**Meet the editor**

Dr. Kruman received her PhD in Cellular Biology from Moscow State University (Russia). She is an Associate Professor in the department of Pharmacology and Neuroscience, Texas Tech University Health Sciences Center, USA. Her research interests focus on mechanisms of apoptosis and DNA damage response in postmitotic neurons. She has authored over 50 scientific publica-

tions, including articles in major peer reviewed journals such as Neuron and Journal Neuroscience, book chapters and invited reviews.

Contents

**Preface XI**

**Part 1 DNA Damage Response 1**

Chuck C.-K. Chao

Chapter 5 **Shared Regulatory Motifs in**

Chapter 6 **Mitochondrial DNA Damage:**

Chapter 4 **The Potential Roles of DNA-Repair**

Ge Wang and Robert J. Maier

Chapter 1 **A Recombination Puzzle Solved: Role for New DNA Repair**

Ekaterina Krutkina, Daniel Klaiman, Tamar Margalit,

Chapter 3 **The Role of DDB2 in Regulating Cell Survival and Apoptosis Following DNA Damage - A Mini-Review 45**

Chapter 2 **RloC: A Translation-Disabling tRNase Implicated in**

Michal Chai-Danino and Alexander Kotlyar

**Proteins in Centrosome Maintenance 57**  Mikio Shimada, Akihiro Kato and Junya Kobayashi

**Promoters of Human DNA Repair Genes 67**  Lonnie R. Welch, Laura M. Koehly and Laura Elnitski

**Proteins: Lessons from BRCA1 and RAD51 Studies 103**  Effrossyni Boutou, Vassiliki Pappa, Horst-Werner Stuerzbecher

**Role of Ogg1 and Aconitase 85**  Gang Liu and David W. Kamp

Chapter 7 **Structure-Function Relationship of DNA Repair**

and Constantinos E. Vorgias

**Systems in** *Helicobacter pylori* **Diversity/Persistence 3**

**Phage Exclusion During Recovery from DNA Damage 21**  Gabriel Kaufmann, Elena Davidov, Emmanuelle Steinfels-Kohn,


**5. Buccal route**

Delivery of proteins or peptides via buccal route involves administration of therapeutics through the mucosal membrane lining the cheeks [152]. In this process, the drug is placed in the mouth between the gum and cheek [153]. Buccal delivery has many advantages such as ease of use, bypassing the GIT, large contact surface area, etc. This method can also be employed to deliver hydrophilic macromolecules [154]. However, this method has some disadvantages such as irritation of the mucosa, low permeability of the peptide, and bitter taste of many buccal drugs. Absorption enhancers and adhesive polymers are being used to overcome some of these problems. Several drugs such as insulin, oxytocin have been successfully delivered via this route. Mucoadhesion (discussed in Section 2.6) is very important during the development of buccal drug delivery systems. Some methods employed for increased drug delivery via this route are described below.

## **5.1 Absorptions enhancers**

Absorption enhancers are essential for delivering protein or peptide therapeutics, which generally show low buccal absorption rates. Some absorption enhancers are aprotinin, benzalkonium chloride, cyclodextrin, polyoxyethylene, sodium EDTA, etc.

## **5.2 pH**

Permeability of acyclovir was investigated in the presence of sodium glycocholate (absorption enhancer) at a pH range of 3.3 to 8.8. The permeability of acyclovir was found to be pH dependent.

## **5.3 Patch design**

Much research have been performed to understand the relationship between the type and amount of support materials and the drug release profile. Results indicated that these two factors are interrelated. It was also shown that single- and multi-layer patches have different drug release profiles.

## **6. Pulmonary route**

The pulmonary route has been used to successfully deliver peptide therapeutics such as desmopressin, calcitonin, human growth hormone, parathyroid hormone,

etc. [155]. There are several barriers to this route of peptide or protein delivery, such as respiratory mucus, mucociliary clearance, pulmonary enzymes, and macrophages that secrete peroxidases and proteases [156]. However, the large surface area, good vascularization, high capacity for solute exchange, and ultra-thinness are some of the attractive features of the alveolar epithelium that can facilitate systemic delivery of proteins or peptides via this route. The passage of large hydrophilic molecules through alveolar epithelium and capillary endothelium is limited. Absorption enhancers and enzyme inhibitors have been used to increase the absorption of peptide or protein therapeutics. However, these can be damaging to lung tissues. Some pulmonary delivery systems are described below.


