Gene Expression and Early Radiation Response of Two Distinct Neuroblastoma Cell Lines.

INTRODUCTION: Neuroblastoma is one of the most common childhood cancers with one of the lowest survival rates, accounting for 15% of childhood cancer mortality. Approximately half of children treated for high-risk neuroblastoma will relapse following remission, while another 15% of patients do not respond to initial treatment. External beam radiation is infrequently used for treatment of pediatric cancer such as neuroblastoma, typically reserved for palliative care in patients with aggressive metastatic disease who fail to respond to alternative treatments. Understanding effects of radiation on neuroblastoma cells could improve efficacy of this final means of therapy to decrease tumor burden and stabilize the disease. METHODS: In this study, we found that two microRNAs with opposite functions were expressed in two neuroblastoma cell lines with marked differences in radiosensitivity. Clonogenic assays were used to evaluate the radiation responses for these 2 cell lines, designated SK-N-AS and SK-N-DZ; cells were then irradiated at doses that cause 90% cell killing based on clonogenic assay and their RNA isolated and subjected to microarray analysis. In addition, cells were transfected with pre-miRNA constructs that led to overexpression of microRNAs miR-34a and miR-1228 to determine possible microRNA regulation of radiation response. RESULTS: Statistically significant differences were detected for expression of several thousand genes when the 2 cell lines were compared with each other. In comparison, radiation exposure resulted in only minor gene expression differences of less than 2-fold at the 1 h postirradiation timepoint in both cell lines. Overexpression of miR-34a and miR-1228 in either cell line did not alter this outcome. DISCUSSION: While these two neuroblastoma cell lines are phenotypically diverse and gene expression differences between them are extensive, we observed that the regulation of gene expression in both cell lines is in a stable equilibrium at early timepoints after exposure to ionizing radiation.

Effect of engineered superparamagnetic iron oxide nanoparticles in targeted cardiac precursor cell delivery by MRI.

A scientific approach is presented describing the fabrication of nanoprobe (GloTrack) that can act as cardiac precursor label to segregate cells from cardiac/non cardiac origins and traced by magnetic resonance imaging (MRI). Signal regulatory protein alpha (SIRPA) and kinase domain receptor (KDR) recognizing antibodies, form a layer on super paramagnetic iron oxide nanoparticle - poly-ethylene glycol (SPION-PEG) complex, and bind to protein expressed on the surface of cardiac muscle cells. Physical attributes size, distribution, labelling efficiency, echocardiogram (ECG) changes and bio-distribution by MRI were analysed. The results indicate that GloTrack has an average size of 471 nm, exhibits negative potential and promotes labelling efficiency. The bio-distribution of GloTrack in in vivo experiments was traceable in 7T MRI showing high accumulation of GloTrack in cardiac muscles as compared to the liver and spleen. ECG data revealed that GloTrack segregated cardiac precursors has the potential benefit in treating heart failure, thereby paving way in the development of minimal cell manipulation with targeted cell delivery approaches.

The tubercular badger and the uncertain curve:- The need for a multiple stressor approach in environmental radiation protection.

This article presents the results of a workshop held in Stirling, Scotland in June 2018, called to examine critically the effects of low-dose ionising radiation on the ecosphere. The meeting brought together participants from the fields of low- and high-dose radiobiology and those working in radioecology to discuss the effects that low doses of radiation have on non-human biota. In particular, the shape of the low-dose response relationship and the extent to which the effects of low-dose and chronic exposure may be predicted from high dose rate exposures were discussed. It was concluded that high dose effects were not predictive of low dose effects. It followed that the tools presently available were deemed insufficient to reliably predict risk of low dose exposures in ecosystems. The workshop participants agreed on three major recommendations for a path forward. First, as treating radiation as a single or unique stressor was considered insufficient, the development of a multidisciplinary approach is suggested to address key concerns about multiple stressors in the ecosphere. Second, agreed definitions are needed to deal with the multiplicity of factors determining outcome to low dose exposures as a term can have different meanings in different disciplines. Third, appropriate tools need to be developed to deal with the different time, space and organisation level scales. These recommendations permit a more accurate picture of prospective risks.

Chemoprevention in gastrointestinal physiology and disease. Natural products and microbiome.

The human intestinal tract harbors a complex ecosystem of commensal bacteria that play a fundamental role in the well-being of their host. There is a general consensus that diet rich in plant-based foods has many advantages in relation to the health and well-being of an individual. In adults, diets that have a high proportion of fruit and vegetables and a low consumption of meat are associated with a highly diverse microbiota and are defined by a greater abundance of Prevotella compared with Bacteroides, whereas the reverse is associated with a diet that contains a low proportion of plant-based foods. In a philosophical term, our consumption of processed foods, widespread use of antibiotics and disinfectants, and our modern lifestyle may have forever altered our ancient gut microbiome. We may never be able to identify or restore our microbiomes to their ancestral state, but dietary modulation to manipulate specific gut microbial species or groups of species may offer new therapeutic approaches to conditions that are prevalent in modern society, such as functional gastrointestinal disorders, obesity, and age-related nutritional deficiency. We believe that this will become an increasingly important area of health research.

