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The damage of DNA chains by environmental factors like radiation or chemical pollutants is a topic that has been frequently explored from an experimental and a theoretical perspective. Such damages, like the damage of the strands of a DNA chain, are toxic for the cell and can induce mutagenesis or apoptosis. Several models make strong assumptions for the distribution of damages; for instance a frequent supposition is that these damages are Poisson distributed. [L. Ma, J. J. Wagner, W. Hu, A. J. Levine and G. A. Stolovitzki, Proc. Natl. Acad. Sci.PNAS 102, 14266 (2005).] Only few models describe in detail the damage and the mechanisms associated to the formation and evolution of this damage distribution [H. Nikjoo, P. O'neill and D. T. Goodhead, Radiat. Res. 156, 577 (2001).] Nevertheless, such models do not include the repair processes which are continuously active inside the cell. In this work we present a novel model, based on a depolymerization process, describing the distribution of damages on DNA chains coupled to the dynamics associated to its repair processes. The central aim is not to give a final and comprehensive model, but a hint to represent in more detail the complex dynamics involved in the damage and repair of DNA. We show that there are critical parameters associated to this repair process, in particular we show how critical doses can be relevant in deciding whether the cell continues its repair process or starts apoptosis. We also find out that the damage concentration is related to the dose via a power law relation.

Ganoderma lucidum (Fr.) Karst is a traditional Chinese herb that has been widely used for centuries to treat various diseases including cancer. Herein, an ethanol-soluble and acidic component (ESAC), which mainly contains triterpenes, was prepared from G. lucidum and its anti-tumor effects in vitro were tested on human breast cancer cells. Our results showed that ESAC reduced the cell viability of MCF-7 and MDA-MB-231 cells in a concentration-dependent manner with IC₅₀ of about 100 μg/mL and 60 μg/mL, respectively. DNA damage was detected by Comet assay and the increased expression of γ-H2AX after ESAC treatment was determined in MCF-7 cells. Moreover, ESAC effectively mediated G1 cell cycle arrest in both concentration- and time-dependent manners and induced apoptosis as determined by Hoechst staining, DNA fragment assay and Western blot analysis in MCF-7 cells. In conclusion, ESAC exerts anti-proliferation effects by inducing DNA damage, G1 cell cycle arrest and apoptosis in human breast cancer cells.

Celastrol is one of the principal active ingredients of Tripterygium wilfordii Hook.f., a toxic Chinese medical herb traditionally prescribed for controlling pain and inhibiting inflammation in various chronic inflammatory diseases, including rheumatoid arthritis (RA). Resistance to apoptosis of fibroblast-like synoviocytes is considered a major characteristic of RA. In this study, we test celastrol's cytotoxic effect and potential mechanisms in human rheumatoid synovial fibroblasts (RA-FLS). In the cytotoxic assay, we found that celastrol dose-dependently decreased RA-FLS viability and increased LDH release. The apoptotic nuclear morphology was observed after celastrol treatment as determined by DAPI fluorescence staining. Flow cytometry analysis with PI and Annexin V revealed that celastrol induced RA-FLS cell cycle arrest in the G2/M phase and apoptosis. Furthermore, celastrol dramatically increased expression of Bax/Bcl-2, proteolytic cleavage of Caspase-3, -9, PARP, and decreased expression of FasR. In addition, celastrol treatment resulted in DNA damage. Collectively, we concluded that celastrol inhibits RA-FLS proliferation by inducing DNA damage, cell cycle arrest, and apoptosis in vitro, which might provide data for its application in RA treatment.

Bufalin is a key component of a Chinese medicine (Chan Su) and has been proved effective in killing various cancer cells. Its role in inducing DNA damage and the inhibition of the DNA damage response (DDR) has been reported, but none have studied such action in lung cancer in detail. In this study, we demonstrated bufalin-induced DNA damage and condensation in NCI-H460 cells through a comet assay and DAPI staining, respectively. Western blotting indicated that bufalin suppressed the protein levels associated with DNA damage and repair, such as a DNA dependent serine/threonine protein kinase (DNA-PK), DNA repair proteins breast cancer 1, early onset (BRCA1), 14-3-3 σ (an important checkpoint keeper of DDR), mediator of DNA damage checkpoint 1 (MDC1), O6-methylguanine-DNA methyltransferase (MGMT) and p53 (tumor suppressor protein). Bufalin could activate phosphorylated p53 in NCI-H460 cells. DNA damage in NCI-H460 cells after treatment with bufalin up-regulated its ATM and ATR genes, which encode proteins functioning as sensors in DDR, and also up-regulated the gene expression (mRNA) of BRCA1 and DNA-PK. But bufalin suppressed the gene expression (mRNA) of p53 and 14-3-3 σ, however, bufalin did not significantly affect the mRNA of MGMT. In conclusion, bufalin induced DNA damage in NCI-H460 cells and also inhibited its DNA repair and checkpoint function.

