic survival of tumor cells was differentially modulated by conditioned media from MCF-10A: decreased in MCF-7 and enhanced in MDA-MB-231 cells. These results signal the relevance of tumor-host interaction in tumor behavior and the response to radiotherapy.

In vivo and in vitro radiotherapy increased dentin enzymatic activity

ObjectivesThe present study investigated the effects of in vitro and in-vivo radiotherapy on endogenous enzymatic activity in dentin using gelatin zymography and in-situ zymography. MethodsGelatin zymographic assays were performed on protein extracts obtained from dentin powder of sound non-irradiated (NRT), in vitro irradiated (VTRT) and in vivo irradiated (VIRT) human teeth. Their proteolytic activities were quantified using band densitometric evaluation. For in-situ zymography, dentin specimens from NRT, VIRT and VTRT were covered with fluorescein-conjugated gelatin and examined with confocal laser-scanning microscopy. Fluorescence intensity emitted by the hydrolyzed fluorescein-conjugated gelatin was quantified and statistically analyzed. In-situ zymography data were statistically analyzed using Kruskal-Wallis ANOVA and Dunn’s multiple comparison procedures (α = 0.05). ResultsNo difference between in vitro and in vivo radiotherapy treatment was found. Both VTRT and VIRT groups showed increase in MMP-9 expression when compared to NRT group. Significant increases (p < 0.05) in gelatinolytic activity (26 % for VTRT; 55 % for VIRT) were observed when compared to the NRT group. ConclusionRadiotherapy increase endogenous enzymatic activity in non-restored dentin.

Post Mastectomy Radiation for Stage II Breast Cancer Patients with T1/T2 Lesions.

Post mastectomy radiation (PMR) is usually recommended for T3 or N2 breast cancer (BC). The role of PMR for stage II BC with T1/T2 lesions remains controversial. The aim of this study was to assess the role of PMR in this subgroup of patients. A retrospective analysis of a prospectively collected database of all stage II BC patients treated with mastectomy at our institution between the years 2005-2008 was performed. Demographics, disease-free survival rates were compared between the patients receiving radiation vs. those who were not irradiated. Eighty-two patients underwent mastectomies for stage II disease with a T1/T2 lesion. Twenty-two of those (27%) received PMR. Loco regional recurrence (LRR) occurred only in the non -irradiated (NR) group. A Kaplan Meier analysis of time to LRR in the NR group was performed. Mean time to local failure was 78.9 months, 6% at 3 years and 13% at 5 years. The time to LRR was significantly lower in the estrogen receptor (ER) negative group compared to the ER positive group (64 vs. 82 months, p=0.029). LRR free rate at 5 years was 100% in low grade tumors vs. 53% in high grade tumors, (p=0.001). In a Cox regression multivariate analysis none of those factors maintained significance. ER negative status, high grade and node negativity were associated with LRR. A prospective trial randomizing stage II BC patients with T1/T2 lesions, negative hormone receptors and high-grade tumors to PMR following mastectomy arm vs. no radiation arm is recommended.

Comparative Analysis of Mitochondrial Proteome Reveals the Mechanism of Enhanced Ram Sperm Motility Induced by Carbon Ion Radiation After In Vitro Liquid Storage.

The aim of this study was to reveal the mechanism of enhanced ram sperm motility induced by heavy ion radiation (HIR) after in vitro liquid storage. Ram semen was stored for 24 hours at 5°C and then irradiated with 0.1 Gy carbon ion radiation (CIR). In comparison to nonirradiated (NIR) sperm, the motility, viability, and adenosine triphosphate content were all higher in CIR sperm, and the reactive oxygen species levels were lower. Moreover, 87 differential mitochondrial protein spots were detected in 2-dimensional gels between CIR and NIR sperm and were identified as 52 corresponding proteins. In addition, 33 differential proteins were involved in a main pathway network, including COX5B, ERAB/HSD17B10, ETFA, SDHB, and SOD2, which are known to be involved in cell communication, energy production, and antioxidant responses. We used immunoblotting and immunofluorescence to analyze the content and localization of these proteins, respectively, and the levels of these proteins in CIR sperm were lower than those in NIR sperm. An understanding of the molecular function of these proteins could provide further insight into the mechanisms underlying high sperm motility induced by HIR in rams.

