**3. Concluding Remarks**

In all the work that has been performed thus far, it has been noted that all the focus has been on FSP as an enhancement technique on aluminium alloys, magnesium, and other alloys. It is also noticed that the common mechanical properties analyzed include the tensile test, fatigue and microhardness. These properties are studied correlatively with the microstructure. Very few works thus far, which considers friction stir processing as a post weld processing technique for tungsten inert gas dissimilar alloy welded joint. There is minimal to no trace of any literature on submerged friction stir processing of tungsten inert gas and friction stir welded dissimilar alloy joints. Therefore, this opens an opportunity for the exploration on the impact of friction stir processing and submerged friction stir processing towards the properties of the friction stir welded and tungsten inert gas welded dissimilar joints.

**Author Contributions:** Both authors have fully contributed equally to all the work produced. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research received no external funding.

**Acknowledgments:** The authors would like to thank the Cape Peninsula University of Technology for allowing this study to be performed.

**Conflicts of Interest:** The authors declare no conflict of interest. The authors declare there is no funding involved in this study.


**1. Introduction**

Cardiovascular diseases are the leading cause of death worldwide [1]. Cardiotoxicity is one of the main causes of withdrawal of drugs from the market [2]. Traditionally, new cardiac drugs and the cardiotoxicity of cardiac and non-cardiac drugs have been tested with rodent cardiomyocytes as well as with transfected non-cardiac cells [3–5]. However, the results of these experiments are not always applicable to humans. Therefore, more accurate human cardiomyocyte models are needed for preclinical analysis of drugs as well as for basic research and disease modeling of human cardiac diseases [6]. Human-induced pluripotent stem (hiPS) cells can be reprogrammed from any somatic cell by introducing the pluripotency factors [7] and these cells can be di fferentiated into functional cardiomyocytes with multiple methods, as recently reviewed [8]. However, these cells have been criticized as being immature and more resembling fetal than adult cardiomyocytes

(CMs) [9]. Compared to adult human CMs, hiPSC-CMs are small in size, round or multi-angular, and typically single-nucleated, whereas adult CMs are rod-like and large, with 25%–57% of the cells multi-nucleated [10,11]. The aspect ratio can be used as an indicator of the cell shape. Due to the round shape, hiPSC-CMs have an aspect ratio of (2–3): 1, whereas adult CMs are clearly longitudinally-oriented with an aspect ratio of (5–9):1. Sarcomeres of the hiPSC-CMs are disorganized and short (<2 μm), the sarcoplasmic reticulum is poorly developed, and the sarcolemma exhibits no transverse tubules [10,12]. The electrophysiological properties and the gene expression of the hiPSC-CMs di ffer from adult CMs [12].

The contractile ability of CMs is enabled by multiple sarcomere units that are integrated in the cytoskeleton of the cell [13]. The e fficiency of the CM contraction relies on the proper orientation and length of the sarcomeres and on the rod-like shape of the cell [14]. Thus, the structural maturation of the cells has been the focus when studying maturation methods for hiPSC-CMs. Multiple methods and strategies have been suggested to improve the maturation of hiPSC-CMs in vitro, including topographical cues, substrate sti ffness, medium additives, mechanical and electrical stimulation, genetic manipulation, and co-culture with other cell types [10,15]. However, a deeper understanding of the maturation process of hiPSC-CMs is still required to develop platforms to promote the maturation of the cells and producing hiPSC-CMs more resembling adult CMs.

Various sca ffolds have been studied to orient hiPS-CMs, such as electrospun-aligned fiber textiles [16,17] and micro-grooved culture substrates [18]. Most of the cell cultures in vitro are coated on flat surfaces, which provide a two-dimensional (2D) environment for the cells. 2D culture forces cell polarization by providing cell-extra cellular matrix (ECM) connections on only one side of the cells [19,20]. Cells in 2D are usually flat because they try to spread out on the surface. By providing a three-dimensional (3D) environment with proper topographical cues and an extracellular matrix, cells can create more cell–ECM connections, which potentially a ffect cell proliferation and even di fferentiation and maturation [20]. Sti ffness of a flat, 2D culture substrate a ffects cell properties, proliferation, and di fferentiation [21]. To increase the cardiac functionality and maturity, hiPS-CMs have been cultured with, for example, endothelial cells, and this has shown to enhance cardiomyocyte proliferation and functionality [22]. Co-culture of hiPS-CMs with cardiac fibroblasts has improved the structural and functional properties of the cells [23,24]. A similar maturation-enhancing e ffect was observed when cardiomyocytes were cultured on top of the vascular-like network produced from endothelial cells and fibroblasts [25,26]. Stem-cell-derived cardiomyocytes aligned according to the vascular structures of the network and their sarcomere structures were more oriented.

Textiles create a 3D culture environment and provide topographical support for di fferent types of cells. Having a highly interconnective porous structure, textiles enable access of media and nutrients to the cells inside the material. Weaving is a conventional and basic textile technique that can also be used to fabricate tissue engineering sca ffolds. Weaving enables the formation of textile structures with controllable properties, such as porosity, orientation, morphology, and mechanical properties. These parameters can be modified, for example, by changing the number of filaments, filament diameter, and weaving patterns. Biostable polyethylene terephthalate (PET) is one of the most used polyesters, and has many applications including in biomedical applications, for example as hernia meshes. It has also been used in many cell culture studies. The raw material of fibers can be changed too, and in the future, biodegradable textiles could be used as a vehicle for implantation of cardiomyocyte sheets for myocardial ischemia or scar repair applications [27–29].

In this study, PET textiles were used as culture substrates for hiPSC-CMs. The aligned textile fibers were hypothesized to provide su fficient topographical cues to improve the maturation state of hiPSC-CMs. The PET textiles had di fferent weaving patterns, including a plain weave and a plain weave derivative, which altered their topography and other properties. They were coated with di fferent biological compounds: GeltrexTM (Thermo Fisher Scientific) and gelatin were used. The CMs were characterized by cell morphology, sarcomere organization, expression of cardiac specific genes, and calcium handling properties.


