The association between new MACE and the variables of interest were investigated by univariable and multivariable analyses. From a source population of 2,184,662 patients, we identified 4072 adults diagnosed with T2DM and CAD . The main comorbidities observed included arterial hypertension, hyperlipidemia, and obesity, with metformin and statins being the treatments most frequently prescribed. MACE development was associated with multivessel (Hazard Ratio (HR) = 2.49) and single coronary vessel disease (HR = 1.71), transient ischemic attack (HR = 2.01), heart failure (HR = 1.32), insulin treatment (HR = 1.40), and percutaneous coronary intervention (PCI) (HR = 2.27), whilst statins (HR = 0.73) were associated with a lower risk of MACE occurrence. In conclusion, we found six risk factors associated with the development of MACE which were related with cardiovascular diseases and T2DM severity, and treatment with statins was identified as a protective factor for new MACE in this study.Patients with Type 2 diabetes mellitus (T2DM) and coronary artery disease (CAD) are at high risk of developing major adverse cardiovascular events (MACE). This is a multicenter, retrospective, and observational study performed in Spain aimed to characterize these patients in a real-world setting. Unstructured data from the Electronic Health Records were extracted by EHRead Diabetes mellitus (DM) is a key public health problem worldwide associated with the development of complications such as cardiovascular comorbidities, which include coronary artery disease (CAD) ,2,3. It Given the clinical burden that cardiovascular complications have on patients with T2DM, there has been an increased focus on the joint management of both entities, T2DM and cardiovascular disease (CVD). Furthermore, during the last decade, there has been increasing pressure from regulatory agencies for antidiabetic treatments to demonstrate cardiovascular safety and benefits, especially for major adverse cardiovascular events (MACE) . However® , a technology that applies Natural Language Processing (NLP) and machine learning, to extract, organize, and analyze the unstructured clinical information that health professionals register in patients’ electronic health records (EHRs) [The present study aimed to use population-based data to provide real-world insights into the clinical characterization and treatment management of diabetic patients who develop CAD in Spain. Moreover, this study investigated the probability of developing MACE after presentation of CAD in T2DM patients and the potential risk or protective factors associated with its occurrence within this population. To do so, we used EHReads (EHRs) ,11,12,13This was a real-world, multicenter, retrospective, and observational study  based on the secondary use of the unstructured data captured in the EHRs. A cross-sectional analysis of all patients was performed at the index date, defined as the first time in the study period when the patient fulfilled all inclusion and no exclusion criteria. At this point, demographics, comorbidities, vital signs, general characteristics of T2DM and CAD and treatments were evaluated. The cumulative incidence of MACE, defined as previously described by the presence of myocardial infarction, stroke, hospitalization for unstable angina, and urgent coronary revascularization , was anaThe data source was the free-text information within the EHRs of 12 representative hospitals from six major regions in Spain: Madrid , Catalonia , Valencia , Balearic Islands , Castilla La Mancha , and Castilla y León . Data entry in outpatient clinical reports, discharge reports, emergency reports, prescriptions, and other medical reports, were collected from all available services and departments in each participating site, including inpatient hospital, outpatient hospital, and emergency room, between 1 January 2014 and 31 December 2018.The source population of the study comprised all adult patients with available EHRs in the participating hospitals within the study period with at least 6 months of follow-up. Patients with T2DM and a diagnosis of CAD were included. These two entities were considered when they were contained in the unstructured, free-text information in the EHRs based on clinical diagnosis. T2DM were also considered when documented ongoing use of glucose-lowering drugs  was found for at least 6 months. CAD was considered if there was evidence of stenosis ≥ 50% of at least 1 coronary artery but without a history of having a previous myocardial infarction or stroke, and without planned coronary, cerebrovascular, or peripheral arterial revascularization. Patients with prior myocardial infarction or stroke, history of liver cirrhosis or liver cancer, intracranial bleeding, renal failure requiring dialysis, or ongoing treatment with anticoagulant medication at index date or non-available follow-up information spanning at least 6 months were excluded from the study.® technology, which uses NLP, machine learning, and deep learning techniques for extracting free text from de-identified and processed EHR and translates it into a study database. EHRead® technology have been described elsewhere [Unstructured clinical data were extracted using the EHReadlsewhere ,17,18,19® includes codes, concepts, synonyms, and definitions used in clinical documentation and is based on SNOMED CT [Using the information obtained from this process, a statistical model was generated to describe the demographic and clinical characteristics of the population with T2DM and CAD. Information was analyzed and expressed by means of concepts that contain the most significant information in the text. The terminology considered by EHReadNOMED CT ,21.® performance was assessed to evaluate the ability of the technology to identify mentions of coronary disease and its related clinical variables within patients’ EHRs [® upon reading that same set of EHRs was carried out independently at each hospital site [® at identifying records that contain mentions of “Coronary disease” in the target population with a precision, recall, and F1-score of 0.89, 0.73, and 0.80, respectively. Moreover, the evaluation yielded a F1-score of above 90% in most of the variables analyzed [EHReadts’ EHRs ,12,13,22tal site . Thus, tanalyzed .p < 0.05. All analyses were performed using “R” software, version 4.0.2 .Categorical variables were described via frequency tables; whereas numerical variables were presented using summary tables that included the mean, standard deviation (SD), median, and quartiles . Missing data were handled according to the nature of the data collection process, thus lack of information  was considered a “true zero” for binary variables  but was treated as missing data for numerical variables . No missing data imputation was performed. Cumulative incidence of new cases of MACE was estimated through the Kaplan–Meier approach. Both univariable and multivariable analyses were performed to investigate the association between the occurrence of new MACE and the variables of interest. Variables with >20% missing values or with zero variance