Numerous evidences have shown that plant flavonoids (naturally occurring substances) have been reported to have chemopreventive activities and protect against experimental carcinogenesis. Kaempferol, one of the flavonoids, is widely distributed in fruits and vegetables, and may have cancer chemopreventive properties. However, the precise underlying mechanism regarding induced DNA damage and suppressed DNA repair system are poorly understood. In this study, we investigated whether kaempferol induced DNA damage and affected DNA repair associated protein expression in human leukemia HL-60 cells in vitro. Percentages of viable cells were measured via a flow cytometry assay. DNA damage was examined by Comet assay and DAPI staining. DNA fragmentation (ladder) was examined by DNA gel electrophoresis. The changes of protein levels associated with DNA repair were examined by Western blotting. Results showed that kaempferol dose-dependently decreased the viable cells. Comet assay indicated that kaempferol induced DNA damage (Comet tail) in a dose-dependent manner and DAPI staining also showed increased doses of kaempferol which led to increased DNA condensation, these effects are all of dose-dependent manners. Western blotting indicated that kaempferol-decreased protein expression associated with DNA repair system, such as phosphate-ataxia-telangiectasia mutated (p-ATM), phosphate-ataxia-telangiectasia and Rad3-related (p-ATR), 14-3-3 proteins sigma (14-3-3σ), DNA-dependent serine/threonine protein kinase (DNA-PK), O⁶-methylguanine-DNA methyltransferase (MGMT), p53 and MDC1 protein expressions, but increased the protein expression of p-p53 and p-H2AX. Protein translocation was examined by confocal laser microscopy, and we found that kaempferol increased the levels of p-H2AX and p-p53 in HL-60 cells. Taken together, in the present study, we found that kaempferol induced DNA damage and suppressed DNA repair and inhibited DNA repair associated protein expression in HL-60 cells, which may be the factors for kaempferol induced cell death in vitro.

Cordyceps militaris is a traditional Chinese medicine frequently used for tonic and therapeutic purposes. Reports from our laboratory and others have demonstrated that extracts of the cultivated fruiting bodies of C. militaris (CM) exhibit a potent cytotoxic effect against many cancer cell lines, especially human leukemia cells. Here, we further investigated the underlying mechanism through which CM is cytotoxic to cancer cells. The CM-mediated induction of PARP cleavage and its related DNA damage signal (γH2AX) was diminished by caspase inhibitor I. In contrast, a ROS scavenger failed to prevent CM-mediated leukemia cell death. Moreover, two signaling molecules, AKT and p38 MAPK, were activated during the course of apoptosis induction. Employing MTT analysis, we found that a p38 MAPK inhibitor but not an AKT inhibitor could rescue cells from CM-mediated cell death, as well as inhibit the cleavage of PARP, formation of apoptotic bodies and up-regulation of the γH2AX signal. These results suggest that CM-mediated leukemia cell death occurs through the activation of the p38 MAPK pathway, indicating its potential therapeutic effects against human leukemia.

In this paper, a p53-Mdm2 mathematical model is analyzed to understand the biological implications of feedback loops in a p53 system. Results show that the model can undergo four types of codimension-3 Bogdanov–Takens bifurcations, including cusp, saddle, focus and elliptic. Specifically, we find new phenomena including the coexistence of four positive equilibria, two limit cycles, the coexistence of three stable states (two stable equilibria and one stable limit cycle, or three stable equilibria), a heteroclinic loop enclosing a smaller stable limit cycle and a larger stable limit cycle. These findings extend the understanding of the complex dynamics of the p53 system, and can provide some potential biological applications.

The study of diseases such as cancer requires the modeling of gene regulations and the loss of control associated with it. Prior work has shown that the genetic alterations in the system can be suitably modeled using different fault models (like stuck-at faults) in the Boolean Network paradigm. By studying the dynamics of the original and the faulty BN, it is possible to design intervention strategies which could drive the system from a diseased state to a less harmful one. In this paper, the method of detecting faults along with the intervention design demonstrated on a couple of real biological pathways (DNA damage pathways and osmotic stress response pathways).