Enhanced survival of lethally irradiated mice given pegylated G-CSF, GM-CSF, and IL-11 with lisinopril via modulation of the hematopoietic cytokines TGF[Symbol]-1 and RANTES

Background and Hypothesis: We have previously shown increased survival in mice given pegylated growth factors, G-CSF, GM-CSF, and IL-11 (triple combo, TC), with lisinopril compared to TC alone in the hematopoietic acute radiation syndrome (H-ARS). This experiment investigated the mechanism behind the survival benefit. We hypothesized lisinopril regulates inflammatory cytokines hence increasing quiescence of primitive bone marrow cells. Experimental Design or Project Methods: C57BL/6 mice were exposed to LD90/30 total body irradiation (TBI). The irradiated mice were given a TC injection on day 1, and lisinopril was administered in drinking water beginning on day 7. Mice were euthanized on day 10. Flow cytometry was used to analyze bone marrow cell cycle and Bioplex was used to analyze cytokines in bone marrow supernatant. Results: Treatment with TC+lisinopril significantly decreased the level of the pro-inflammatory cytokine RANTES compared with vehicle mice (Veh) and TC alone (TC+Liso vs. Veh: 3.6 vs. 7.8 pg/mL, p&lt;0.01; TC+Liso vs. TC: 3.6 vs. 5.2 pg/ml, p&lt;0.01). TC+lisinopril increases levels of the pro-quiescence cytokine TGF-[Symbol]1, compared with Veh and TC alone (TC+Liso vs. Veh: 92.8 vs. 45.6 pg/mL, p&lt;0.05; TC+Liso vs. TC: 92.8 vs. 44.7 pg/ml, p&lt;0.01). TC alone had lower percentages of quiescent Lin- primitive bone marrow cells (G0+G1) than non-irradiated (NI) mice (75.7 vs. 85.2%, p&lt;0.05), whereas there is no difference between TC+Liso and NI mice (83.0 vs. 85.2%, p=0.44). Conclusion and Potential Impact: The increased survival of H-ARS mice given lisinopril might be due to the regulation of TGF-[Symbol]1 and RANTES levels in the bone marrow leading to quiescence of primitive bone marrow cells.

Effect of irradiation energy and residence time on decomposition efficiency of polycyclic aromatic hydrocarbons (PAHs) from rubber wood combustion emission using soft X-rays

This research aims to investigate the effects of irradiation energy and residence time of soft X-ray irradiation in the decomposition of sixteen polycyclic aromatic hydrocarbons (PAHs) in smoke particles emitted from the Para rubber wood burning. The burning process was carried out in a tube furnace and the soft X-ray radiation used had a wave length of 0.13–0.41 nm. The irradiated (IR) and non irradiated (N-IR) smoke particles were collected simultaneously using a 10-stage Andersen sampler equipped with an inertial-filter stage (ANIF), in order to determine the physicochemical characteristic of both IR and N-IR smoke particles, including particle size distribution and concentration, and particle-bound PAHs concentration. Results show that the nano-size smoke particles contained the highest amount of PAHs and of carcinogenic potency equivalent (BaP-TEQ). About 75% of PAH compounds on the total smoke particles were decomposed at the highest irradiation energy. Moreover, 4–6 ring PAHs in nanoparticles (<70 nm) were decomposed of up to about 91% at the highest irradiation energy. The decomposition efficiency of PAHs was influenced by particle size, PAHs boiling temperature and irradiation energy. It was higher for PAHs with lower boiling temperature and smaller size particles, while the effect of residence time was not significant.

Glass Ionomer Cements can be used for Bonding Orthodontic Brackets After Cancer Radiation Treatment?