were excluded. In the univariable analysis, a Cox proportional hazard (PH) regression model was fitted to the study population for each variable at baseline. In the multivariable approach, a data-driven variable and model selection was performed. First, a Cox PH model was fitted to the study population using all eligible study variables. Then, we carried out backward stepwise selection . We used the Bayesian information criterion (BIC) to reach a model with the optimal explanatory variables. Significant differences were considered when This study was classified as a “non-post-authorization study” by the Spanish Agency of Medicines and Health Products (AEMPS) and was approved by the Institutional Review Board (IRB) of each participating hospital. All methods and analyses were conducted in compliance with local legal and regulatory requirements, as well as generally accepted research practices described in the Helsinki Declaration in its latest edition, and Good Pharmacoepidemiology Practices. Data were analyzed from de-identified EHRs, which were aggregated in an irreversibly dissociated manner. Thus, individual patient consent was not required in the study.The source population of the study comprised a total of 2,184,662 adult patients with available EHRs in the hospitals of interest including 217,632 (10%) patients with a confirmed diagnosis of T2DM. The study population only included adult patients with T2DM and stable CAD diagnosis with no previous history of myocardial infarction and stroke, which yielded to a final population of 4072 patients. In total, 37.4% of the patients presented insulin-dependent diabetes. Type of CAD was identified in 60% of the patients. Of which, 833 of the patients (34.21% of the patients with CAD type identified within the EHR) had single coronary vessel disease and 1,602 (65.79%) multivessel coronary disease. Patients were followed up for a median of 33.6 months (IQ1-3: 19.5–47.1).The sociodemographic and comorbidities of the study population at index date are summarized in The most common cardiovascular comorbidities reported in the target population included arterial hypertension (84.65%), and heart valve disease (38.51%) A. In regAs shown in Treatment strategies in our study population at index date included pharmacological management and interventional approaches.Data regarding pharmacological treatment for the management of patients diagnosed with T2DM and CAD are shown in Metformin was identified as the most frequently prescribed antidiabetic drug (77.6%), followed by insulin/analogs (25%), sulfonylureas (21.64%), dipeptidyl peptidase-4 inhibitors (iDPP4) (20.83%), and glinides (12.45%). Other treatments such as statins (79.15%) and antiplatelet therapy, mainly ASA (64.0%), were commonly prescribed according to the available information within the EHRs.Regarding interventional procedures, we observed that a total of 1574 patients (38.8%) were revascularized with percutaneous coronary interventions (PCI) and 585 patients (14.4%) were treated with coronary artery bypass graft (CABG).We evaluated the cumulative incidence of MACE, during the study follow-up.As shown in Cumulative incidence curves of the first MACE occurrence after study entry and detailed probability of individual components of interest over study follow-up are shown in p < 0.001)  .p < 0.001), some hypoglycemic agents including sulfonylureas (p = 0.008) and alpha-glucosidase (p = 0.049), anticoagulant and antiplatelet therapy , beta blockers, calcium channel, nitrates, ranolazines, lipid lowering drugs (p < 0.001 each), diuretics (p = 0.012), and statins (p = 0.003) (p < 0.001).Regarding comorbidities, in general, the presence of all cardiovascular disorders was associated with the occurrence of the first MACE detected during the follow-up. Likewise, diabetic retinopathy, hypothyroidism, and COPD/asthma were found to be associated with the appearance of new MACE in the study . In addi= 0.003) . Revascup < 0.001), and , transient ischemic attack , and heart failure . Pharmacological treatment with insulin and interventional therapies as PCI were also found to be independently associated with the occurrence of new MACE . On the contrary, treatment with statins was found to be inversely associated with MACE during follow-up  T2DM patients who develop CAD are predominantly male with arterial hypertension, hyperlipidemia and obesity as the most prevalent comorbidities; (2) these patients present an increasing probability of developing MACE during the study follow-up; and (3) in this population the risk factors associated with the development of MACE are related with cardiovascular diseases  and T2DM severity (insulin treatment), whilst lipid lowering control strategies (treatment with statins) seems to be a protective factor. The described methodology has been recently used in the same cohort of patients to describe medical management in coronary T2DM patients with previous percutaneous coronary intervention in Spain, where the EHReadis shown .Prior studies have described that around 7.5–15% of the patients with T2DM also had a diagnosis of CAD ,24, and Due to the chronicity of the diseases and the different comorbidities associated, the management of the patients with diabetes and CAD is challenging, and usually requires a multidisciplinary approach. In 2012, the ACCF/AHA/ACP/AATS/PCNA/SCAI/STS published their guideline for the diagnosis and management of stable CAD with a few specific recommendations for those patients with diabetes . Since tDespite disease management, patients with T2DM and CAD are at high risk of experiencing MACE . In ordeIn line with previous publications, we found that insulin treatment was associated with MACE occurrence ,33. AlthThere are several limitations to this study. First, our results are based on data captured directly from the unstructured, free-text narratives in patients’ EHRs. Thus, the findings are limited by the availability and accuracy of EHRs, and by the information reported by physicians in their routine practice. Second, because this is a retrospective study based on real-world data, the findings are only hypothesis generating. Moreover, it was not designed to collect variables in a strict, a priori fashion. Thus, there were some interesting variables that were not properly documented and therefore not analyzed. In this regard, the presence of cardiomyopathy at baseline was not analyzed and patients with an implantable cardioverter defibrillator (ICD) were not considered, both possibly related with MACE development ,35,36,37In conclusion, this study showed that CAD appears in mainly male T2DM patients with other cardiovascular related comorbidities. When it happens, these patients need to be closely monitored because there exists an increasing risk of MACE occurrence. While independent risk factors associated with MACE are difficult to modify since they are intrinsic to the underlying diseases , here we identified statins used as a protective factor that could be used by physicians in clinical practice. Further research is warranted to confirm these findings in future studies.


