Roles Of DNA Damage in Renal Tubular Epithelial Cells Injury Ⅱ

4 DNA repair in renal tubular epithelial cells 

In eukaryotes, histone proteins serve as “packing” agents for DNA (Uckelmann and Sixma, 2017; Gong and Miller, 2019). The impacts of DNA damage might affect the superstructures. Damage to DNA mostly affects the double helix’s essential structure. There are several checkpoints built into the cell cycle to make sure the cell is ready to move on to mitosis. The G1/S, G2/M, and spindle assembly checkpoints are the three primary checkpoints that control the passage through anaphase. The G1 and G2 checkpoints are responsible for detecting the presence of DNA damage, with the S phase being the stage of the cell cycle where DNA damage is most likely to occur. At the G2 checkpoint, DNA replication completion and the presence of DNA damage are both examined (O’Hagan et al., 2008).

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As we previously stated, environmental influences as well as regular metabolic processes can damage DNA in human cells. These lesions can result in structural damage to the DNA molecule, potentially leading to harmful mutations in the cell’s genome that can affect the survival of daughter cells following mitosis. The DNA repair mechanism is always active in response to damage to the DNA structure. DNA repair refers to the biochemical and molecular biological procedures for removing DNA damage and restoring genomic integrity (Huang and Zhou, 2021). These procedures include DNA damage sensing and signaling, recruitment of repair machinery proteins to the sites of damage, and step-by-step release of these proteins to restore the integrity of the genome. The study of DNA damage repair has quickly branched out into fields including cancer biology, photobiology, and radiobiology. Tomas Lindahl, Paul Modrich, and Aziz Sancar received the 2015 Nobel Prize in Chemistry for their research on the molecular principles underlying DNA repair procedures.

Cells can recognize DNA damage because it changes the spatial organization of the double helix. As soon as the damage is located, specific DNA repair molecules bind to it or the area around it, allowing other molecules to bind and form complexes that allow the actual repair to occur. DNA repair involves numerous standard processes as follows (Figure 1).

4.1 Direct reversal 

DNA damage can be repaired by cells using chemical reversals. This direct reversal method does not entail a break in the phosphodiester backbone and is specific to the type of DNA damage. Although there is no need for a template for this repair, the kind of damage that is fixed can only happen in one of the four bases. In bacteria, fungi, and most animals, the light reactivation process catalyzed by photolysis enzyme reverses this damage. In human cells, nucleotide excision repairs damage caused by ultraviolet rays. Pyrimidine dimer formation under UV irradiation results in abnormal covalent bonds formed between adjacent pyrimidine bases (Lucas-Lledo and Lynch, 2009). The protein methylguanine methyltransferase can directly undo another sort of damage, guanine-based methylation (MGMT). The third type of DNA damage that cells can repair is specific methylation of the nucleotide’s cytosine and adenine.

4.2 Single-strand damage 

The other strand of a double helix can be utilized as a template to direct the repair of the damaged strand when just one of the two strands is flawed. To repair damage to one of the two paired molecules of DNA, several excision repair methods remove the damaged nucleotide and replace it with an undamaged nucleotide that is complementary to the undamaged DNA strand. 

4.3 Base excision repair (BER) 

Most frequently, single base or nucleotide damage is fixed by deleting the offending base or nucleotide and replacing it with the proper base or nucleotide. During base excision repair, a glycosylase enzyme eliminates the damaged base from the DNA by cleaving the bond between the base and the deoxyribose. These enzymes produce an apurinic or apyrimidinic site by removing a single base (AP site). At the AP location, AP endonucleases snip the DNA’s broken backbone. After using its 5′to 3′exonuclease activity to remove the damaged area, the complementary strand serves as a template for the DNA polymerase as it correctly synthesizes the new strand. The DNA ligase enzyme then closes the gap (Zhao et al., 2021). A 2015 study found that the degree of kidney fibrosis was positively connected with higher MUTYH expression, a DNA repair enzyme that starts a BER by identifying and deleting 8-Oxoguanine (8- oxo) and its matched adenine (Lu et al., 2015). 