The mechanism of DNA damage caused by the isomerization of purine base is studied with density functional theory calculations at the B3LYP/6-311+G(d,p) level. The transition states of all the isomerizations are obtained, and the intrinsic reaction coordinate (IRC) analyses are performed to identify these transition states further. The isomerizations of purine bases can be classified into two types. The first is the hydrogen transfer between atoms, whose transition state includes a four-member ring. The second is the bond N–H rotation about the double bond N=C, and the plane CNH is perpendicular to the molecular plane in its transition state. The hydrogen transfer has higher reaction potential barrier, larger tunnel effect, and smaller equilibrium constant and rate constant than that of the N–H rotation. Effects of the hydration are considered in the framework of the polarizable continuum model (PCM) in SCRF method at the B3LYP/6-311+G(d,p) level. The isomerizations which result in the configuration changes of purine base and bring directly the DNA damage are endothermic and thermodynamic nonspontaneous processes. The probability of DNA damage caused by the guanine isomerization is larger than that by adenine.

Ionizing radiation (IR) causing damages to Deoxyribonucleic acid (DNA) constitutes a broad range of base damage and double strand break, and thereby, it induces the operation of relevant signaling pathways such as DNA repair, cell cycle control, and cell apoptosis. The goal of this paper is to study how the exposure to low dose radiation affects the human body by observing the signaling pathway associated with Ataxia Telangiectasia mutated (ATM) using Reverse-Phase Protein Array (RPPA) and isogenic human Ataxia Telangiectasia (A-T) cells under different amounts and durations of IR exposure. In order to verify which proteins could be involved in a DNA damage-caused pathway, only proteins that highly interact with each other under IR are selected by using correlation coefficient. The pathway inference is derived from learning Bayesian networks in combination with prior knowledge such as Protein–Protein Interactions (PPIs) and signaling pathways from well-known databases. Learning Bayesian networks is based on a score and search scheme that provides the highest scored network structure given a score function, and the prior knowledge is included in the score function as a prior probability by using Dempster–Shafer theory (DST). In this way, the inferred network can be more likely to be similar to already discovered pathways and consistent with confirmed PPIs for more reliable inference. The experimental results show which proteins are involved in signaling pathways under IR, how the inferred pathways are different under low and high doses of IR, and how the selected proteins regulate each other in the inferred pathways. As our main contribution, overall results confirm that low dose IR could cause DNA damage and thereby induce and affect related signaling pathways such as apoptosis, cell cycle, and DNA repair.

Porphyrins have been studied as photosensitizers in photodynamic therapy. DNA is one of the most important targets of the sensitizer. In the present study, we have examined the photosensitized DNA damage caused by dihydroxoP(V) tetraphenylporphyrin (P(V)TPP), a cationic water-soluble porphyrin. P(V)TPP photosensitized guanine-specific damage to the DNA fragment. P(V)TPP induced severe photodamage to single-stranded rather than to double-stranded DNA. High performance liquid chromatography measurements confirmed the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-G), an oxidized product of 2'-deoxyguanosine, and showed that the content of 8-oxo-G in single-stranded DNA is larger than that in double-stranded DNA. The effects of reactive oxygen scavengers on DNA damage suggested the involvement of singlet oxygen (¹O₂). Photosensitized ¹O₂ formation was confirmed by near-infrared emission measurements. The results showed that ¹O₂ formation mainly contributes to the mechanism of DNA photodamage by P(V)TPP. Absorption spectrum measurements showed the interaction between P(V)TPP and DNA. This interaction is expected to enhance the ¹O₂-mediated DNA damage since the lifetime of ¹O₂ in a cell is very short. On the other hand, P(V)TPP induced DNA damage at the consecutive guanines in double-stranded DNA. Because the consecutive guanines act as a hole trap, this DNA-damaging pattern suggests the partial involvement of photo-induced electron transfer. The fluorescence of P(V)TPP was quenched by DNA, supporting the electron transfer mechanism. However, DNA damage by electron transfer was not a main mechanism possibly due to reverse electron transfer. In conclusion, P(V)TPP binds to DNA and induces guanine-specific, photo-oxidation mainly via ¹O₂ generation.