Chromatin maintenance and remodeling are processes that take place alongside DNA repair, replication, or transcription to ensure the survival and adaptability of a cell. The environment and the needs of the cell dictate how chromatin is remodeled; particularly where and which histones are deposited, thus changing the canonical histone array to regulate chromatin structure and gene expression. Chromatin is highly dynamic, and histone variants and their chaperones play a crucial role in maintaining the epigenetic regulation at different genomic regions. Despite the large number of histone variants reported to date, studies on their roles in physiological processes and pathologies are emerging but continue to be scarce. Here, we present recent advances in the research on histone variants and their chaperones, with a focus on their importance in molecular mechanisms such as replication, transcription, and DNA damage repair. Additionally, we discuss the emerging role they have in transposable element regulation, aging, and chromatin remodeling syndromes. Finally, we describe currently used methods and their limitations in the study of these proteins and highlight the importance of improving the experimental approaches to further understand this epigenetic machinery. Chromatin protects and regulates the DNA; it is also tightly involved in nuclear processes that ensure the proper functionality of the cell. The basic unit of the chromatin is the nucleosome, composed of an octamer of histones  that surround 146 base pairs (bp) of DNA . The rolAlthough during the last 10 years the study of histone variants, particularly of H3 and H2A, and their chaperones has increased considerably, the majority of these studies have focused on a few cellular processes and pathologies, resulting in an imbalanced knowledge and scattered information. Given this, in the present work, we aim to focus on the involvement of histone variants, particularly H3 and H2A, and their chaperones in distinct processes, highlighting their adaptiveness as an important component of replication, transcription, DNA repair and regulation of transposable elements . For morIn order to maintain the structure of the chromatin, histones play a critical role. Two broad types of histones have been described. The first type comprises what are commonly known as canonical histones , and the second class encompasses histone variants . Replication-coupled histones’ genes are encoded in clusters , they are intronless, their messenger RNAs (mRNAs) have a special 3′-loop end and are especially transcribed and deposited during the DNA synthesis (S) phase . Meanwhiin vitro without the need for ATP  complex and the MCM2 (minichromosome maintenance complex component 2). Different studies have shown that the SPT16 (suppressor of Ty 16) subunit of the FACT complex displaces H2A-H2B dimers, while the other subunit, the structure-specific recognition protein 1 (SSRP1), stabilizes the H3-H4 tetramer . The abiThe cell differentiates between transcription states not only by the histone PTMs, but also by differentially depositing histone variants at particular genomic regions . This isHistone variants H3.1 and H3.2 show a high similarity to H3.3; however, one of the unique PTMs of H3.3 is phosphorylation of serine 31 (S31ph), which has previously been linked to processes like mitosis . This unAnother histone variant that has been broadly linked to transcriptional regulation is H2A.Z. This histone was originally linked to transcriptional activation . MoreoveA third variant involved in the transcription of genes in brain and testes is H2A.B. Cells are constantly exposed to DNA damage insults, and in order to counteract them, they have developed different DNA damage response (DDR) pathways. Importantly, distinct histone variants and their chaperones are implicated in some of these pathways . Once thIt was previously reported that H3.3 can be deposited by HIRA at UVC-damaged chromatin , howeverAnother H3 histone variant, whose role in DNA repair has not been clearly established, is CENPA . RecentlThe role of histone variants and chaperones in replication, transcription, and DNA damage repair are the most common studied molecular mechanisms. However, recent findings about their importance in genomic stability and chromatin structure related to transposable elements is starting to unravel, and we discuss the findings regarding this topic in the next sections.Mobile genetic elements (MGEs) are any type of genetic material that can move within a genome or between cells . The setTransposons contribute to the variation, adaptation, and evolution of genomes through the duplication or deletion of genes or their regulatory elements . TEs canThe CAF-1 complex is an H3-H4 chaperone conserved among all eukaryotes. It has important roles during DNA replication and repair, regulation of gene expression, and maintenance of chromatin accessibility . The CAFvia H3K9me3 and H4K20me3 marks. Specifically, CHAF1A reinforces transcriptional repression through its interaction with members of the NuRD complex  and ESET, while SUMO2 orchestrates the provirus repressive function of the canonical zinc finger (ZNF) protein 809 (ZFP809)–TRIM28–ESET machinery by sumoylation of TRIM28.The CAF-1 complex has several interactors. Another important histone chaperone is the FACT complex, which mediates the deposition of H2A/H2B histones and binds to H3-H4 dimers simultaneously . This coAltogether, these results supported the mechanism that the SSRP1 subunit of the FACT complex recruits USP7 to repress MERVL and MERVL-fused genes in ESCs by impeding the ubiquitination of H2Bub. However, it is possible that other FACT-associated proteins may contribute to the repression of MERVL and MERVL-fused genes or that other pathways repressing cryptic transcription initiation may be regulated by the FACT complex itself .As we addressed previously, the histone variant H3.3 can be incorporated at distinct regions of the chromatin by either the HIRA or ATRX–DAXX histone chaperone complexes . While HIn this context, two models have been proposed for the mechanism by which DAXX and its interactors repress retrotransposons. Both models rely on the fact that ERVs are silenced through H3K9me3 by ESET  and TRIMOn the one side, a study by On the contrary, a structural and biochemical study by The set of elements of the DAXX-mediated transposon regulation mechanism is continuously being elucidated. Recently, The specific activity of HIRA lies in its ability to recruit and form complexes with histone-modifying proteins such as histone acetyltransferases and HDACs and to help regulate specific histone variant depositions . In geneFor the proper functioning of cellular processes, fine-tuning of the chromatin must be assured through the previously described mechanisms and genomic components that involve a balance of histones and their chaperones. Nevertheless, in health and disease the balance and function of the histones and chaperones changes, as we discuss in the following sections. Aging is a natural process, where recently the role of these epigenetic components is being elucidated. On the other hand, diseases, such as cancer, alterations, and mutations in both histones and their chaperones have been described. We will not review the information on cancer, but we invite the reader to refer to the following literature on the topic: Aging is a life-lasting event that results from the accumulation of damage and different molecular and cellular alterations through time, leading to a deterioration of physiological functions that are necessary for survival. Epigenetic alterations are a major contributor to cellular senescence, including the exchange of replication coupled histones for variants, and their concomitant accumulation. Given this, it is important to address the latest findings about the role of histone variants in cellular senescence and how they can contribute to altered gene expression and overall aging in proliferative vs. non-proliferative cells .Cellular senescence can be defined as a permanent or highly stable state of cell cycle arrest . In