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**A Program Management Information System**

The urban renewal project is an activity to regenerate existing city areas that have become run-down. In South Korea, urban renewal is led by the government, which perceived the importance of reducing environmental burden and enhancing the efficiency of use of existing urban infrastructure (Bae et al., 2008). Managing these projects is challenging for the government because of its insufficient experience with projects of a such massive scale and input resources (Lee, 2000). Furthermore, since the locations designated to be renewed are

On the one hand, information technology and information systems (IT/IS) in urban planning and construction have advanced with the advance of IT in general. In South Korea, in particular, one of the leading countries for IT, there are moves to introduce IT/IS into

To encourage urban renewals, the South Korean government has funded research and development (R&D) projects. An organization named the Korea Urban Renaissance Center (KURC) was established to manage these R&D projects. The center aims to make an adjustment to urban renewals to follow the localized model in South Korea and create a long- or mid-term strategy, reinforcing the national capacity for urban renewal, analyzing the effect or performance of urban renewal projects, and implementing test beds and

Launched in 2007, the R&D project began with 41 sub-projects. One of these, "Development of Intelligent Program Management Information Systems" (i-PgMIS) aims to develop information systems to support decision making and information management for urban renewal mega projects (Kim et al., 2009). The authors have involved some of the research group, developing a framework of i-PgMIS according to their requirements. Integrating unit systems for functions such as cost/time management into i-PgMIS is also included. Unit systems are also being researched by other participants in the same R&D

This chapter introduces the framework of i-PgMIS managing the first version of a completed

dispersed over the whole territory, supporting tools are necessary.

management of megaprojects such as urban planning (Oh, 2001).

commercializing the output of R&Ds (KURC, 2007b).

system and its development background.

**1. Introduction**

project.

**for Managing Urban Renewals**

Hyeon-Jeong Choi and Ju-Hyung Kim

*Hanyang University Republic of Korea*


**5. Conclusions**

We have identified multiple patient-identified barriers to living kidney donation in the UK Black, Asian and minority ethnic group population, which should be further investigated and addressed to reduce the ethnic inequity in living-donor kidney transplantation in the UK.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2077-0383/9/11/3751/s1, Supplementary Study sites, Supplementary Interactions, Table S1. Responders and non-responders, Table S2. Responders compared to national denominator population, Table S3. Missing data analysis, Table S4. Participant demographics—comparison of White and Black, Asian and minority ethnic group respondents, Table S5. Multivariable logistic regression analysis comparing reasons potential donor unsuitable between White and Black, Asian and minority ethnic group participants—missing explanatory variables imputed, Table S6. Characteristics of Black, Asian and minority ethnic group participants providing qualitative responses. Table S7. Themes and illustrative quotes from thematic analysis of White participant responses.

**Author Contributions:** Conceptualization, P.B., Y.B.-S., C.R.T.; methodology, P.B., Y.B.-S., S.M.; software, K.W., P.B.; formal analysis, K.W., P.B., S.M., A.O.-S.; investigation, K.W., P.B.; data curation, P.B.; writing—original draft preparation, K.W., P.B.; writing—review and editing, K.W., P.B., A.O.-S., S.M., F.J.D., Y.B.-S., C.R.T., F.C., D.I., S.B.; supervision, A.O.-S., Y.B.-S., C.R.T., F.C.; project administration, P.B.; funding acquisition, P.B. All authors have read and agreed to the published version of the manuscript.

**Funding:** This report is independent research arising from a Kidney Research UK Project Grant (Reference RP\_028\_20170302). Neither Kidney Research UK nor the University of Bristol had any role in study design, data collection, analysis, interpretation, manuscript preparation of the decision to submit the report for publication. K.W. is supported by the Elizabeth Blackwell Institute for Health Research, University of Bristol and the Wellcome Trust Institutional Strategic Support Fund. P.K.B. is funded by a Wellcome Trust Clinical Research Career Development Fellowship. Y.B.-S. is supported by the NIHR Applied Research Collaboration West (NIHR ARC West) as data science lead. The views expressed in this article are those of the author(s) and not necessarily those of the NIHR or the Department of Health and Social Care or any of the other funders.

**Acknowledgments:** The authors would like to thank all the study participants, the participating centre research nurses and coordinators (Hugh Murtagh, Nina Bleakley, Mary Dutton, Kulli Kuningas, Cecilio Bing Andujar, Ann-Marie O'Sullivan, Nicola Johnson, Kieron Clark, Thomas Walters, Mary Quashie-Akponeware, Jane Turner, Gillian Curry, Hannah Beer, Lynn.D Langhorne, Sarah Brand, Maria Weetman, Molly Campbell, Megan Bennett, Sharirose Abat, and Agyapong Kwame Ansu) and the local collaborators who facilitated the study (Sarah Heap, Mysore Phanish, Shafi Malik, Aisling Courtney, Adnan Sharif, Nicholas Torpey, Refik Gökmen, Michael Picton, Linda Bisset, Edward Sharples, and Simon Curran).