2-deoxy-D-glucose mitigates Citrobacter rodentium and dibenzazepine-induced gastrointestinal damage and colitis: novel implications of 2-DG polypharmacopea.

PURPOSE: Citrobacter rodentium (CR) infection coupled with blocking Notch/Wnt signaling via γ-secretase inhibitor dibenzazepine (DBZ) disrupts the gastro-intestinal (GI) barrier and induces colitis, akin to ionizing radiation (IR)-induced GI-injury. We investigated the effects of 2-deoxy-D-glucose (2-DG) to ameliorate the CR-DBZ-induced GI damage. MATERIALS AND METHODS: NIH:Swiss outbred mice were inoculated with 109CFUs of CR orally. DBZ was administered intraperitoneally (10 µM/kg b.wt; for 10 days 2 days post-CR infection). Mice were fed with 0.4% 2-DG (w/v) daily in drinking water. For microbiota depletion, antibiotics (Abx), 1 g/l metronidazole, and 0.2 g/l ciprofloxacin were administered for 10 days in drinking water. Oxidative stress, survival assay, colonic crypt hyperplasia, Notch/Wnt downstream signaling, immunomodulation, and bacterial dysbiosis were measured. RESULTS: We show that real-time visualization of reactive oxygen species (ROS) is similar during CR-induced colonic infection and IR-induced GI-damage. The histology revealed that dietary 2-DG mitigates CR + DBZ-induced colitis and improves survival compared with CR + DBZ alone. These changes were phenocopied in Abx-treated mice. Both 2-DG and Abx reduced dysbiosis, increased proliferation, inhibited pro-inflammatory response, and restored Hes-1 and ß-catenin protein levels, in the crypts. CONCLUSION: The energy disruptor 2-DG mitigates bacterial infection and its responsive hyperplasia/colitis, indicating its utility as a mitigator of infection/IR-induced GI-damage.

Technological advancements in cancer diagnostics: Improvements and limitations.

BACKGROUND: Cancer is characterized by the rampant proliferation, growth, and infiltration of malignantly transformed cancer cells past their normal boundaries into adjacent tissues. It is the leading cause of death worldwide, responsible for approximately 19.3 million new diagnoses and 10 million deaths globally in 2020. In the United States alone, the estimated number of new diagnoses and deaths is 1.9 million and 609 360, respectively. Implementation of currently existing cancer diagnostic techniques such as positron emission tomography (PET), X-ray computed tomography (CT), and magnetic resonance spectroscopy (MRS), and molecular diagnostic techniques, have enabled early detection rates and are instrumental not only for the therapeutic management of cancer patients, but also for early detection of the cancer itself. The effectiveness of these cancer screening programs are heavily dependent on the rate of accurate precursor lesion identification; an increased rate of identification allows for earlier onset treatment, thus decreasing the incidence of invasive cancer in the long-term, and improving the overall prognosis. Although these diagnostic techniques are advantageous due to lack of invasiveness and easier accessibility within the clinical setting, several limitations such as optimal target definition, high signal to background ratio and associated artifacts hinder the accurate diagnosis of specific types of deep-seated tumors, besides associated high cost. In this review we discuss various imaging, molecular, and low-cost diagnostic tools and related technological advancements, to provide a better understanding of cancer diagnostics, unraveling new opportunities for effective management of cancer, particularly in low- and middle-income countries (LMICs). RECENT FINDINGS: Herein we discuss various technological advancements that are being utilized to construct an assortment of new diagnostic techniques that incorporate hardware, image reconstruction software, imaging devices, biomarkers, and even artificial intelligence algorithms, thereby providing a reliable diagnosis and analysis of the tumor. Also, we provide a brief account of alternative low cost-effective cancer therapy devices (CryoPop®, LumaGEM®, MarginProbe®) and picture archiving and communication systems (PACS), emphasizing the need for multi-disciplinary collaboration among radiologists, pathologists, and other involved specialties for improving cancer diagnostics. CONCLUSION: Revolutionary technological advancements in cancer imaging and molecular biology techniques are indispensable for the accurate diagnosis and prognosis of cancer.

Technological Advancements in External Beam Radiation Therapy (EBRT): An Indispensable Tool for Cancer Treatment.