Patients undergoing radiotherapy treatment present more susceptibility to dental caries and the use of an orthodontic device increases this risk factor due to biofilm accumulation around the brackets. The objective of this study was to evaluate the shear bond strength to irradiated permanent teeth of orthodontic brackets bonded with conventional glass ionomer cement and resin-modified glass ionomer cement due to the fluoride release capacity of these materials. Ninety prepared human premolars were divided into 6 groups (n=15), according to the bonding material and use or not of radiation: CR: Transbond XT composite resin; RMGIC: Fuji Ortho LC conventional glass ionomer cement; GIC: Ketac Cem Easymix resin-modified glass ionomer cement. The groups were irradiated (I) or non-irradiated (NI) prior to bracket bonding. The specimens were subjected to a fractioned radiation dose of 2 Gy over 5 consecutive days for 6 weeks. After the radiotherapy, the brackets were bonded on the specimens with Transbond XT, Fuji Ortho LC and Ketac Cem Easymix. After 24 h, the specimens were subjected to shear bond strength test. The image of enamel surface (classified by Adhesive Remnant Index - ARI) was also evaluated and its frequency was checked among groups/subgroups. The shear bond strength variable was evaluated with ANOVA and Tukey's post-hoc test. GIC group showed the lowest adhesion values among the groups (p<0.05). There was no statistically significant difference among non-irradiated and irradiated groups (p>0.05). As for the ARI, the CR-I group showed the highest material retention on enamel surface among the irradiated groups. RMGIC group showed the highest values for shear bond strength and presented ARI acceptable for clinical practices.

Effect of ionizing radiation on properties of restorative materials

ObjectiveTo evaluate the effect of ionizing radiation from high energy X-ray on properties of restorative materials. MethodsStudy materials (3M-ESPE) were: Z250—microhybrid resin-based composite (Filtek Z-250); Z350—nanofilled resin-based composite (Filtek Z-350XT); VIT—resin-modified glass ionomer cement (Vitremer); and KME—conventional glass ionomer cement (Ketac Molar Easymix). Sixty bar-shaped and cylinder-shaped specimens were fabricated from each material. Specimens were light activated (980mW/cm2, Radii, SDI) for 60s (3×20s for Z250 and Z350) and 120s (3×40s for VIT) and thirty specimens from each shape were irradiated (IR) with 1.8Gy/day for 39days (total IR=70.2Gy). IR and non-irradiated (NI) specimens were evaluated for flexural strength (σ, n=30) followed by fractography (SEM), diametral tensile strength (DTS, n=30), hardness (H, n=10), surface roughness (Ra, n=10) and chemical composition (n=3). The IR effect on each material property was statistically analyzed using Student’s t test (α=0.05). Data from σ and DTS were also analyzed using Weibull statistics. ResultsIR significantly increased the mean σ values of VIT and KME and the mean DTS value of VIT (p<0.05). IR increased Ra and H values for VIT and decreased H value for Z-250 (p<0.05). The remaining materials and properties were not significantly affected by IR (p>0.05). There was no significant change on materials composition after IR. SignificanceThe recommended radiotherapy protocol for head and neck cancer altered some material properties, mainly for glass ionomer cements. Such variations on material properties are not related to chemical composition changes.

Effects of gamma irradiation on the biomechanical properties of peroneus tendons.