prolAs mentioned, senescence involves changes at molecular and cellular level, which can be related to organelle function, gene expression, epigenetic regulation, disruption in the energy metabolism, etc. Given this, several biomarkers have been established to characterize the senescent phenotype. A senescence-associated secretory phenotype (SASP) of several proinflammatory molecules, like cytokines and some other molecules, such as growth factors and proteases, is seen . BesidesIt is interesting to highlight that there are numerous epigenetic alterations seen as important elements of the senescent phenotype. Yet, only a few histone variants, such as macroH2A and γH2AX, have been studied, while others, like H3.3, have started to become widely explored, particularly because of their role in chromatin regulation and gene expression.In an adult organism, proliferative cells are expected to accumulate H3.3 in their chromatin due to the replication-independent kinetic deposition of the variant compared to the canonical H3.1/H3.2 histones. Some authors hypothesize that the accumulation of H3.3 in certain regions can be a strategy of the cell to follow a semiconservative replication and to “hold/mark” the sites where H3.1/H3.2 need to be deposited once the cell replicates; since eventually these cells stop replicating, H3.3 stays in those regions substituting the canonical histone . This H3The PTMs in canonical or histone variants add another level of complexity to the mechanisms of H3.3 accumulation through the aging process. Enzymes in charge of establishing PTMs selectively identify protein sequences, thus affecting the outcome: the functional readout. In this sense, the ZMYND11 protein specifically recognizes H3.3K36me3 in the body of genes where it regulates the elongation rate of the RNA pol II . AdditioChromatin reorganization is an important feature of senescent cells. When seen under a microscope with 4′-6-diamino-2-phenylindole (DAPI) staining, senescent cells show punctate DNA foci instead of the characteristic heterogeneous distribution of the staining that proliferating cells have. These are the SAHF, and they are enriched with macroH2A. The deposition of this variant is dependent on the HUCA histone chaperone complex activity , the samIL1B, CCL2, and CXCL5, as seen in aged tissues like the liver, kidney, and brain . This PTM is an important regulator of the HIRA complex: when absent, H3.3 deposition is reduced . InteresMost of the works and reviews show evidence of how epigenetic alterations are related to senescence through the perspective of cellular replication. Meaning, most of them focus on available reports on proliferative cells, such as fibroblasts . NeverthDuring the neurogenic process in embryogenesis, there is a differential deposition of histone variants. H2A.1 is enriched twice as much as H2A.2 both in neurons and neuroblasts. Nevertheless, during postnatal development H2A.1 decreases, suggesting that H2A.2 has more stability. On the other hand, another H2A variant, H2A.Z presumably deposited by the INO80 subfamily chromatin remodelers , accumulNanog and Oct4, the H3.3 enriched in the gene bodies is lost  showed that when the expression of pluripotency genes decreases, such as  is lost ; while t is lost . Further is lost . Additio is lost . This plBdnf Exon IV, c-Fos, Dusp6, among others. These genes, which are known as immediate early genes (IEGs) because they are transiently and rapidly activated upon neuronal activation, need a rapid chromatin remodeling, in part mediated by DAXX, which loads H3.3 at their regulatory elements  of human postmortem brains diagnosed with major depressive disorder  have been described . These arsk-2 gene in Coffin-Lowry syndrome, affecting phosphorylation of histone H3  4 syndrome, characterized by immunodeficiency, neurologic defects, and reduced growth  histone H3 (H3K4), leading to reduced histone methylation . Anotherstone H3 . As mentant H3.3 . This sud growth . TherefoHIRA is included. As it was previously mentioned, HIRA chaperone, when forming a four-subunit complex, is in charge of the deposition of H3.3 in many regions of the genome; for example, DNA damage sites and also bivalent genes in ESCs. For hematopoietic stem cells (HSCs) to function properly, epigenetic regulation needs to be carried out, including chromatin remodeling and histone PTMs. Therefore, aberrant epigenetic modifications may lead to impaired HSC development , it was proven that HIRA KO restricts the access to chromatin affecting regions crucial for the transcription of HSC-specific genes. ir genes .The phenotype seen in these patients is very heterogeneous, including heart defects, parathyroid hypoplasia, immune deficiency, and hypocalcemia . Given tATRX is a chromatin remodeler named after the developmental syndrome on which it was found mutated, the a-thalassemia/intellectual disability syndrome, X-linked . ThroughWith a DNA fiber analysis after HU-mediated replication fork stalling, one study evaluated the proportion of restart-competent replication forks between DAXX, FANCD2 and DAXX/FANCD2 double-deficient cells and found out they were similarly reduced. These results suggest that, when replication stress occurs, FANCD2 is ubiquitinated and locates to stalled replication forks to recruit homologous recombination factors and promote a replication fork start, while suppressing new replication origins; cooperating with the ATRX/DAXX complex, most likely including its H3.3 chaperone activity .Drosophila it was found that the clusters of genes of the linker histone H1 and the core histones  could be distinguished by the transcription factors that bind to the respective promoters. While TBP (TATA-box-binding protein)-related factor 2 (TRF2) regulated the H1 gene promoter, the core histones genes were controlled by TBP/TFIID  . This prBP/TFIID . This diin vivo: SNAP-tag System.At the protein level, a collection of methods and techniques have evolved to study the interactions of histones and chaperones. For a detailed revision on this topic, based on H3 but applicable to all histones, please refer to One of the first methods to isolate histones was through acid extraction of whole protein extract . This me7 cells.To study the deposition pattern of histones and their chaperones in chromatin, one of the gold standard methods is ChIP . This teNowadays, there are also several variations of the technique that grants different approaches, like the chromatin immunoprecipitation followed by selective isolation of chromatin-associated proteins (ChIP-SICAP). This ChIP-based technology allows differentiating stable from labile protein interactions and capture the stable ones using DNA biotinylation . Chromatin vivo tracking of newly synthesized proteins via a fluorescent microscope. Based on this method, the “quench-chase-pulse” assay was developed to follow and distinguish between new and old histones. in vivo dynamics, order of deposition, chaperones involved, and the distribution pattern in the nucleus. All of which provide a better understanding of these essential proteins.Finally, another technique that has been widely used to study histones and chaperones is the SNAP-tag system developed by During the last few years, the study of histone variants and their chaperones has increased considerably. However, most of these studies have focused on only a few variants and chaperones, resulting in scarce information. Aside from H3, H2A, and its variants, further studies addressing the rest of the histones are needed to have a bigger picture of how they influence chromatin regulation. Nevertheless, new emerging technologies are improving the experimental approaches for their study. Given this, we highlight the importance of further research about histone variants and their chaperones in the associated syndromes, physiological processes like aging, and their role in molecular processes, such as replication, transcription, DNA damage and their contribution to the regulation of transposable elements. This will further shed light on their involvement as important regulators of the ever-changing chromatin.