4.4 Nucleotide excision repair (NER)

NER is typically utilized to fix bulky, helix-distorting damage initiated by UV light, such as pyrimidine dimerization. Nearly all eukaryotic and prokaryotic cells use NER, a highly evolutionarily conserved repair process. Uv proteins in prokaryotes mediate NER. Many additional proteins are involved in eukaryotes. The DNA polymerase uses the complementary strand as a template to properly create the new strand (Reardon and Sancar, 2006).

According to a study that compared the stress responses of rat kidney tubular (NRK-52E) and glomerular (RGE) cells after exposure to cisplatin, RGE performed the best in repairing Pt- (GpG) intrastrand crosslinks, as evidenced by the high mRNA expression of NER factors (Kruger et al., 2015).

4.5 Mismatch repair (MMR) 

Almost all cells include mismatch repair mechanisms to fix faults that are not caught by proofreading. At least two proteins make up these systems. The first recognizes the mismatch, and the second calls in an endonuclease to break the freshly manufactured DNA strand adjacent to the area that has been damaged. Following this, an exonuclease removes the damaged area, followed by DNA polymerase resynthesis and DNA ligase nick sealing.

4.6 Double-strand breaks 

The most serious and fatal form of DNA damage to cells is DNA DSBs. Histone proteins can shield a single strand of damaged DNA from additional damage and chemical attack. The end of the DNA is exposed as a result of DNA DSBs. The intracellular DDR system will become active if this scenario persists without prompt treatment. One of the effects is to halt cell growth and division or to start apoptosis, inducing the cell’s eventual demise. Fortunately, cells have evolved several defenses against DNA DSBs breaks since the eukaryotic stage (Liang et al., 2005).

4.7 Homologous recombination (HR) 

The cell can access and duplicate intact DNA sequence information in trans through homologous recombination, which is particularly useful for repairing DNA damage that affects the two strands of the double helix. It uses the corresponding feature of the chromosomes in cells. If one of the double-stranded DNA on a chromosome breaks, the other corresponding DNA sequences on a chromosome can serve as a fixed template to respond before breaking the sequence, which is why under certain circumstances homologous recombination is also known as gene conversion. Cell cycle progression is crucial for the homologous recombination HR repair process. Homologous chromosomes are the sole template that HR can employ in the G1 phase when there are 2n chromosomal sets. The number of chromosomal pairs doubles to 4n with the insertion of a sister chromatid in the S/G2 phase, giving the HR mechanism extra repair templates to choose from. It is widely accepted that HR maintenance operations are more active during the S/G2 period.

4.8 Non-homologous end joining (NHEJ) 

The most significant difference between NHEJ and HR is that the repair proteins in NHEJ can directly bring the broken ends closer to each other without the help of any template, and then re-join the broken ends with the help of DNA ligase. The mechanism of NHEJ is both simple and templating-independent. In organisms with more complex gene bodies and more junk DNA, NHEJ is more active than HR. However, in organisms with simpler gene bodies, especially single-cell organisms, NHEJ is more likely to break the original sequence integrity.

Double-strand breaks and DNA cross-linkages may result in permanent DNA damage when typical repair mechanisms fall short and cellular death does not take place. The type of cell, the cell’s age, and the extracellular environment are only a few of the variables that affect how quickly DNA is repaired. For a cell that has considerable DNA damage or is no longer able to repair DNA damage correctly, one of three scenarios is possible: Senescence, an irreversible state of dormancy, apoptosis, or programmed cell death, and uncontrolled cell proliferation, which can result in the growth of a malignant tumor. These are three common consequences of senescence. To maintain the integrity of its genome and, by extension, the normal functioning of the organism, a cell’s ability to repair damaged DNA is essential. Many genes that were first thought to affect lifespan have since been found to play a part in DNA damage repair and defense.