PurposeThis study was designed to investigate the biomechanical properties of nonirradiated (NI) and irradiated (IR) peroneus tendons to determine if they would be suitable allografts, in regards to biomechanical properties, for anterior cruciate ligament reconstruction after a dose of 1.5–2.5 Mrad.MethodsSeven pairs of peroneus longus (PL) and ten pairs of peroneus brevis (PB) tendons were procured from human cadavers. The diameter of each allograft was measured. The left side of each allograft was IR at 1.5–2.5 Mrad, whereas the right side was kept aseptic and NI. The allografts were thawed, kept wet with saline, and attached in a single-strand fashion to custom freeze grips using liquid nitrogen. A preload of 10 N was then applied and, after it had reached steady state, the allografts were pulled at 4 cm/sec. The parameters recorded were the displacement and force.ResultsThe elongation at the peak load was 10.3±2.3 mm for the PB NI side and 13.5±3.3 mm for the PB IR side. The elongation at the peak load was 17.4±5.3 mm for the PL NI side and 16.3±2.0 mm for the PL IR side. For PL, the ultimate load was 2,091.6±148.7 N for NI and 2,122.8±380.0 N for IR. The ultimate load for the PB tendons was 1,485.7±209.3 N for NI and 1,318.4±296.9 N for the IR group. The ultimate stress calculations for PL were 90.3±11.3 MPa for NI and 94.8±21.0 MPa for IR. For the PB, the ultimate stress was 82.4±19.0 MPa for NI and 72.5±16.6 MPa for the IR group. The structural stiffness was 216.1±59.0 N/mm for the NI PL and 195.7±51.4 N/mm for the IR side. None of these measures were significantly different between the NI and IR groups. The structural stiffness was 232.1±45.7 N/mm for the NI PB and 161.9±74.0 N/mm for the IR side, and this was the only statistically significant difference found in this study (P=0.034).ConclusionOur statistical comparisons found no significant differences in terms of elongation, ultimate load, or ultimate stress between IR and NI PB and PL tendons. Only the PB structural stiffness was affected by irradiation. Thus, sterilizing allografts at 1.5–2.5 Mrad of gamma irradiation does not cause major alterations in the tendons’ biomechanical properties while still providing a suitable amount of sterilization for anterior cruciate ligament reconstruction.

Activation of the JNK-c-Jun pathway in response to irradiation facilitates Fas ligand secretion in hepatoma cells and increases hepatocyte injury.

BackgroundIt is well established that some irradiated liver non-parenchymal cells secrete pro-inflammatory cytokines to facilitate the development of radiation-induced liver disease. However, little is known on whether the irradiated hepatoma cells-mediated non-irradiated hepatocyte injury occurs in HCC patients. Here, we elucidated the roles of the irradiated hepatoma cells in driving non-irradiated hepatocyte injury and its underlying mechanism.MethodsSMMC7721 cells were cultured and divided into irradiated (4-Gy X-ray, R) and non-irradiated (NR) groups. At 24th hour after irradiation, conditioned medium (CM) from these cultures was mixed with normal culture medium in specific proportions, and termed as 7721-R-CM and 7721-NR-CM. Following incubation with these CM compound, the biological characteristics of L02 cells related to liver cell injury including viability, apoptosis and liver dysfunction indices were comparatively analyzed. Simultaneously, the levels of proliferation- and apoptosis-related cytokines in irradiated and non-irradiated SMMC7721 cells were also measured. FasL as a cytokine with significantly differential expression, was selected to clarify its effects on L02 apoptosis. Subsequently, FasL expression following irradiation was examined in SMMC7721 and other HCC cells with varying malignant potentials, as well as in HCC tissues, the related mechanism of higher expression of FasL in irradiated HCC cells was further investigated.ResultsApoptosis and liver dysfunction indices were all significantly enhanced in L02 cells treated with 7721-R-CM, whereas viability was suppressed, compared to those with 7721-NR-CM stimulation. FasL was identified as a leading differential cytokine in the irradiated SMMC7721 cells. Higher proportion of apoptosis was also found in L02 cells following FasL incubation. A recombinant Fas-Fc protein, which blocks Fas-FasL interaction, ameliorated 7721-R-CM-induced apoptosis in L02 cells. FasL was highly expressed in a dose-dependent manner, and peaked at the 24th hour post-irradiation in different HCC cells and their culture supernatant. Meanwhile, phosphorylation levels of JNK, ERK, Akt, and p38 were all upregulated significantly in irradiated HCC cells. But, only JNK inhibition was validated to block radiation-induced FasL expression in HCC cells. c-Jun, the target transcription factor of JNK, was also activated.ConclusionIn HCC cells, the JNK-c-Jun pathway plays an important role in mediating irradiation- induced FasL expression, which may be critical in determining non-irradiated hepatocyte injury.Electronic supplementary materialThe online version of this article (doi:10.1186/s13046-016-0394-z) contains supplementary material, which is available to authorized users.