**9. Extendable mass-Analyzer for meter-scale ribbon beams**

Conventional magnetic mass analysis of ribbon beams broader than 500 mm is difficult and does not scale well. The gap between the poles of a conventional dipole magnet is much smaller than the bending radius of the trajectories in the magnet, to avoid enormous aberrations, stray fields and related difficulties. As the beam breadth is increased, the cost, weight, power of the magnet all increase more steeply than with the square of the beam breadth. The fringe field grows in proportion to the pole gap and is a source of aberrations and crosstalk with other system components, which also increase in magnitude with the square of the pole gap.

A number of partial solutions have been developed, as described above, but these have been incremental improvements with significant tradeoffs. Aitken proposed a radically new analyzer in 2002 [20] which used a new approach, from which the present design has taken inspiration; however, it was more complex and larger than the present device for given beam parameters. I attempted a design using an E-shaped magnetic yoke [21], which was successfully tested, but the present design is simpler and lighter, with better resolution. In a conventional magnet the focal length scales with the bending radius, and the bending radius of necessity scales with the gap between the poles, so for a beam 2 m broad, one can expect the focal length to be several meters, while the aberrations would be severe, and the weight to exceed a hundred tons.

The magnet design presented here addresses these issues, and differs from conventional dipole magnet configurations in the following ways:

A system meeting these goals can have no component of magnetic field in the beam breadth direction, or it must violate these requirements. But if a field component normal to the breadth deflects the beam, then after a finite short distance the opposite field restores the beam direction, there must be a component of field in the gap between these zones in which a component of field is orthogonal to the s-shaped beam trajectories. Thus the solution is a magnet which deflects initially in the breadth direction, and produces a 3D bent S-shaped set of trajectories, hence the acronym U3DS [22].


**Preface to "Current Challenges and Advances in Atherosclerosis"**

Atherosclerosis is a systemic disorder which is of major interest in both basic science and clinical medicine in order to achieve healthy longevity worldwide. Atherosclerosis generally develops without any signs and symptoms, eventually resulting in various kinds of catastrophic cardiovascular events, such as ischemic heart disease, ischemic stroke, and rupture of major vessel aneurysms. Scientists working on the biological facets of atherosclerosis have recently focused on the persistent inflammation in vessel walls from intimal to adventitial tissues. Although inflammation is indeed a dominant pathological phenomenon in atherosclerosis, it has been recognized that there is no simple answer to the question of whether inflammation promotes or delays atherosclerosis. Atherosclerosis is a complex disease that involves several different cell types, such as lymphocytes, macrophages, monocytes, dendric cells, and their molecular products. Chronic inflammation in atherosclerosis can be accompanied by immunological changes, including auto-immune disorders. For example, our recent studies have shown that multiple auto-antibodies are found in the sera of subjects with atherosclerosis via inventory high-throughput auto-antibody screening techniques using cell-free technologies. Nevertheless, the immune system is even more complicated, with many cell types, hundreds of cytokines, and literally millions of different antigens, making research into atherosclerosis more complicated. Macrophages, which are differentiated from monocytes, one of the components of blood cells, phagocytize denatured LDL cholesterol on the vascular wall, become foamy cells, accumulate under the vascular endothelium, and cause atherosclerotic plaque formation. This forms the primary lesion, the fatty streak. Some of the aggregated foam cells undergo cell death over time, release denatured lipids in the cytoplasm, and increase in volume while forming a lipid core (necrotic core). The lipid core is surrounded by immune response cells, such as T cells, monocytes, macrophages, dendritic cells, and fibroblasts, to form atherosclerotic plaques. Enlarged lipid cores are at risk of rupture due to stresses on the vascular endothelium, such as blood pressure and shear stress from blood flow. The vulnerability of atherosclerotic plaques is thought to be determined by the formation of a fibrous cap around the plaque and the antagonism of degradation of fibrous components by protease, peptidase, etc., secreted by cells.