In this paper, a delayed mathematical model was developed based on experimental data to understand how the time delays required for transcription and translation in Mdm2 gene expression affect the kinetic behavior of the p53-Mdm2 network. Taking the time delays as the main research parameters, the stability of the system at the positive equilibrium was studied by using the theoretical method of delay differential equation. We found that such delays can induce oscillations by undergoing a supercritical Hopf bifurcation. Then, we used the normal form theory and the center manifold reduction to study the direction and stability of the bifurcation in detail. Furthermore, we also studied the effects of the length of time delays and the model parameters by numerical simulations. We found that time delays in Mdm2 synthesis are required for p53 oscillations and the length of such delays can determine the amplitude and period of the oscillations. In addition, the model parameters can also change the stability of the system. These results illustrate that the repair process after DNA damage can be regulated by varying time delays and the model parameters.

It is well documented that various particulate matter — either incidental or engineered — are known to generate reactive oxygen species (ROS) in living cells. In circumstances where these reactive species are generated, antioxidant production is often increased. This balance in the biological reduction/oxidation (a.k.a. redox) state within the cell has not been thoroughly studied in exposures involving engineered nanoparticles. However, nanoparticle exposure has been postulated to induce a DNA damage cascade. In this study, we examined primary human dermal fibroblasts (HDF) exposed to three different, but commonly used engineered nanoparticles (i.e., cerium dioxide (CeO₂), titanium dioxide (TiO₂) and zinc oxide (ZnO)) in an attempt to determine the potential DNA damaging effects through the analysis of ROS generation, relevant protein upregulation response and single and double DNA strand breaks. Cell death was most elevated with exposure to ZnO, followed by TiO₂ and CeO₂. ROS generation was measured at 1 h, 6 h and 24 h after exposure to particles via a cell-based DCFH-DA (2′, 7′-dichlorfluorescein-diacetate) assay and indicated that ZnO generated the most significant amount of ROS. ZnO also caused upregulation of oxidative stress protein, heme oxygenase-1 and phosphorylation of p38; whereas CeO₂ caused upregulation of superoxide dismutase. Results from the comet assay indicated that ZnO triggered significant DNA damage in cells at relatively low dosing concentrations (20 ppm). Immunocytochemistry with ZnO-treated cells revealed notable DNA double strand breaks evidenced by a marked increase in the presence of γ-H2AX foci. This finding was also indicated by western blot, as well as cell cycle arrest by the phosphorylation of cyclin-dependent kinase 1. These data suggest that the three particle-types induce different degrees of DNA damage. And, of the three particle-types tested, exposure to ZnO nanoparticles may cause the most significant DNA damage.

In this study, a new series of 1,3,4-oxadiazole derivatives (3a– 3h) was synthesized, characterized using various analytical techniques (FT-IR, ¹H- and ${}^{13}$C-NMR, mass spectrometry), and tested for their effectiveness against Ehrlich’s Ascites Carcinoma (EAC) cell lines in vitro. After 48 h of exposure to these test compounds, the EAC cells exhibited a dose-dependent reduction in their viability. Among the tested compounds, 3b and 3e demonstrated the most potent anticancer effects, with IC${}_{50}$ values of 352.69 $\mu$M and 177.44 $\mu$M, respectively. Consequently, these compounds were chosen for further investigation into their mechanisms of action on EAC cell lines. The assessment included the induction of apoptosis and the analysis of DNA damage, which were evaluated using fluorescence staining and the comet assay. These assessments revealed distinctive apoptotic characteristics such as nuclear fragmentation, cytoplasmic shrinkage and DNA damage. As a result, these compounds hold promise as potential anticancer agents. The study also delved into the binding affinities of these compounds through molecular docking analysis, and the findings showed that compounds 3b and 3e exhibited a strong binding affinity with the receptor Transforming Growth Factor-Beta Receptor I (TGF-$\beta$RI) kinase (PDB ID: 1PY5), surpassing the reference compound 5-fluorouracil. Additionally, calculations related to Molecular Mechanics Generalized Born Surface Area (MM-GBSA) indicated favorable free binding energy. The compounds also displayed acceptable ADMET properties. To validate the stability of the bond between compounds 3b and 3e with the 1PY5 receptor, a molecular dynamics simulation lasting 100 ns was carried out.

The investigation of fragment length distributions of plasmid DNA following ion irradiation leads to a better understanding of the induction of DNA damage, particularly the clustering of double strand breaks. We present a model that calculates the fragment distributions of plasmid DNA following heavy ion irradiation by combining the Local Effect Model with a statistical model initially developed for X-rays. The integration of experimental constraints into the model calculations changes the resulting distributions strongly. We find a good agreement of our simulations with experimental fragment distributions based on atomic force microscopy studies. The model provides the means to rapidly predict the results for all ions. It may thus help to find the best ion species in order to demonstrate the impact of the localized energy distribution of particles.