5 DDR in renal tubular epithelial cells 

Some of the processes discussed above can function on their own to repair simple lesions. The DNA damage response, however, controls the repair of more complicated lesions requiring numerous DNA processing steps. The DDR may be crucial for a successful restoration of the lesions that are the hardest to treat. When different types of DNA lesions activate DNA-damage response proteins, DNA repair begins. By phosphorylating repair proteins to change their activities, by initiating a complex chain of modifications to the local chromatin structure close to the damage site, and by altering the cellular environment generally to make it more conducive to repair, these kinases improve the effectiveness of DNA repair (Sirbu and Cortez, 2013). The signaling pathways that these proteins then activate include those for DNA repair, cell cycle arrest, apoptosis, and senescence. In the context of chromatin close to the damage site, complex repair mechanisms take place. Cancers result from dysregulation of DNA damage response and repair, and several of these proteins offer potential therapeutic targets (Figure 1).

The phosphatidylinositol 3-kinase-related kinases (PIKK) oversee the DDR kinase signaling cascades, ataxia telangiectasiamutated (ATM), DNA-dependent protein kinase (DNA-PKcs), and ATM and Rad3-related (ATR) are included. ATR reacts to a variety of DNA lesions, including those related to DNA replication, while DNA-PKcs and ATM are largely engaged in DSB repair (Cimprich and Cortez, 2008). Because of its adaptability, ATR is crucial for the survival of reproducing cells in both mice and humans (Brown and Baltimore, 2000; de Klein et al., 2000; Cortez et al., 2001).

Three levels of DNA repair are regulated by DDR kinases. First, they directly control DNA repair enzymes’ activity through post-translational changes. When complicated lesions are repaired and when replication forks are halted, these alterations seem to be especially crucial. Second, DDR kinases modify the chromatin around DNA damage to create an environment that is conducive to repair. Additionally, this chromatin response serves as a scaffold to draw in other DDR components that control signaling and repair. Finally, DDR kinases work on the nucleus or possibly the entire cell, creating a biological milieu favorable to repair. This overall response involves alterations in transcription, cell cycle, chromosomal mobility, and deoxynucleotide (dNTP) levels. When damage is persistent, control of these mechanisms might be particularly crucial for repair (Sirbu and Cortez, 2013).

DDR has been the subject of further research lately, including studies on its function in renal tubular epithelial cells. Cellular communication factor 2 (CCN2, also known as CTGF) is a significant factor in the development of CKD and was found to exacerbate DNA damage and the ensuing DDR-cellular senescence-fibrosis sequence after renal IRI. By lowering DNA damage caused by oxidative stress and the ensuing DDR, CCN2 inhibition may lessen the risk of AKI (Valentijn et al., 2021). According to M. Uehara et al., ATM inhibition worsens tubular injury via upregulating p53-dependent pro-apoptotic signaling and does not enhance DNA repair following cisplatin-induced DNA damage. When ATM inhibitors are made available in clinical practice in the future, acute renal injury must be closely monitored (Uehara et al., 2020). After kidney epithelial cell damage, Kishi et al. show that DNA repair—rather than cell proliferation—plays a key role in recovery and lifespan by lowering apoptosis, G2/M cell-cycle arrest, and subsequent fibrosis (Molitoris, 2019). By blocking the DNA damage response, p53, MAP kinases, and oxidative/nitrosative stress pathways, JQ1 which is one of the most characterized BET inhibitors with high specificity against BET proteins may protect against the nephrotoxic effects of cisplatin, according to Sun et al. (2018).

According to a different study, lovastatin effectively blocks the pro-apoptotic signal pathways of the DDR of injured tubular epithelial cells caused by cisplatin (Krüger et al., 2016).

An essential step in the direct religation of the DSB by NHEJ is the autophosphorylation of DNA-PKcs. The substrates involved in DNA repair, checkpoint signaling, and cell fate decisions including apoptosis and senescence are both distinct and shared by ATM and ATR. PLK3 (polo-like kinase 3) is implicated in oxidative stress-induced DNA damage and TEC apoptosis in renal I/R injury, and PLK3 suppression reduces TEC death after I/R injury by obstructing the ATM/P53-mediated DDR, according to research on the enzyme’s function in renal I/R injury (Deng et al., 2022). Another study found that decreasing the expression of the ATM gene in HK2 cells reduced the amount of LPS-induced inflammation and autophagy in sepsis-induced AKI in vitro. This data suggests that the ATM pathway may be used by LPS to cause autophagy in HK2 cells, which would boost the production of inflammatory markers (Zheng et al., 2019). The development of AKI may be influenced by cytoplasmic DNA-PKcs and phosphorylated Fis1 (Wang et al., 2022). For the past several years, our team has been researching the function and mechanism of chronic kidney disease.