Fatal and non-fatal myocardial infarctions and cerebral infarctions, collectively referred to as cardiovascular events, result from accidental plaque rupture. As a result of plaque rupture, plaque components leak into the vascular lumen, causing local thrombus formation and subsequent activation of the fibrinolytic system. When the force of thrombus formation brought about by the qualitative and quantitative abnormalities of denatured lipids leaking from the lipid core exceeds the force of the endogenous fibrinolytic system, the thrombus occupies the finite space of the vascular lumen. As a result, blood flow with the thrombus is interrupted, and infarction of important organs occurs. Plaque rupture is accidental and unpredictable, and medical treatment starting from plaque rupture is a post-event response. Regarding the occurrence of cardiovascular events, there is only a therapeutic strategy that controls the risk and lowers the probability of atherosclerotic accidents, and a fundamental, radical therapeutic approach is required.

Among the immune cells and inflammatory cells involved in atherosclerosis, B cells have not necessarily been recognized to play a central role so far. Looking back at the history of atherosclerosis research, we can see that research on B cells is progressing. Arteries have a three-layered structure consisting of the intima, media, and adventitia. In 1915, at the very early stage of atherosclerosis research, Sir Clifford Allbutt investigated the relationship between atherosclerosis and B cells infiltrating the tunica adventitia, which can be traced back to the description of "round cell growth in the adventitia in atherosclerosis is correlated with absorption of depraved matter from the diseased intima." ([Diseases of the Artery]). The "round cells" infiltrating the adventitia described in this article were also reported in 1956 and 1962, and were found to be mainly B cells. Recent studies have shown that the activation of B cells in the adventitia is important in controlling atherosclerosis. Hamze et al. used microdissection techniques to analyze individual lymphocytes in coronary arteries. (J Immunol. 2013;191:3006-16.) They found that the majority of B cells reside in the adventitia of coronary arteries. In human and rodent models of atherosclerosis, B cells are present in the adventitia and form the tertiary lymph node tissue (ATLO, Artery Tertiary Lymphoid Organ). PVAT (perivascular adipose tissue), which is adjacent to the adventitia of arteries, contains complex and diverse cellular components, such as macrophages, T cells, and B cells, named FALCs (Fat-Associated Lymphoid Clusters). FALCs are in close proximity to the adventitial ATLO and may play a role in the establishment of atherosclerosis. Inflammation in atherosclerosis is not a transient inflammation. We reported that as a result of the activation and conditioning of B2 cells in the spleen, they infiltrate PVAT, and become the starting point of the pathology related to the onset of atherosclerosis. (Chen L, Ishigami T, et al. eBioMedicine, 2016) A high-fat, high-calorie diet causes changes in intestinal bacteria, dysbiosis, a decrease in intestinal barrier function, and activation of splenic B2 cells. Activated B2 cells can serve as a hypothetical biological model of atherosclerosis, in which they infiltrate the aortic adipose tissue and secrete IgG/IgG3 antibodies to promote atherosclerosis.

B cells infiltrate the PVAT from the very early stage of atherosclerosis, form FALCs, and form ATLO, a tertiary lymphoid tissue, in the adventitia. This may be the origin of immune response in atherosclerotic plaques in the endothelium/intima. The humoral immune response mediated by atherosclerotic B cells derives from antibody production and specific activation of B cell subtypes in the spleen. If this reaction, which can be said to be the biological starting point of atherosclerosis, can be controlled, it may become an option for the treatment of atherosclerosis.

This Special Issue, entitled "Current Challenges and Advances in Atherosclerosis", will focus on recent challenges and advances in the field of atherosclerosis by global expert scientists and clinicians to elucidate the complicated features of atherosclerosis and translational attempts for both medical and scientifical frontiers.