Photosystem II is a light-driven water-plastoquinone oxidoreductase present in cyanobacteria, algae and plants. It produces molecular oxygen and protons to drive ATP synthesis, fueling life on Earth. As a multi-subunit membrane-protein-pigment complex, Photosystem II undergoes a dynamic cycle of synthesis, damage, and repair known as the Photosystem II lifecycle, to maintain a high level of photosynthetic activity at the cellular level. Cyanobacteria, oxygenic photosynthetic bacteria, are frequently used as model organisms to study oxygenic photosynthetic processes due to their ease of growth and genetic manipulation. The cyanobacterial PSII structure and function have been well-characterized, but its lifecycle is under active investigation. In this review, advances in studying the lifecycle of Photosystem II in cyanobacteria will be discussed, with a particular emphasis on new structural findings enabled by cryo-electron microscopy. These structural findings complement a rich and growing body of biochemical and molecular biology research into Photosystem II assembly and repair. It is the enzyme responsible for splitting water, providing the oxygen that fuels life on Earth as we recognize it. PSII performs this demanding photochemical reaction through sequential oxidation of a catalytic Mnxidation  before oSynechocystis sp. PCC 6803 (S6803), Thermosynechococcus elongatus, and Thermosynechococcus vulcanus. The mesophilic S6803 has been utilized for physiological studies and studies that require genetic manipulation of photosynthetic proteins, as it is a facultative heterotroph. The ability to grow S6803 on glucose allows for mutations and deletions to be made in genes necessary for photoautotrophic growth. PSII from either T. elongatus or T. vulcanus, thermophilic cyanobacterial strains, has been utilized for all high-resolution X-ray diffraction structures of PSII. Proteins from thermophilic organisms are often used for structural studies, as these proteins are more stable than their mesophilic homologs. Currently, due to these structural studies, our ‘static’ picture of active PSII is quite detailed. Atomic-resolution X-ray diffraction structures are available from T. elongatus and T. vulcanus [4CaO5 cluster up to S3 [Cyanobacteria, oxygenic photosynthetic prokaryotes, have frequently been used as model organisms to study oxygenic photosynthetic processes due to their relatively fast growth and ease of genetic manipulation as compared with plants. For study of Photosystem II, three organisms have been widely utilized: vulcanus ,3,4 and The biogenesis and repair (PSII lifecycle) of this membrane-protein-pigment complex are under active investigation. The assembly of PSII, a 20+ subunit membrane-protein-pigment complex, is an intricate process, involving cellular localization of protein subunits, pigment insertion, subunit assembly and processing, and cofactor assembly. Furthermore, under normal growth conditions, PSII is routinely damaged by reactive oxygen species that form within the protein and in its immediate vicinity ,8. To maIn this review, we will discuss what is known about PSII assembly and repair in cyanobacteria, with an emphasis on new findings since the topic was last reviewed ,15,16,17Active PSII has a modular architecture, consisting of what can be described as four ‘modules’ . These iA and QB and the catalytic Mn4CaO5 water-oxidizing complex (WOC or OEC for oxygen evolving center). The PsbI subunit and cytochrome b559 (made up of the PsbE and PsbF subunits) complete the reaction center. This reaction center complex has been isolated independently and is the smallest PSII sub-complex capable of charge separation [The PSII reaction center (RC) is localized on the D1 and D2 protein subunits. These subunits bind the P680 reaction center chlorophyll special pair as well as the primary electron transfer chain, including plastoquinones Qparation ,20,21. IThe two antenna modules bind chlorophyll to absorb light energy that is funneled into the reaction center. These modules consist of the CP47 and CP43 proteins and their associated low molecular weight subunits . PsbH, PsbL, PsbM, PsbT, PsbX, and PsbY bind to CP47. CP43 is associated with the small transmembrane subunits PsbJ, PsbK, and PsbZ and Psb30 (Ycf12). These low-molecular weight subunits can be individually deleted without a loss of photosynthetic capacity, but stabilize and optimize PSII photochemistry, and some may play a regulatory role in the PSII lifecycle.The last module consists of membrane-extrinsic proteins which bind PSII on the lumenal side of the thylakoid membrane and contribute to PSII stability and optimal performance. This module is the site of greatest variability between and among plants, algae, and cyanobacteria ,23,24,25In addition to the active PSII subunits, there are assembly factors and accessory proteins involved in de novo biogenesis and repair following photodamage that are not part of the active PSII complex. Knowledge of these is almost certainly not complete, and they number as many if not more than the protein subunits of the active complex. These proteins, especially the Psb27 subunit, have been the focus of much recent research effort.CtpA (see 4CaO5 cofactor [psbA (gene coding for D1) transcript, insertion of the nascent peptide into the membrane, and chlorophyll binding to the newly inserted peptide. One such protein, PratA, is involved with the insertion of pD1 to the membrane and coordination of an initial Mn2+ ion to D1 [RubA [Chlamydomonas reinhardtii [PSII assembly is thought to follow a highly ordered series of steps. The first step in PSII assembly is the formation of two pre-complexes, one containing pD1 and PsbI ,27,28, acofactor ,33. The on to D1 ,35,36. Ion to D1 ,37. The D1 [RubA ,31,38 alHliC (ScpB) and HliD (ScpE), associate with D1 co-translationally [Pam68, along with Ycf48 and Ycf39, is implicated in co-translational insertion of chlorophyll to the reaction center proteins D1 and D2 [Knoppova et al. found that Ycf39 and two HLIP family chlorophyll-binding proteins [tionally . The HLItionally . Ycf39, tionally ,42. Pam61 and D2 ,37,43,441 and D2 ,47,48,491 and D2 , providePsbP (CyanoP), which binds to the D2 pre-complex [PsbN as well as the protein products of slr0575 and slr1470 [The reaction center forms A 29,30,,30,50,51-complex , is thou-complex ,53,54. R slr1470 . This coPsb35 also associates with the CP47 pre-complex and