**Conflicts of Interest:** The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.


**Radiation Damage in Materials: Helium Effects**

Special Issue Editors

**Yongqiang Wang Khalid Hattar**

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*Special Issue Editors* Yongqiang Wang Los Alamos National Laboratory USA

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*Editorial* **Special Issue: Synthesis, Application, and Biological Evaluation of Chemical Organic Compounds**

**Stanimir Manolov \* and Iliyan Ivanov \***

Department of Organic Chemistry, Faculty of Chemistry, University of Plovdiv, 24 Tsar Assen Str., 4000 Plovdiv, Bulgaria

**\*** Correspondence<EMAIL_ADDRESS>(S.M<EMAIL_ADDRESS>(I.I.)

This Special Issue of *Processes*, entitled "Synthesis, Application, and Biological Evaluation of Chemical Organic Compounds", gathers the most recent work of leading researchers in a single forum. The contents include a broad range of synthesized or new organic compounds and nanostructures, the extraction of active components of plant and animal origin, the in silico analysis of such compounds, and biological assessment using various methodologies. In one of the presented studies, the authors [1] report the synthesis of chitosan/Fe(III)/deferoxamine nanoparticles. The developed nanoparticles have demonstrated exceptional antibacterial activity in vivo and in vitro, outperforming the conventional drugs ampicillin and gentamicin. Furthermore, the authors assert that the nanoparticles are risk-free regarding their potential toxicity. They discovered that adding iron ions to the chitosan matrix improves the ability of the resultant nanoparticles to damage the integrity of microbe membranes when compared to pure chitosan. The addition of deferoxamine to the produced nanoparticles significantly increases their ability to destroy the bacterial membrane. Ahmad and co-workers [2] reported the synthesis of poly(*o*-anisidine)/BaSO4 nanocomposites via the oxidative polymerization of *o*-anisidine monomer with BaSO4 filler for the composite materials' possible antibacterial capabilities. To attain optimal and controllable characteristics in the nanocomposites, the BaSO4 filler ratio was varied between 1% and 10% with respect to the matrix. The FTIR measurements demonstrated a considerable interaction between POA and barium sulfate, as well as good UV-visible behavior of absorption. By altering the percentage load of the BaSO4 filler, the conducting characteristics may be controlled. Furthermore, distinct bacterial strains, *Pseudomonas aeruginosa* and *Staphylococcus aureus*, were employed to assess the antibacterial activity of the POA/BaSO4 nanocomposites. For *Staphylococcus aureus* and *Pseudomonas aeruginosa*, the highest inhibition zones of 0.8 and 0.9 mm were achieved using 7% and 10%, respectively.

In this Special Issue, a few articles have been published reporting the synthesis, full characterization, and in vitro and in silico biological assessment of novel compounds. A very interesting article was presented by Apostol and her colleagues [3]. They describe the synthesis and design of new compounds with an *L*-valine fragment and a 4-[(4-bromophenyl)sulfonyl]phenyl skeleton, which belong to *N*-acyl-*α*-amino acids, 4*H*-1,3-oxazol-5-ones, 2-acylamino ketones, and 1,3-oxazoles chemotypes. Antibacterial activity against bacterial and fungal strains, antioxidant activity utilizing DPPH, ABTS, and ferricreducing power assays, and toxicity against the freshwater cladoceran Daphnia magna Strauss were all assessed. In addition, in silico investigations of the possible antibacterial action and toxicity were carried out by the authors. The findings regarding the antibacterial activity, antioxidant effect, and toxicity experiments, as well as in silico analysis, demonstrated the potential of *N*-{4-[(4-bromophenyl)sulfonyl]benzoyl}-*L*-valine and 2-{4- [(4-bromophenyl)sulfonyl]phenyl}-4-isopropyl-4*H*-1,3-oxazol-5-one for the development of new antimicrobial drugs to combat Gram-positive bacteria, particularly *Enterococcus faecium* biofilm-associated illnesses. Manolov and co-workers [4] reported the synthesis of a series

**Citation:** Manolov, S.; Ivanov, I. Special Issue: Synthesis, Application, and Biological Evaluation of Chemical Organic Compounds. *Processes* **2023**, *11*, 2802. https:// doi.org/10.3390/pr11092802

Received: 1 September 2023 Accepted: 19 September 2023 Published: 21 September 2023

**Copyright:** © 2023 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

of novel compounds between ketoprofen and nitrogen-containing heterocyclic compounds (piperidine, pyrrolidine, 1,2,3,4-tetrahydroquinoline, and 1,2,3,4-tetrahydroisoquinoline). The compounds were investigated for their anti-inflammatory and antioxidant properties in vitro. The hybrids' lipophilicity was assessed, both theoretically (*cLogP*) and empirically (RM). The compounds' affinity for human serum albumin was estimated in silico using two software tools, and the stability of the predicted complexes was determined using molecular dynamics analysis. All new hybrids outperformed the reference compound, quercetin, in terms of HPSA activity. The in vitro results were validated by molecular docking. The maximum affinity for albumin is shown by hybrid molecule **3c** and compound **3d**. They are more potent anti-inflammatory agents than their forerunner, ketoprofen, as well as the regularly used ibuprofen. The same scientific group, from Plovdiv, Bulgaria [5], reported the synthesis of new hybrid molecules between divinylene oxide and the same nitrogen-containing heterocyclic compounds mentioned above. The anti-inflammatory, anti-arthritic, antioxidant, reducing, and chelating activities of the synthesized compounds were evaluated. When compared to the utilized ketoprofen (720.57 ± 19.78) standard, the less lipophilic molecules, H2 (60.1 ± 8.16) and H4 (62.23 ± 0.83), had an ATA that was approximately 12 times greater. The inhibition of albumin denaturation resulted in the newly produced hybrids being considered as promising anti-inflammatory medicines, as the expressed values were greater than the ketoprofen standard (126.58 ± 5.00), with the exception of H3 (150.99 ± 1.16). All reported compounds had high activity in terms of in vitro biological activities, making them excellent candidates for possible future medications. Elkaeed and colleagues investigated the anticancer and VEGFR-2 inhibitory properties of a novel 1H-indole derivative in vitro and in silico [6]. A novel 1H-indole derivative was designed to correspond to the reported properties of anti-VEGFR-2-approved drugs. The new compound's inhibitory potential was revealed by a molecular docking experiment that identified the pertinent binding sites. After that, six studies of MD simulation were carried out by the authors for 100 ns to verify the precise binding and optimal energy. Furthermore, MM-GBSA demonstrated flawless binding with a total exact energy of −40.38 Kcal/Mol. Using binding energy decomposition, the MM-GBSA tests identified the important amino acids in the protein–ligand interaction, revealing the diversity of interactions of compound **7** inside the VEGFR-2 enzyme. Because chemical **7** is novel, the authors' DFT experiments were used to optimize its molecular structure. In silico ADMET experiments revealed that compound **7** had a high drug-likeness value. It is interesting to note that compound **7** has a higher experimental in vitro prohibitory capacity than sorafenib, with an IC50 value of 25 nM. It is noteworthy that compound **10** showed potent inhibitory actions against two cancer cell lines (MCF-7 and HCT 116) with IC50 values of 12.93 and 11.52 μM, revealing good selectivity indices of 6.7 and 7.5, respectively.