6 DNA damage and aging 

Senescence is a crucial biological process that helps to support embryo development, avoid the growth of tumors, and minimize tissue harm. Senescent cells do, however, play a role in the emergence of age-related illnesses as they accumulate in organs with aging (Shmulevich and Krizhanovsky, 2021). Aging is caused by a variety of variables, including the time-dependent buildup of macromolecular harm, which includes DNA damage and incomplete DNA repair (Yousefzadeh et al., 2021). To escape replicating the damaged genome, sustained DNA damage (genotoxic stress) sets off a cascade of signals that cause apoptosis or senescence. At the same time, these processes encourage cellular senescence (Babbar et al., 2020; Yousefzadeh et al., 2021). A growing body of data points to inflammation as another significant side effect of DNA harm. One of the signs of aging and the primary cause of many age-related illnesses is inflammation (Zhao et al., 2023). The buildup of age-related DNA damage, activation of transposons, cellular senescence, and accumulation of persistent R-loops are thought to be the triggers of these signaling cascades, which are activated by engaging the cGAS-STING axis or activating NF-kappaB via ATM. Meanwhile, the shift of heterochromatin components mediated by epigenetic modifications generated by DNA harm can result in inflammation and aging (Zhao et al., 2023). AKI and the aging activation pathway are closely linked. Tubular cells suffer damage as a result of kidney toxicity or ischemia-reperfusion injury, which is primarily expressed as cell membrane damage, cytoskeleton damage, and DNA degradation. These damages eventually cause tubular cell necrosis, apoptosis, and mortality (Andrade et al., 2018). Age-related processes like interstitial fibrosis, renal tubule atrophy, and scant capillaries make it difficult for the kidneys to recoup from structural and functional damage, which raises the possibility that AKI and the development of chronic kidney disease are closely related (Andrade et al., 2018; Kim et al., 2021). Studies in recent years have also concentrated on the connection between chronic renal illnesses like diabetic nephropathy and cell aging. Diabetic nephropathy is caused and developed by DNA damage, epigenetic change, and mitochondrial malfunction (Xiong and Zhou, 2019).

7 Gender differences in DNA damage and repair 

Clinical epidemiological studies have shown that while CKD is more common in women than in men, end-stage renal disease, including dialysis and kidney transplantation, is more common in men (Carrero et al., 2018). There is mounting evidence that the way each gender responds to kidney injury differs significantly. The tolerance for renal ischemia-related impairment varies by gender. Although recent research suggests that male hormones also play a significant role in these differences, it has long been believed that the protective effects of estrogen account for sex differences in disease susceptibility. Gender and sex hormones appear to have an impact on vascular factors like endothelin, nitric oxide, and angiotensin II (Metcalfe and Meldrum, 2006). Additionally, inflammatory mediators with different expression and activity based on sex and the presence of sex steroids included TGF-β1, TNF-α, and p38 mitogen-activated protein kinase. Studies have revealed that the renal toxicity of cisplatin varies significantly by gender (Pezeshki et al., 2017; Valdivielso et al., 2019). Proteomic research on the disruption of dihydrotestosterone metabolism in human primary proximal tubule epithelial cells raises the possibility that changed tubular energy metabolism may be connected to the harmful effects of androgens on the kidney (Valdivielso et al., 2019).

There are also gender differences in DNA damage and repair especially DSB, according to a growing body of studies. One of the studies has shown that the chance of developing Alzheimer’s disease (AD) may be influenced by the presence of more DSB markers in the hippocampus of female E4 mice (Boutros et al., 2023). Males and females experience different age-related changes in DSB repair, according to research that examined DSB repair in peripheral blood lymphocyte (PBL) circulation in male and female donors of various ages (Rall-Scharpf et al., 2021). According to one research, the new tank tests had a greater impact on female zebrafish, who displayed constrained locomotion and exploratory behavior, decreased mitochondrial enzyme activity, increased DNA damage, and cell death after hypoxia damage (Das et al., 2019). The effect of the aromatic amine 2-acetylaminofluorene (AAF) on hepatocellular carcinoma induction in male rats was more pronounced than in female rats, demonstrating that the AAFDNA adduct in male rats was approximately twice as large as that in female rats, and the DNA repair level in male rats cultured with AAF dosage was approximately three times greater than that in female rats (Williams et al., 2016).