Protein expression of nucleophosmin, annexin A3 and nm23-H1 correlates with human nasopharyngeal carcinoma radioresistance in vivo.

Radioresistance is a significant obstacle in the treatment of endemic nasopharyngeal carcinoma (NPC). The present study aimed to identify proteins associated with radioresistance in NPC in vitro and in vivo. Proteomics analyses were conducted to screen for differentially-expressed proteins (DEPs) in parental CNE-2 cells and CNE-2R cells. Using proteomics approaches, 16 DEPs were identified. Of these DEPs, nucleophosmin (NPM1), annexin A3 and nm23-H1, were verified using western blot analyses. The tumorigenicity was investigated using mouse xenograft tumorigenicity assays, and tumor growth curves were generated. The protein expression of NPM1, annexin A3 and nm23-H1 was examined by immunohistochemically staining tumor tissues. NPM1 and annexin A3 protein levels were downregulated in the CNE-2R cells, whereas nm23-H1 expression was upregulated. In vivo tests showed that compared with the CNE-2 tumors, CNE-2R tumor growth was significantly retarded (P<0.05). CNE-2 tumor progression was inhibited by irradiation, but CNE-2R tumor progression was not, indicating that the CNE-2R cells were also radioresistant in vivo. NPM1 and annexin A3 expression was significantly lower in non-irradiated (NIR)-CNE-2R tumors compared with NIR-CNE-2 tumors (P<0.01). However, Nm23-H1 protein levels were significantly higher (P<0.05). Overall, the present study established comparable radioresistant and radiosensitive tumor models of human NPC, and identified candidate biomarkers that may correlate with radioresistance. The data showed that dysregulation of NPM1, annexin A3 and nm23-H1 expression correlated with the cellular and tumor radioresponse. These proteins are involved in the regulation of intracellular functions, including stress responses, cell proliferation and DNA repair. However, further clinical evaluations are required.

Effect of gamma irradiation on cytokines released by platelets during storage

Most published gamma irradiation studies are on the cytokines secreted by leucocytes than platelet cytokines. If cytokines secretion is suppressed by irradiation then it will be an important application of irradiation. To know this we planned to measure cytokines like Regulated upon activation normal T cells expressed and secreted (RANTES), platelet factor 4 (PF-4), Transforming growth factor βeta1 (TGF-β) and βeta Thromboglobulin (β TG) during the storage of irradiated (IR) and non-irradiated (NI) platelets. Ten platelet concentrate (PCs) were prepared using platelet rich plasma and buffy coat methods each and 10 by apheresis on Trima Cell separator. Each PC was transferred in a transfer bag and IR at about 25 Gy and stored at 22 °C in platelet agitator. Samples were taken from NI and IR bag each on 0 day, 3rd day and 5th day from supernatant for cytokine analysis. The samples were stored below −50 °C for cytokines assays using commercial ELISA kits. RANTES levels were in the range of 12–400 ng/ml on 0 day and increased to 108–800 ng/ml on 5th day in NI platelets and 104–800 ng/ml in IR platelet. PF4 increased from 0 day to 5th day showing levels 300–1500 ng/ml in both types of PC. β-TG range was 748–5258 ng/ml in NI and 878–4638 ng/ml in IR platelets on 5th day. TGF-β1 increased up to 780–38431 pg/ml in NI and 461–50,000 pg/ml in IR PC. The study showed that comparison of cytokine levels during the storage in NI and IR platelets was not significant by Mann–Whitney U- test (p > 0.05). Significant increase was observed in these cytokine levels from 0 day to 3rd and 5th day in NI as well as IR samples by Wilcoxon Signed- Rank test (p < 0.05).