Scientists from the Republic of Korea [7] set out to investigate and compare the coumarin-based compounds found in *G. littoralis* extracts, as well as the antioxidant and anti-osteoporotic properties of various *G. littoralis* extracts (leaf and stem, fruit, whole plant, and root extracts) on bone metabolism. In their research, they looked into how *G. littoralis* extract affected the growth and osteoblastic differentiation of MC3T3-E1 osteoblasts. The highest concentrations of scopoletin (53.0 mg/g) and umbelliferone (1.60 mg/g) were found in stem extracts when compared to the other samples. According to the findings presented here, ethanolic extracts of *G. littoralis* are an efficient osteoporosis preventative.

In their article [8], Rahman and colleagues describe the development of charge transfer (CT) complexes between organic and/or bioactive compounds, which is a crucial component in understanding molecule–receptor interactions. They created two novel CT complexes, procainamide–chloranilic acid and procainamide–2,3-dichloro-5,6-dicyano-1,4 benzoquinone, using the electron donor procainamide, electron acceptor chloranilic acid, and 2,3-dichloro-5,6-dicyano-1,4-benzoquinone. The stability of each complex was examined for the first time by utilizing spectroscopic properties such as the formation constant, molar extinction coefficient, oscillator intensity of the ionization potential, dipole moment, and standard free energy. Density functional theory (DFT) calculations were carried out, utilizing the ωB97XD/6-311++G(2d,p) level of theory to comprehend the complexes' noncovalent interactions. Both the DFT-computed interaction energies (ΔIEs) and the Gibbs free energies (ΔGs) matched, as observed experimentally. The DFT results strongly support the experimental findings.

Sheep placenta extract (SPE) is widely used in traditional medicine for its physiological effects, such as its wound-healing, antioxidant, and anti-inflammatory characteristics. However, the antiaging effects of SPE are uncertain. Chou and colleagues [9] studied the effect of SPE on aging using the senescence-accelerated mouse prone 8 (SAMP8) strain. After assessing age index characteristics, such as skin glossiness, spine lordosis, and kyphosis, it was discovered that SPE therapy reduced the aging index. Furthermore, they discovered that biochemical indicators such as lactic acid, glucose, ketone bodies, free fatty acids, tumor necrosis factor-alpha (TNF-α), and interleukin 6 (IL-6) did not change in the SPE-treated experimental group after 13 weeks. They discovered that lipid peroxidation (LPO) was reduced, while catalase and superoxide dismutase (SOD) activity was dramatically elevated in the brain tissues of SPE-treated male and female mice. SPE supplementation decreased the aging index and minimized the oxidative stress brought on by the aging process in mice without creating any harmful effects, suggesting the potential of SPE as a potent antiaging remedy.

Additionally, Rosales Martínez and Rodríguez-García shared their thoughts on how hydrogen/deuterium exchange in ambrox could lengthen the aroma of perfumes and extend their shelf lives [10]. Ambrox is a common ingredient in high-end perfumery since it is a natural marine compound with a wonderful ambergris-like scent. Improving the long-term aroma and shelf life of perfumes is a primary objective in the fragrance industry. To the best of the authors' knowledge, the exchange of hydrogen for deuterium to minimize the volatility of smell components has not yet been researched. In their opinion-type article, they share a new use of deuteration to synthesize deuterated ambrox in order to reduce volatility, improve long-term smell, and extend the shelf-life of perfumes.

The included articles demonstrate the importance and versatility of the search for new potential biologically active structures and their applications. All contributors show an impressive passion and professional abilities in the pursuit of new synthetic methods for producing new structural hybrid molecules, as well as those extracted from plant and animal sources. They use novel, proven, and dependable methods and technology to attain their goals. The new synthetic compounds and extracts described here have the potential to be used as new therapeutic formulations due to the variety of actions they exhibit, some of which are even more active than the standards with which they are compared.

As we reflect on the collective efforts and accomplishments showcased in this special issue, it is clear that the synthesis, application, and biological evaluation of chemical organic compounds represent a thriving and ever-evolving field of research. The potential for discovery remains boundless, and the pursuit of innovative solutions to global challenges continues to inspire scientists worldwide.

We extend our sincere gratitude to all the authors, reviewers, and contributors who have made this special issue possible. Your dedication to advancing the frontiers of knowledge is truly commendable. We hope that the research presented here serves as a source of inspiration and a catalyst for further exploration in the fascinating world of chemical organic compounds.

In the spirit of scientific inquiry and collaboration, we look forward to the continued advancement of this field and the remarkable discoveries that lie ahead. Thank you for joining us on this enlightening journey, and may our collective efforts continue to shape a brighter, more sustainable future.

We appreciate all of the contributors and the Special Issue's Editor-in-Chief, Mr Scott Pan, for their enthusiasm, as well as the editorial staff of *Processes*. Thank you all for the support.