was shown to stabilize it and its association with HliA and HliB (ScpC and SpcD) [The CP47 pre-complex contains PsbH, PsbL, PsbT, and possibly PsbM, PsbX, and PsbY ,56. The nd SpcD) . Psb35 ind SpcD) .Psb28 [In the next step of PSII assembly, the RC joins together with the CP47 pre-complex to form the RC47 complex B. RC47 iPsb28 ,62,63,64Psb27 assembly protein, PsbK, Psb30, and possibly PsbZ [Psb34 (Ssl1498), an HLIP-family protein with no pigment binding site, is also suggested to be involved in RC47 binding to CP43 [C. gracilis. However, the sequence of that subunit was unable to be determined. Therefore, we defer to more recent publications that have assigned Psb34 to a PSII subunit with a known sequence and gene, Tsl0063 in T. elongatus or Ssl1498 in S6803, which is in a different position in the PSII structure from the unknown subunit.RC47 binds to the CP43 pre-complex to form Psb27-PSII C. The CPbly PsbZ ,70,72,87bly PsbZ . Psb34  association of the CP43 module and the existence of the RC47 complex, that the CP43 module dissociates, allowing for proteolytic degradation of D1 and re-synthesis and insertion at the RC47 level ,97. It iRecently, a complex that was a combination of the CP47 module and CP43 modules, called NRC (for No Reaction Center) , was isoCryo-electron microscopy (cryo-EM) has brought forth some of the most significant advances in our understanding of PSII assembly and function in recent years. An area of intense research focus and progress has been the elucidation of PSII structures from S6803 and also intermediate PSII complexes. Cryo-EM has provided an excellent means of studying such membrane protein complexes, as the technique does not require protein crystallization and allows for greater heterogeneity within samples. Protein crystals of PSII from thermophilic cyanobacteria have been extensively characterized by X-ray crystallography. However, the structure of PSII from the mesophilic S6803, on which many physiological and biophysical studies have been performed, had been elusive. Gisriel et al. ,91, but T. vulcanus and determined the cryo-EM structure to 3.78 Å. The complex was isolated from a deletion mutant of the PsbV subunit, as increased assembly and repair intermediates are present in that strain. This dimeric Psb27-PSII does not have extrinsic proteins bound and it does not have an assembled Mn4CaO5 cluster. Psb27 binds to the E-loop of CP43, as evidenced previously by mass-spectrometry cross-linking data [4CaO5 cluster to occur by preventing binding of extrinsic subunits [Many intermediate PSII complexes have been isolated and characterized, but until recently, structural details of these complexes and their implications for PSII function were unknown. To better understand the role of Psb27 in the PSII lifecycle, Huang et al.  isolateding data ,72,87. Tsubunits . AdditioThe dimeric Psb27-PSII in  may be tT. elongatus, based on the observation from [2+ or a Ca2+ cation. In addition to the difference in the D1 C-terminus described above, there is a slight discrepancy with the finding in the Huang Psb27-PSII structure in that they do not find a direct steric conflict with PsbO binding. PsbO has been found to be associated with Psb27-PSII in [B binding site and the non-heme iron hydrogen bond network are altered from the active PSII structure, resulting in the absence of the bicarbonate ion and QB. They found that these changes lead to changes in the PSII electron transfer pathway that reduce singlet O2 formation (discussed below). These findings suggest that Psb28 binds to PSII during its assembly to protect PSII from photodamage. Interestingly, Eaton-Rye found that PsbT interacts with the D1 DE loop in active PSII, and PsbT deletion causes increased photodamage [Zabret et al.  purifiedion from  that thi-PSII in , so perhtodamage ,101,102,T. vulcanus. Both the 28-PSII and the RC47 structure have Psb28, Psb34, and an unknown subunit bound that they were not able to model. This additional subunit was not found in the similar PSII-I structure in [Xiao et al. also solved the structure of a Psb28 bound-PSII, as well as a Psb28-bound RC47 , to 3.14cture in , but maycture in ,103, whicture in , althougB binding pocket and bicarbonate and heme iron binding site in the Psb28-bound PSII. These structures are consistent with the position of Psb28 that was identified by [A/QA−•, as proposed by Brinkert et al. [Em of QA/QA−• to a more positive value. This positive shift is predicted to increase the potential between P680+•/QA−• and P680+•/Pheo−•, which may favor direct recombination between P680+•/QA−• decrease back-reactions through P680+•/Pheo−• recombination, reducing the probability of the formation of an RC triplet state [2, consistent with the findings in Zabret et al. Additionally, photoactivation is also thought to decrease the potential of QA/QA−• [Xiao  and Zabrified by , and thet et al. , who shoet state ,106,107. QA/QA−• , so inteHigher levels of Psb27-PSII are also associated with higher NPQ . This fi4CaO5 cluster takes place through sequential binding and oxidation of Mn2+ ions, as well as the binding of Ca2+ and the deprotonation of water, using the same photochemical machinery for charge separation as active PSII. Photoactivation must take place during both de novo PSII assembly and repair, when the D1 subunit is replaced. This phenomenon has long been known and studied [Assembly of the Mn studied ,113,114,4CaO5 cluster) from T. elongatus using X-ray crystallography and found that it aligned closely to the active PSII structure, but without electron density corresponding to the OEC. The authors in this study intentionally removed the Mn4CaO5 cluster by treating crystals with hydroxylamine and EDTA, but the extrinsic PSII subunits were maintained. They propose that the lack of structural alteration of the OEC ligands in its absence means that the protein scaffold is already ‘pre-organized’ prior to Mn4CaO5 cluster assembly, and significant rearrangement does not need to take place for it to form. However, in another study, Gisriel et al. solved a structure of apo-PSII from S6803 [4CaO5 cluster during sample preparation, unlike the Psb28-and Psb27-PSII structures described above, which are most likely assembly complexes. However, the study does provide insight into photoactivation, as the S6803 apo-PSII lacks the OEC and extrinsic PSII subunits and CP43 is shifted, as in [In one such structural study, Zhang et