> **Tomoaki Ishigami** *Editor*


*2.2. VOO Samples*

Each sensory panel (EU, IT, IP, UN, UP/ZRS, UZ) was responsible for the sampling (two years: 2016–2017 and 2017–2018 olive harvest seasons) of a possibly balanced number of extra virgin (EV), virgin (V), and lampante (L) samples defined by sensory evaluation, according to EU Regulations [8] and later modifications.

These samples were collected to be representative of the most common olive cultivars, different geographical origins (without restrictions to the national market of each sensory panel), different sensory profiles and, especially, the main sensory defects perceived. Samples were directly requested from olive oil companies under a nondisclosure agreement containing information related to responsibility and confidentiality of data. The selection of the sample set for each year was based on sensory screening: each panel leader, assisted by his/her deputy panel leader, was responsible for applying the official procedure for assessing the organoleptic characteristics of VOOs (according to [8] and later modifications). At the end of the sensory screening, panel leaders sent the results of sample screening to the UNIBO panel leader.

Each olive oil company that agreed to participate in the sampling phase had to provide information related to each commercial sample furnished (date of sampling, geographical origin of olives, olive variety/varieties, PDO, PGI, sanitary state of olives, time and storage conditions of olives before milling, mill location, technology parameters, date of production of the oil, date of start of oil storage, type of storage tank/bottles, oil storage temperature). The need to collect all available information on each sample was the reason why samples were requested from companies, avoiding collection directly on the market. However, the oils collected were representative of possible commercial samples and also so-called borderline samples that can be the object of disagreement between panels in terms of sensory characteristics. The olive oil company indicated a person who was responsible for oil sampling among its employees, who followed and applied guidelines for oil sampling [23,24].

For each sample, a volume of 7 L was requested from the olive oil company and collected inside adequate tins or bottles. In case of a batch produced and packed (oil already bottled), the responsible person selected by the olive oil company had to collect the volume required (7 L) taken with a random selection of bottles. The panel leader assisted by his/her deputy panel leader was responsible for managing the laboratory samples. The olive oil company dispatched the packaged samples to the panel leader, who organized the preparation of laboratory samples (after proper homogenization, using a 0.5 L tin), their label codes, and shipment to all the OLEUM partners involved (carried out in the shortest time possible by tracked shipments). Sample codes summarized the basic information: partner acronym (responsible for the sampling) and number (progressive for two years, related in unique way for each sample).

The samples collected for each year (first year, 180 oils; second year, 154 oils) were divided into four subgroups and their sensory evaluation as well as sensory results were planned over time by the UNIBO panel. All samples, stored in the lab at 10–12 ◦C, were reconditioned at room temperature for 6–8 h before preparing samples for sensory analysis.

## *2.3. Sensory Analysis*

The panel test method was carried out using six OLEUM panels. Positive and negative descriptors were evaluated according to the official procedure ([8] and later modifications). The intensity of each attribute was graded by assessors using a continuous unstructured line scale of 10 cm. Each 15 mL sample was tasted at 28 ± 2 ◦C in a tasting booth, regulated in terms of shape and equipment [4]. Each panel leader collected the profile sheets completed by each taster (8–12) from his/her panel, reviewed the intensities assigned to the different attributes, and inserted the sensory data in the IOC Excel program for statistical elaboration based on the calculation of the median. The robust coefficients of variation (CVr%) were calculated and validated ([8] and later modifications).

Moreover, with the aim to monitor and possibly improve the performance of panels, after elaboration of each subgroup of sample data, the UNIBO panel, being responsible for the sensory activities, adopted and applied the quality control procedures to check the validity of the results obtained by OLEUM panels in agreement with IOC guidelines for the quality control of virgin olive oil panels revised in 2018 [25], specifically: (i) z-score estimation was conducted for each sensory panel (IOC z-score and OLEUM z-score) to estimate the panel's trueness; (ii) available IOC standards and other materials characterized (samples from previous IOC proficiency tests) were provided to each panel for training purposes; (iii) replicate analysis of three samples selected between the entire set of samples, to estimate panel precision, was performed.