**Conflicts of Interest:** The authors declare no conflict of interest.


**Chapter 10**

*Bioactive Compounds - Biosynthesis, Characterization and Applications*

Dias, D., Pinto, E., … Colepicolo, P. (2017). Extraction of sterols in brown macroalgae from Antarctica and their identification by liquid chromatography coupled with tandem mass spectrometry. Journal of Applied Phycology, *29*(2), 751-757. https://doi. org/10.1007/s10811-016-0905-5

[43] D'Adamo, S., Schiano di Visconte, G., Lowe, G., Szaub-Newton, J., Beacham, T., Landels, A., … Matthijs, M. (2019). Engineering the unicellular alga Phaeodactylum tricornutum for high-value plant triterpenoid production. Plant Biotechnology Journal, *17*(1), 75-87. https://doi.org/https://doi.org/10.1111/

pbi.12948

**152**


**1.  This disease increases the risk of non-alcoholic hepatitis (NASH), chronic interstitial hepatitis, hepatic failure, and even hepatocellular carcinoma [1–3]. One of the characterizations of NAFLD is increased levels of reactive oxygen species (ROS) [4]. Some studies have shown that high levels of ROS promote the development of NAFLD/NASH and hepatocellular carcinoma by inducing ER stress [5] or regulating the AMPK signaling pathway [6]. There are several antioxidant enzymes such as superoxide dismutase (*SOD*), catalase (*CAT*), and glutathione peroxidase (*GPX*) responsible for scavenging cellular ROS. Meanwhile, the expression levels of *SOD* and other antioxidant enzymes are decreased in NAFLD/NASH [4]. Interestingly, lipid droplets (LDs) have been shown to be involved in the cellular stress response process. Bailey et al. demonstrated that lipid droplets can act as antioxidant organelles that protect *Drosophila* neural stem cells from hypoxia-triggered ROS [7], by allowing neuronal stem cells to keep proliferating under hypoxic conditions, and protection likely involves sequestering vulnerable membrane lipids away from ROS [8]. Furthermore, LDs also respond to starvation-induced stress by increasing their contact with mitochondria and lysosomes, which could

consist in the role of these contacts in transferring fatty acids from LDs to mitochondria or lysosomes for energy supply [9]. Moreover, the formation of nuclear LDs is related to the stress induced by phospholipid shortages [10,11]. Our previous study has shown that hydrogen peroxide promoted the formation of cellular LDs [12]. However, whether the increased cellular LDs play a role as anti-oxidants is largely unknown.

*Perilipin 5* (*PLIN5*) is one of the conserved LD proteins, which belongs to the PAT (*perilipin, adipophilin,* and *TIP47*) protein family [13]. Oxidative tissues such as skeletal muscle, liver, and brown fat have high expression levels of *PLIN5*, indicating that *PLIN5* plays an important role in lipid storage and LD function [14–16]. Previous studies identified that *PLIN5* regulated triglyceride contents in hepatocytes [17] and skeletal muscle [18]. Overexpression of *PLIN5* in skeletal muscle promotes oxidative gene expression and lipid content [19]. Recently, *PLIN5* was reported to be the key factor that regulated LD contacting mitochondria [20,21]. The N-terminal (1-188aa) of *PLIN5* is the conserved PAT domain, and 189-391aa is the domain contacting with patatin like phospholipase domain containing 2 (*PNPLA2, ATGL*). The C-terminal of *PLIN5* (443-463aa) is the key sequence related to mitochondrial recruiting [22]. LD–mitochondria contact is important for the energy supply during starvation stress, which promotes lipid β-oxidation [9,23], and the transfer process of fatty acids from LDs to mitochondria was also observed by probe imaging [24,25]. Recently, a study also found that LD–mitochondria contact contributed to lipid synthesis and LD expansion [22]. Furthermore, LDs are able to protect against cellular apoptosis by clearing harmful proteins from outer mitochondrial membranes [26]. Moreover, *PLIN5* has been shown to limit fatty acid toxicity [27]. These studies suggested that *PLIN5* was involved in the process of cellular anti-oxidation.

In the present study, we found that hydrogen peroxide- or lipopolysaccharide (LPS)-induced oxidative stress up-regulated both mRNA and protein levels of *PLIN5*. The overexpression of *PLIN5* increased the cellular LD content, promoted LD–mitochondria contact, reduced cellular ROS level, and enhanced mitochondrial function-related gene expression, whereas knockdown *PLIN5* indicated opposite phenotypes. Moreover, we identified that the promoter region of *PLIN5* contained the binding sites of *JUN, ATF1, ATF3,* and *ATF4*, and therefore *PLIN5* expression was activated by the JNK-p38-ATF pathway. By bioinformatic analysis, it has been found that *PLIN5* has a high expression in liver hepatocellular carcinoma (LIHC), and additionally, low expression of *PLIN5* is correlated with poor prognosis in LIHC. Therefore, *PLIN5* can be a potential therapeutic target in NAFLD and NAFLD-induced LIHC.


**2.6 Marker assisted characterization of germplasm**

In the twenty-first century, food, water and land are biggest challenges for increasing population. Agricultural activities need to expand, become sustained, and be more adaptive to climate change. To improve sustainability in agricultural systems, new paradigms are required, to explore the genetic potential of the huge although

unfortunately underutilized resources of genetic diversity available for different crops. For breeding of climate-resilient varieties, a better understanding of evolutionary genetic variability is essential. Genetic diversity is the precious wealth for any crop improvement programme but due to climatic changes, it is reducing continuously. In last century, 75% decline in genetic diversity was witnessed in farmers' fields and it would further decline by about 20% by 2050.

Besides that, genetic resources can be excellent breeding material to develop superior variety in future breeding programmes. They can also be used in different breeding programs in order to increase the genetic base of cultivated crop varieties.

It has been observed that wild relatives have several desirable characteristics like resistance to biotic and abiotic streses, nutritional characteristics, cleistogamy, photo insensitivity and cytoplasmic male sterility (CMS).

In the past few decades, revolutionary approaches and systems have offered a great wealth of genetic and genomics resources that revolutionized research in both model and crop legumes.

A recent study on chickpea presents evidence of severe domestication bottleneck. Efficiency of cultivated population of chickpea is 100 times lesser than that of wild chickpea (Cicer reticulatum and C. echinospermum). In legume crops, study on landraces and wild relatives are significantly benefited by advanced technologies of genomics, phenotyping and computational biology.