al.  solved tom S6803  using crd, as in ,62,74. T2+ oxidation [Computational modeling can complement structural studies and provide insight into structural perturbations and dynamics. Narzi et al. used a molecular dynamics simulation to find that structural rearrangements do need to occur after the first Mnxidation  for the xidation ,74,116, xidation ,114,118.2+ and is known to have a much lower quantum efficiency than photochemistry in active PSII leading to O2 evolution. Low quantum efficiency of photoactivation has been explained by the need for rearrangement of the ligand shell after the first Mn2+ oxidation, but also the need for acceptor side rearrangement to downshift QA/QA−• potential [- is necessary for photoactivation as well. Vinyard and coworkers found, by tracking photoactivation using EPR, that it tunes the apK of residues around the OEC site, allowing for the necessary deprotonation [2+/Mn2+, the quantum efficiency of photoactivation also increased in OE27, implicating a role for Psb27 beyond just blocking binding of extrinsic proteins as previously proposed [The conditions necessary for optimal photoassembly, or photoactivation, of the manganese cluster have been the focus of recent work ,119,120.otential . Cl- is proposed . The autproposed ,62. How CRISPR and its associated techniques have emerged as powerful tools to manipulate genes and gene expression in cyanobacteria ,124,125.S. elongatus UTEX 2973 as part of a CRISPRi system to target Photosystem I for inhibition [psbD gene) protein, and it was fully reversible. These inhibition strategies introduce a new paradigm for the study of membrane protein complexes, in particular PSII, whereby its assembly can be reversibly halted at varying stages, depending on the gene targeted for inhibition.Of recent interest in terms of Photosystem II research is CRISPR-interference, whereby a DNase-dead CRISPR-associated protein is targeted to a specific genomic locus (or loci) to block transcription. CRISPRi has emerged as a strategy to selectively inhibit photosynthetic genes. CRISPRi enables a particularly intriguing strategy for manipulation of photosynthetic proteins due to its reversibility and its potential to multiplex targets through incorporation of multiple guide RNAs ,127. In hibition . Additiohibition  to inhibhibition  introducA common point that has been further emphasized in recent studies is that subtle perturbations in PSII structure by either assembly factors or low-molecular-weight peripheral subunits have a significant impact on photochemistry. These perturbations can lead to overall efficiencies on a cellular level by protecting assembly intermediates from photodamage when necessary and optimizing photochemistry in active PSII. Along that theme, the impact of detergent solubilization on our current understanding of PSII function is not well described. A few alternate solubilization techniques that may preserve a more native membrane lipid arrangement, including lipid/protein nanodiscs ,132,133 A more complete structural and functional picture of the lifecycle of PSII is coming into focus, but there are most likely PSII assembly and repair factors that have not yet been identified. Open questions remain about several aspects of the lifecycle. These aspects include the mechanism of cofactor insertion into nascent PSII subunits and what signals that PSII is damaged and should be repaired ,137. Add2O2) to create a BP radical that covalently attaches to proteins, fused to a known thylakoid lumen protein, PsbU, to selectively biotinylate proteins localized in the thylakoid lumen of Synechococcus sp. PCC 7002. Approaches such as this have great potential for answering questions of cellular localization and reinforcing findings by other methods.While knowledge about structures of isolated PSII intermediates has been expanded in recent studies, an open area of investigation is the cellular localization of PSII biogenesis and repair in cyanobacteria. The topic of localization of PSII biogenesis and repair is related to the question of the structure of the cyanobacterial membrane system. Liberton et al.  found thThermosynechococcus elongatus, Synechococcus sp. PCC 7002, and Synechocystis sp. PCC 6803 suggest that there are defined regions of the thylakoids that are differentiated by the arrangement of photosynthetic and respiratory complexes in cyanobacteria. This finding is corroborated by Casella et al. [Synechococcus elongatus PCC 7942. The purpose of these differentiated membrane regions is yet to be determined, but the arrangement certainly has an impact on the diffusion of photosynthetic reactants and intermediates, such as quinones, as well as the spatial organization of PSII biogenesis and repair. However, it should also be noted that thylakoid arrangement varies drastically among cyanobacterial species [Atomic force microscopy and hyperspectral confocal  studies a et al.  in Synec species , and so  species . New tec


RMI2) is an essential component of the BLM-TopoIIIa-RMI1-RMI2 (BTR) complex. However, the mysterious veil of the potential immunological relationship of RMI2 in tumorigenesis and development has not been revealed.Background: RecQ mediated genome instability 2 , microsatellite instability (MSI), tumor microenvironment (TME), tumor immune-infiltrated cells (TILs), immune checkpoints (ICP), mismatch repairs (MMRs) -related genes, m6A-related genes, DNA methylation-related genes. Finally, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional networks were also performed for annotation via gene set enrichment analysis (GSEA).Methods: We conducted the differential expression (DE) analysis of the RMI2 expressed remarkably high in most cancer types compared to cancer adjacent normal tissues (P < 0.05). High expression of RMI2 was linked to unfavorable prognosis and advanced stage of disease, especially in LIHC and PAAD. RMI2 expression was related to TMB in 16 cancer types and MSI in 8 cancer types. Furthermore, it is significant positive correlations between RMI2 and stromal and immune cells, ICP-related genes, MMRs-related genes, m6A-related genes, and DNA methylation-related genes. Finally, GSEA analysis revealed that RMI2 was engaged in a variety of signaling pathways in pan-cancers.Results: The RMI2 may serve as a potential biological target and probably assume a crucial part in tumorigenesis and progression.Conclusions:  RMI2 is a part, maintains genome