Recent technological advancements have increased the efficacy of radiotherapy, leading to effective management of cancer patients with enhanced patient survival and improved quality of life. Several important developments like multileaf collimator, integration of imaging techniques like positron emission tomography (PET) and computed tomography (CT), involvement of advanced dose calculation algorithms, and delivery techniques have increased tumor dose distribution and decreased normal tissue toxicity. Three-dimensional conformal radiotherapy (3DCRT), intensity-modulated radiotherapy (IMRT), stereotactic radiotherapy, image-guided radiotherapy (IGT), and particle therapy have facilitated the planning procedures, accurate tumor delineation, and dose estimation for effective personalized treatment. In this review, we present the technological advancements in various types of EBRT methods and discuss their clinical utility and associated limitations. We also reveal novel approaches of using biocompatible yttrium oxide scintillator-photosensitizer complex (YSM) that can generate X-ray induced cytotoxic reactive oxygen species, facilitating X-ray activated photodynamic therapy (XPDT (external beam) and/or iXPDT (internal X-ray source)) and azido-derivatives of 2-deoxy-D-glucose (2-DG) as agents for site-specific radiation-induced DNA damage.

Overcoming Barriers to Radiopharmaceutical Therapy (RPT): An Overview From the NRG-NCI Working Group on Dosimetry of Radiopharmaceutical Therapy.

Radiopharmaceutical therapy (RPT) continues to demonstrate tremendous potential in improving the therapeutic gains in radiation therapy by specifically delivering radiation to tumors that can be well assessed in terms of dosimetry and imaging. Dosimetry in external beam radiation therapy is standard practice. This is not the case, however, in RPT. This NRG (acronym formed from the first letter of the 3 original groups: National Surgical Adjuvant Breast and Bowel Project, the Radiation Therapy Oncology Group, and the Gynecologic Oncology Group)-National Cancer Institute Working Group review describes some of the challenges to improving RPT. The main priorities for advancing the field include (1) developing and adopting best practice guidelines for incorporating patient-specific dosimetry for RPT that can be used at both large clinics with substantial resources and more modest clinics that have limited resources, (2) establishing and improving strategies for introducing new radiopharmaceuticals for clinical investigation, (3) developing approaches to address the radiophobia that is associated with the administration of radioactivity for cancer therapy, and (4) solving the financial and logistical issues of expertise and training in the developing field of RPT.

Increased sensitivity in antigen detection with fluorescent latex nanosphere-IgG antibody conjugates.

IgG antibodies were conjugated to Kodak X-Sight nanospheres to develop fluorescent-labeled antibodies using two different synthetic routes: one involving the DTT reduction method, and the other involving Traut's Reagent modification method. These two methods result in different conjugation efficiencies and different performances in antigen detection. Western blotting shows that the nanosphere-IgG antibody conjugates synthesized using the DTT reduction method are more immunospecific than the conjugates synthesized using Traut's Reagent modification method. In addition, the conjugates synthesized using DTT reduction also show higher antigen detection sensitivity than other commercially available fluorescent-IgG antibody conjugates, including Alexa Fluor, Qdot, and CyDye conjugates.

A combinatorial approach of a polypharmacological adjuvant 2-deoxy-D-glucose with low dose radiation therapy to quell the cytokine storm in COVID-19 management.

COVID-19, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a pandemic disease and is the major cause of deaths worldwide. The clinical complexities (inflammation, cytokine storm, and multi-organ dysfunction) associated with COVID-19 poses constraints to effective management of critically ill COVID-19 patients. Low dose radiation therapy (LDRT) has been evaluated as a potential therapeutic modality for COVID-19 pneumonia. However, due to heterogeneity in disease manifestation and inter-individual variations, effective planning for LDRT is limited for this large-scale event. 2-deoxy-D-glucose (2-DG) has emerged as a polypharmacological agent for COVID-19 treatment due to its effects on the glycolytic pathway, anti-inflammatory action, and interaction with viral proteins. We suggest that 2-DG will be a potential adjuvant to enhance the efficacy of LDRT in the treatment of COVID-19 pneumonia. Withal, azido analog of 2-DG, 2-azido-2-DG can produce rapid catastrophic oxidative stress and quell the cytokine storm in critically ill COVID-19 patients.

Taurine and guanidinoethanesulfonic acid (GES) differentially affects the expression and phosphorylation of cellular proteins in C6 glial cells.

Effects of taurine and guanidinoethanesulfonic acid (GES), a taurine transport inhibitor, on the expression and phosphorylation of cellular proteins in C6 glial cells were examined using two-dimensional (2-D) gel electrophoresis and 2-D immunoblots. 2-D gels stained with Coomassie Blue or SYPRO Ruby showed differential distribution patterns of cellular proteins in the control, taurine-supplemented and GES-supplemented cells. 2-D immunoblot analysis using the anti-phosphotyrosine antibody recognized only few immuno-reactive proteins in all three samples, and their distribution patterns were different. On the other hand, 2-D immunoblot analysis using the anti-phosphothreonine antibody recognized many immuno-reactive proteins with distinctly different distribution patterns in the control, taurine-supplemented and GES-supplemented cells. In GES-supplemented cells, the relative number of the anti-phosphotyrosine immuno-reactive protein spots increased modestly, whereas the relative number of the anti-phosphothreonine immuno-reactive protein spots decreased markedly than those in the control and taurine-supplemented cells.