The Vavilov Institute of Plant Genetic Resources (VIR), which houses a special genebank in St. Petersburg, Russia, using a mixture of genomics, computational biology and phenotyping to classify the 147 accessions of chickpea from Turkey and Ethiopia. The combination of high-density genotyping data with historical phenotypic information on these VIR landraces allowed chickpea genomes to enter 'agro islands' or 'domestication islands' that display significant associations with multiple phenotypes. These "genomic gems" have also been identified in chickpeas containing co-adapted and co-localized gene complexes. These are LG4 and LG2 in chickpea containing multiple genes/ QTLs related to drought and disease resistance, respectively. WGRS/RADSeq of 90 Cicer accessions, including cultigens, landraces and wild accessions, previously identified a wide collection of 54 genes on LG3 that could have been targeted during modern breeding efforts to manipulate salient characteristics such as flowering time.

Similarly, a genomic segment with an excess of MTAs for agronomically significant traits was observed on LG9 after re-sequencing of 292 accessions in pigeonpea [13].

In a recent study, to understand the genetic relationships between various lentil species/subspecies, a lentil collection comprised of 467 wild and cultivated genotypes originating from 10 different geographical regions was evaluated. A total of 422, 101 high-confidence SNP markers were identified against the reference lentil genome (cv. CDC Redberry). Phylogenetic analysis clustered the germplasm collection into four groups, namely, Lens culinaris/Lens orientalis, Lens lamottei/Lens odemensis, Lens ervoides, and Lens nigricans. Results of this study indicated that L. nigricans is most distantly related to L. culinaris and major differences were observed in six genomic regions with the largest being on Chromosome 1 (c. 1 Mbp) and further additional structural variants are likely to be identified from genome sequencing studies. In order to improve germplasm and for introgression of novel genes, this will provide further insights into the evolutionary relationship between cultivated and wild lentil germplasm.

Guar (Cyamopsis tetragonoloba (L.) Taub.) is primarily grown as an industrial crop due to its high-quality galactomannan gum used as a thickener, flocculant,

emulsion stabiliser and gelling agent. Therefore, the novel set of molecular markers (nSSR) could be adopted as a useful tool to characterize the guar accessions for future breeding programmes.

#### **2.7 Marker assisted backcrossing (MABC)**

Research on legumes has greatly benefited from different available molecular markers in crop plants. Association between molecular markers and plant traits in these crops has introduced a novel approach to breeding that is based on the crossing of selected genotypes and selection of suitable progenies based on associated markers/QTL(s) rather than depending solely on phenotypes. Over the past three decades, the advancement and development of molecular marker technologies have been steady, such as low-throughput restriction fragment length polymorphisms (RFLPs) in the 1980s, high-throughput array-based markers in the 2000s and next-generation ultrahigh-throughput sequence-based marker systems in the 2010s. RFLP, RAPD, AFLP and SSR markers are low-throughput marker systems and are also considered past molecular markers. Besides these, next-generation sequencing (NGS) and genotyping by sequencing (GBS) are high and ultrahigh-throughput marker systems. These are based on low-cost and high-throughput sequencing technologies and are considered as present marker systems.

In cereals, so many outstanding achievements of marker-assisted breeding are available, but in legumes, negligence and lack of genomic resources adversely affected their initial establishment in the field of molecular breeding. Now recent advances in pulse genomics have led to the launch of several marker-assisted breeding projects.

RAPD and RFLP markers were used in five wild lentil taxonomic groups to understand their genetic makeup [47]. A genetic linkage map was also constructed in lentils with RAPD, AFLP and RFLP markers [48].

For shielding the varieties against various biotic and abiotic stresses and for ensuring crop productivity; gene mapping, tagging and marker-assisted selection have vital importance. Identifying and deploying molecular markers/QTLs in a desired background would be a priority. Marker-aided selection (MAS) greatly reduce the time and effort required to recover high levels of resistance from the donor and simultaneously recover the genomes of the recurrent parent. It has become more easier to transfer desirable genes/QTLs from wild relatives to existing cultivars due to MAS and transgenics.

Fusarium wilt (FW) and Ascochyta blight (AB) are two major constraints in chickpea (Cicer arietinum L.) production. The most affordable approach for longterm control of ascochyta blight and fusarium wilt in chickpeas is known to be the use of varieties with high resistance levels.

The availability of molecular markers associated with QTL for ascochyta blight resistance provided an opportunity to introgress the traits into adapted chickpea cultivars. backcrossing between moderately resistant donors (CDC Frontier and CDC 425-14) and the adapted varieties (CDC Xena, CDC Leader and FLIP98-135C) resulted in a variety with improved resistance to ascochyta blight [49].

More recently, five resistant lines representing foc2 gene introgressed into the background of Pusa 256 were reported with the help of foreground selection aided by two SSR markers (TA 37 and TA110). Cultivar Vijay was used as a donor of foc2 gene [50]. Annigeri 1 and JG 74 are elite high-yielding desi cultivars of chickpea, in Karnataka and Madhya Pradesh. in recent years, have become susceptible to race 4 of Fusarium wilt (FW).

A widely grown cowpea variety in Africa was improved by adding drought tolerance, striga and root knot nematode resistance QTLs using SNP markers. The major QTL region on LG 8 was introgressed from cultivar V-16 into the bacterial leaf blight susceptible variety C-152 through marker-assisted backcrossing (MABC) [51]. Similarly, By backcrossing resistance to CpMv gene was transferred into variety C-152. Cowpea mosaic virus (CpMV) was responsible for 80–100% yield loss in cowpea. SSR markers were used for linkage map construction and indicated that two markers MA15 and MA 80 were linked to CpMV resistance.