The question of whether DNA amplification, DNA repair, and DDR are gender-related in kidney disease has received very little attention in studies and articles up to this point. It is beneficial for clinicians and researchers to have a deeper understanding of the specific role of DNA damage in kidney disease as it may provide a more accurate treatment direction for the prevention of kidney disease and kidney injury in the future. This will help to improve the understanding of sex and sex-specific differences in the etiology, mechanism, and epidemiology of kidney disease, as well as to combine the factors related to DNA damage and sex.

8 Interventions targeted DNA damage in renal diseases 

As discussed earlier, DNA damage is a common occurrence in a variety of renal diseases. Numerous studies have focused on DNA damage in kidney disorders, including treatments that target DNA damage specifically. According to one study, miR-155 deficiency dramatically lessens pathology and mortality in cisplatin-induced AKI by preventing DNA damage (Yin et al., 2022). Another study has demonstrated that when compared to normoxia-cultured mesenchymal stem cells, hypoxic mesenchymal stem cells (HMSCs) demonstrated a superior anti-oxidative impact in I/R-damaged rat kidneys via reducing DNA damage (Tseng et al., 2021). A study on the prevention of chronic fibrosis revealed that NMN could drastically reduce DNA damage, senescence, and inflammation in tubular cells. As a result, giving NMN may be an effective way to stop or cure renal fibrosis after AKI (Jia et al., 2021). Additionally, M. A. Mohammed et al. have demonstrated that vitamin D protects against oxidative stress and DNA damage to lessen the acute renal damage caused by gentamicin (Mohammed et al., 2019). Therefore, focused approaches to DNA damage may be effective methods for kidney protection in renal diseases.

9 Perspective 

DNA damage alters the structure of genetic materials, which makes it impossible for the replication mechanism to work properly. In response to the presence of DNA lesions, cells could either be repaired or eliminated by triggering cell death after DNA repair fails. When DNA is damaged, DNA repair proteins are frequently activated or induced. However, if the level of DNA damage is too severe to repair, the organism has acquired the further option of initiating an apoptosis program that can stop excessively damaged cells from mutating and developing into cancers. The complicated signal transduction network known as DDR detects when DNA is damaged and starts the cellular response to the damage. While most of the DNA damage can be repaired, such a repair function is not 100% successful all the time. Since DNA damage has been implicated in a variety of kidney injuries, its early and sensitive detection in kidney disease by specific assays may become a new target for early diagnosis or prognosis of the disease. In addition, targeting DNA damage and DNA damage response may well be a successful kidney protection technique in AKI treatment.

References 

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., Cervelli, T., Galli, A., and Andreassi, M. G. (2013). DNA modifications in atherosclerosis: From the past to the future. Atherosclerosis 230, 202–209. doi:10. 1016/j.atherosclerosis.2013.07.038 Boutros, S. W., Zimmerman, B., Nagy, S. C., Unni, V. K., and Raber, J. (2023). Age, sex, and apolipoprotein E isoform alter contextual fear learning, neuronal activation, and baseline DNA damage in the hippocampus. Mol. Psychiatry. doi:10.1038/s41380- 023-01966-8 Brown, E. J., and Baltimore, D. (2000). ATR disruption leads to chromosomal fragmentation and early embryonic lethality. Genes Dev. 14, 397–402. doi:10.1101/ gad.14.4.397 Cadet, J., Loft, S., Olinski, R., Evans, M. D., Bialkowski, K., Richard Wagner, J., et al. (2012). Biologically relevant oxidants and terminology, classification and nomenclature of oxidatively generated damage to nucleobases and 2-deoxyribose in nucleic acids. Free Radic. Res. 46, 367–381. doi:10.3109/10715762.2012.659248

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