We discuss different physical processes which may be responsible for biological damage induced by an ion beam propogating through living tissue. The creation of free radicals and an electron plasma, dissociative attachment of low-energy electrons, and local heating mechanisms are considered. Numerical estimates are performed for the case of carbon ion beams. We conclude that all three mechanisms are capable of inflicting single and double strand breaks in DNA molecules and thus can be responsible for the observed enhanced biological effectiveness of ion beams in cancer therapy.

Apart from biochemical cues, physical forces (stiffness, topography, shear forces, etc.) attributed to the extracellular matrix (ECM) play a crucial role in regulating cell and tissue homeostasis. ECM stiffness is one such physical factor, alterations of which can cause diseases including cancer and progeria. The ECM constitutes a dynamic microenvironment for stem cells to proliferate, migrate, self-renew, and differentiate, which are coordinated through a complex network of cell–matrix interactions. In the in vitro context, polyacrylamide (PA) hydrogels and cell-derived matrices (CDMs) (mimic the biophysical properties of stem cell microenvironment) are widely used to study cell–matrix interactions and their fate. Studies have shown the ability of cells to “sense” varying matrix stiffness via integrins to the nucleus, thus activating mechano-signaling cascades that lead to modulation of gene expression and direct stem cell lineage specifications. These force signals transmitted to the nucleus are also known to cause nuclear deformations and nuclear envelope rupture, causing a loss in pluripotency and genome integrity of embryonic stem cells (ESCs). While stem cell therapies hold immense potential for treating several genetic disorders, ESCs accumulate aneuploidy and greater DNA damage during prolonged culture on stiff cell culture dishes as compared to those when cultured on softer CDMs, thus motivating investigators to study biophysical regulation of stem cell fate and genome integrity.

The following sections are included:

• Aging is Our #1 Health Problem

• Degenerative Effects of Aging are Universal

• Why do Humans Live So Long? It is in Our Genes

• What is Oxidative Stress?

• Measuring Oxidative Damage
  • Alkenals
  • Hydroperoxides (Aqueous)
  • Hydroperoxides (Lipid)
  • 8-Hydroxydeoxyguanosine (8-OHdG)
  • 8-Epi-Prostaglandin F2α (8-epi-PGF2α)(Isoprostane)
  • Urine Creatinine

• Measuring Prooxidants
  • Antimony
  • Arsenic
  • C-Reactive Protein
  • Cadmium
  • Chromium
  • Chlorine
  • Iron
  • Ferritin
  • Copper
  • Homocysteine
  • Lead
  • Manganese
  • Mercury
  • Nickel

• Measuring Total Antioxidant Capacity
  • Lipid Peroxidation Inhibition Capacity (LPIC)* Assay
  • Oxygen Radical Absorption Capacity (ORAC) Assay
  • ORAC (Aqueous Soluble)
  • ORAC (Lipid Soluble)

• Measuring Primary Antioxidants
  • Available Iron Binding Capacity (AIBC)
  • % Iron Saturation (Total Iron/TIBC)*100%
  • Ceruloplasmin

• Measuring Secondary Antioxidants

• Aqueous Soluble Antioxidants
  • Ascorbic Acid (vitamin C)
  • Bilirubin (Conjugated and Total)
  • Thiols
  • Uric Acid

• Lipid Soluble Antioxidants
  • α-Carotene
  • β-Carotene
  • β-Cryptoxanthin
  • Lutein
  • Lycopene
  • Retinol (vitamin A)
  • Retinyl Palmitate
  • α-Tocopherol (vitamin E)
  • δ-Tocopherol (vitamin E)
  • γ-Tocopherol (vitamin E)
  • Tocopherols/(Chol. + Trig.)
  • Ubiquinol (Coenzyme Q10)
  • Zeaxanthin

• Antioxidant Supporting Factors
  • Albumin
  • Total Protein
  • Albumin/Globulins Ratio
  • Magnesium
  • Selenium
  • Sulfur
  • Zinc

• Clinical Interpretation and Use of Oxidative Stress Biomarkers for Metabolic Optimization

• References

In this chapter, radiation quantities and units and the induction and repair of radiation-induced DNA damage are briefly reviewed. Results of the authors' theoretical studies of DNA damage induction and outcomes from the excision repair of clustered DNA lesions are reported for selected types of low- and high-LET (linear energy transfer) radiation. Models for the conversion of clusters into small-scale mutations and aberrations are presented and used to illustrate trends in mutagenesis as a function of dose and particle LET.