At ICRISAT in hybrid pigeonpea programmes, markers associated with fertility restoration and CMS are being used. This improved the selection efficiency of hybrid breeding and accelerated the breeding work [52]. In addition, a range of markers, including SSRs and SNPs are now available to enable genetic purity testing of pigeonpea hybrids and their parents. Recently, ICRISAT has launched a collaborative effort with ICAR-IIPR and other NARS institutions/universities to accelerate and target the improvement of ruling mega varieties of pigeonpea in India.

In groundnut breeding, the use of molecular markers in backcross breeding programme accelerated selection of recombinant progenies bearing nematode resistance and high oleic acid. Selection for high oleic acid content in groundnut was facilitated by one CAPS marker along with gel-free SNP assay using HybProbe design for the selection of nematode resistance SCAR, SSR and CAPS marker were used.

Recently in peanuts, two ahfad2 alleles from SunOleic 95R were introgressed into ICGV 05141 using marker-assisted selection. Marker-assisted breeding effectively increased oleic acid and oleic to linoleic acid ratio in recombinant lines up to 44% and 30%, respectively as compared to ICGV 05141. Subsequently, In the marker-assisted backcrossing-introgression lines, a 97% increase in oleic acid, and a 92% reduction in linoleic acid content were observed in comparison to the recurrent parent [53].

As opposed to traditional breeding, gene stacking or pyramiding is a useful strategy for transferring multiple desired genes or QTLs from various parents into a single genotype in the shortest possible time (two to three generations). Molecular markers that may be beneficial for marker-assisted selection and gene pyramiding have been identified through genetic linkage analyses and QTL mapping. The most effective and inexpensive means of combating plant diseases is the use of genetic resistance. Gene pyramiding is thus a sensible approach to creating multiple and enduring resistance. Most successful approach in common bean for wide spectrum control of common mosaic virus is to combine I, bc-u, bc-12, bc-22, and bc-3 genes. SCAR marker was used for MAS. In lentils, molecular marker-assisted gene pyramiding was used for resistance to ascochyta blight and anthracnose. In this research, two genes for resistance to ascochyta blight and the gene for anthracnose resistance in lentil breeding lines were pyramided using linked RAPD marker [49].

#### **2.8 Genome editing**

Crop plant genome editing is a faster-growing technique for inserting specified changes into the genome precisely and with great accuracy. Genome editing has emerged as an alternative approach to conventional plant breeding, and transgenic (GMO) approaches to improve food legumes and their sustainable production. Instead of spontaneous non-specific changes caused by radiation or chemical mutagenesis, crop researchers have long required mutations at specific sites in the genome. This

#### *Recent Advancements in Genetic Improvement of Food Legume Crops DOI: http://dx.doi.org/10.5772/intechopen.106734*

approach allows for site-specific DNA insertion, deletion, modification, or replacement in a living organism's genome. The plant research community has not been widely involved with earlier SSN-specific (sequence-specific) genome-based editing technologies, because of the complex design and labour-intensive assembly of particular DNA binding protein for each target gene. A relatively new and comparatively easier technique for genome editing is CRISPR (clustered regularly interspaced short palindromic repeats) technique which is based on a simplified version of the bacterial CRISPR-Cas9 antiviral defence system. CRISPR genome editing technique is based on Cas9 protein which is an endonuclease. This endonuclease induces double-strand breaks using a guide RNA that is complementary to a target gene [54]. In order to create mutants for inaccessible genes, CRISPR-Cas9 would be a very useful technique. It can mutate multiple loci and make large deletions, thereby speeding the plant breeding without directly adding any transgene. The sequence-specific nucleases-based plant genome editing has a great potential to develop modified crops which can address the increased global food requirements and sustainable agriculture production. CRISPR/ Cas9 was applied first in model legume plants to induce targeted mutagenesis.

A web tool was designed to identify potential CRISPR/Cas9 target sites and also a soybean codon-optimized CRISPR/Cas9 platform to induce mutation at target sites in somatic cells of Glycine max and Medicago truncatula [55]. In a recent study, an efficient CRISPR/Cas9 system was developed for targeted gene mutations in the model legume M. truncatula. A specific sgRNA was designed that targeted medicago phytoene desaturase (MtPDS) gene involved in the carotenoid biosynthesis. Very recently in Cowpea, the representative SNF gene has been effectively disrupted with an efficient CRISPR/Cas9-mediated genome editing. Guide RNAs (gRNAs) for the symbiosis of receptor-like kinase (SYMRK), reached ~67% mutagenesis efficiency in plants with hairy roots, and the formation of nodules in both mutants was totally prevented [56]. Conventional breeding is based on natural genetic variation and rigorous back-crossing systems are needed to incorporate the selected traits into an elite genotype. Unlike conventional breeding techniques, the present diversity does not limit CRISPR because it can directly integrate new mutations. This approach will benefit particularly those crops which have narrow genetic diversity and low variability for traits. Therefore, genome editing can speed up plant breeding programmes by inserting correct and predictable modifications directly in desirable backgrounds. The CRISPR/Cas9 system is especially beneficial because multiple traits can be modified simultaneously.


**About the Editor**

#### **Bhaskar K Somani**

Bhaskar K Somani has been involved in clinically innovative patient-centred treatments. His research includes minimally invasive surgical techniques in the management of kidney stone disease, enlarged prostate (BPH), renal cancer and urinary tract infections. He is the Clinical Director of 'South Coast Lithotripter Services'. He is an active member of BAUS Endourology sub-section and is the Wessex Clinical Research Network and Simulation Lead for Urology. He is a board member of the European School of Urology (ESU), ESU Training and Research group and EAU section of the uro-technology (ESUT) endourology group. He is also the chosen representative for the UK in the Endourology society and is a member of the EAU urolithiasis guidelines panel. He coordinates the largest hands-on-training simulation course for urology in the world (EAU-EUREP). He has published over 550 scientific papers and has been invited as a speaker to perform live surgery for moderations in more than 30 countries worldwide. He has raised a grant income of GBP 3.2 million. He has published excellent clinical outcomes; his outcomes and research translate into benefits for patients. For his work, he has won the Endo Society 'Arthur Smith' award in 2020, BAUS 'Golden Telescope' in 2021 and Zenith Global Health Special Recognition award in 2021.