A realistic utilization of nanotechnology in molecular imaging and targeted radiotherapy of solid tumors.

Precise dose delivery to malignant tissue in radiotherapy is of paramount importance for treatment efficacy while minimizing morbidity of surrounding normal tissues. Current conventional imaging techniques, such as magnetic resonance imaging (MRI) and computerized tomography (CT), are used to define the three-dimensional shape and volume of the tumor for radiation therapy. In many cases, these radiographic imaging (RI) techniques are ambiguous or provide limited information with regard to tumor margins and histopathology. Molecular imaging (MI) modalities, such as positron emission tomography (PET) and single photon-emission computed-tomography (SPECT) that can characterize tumor tissue, are rapidly becoming routine in radiation therapy. However, their inherent low spatial resolution impedes tumor delineation for the purposes of radiation treatment planning. This review will focus on applications of nanotechnology to synergize imaging modalities in order to accurately highlight, as well as subsequently target, tumor cells. Furthermore, using such nano-agents for imaging, simultaneous coupling of novel therapeutics including radiosensitizers can be delivered specifically to the tumor to maximize tumor cell killing while sparing normal tissue.

The DNA damage effector Chk1 kinase regulates Cdc14B nucleolar shuttling during cell cycle progression.

Chk1 is a critical effector of DNA damage checkpoints necessary for the maintenance of chromosome integrity during cell cycle progression. Here we report, that Chk1 co-localized with the nucleolar marker, fibrillarin in response to radiation-induced DNA damage in human cells. Interestingly, in vitro studies using GST pull down assays identified the dual-specificity serine/threonine nucleolar phosphatase Cdc14B as a Chk1 substrate. Furthermore, Chk1, but not a kinase-dead Chk1 control, was shown to phosphorylate Cdc14B using an in vitro kinase assay. Co-immunoprecipitation experiments using exogenous Cdc14B transfected into human cells confirmed the interaction of Cdc14B and Chk1 during cell cycle. In addition, reduction of Chk1 levels via siRNA or UCN-01 treatment demonstrated that Chk1 activation following DNA damage was required for Cdc14B export from the nucleolus. These studies have revealed a novel interplay between Chk1 kinase and Cdc14B phosphatase involving radiation-induced nucleolar shuttling to facilitate error-free cell cycle progression and prevent genomic instability.

Analytical radiography for planar radiographic images implemented with a multi-modal system.

A radiographic system is optimized for the contrast inherent to small animals and is developed for a multi-modal imaging system devised for in-vivo studies. The range of X-ray energies utilized (generally considered "soft X-rays") enables enhanced spatial resolution and superior contrast for detailed study of the mouse anatomy and smaller specimens. Despite the difficulties presented by the complicated energy spectrum of soft X-rays, relevant system calibrations for bone measures are described in detail and applied to the mouse. Further, long-bone symmetry modeling using a cylindrical projection is applied to the planar density image, providing convenient bone density estimates that are consistent with other methodologies.

Suramin interacts with the calmodulin binding site on the ryanodine receptor, RYR1.

Apocalmodulin and Ca(2+) calmodulin bind to overlapping sites on the ryanodine receptor skeletal form, RYR1, but have opposite functional effects on channel activity. Suramin, a polysulfonated napthylurea, displaces both forms of calmodulin, leading to an inhibition of activity at low Ca(2+) and an enhancement of activity at high Ca(2+). Calmodulin binding motifs on RYR1 are also able to directly interact with the carboxy-terminal tail of the transverse tubule dihydropyridine receptor (DHPR) (Sencer, S., Papineni, R. V., Halling, D. B., Pate, P., Krol, J., Zhang, J. Z., and Hamilton, S. L. (2001) J. Biol. Chem. 276, 38237-38241). Suramin binds directly to a peptide that corresponds to the calmodulin binding site of RYR1 (amino acids 3609-3643) and blocks the interaction of this peptide with both calmodulin and the carboxyl-terminal tail of the DHPR alpha(1)-subunit. Suramin, added to the internal solution of voltage-clamped skeletal myotubes, produces a concentration-dependent increase in the maximal magnitude of voltage-gated Ca(2+) transients without significantly altering L-channel Ca(2+) channel conducting activity. Together, these results suggest that an interaction between the carboxyl-terminal tail of the DHPR alpha(1)-subunit with the calmodulin binding region of RYR1 serves to limit sarcoplasmic reticulum Ca(2+) release during excitation-contraction coupling and that suramin-induced potentiation of voltage-gated Ca(2+) release involves a relief of this inhibitory interaction.