## *Editorial* **Minimally Invasive Urological Procedures and Related Technological Developments—Series 2**

**Bhaskar Somani**

Department of Urology, University Hospital Southampton NHS Foundation Trust, Southampton SO166YD, UK<EMAIL_ADDRESS>
The world of minimally invasive urology has experienced enormous growth in recent decades with technological innovations related to new techniques and equipment, better training, and the clinical adoption of translational research. There has been a substantial increase in studies related to the application of lasers both for the treatment of stones and enlarged prostates. This Special Issue in the *Journal of Clinical Medicine* (JCM) is dedicated to a collection of eleven high-quality scientific contributions.

The first paper looked at a systematic review and meta-analysis of randomised trials on the use of coated versus non-coated urethral catheters for the prevention of catheter-associated urinary tract infections (CAUTI) [1]. A meta-analysis of 12 studies and 36,783 patients showed no significant difference in patients with long-term catheterization, and this benefit of coated catheters should be balanced against the cost to healthcare. The second paper compares the mineral content of tap water in UK and whether this was relevant to kidney stone disease (KSD) [2]. Data from 66 UK cities showed a significant variation, and depending on where someone lived, drinking 2–3 L of tap water could contribute over one-third of the recommended daily calcium and magnesium, with possible implications for KSD incidence and recurrence.

Lasers have evolved with the advent of high-power holmium lasers, thulium fiber laser (TFL), and pulse modulation such as Moses technology [3–5]. There are four papers involving the use of lasers in endourology [6–10]. Laser efficiency and safety were compared between Holmium YAG and TFL lasers, with the latter showing higher efficiency [6]. Laser lithotripsy during ureteroscopy and stone fragmentation with comparative outcomes between ≤10 years and ≥80 years were examined by the second paper [7]. The results showed that while the former group had a higher incidence of repeated procedure, there was no difference in the overall stone-free rate (SFR) and complications between the groups. The third paper looks at low- vs. high-power lasers during the Holmium laser enucleation of the prostate (HoLEP) procedure [8]. Current evidence shows no difference in outcomes between the two and that low-power HoLEP is safe, feasible, and effective. The last paper looked at a comparison of the safety and efficacy of laser vs. pneumatic intracorporeal lithotripsy for the treatment of bladder stones in children [9]. In this prospective randomised study of 64 children, while the stone treatment was similar between groups, pneumatic lithotripsy was associated with a significantly greater risk of having at least 1 adverse effect.

Fluoroless endourology has been on the rise to minimise radiation doses. The total X-ray-free ultrasound-guided mini-PCNL in a Galdakao-modified supine Valdivia position was reported in the next paper from Taiwan on 150 patients [10]. The outcomes show that complete X-ray procedures are feasible, safe, and effective. Intrarenal pressure monitoring is also postulated to help minimise complications, but so far, there was a lack of pressure monitoring devices [11,12]. Measuring and minimising pressure would lower the risk of sepsis and other related complications [13]. Real-time intrarenal pressure control during flexible ureteorenoscopy was demonstrated in the next study using a vascular wire [14]. In this pilot study, a pressure wire was placed in the renal cavities to measure the intrapelvic

**Citation:** Somani, B. Minimally Invasive Urological Procedures and Related Technological Developments—Series 2. *J. Clin. Med.* **2023**, *12*, 2879. https://doi.org/ 10.3390/jcm12082879

Received: 5 April 2023 Accepted: 10 April 2023 Published: 14 April 2023

**Copyright:** © 2023 by the author. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/).

pressure (IPP), with results showing the feasibility of this technique and the monitoring of IPP to identify and avoid high IPP, thereby avoiding complications.

In the world of smart apps [15], there is a rise in wearable device technology with an annual growth rate of nearly 26% projected for India. A cross-sectional web-based survey of 495 responders exhibited a significant correlation with the adoption and acceptance of wearable devices for healthcare management in the Indian context [16]. Minimally invasive surgical therapies (MISTs) for benign prostate enlargement (BPE) have experienced a revolution with new technologies that are now on the market [17–19]. However, the transurethral resection of prostate (TURP) is still practiced as the primary modality of treatments in many centres. In the context of cardiovascular morbidity, patients often have to remain with the choice of preoperative antithrombotic therapy. A single-centre study from China examined venous thromboembolism (VTE) and bleeding after TURP in patients with preoperative antithrombotic therapy [20]. In a cohort of 31 patients, the authors conclude that the short-term preoperative discontinuation of therapy may help patients obtain a relatively safe opportunity for TURP surgery. This must be balanced against the risks of VTE, perioperative bleeding, and serious cardiovascular and cerebrovascular complications.

Artificial intelligence (AI) is now used in various urological conditions, including urolithiasis, benign prostate hyperplasia (BPH), and uro-oncology [21]. Similarly, radiomics is increasingly applied to the diagnosis, management, and outcome prediction of various urological conditions [22]. In a systemic review of the role of radiomics in urolithiasis, seven studies were included [23], with radiomics used to identify calculi, differentiate phleboliths, and classify stone types and compositions pre-operatively. It has also been utilized to predict outcomes and complications after endourological procedures and, hence, has great future potential.

This Special Issue has several interesting papers that will help clinicians in decision making and treatment choices. While there is a spectrum of papers from tap water to catheters, lasers and its use in BPH and stone surgery, wearable devices, and the role of radiomics, perhaps more needs to be performed to address other aspects of a patient's journey, such as cost and the quality of life in the management of these patients [24,25]. I am thankful to the reviewers for their professional comments and to the *JCM* team for their ongoing support with this Special Issue. Lastly, I want to thank all authors for their valuable contributions.

**Conflicts of Interest:** The authors declare no conflict of interest.

#### **References**

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