stability and has a significant impact on DNA replication and its damage repairs [RMI2 works to synergizes with RMI1 and topoisomerase III alpha to maintain replication fork stability. At the same time, it can also dissolve double Holliday junctions to prevent genome instability. It has been reported that the downshift of RMI2 can cause mild Bloom syndrome disease features [The BTR complex, of which  repairs . RMI2 woRMI2 and cancers. RMI2 can affect cell function by up-regulating runt-related transcription factor 2(RUNX2). In parallel, it can also increase downstream of RUNX2 downstream molecule SLUG by regulating EMT, thereby regulating the invasion and migration of lung cancer [RMI2 served a positive impact on the growth of hepatocellular carcinoma (HCC) and inhibits its apoptosis by regulating the p53 signaling pathway [RMI2 will result in a significant down-regulation of p53, p21, PUMA, and Gadd45. Liu et al. found that the high expression of RMI2 affects the tumorigenesis of cervical cancer in a variety of ways [RMI2 has implications on DNA replication, repair, and cell metabolism. These studies suggest that the expression level of RMI2 is closely associated with tumor appreciation, invasion, and migration.In recent years, numerous studies have emerged to reveal the relationship between g cancer . It was  pathway . OverexpRMI2 and immunology in cancer is still vague. Further, exploring the immune-related relationship of RMI2 and tumors is urgently required. In this research, comprehensive, analyzed the survival relationship of RMI2 expression in multiple cancer types. And also we explored the correlation of RMI2 expression with TMB, MSI, tumor microenvironment, tumor- and immune-related genes, methyltransferases, and MMRs-related genes. Finally, we explored the possible functions and pathways of RMI2 via GSEA.In recent years, TME and immune infiltration are the focus of tumor research, which is extremely important for understanding the oncogenesis and developing corresponding immunotherapy. However, the relationship between RMI2 mRNA is significantly differentially expressed in normal and cancerous tissues in pan-cancerous. As shown in RMI2 was correlated with 10 cancers  and significantly lower expression only in leukemia. Next, the TIMER was utilized to validate the expression of the RMI2 gene in 33 cancers. Not surprisingly, RMI2 was significantly highly expressed in 17 cancer types  compared to normal tissues and was only low expressed in one cancer type, SKCM . Since some tumor types of the above figure did not have corresponding normal samples, we further used the GEPIA database to match TCGA tumor and GTEx normal data, in order to ensure the comprehensiveness of the study. The results of GEPIA analysis indicate that RMI2 was also highly expressed in five tumors   . In combRMI2 expression and OS by analyzing gene expression transcripts and clinical data. KM survival analysis results revealed that high RMI2 expression was related to low OS time in 7 cancers, including ACC, GBM, KIRP, LGG, LIHC, MESO, and PAAD. And only in CESC, RMI2 low expression was associated with low OS time , and low risk of survival in 4 cancers   .RMI2 expression and DSS in TCGA 33 cancers. KM analysis revealed that high expression of RMI2 was associated with low OS time in 7 cancers, including ACC, GBM, KIRC, LGG, LIHC, MESO, PAAD. Only in CESC and UVM, the high expression of RMI2 was related to the prolongation of DSS time  increasing the DSS of patients  (P<0.05). The expression of RMI2 increased with rising tumor grades in most tumor types.Overall, Usually, patients with late clinical-stage of tumor have poorer survival prognosis We analyzed the relevance between the expression of RMI2 expression and TMB. The detailed information is shown in RMI2 was significantly correlated with 16 cancer types, of which 14 cancer types  were positively correlated, but negatively correlated with THYM and CESC  with a strong correlation with m6A related genes , and the late clinical stage is related to the high expression level. Meanwhile, the expression of RMI2 has closely associated with TMB, MSI, MMR, and DNA methylation. Furthermore, this study demonstrates that the expression of RMI2 in many tumors  was negatively correlated with the immune score and stromal score. The expression level of RMI2 is related to a variety of immune infiltrating cells. To sum up, RMI2 may become a biomarker and provide some new ideas about tumor immunotherapy of cancer.In the current research, we comprehensively demonstrated, the expression level of RMI2 is related to DNA methylation in multiple types of cancer, among which BRCA, STAD, UCEC is associated with four DNA methylation genes . N6-methyladenosine (m6A) is widely involved in the internal modification of RNA in eukaryotic cells and plays a crucial role in RNA metabolism and a variety of biological processes. In acute myeloid leukemia (AML), METTL14 regulates MYB and MYC via M6A modification, and plays a carcinogenic role in regulating cell self-renewal and inhibiting bone marrow differentiation [RMI2 expression and the genes related to m6A. Our study found that the RMI2 gene is associated with multiple M6A genes in BLCA, ESCA, HNSC, LIHC, LUSC, OV, SARC, UCEC, which is helpful for the development of drugs aimed at M6A related genes. However, further experiments are needed to study the relationship between RMI2 expression and M6A. Suppressing ICPs related to immune escape is one of the methods of immunotherapy [RMI2 is related to a variety of ICP genes. Especially in cancer types such as HNSC, KIRC, LIHC, PRAD, and THCA.According to previous studies , 18, DNAntiation . Studiesntiation . Howeverotherapy . In receotherapy , 24. Somotherapy . In addiRMI2 was mainly negatively related to the scores of the immune score and stromal score of TME, such as ACC, GBM, KIRP, LUSC, UCEC, and other cancer types. This shows that the higher the expression of RMI2, the fewer immune cells and stromal cells in TME. RMI2 may inhibit or promote the progression of cancer by aggregating and regulating immune infiltrating cells. In breast cancer [RMI2 and immune infiltration-related cells can promote tumor immunity and drug development and therapy. Our current study found that RMI2 expression was significantly associated with TILs in BRCA, HNSC, LUAD, THYM, such as B cells, CD4+ T cells, CD8+ T cells, T cells follicular helper, T cells regulatory (Tregs), NK cells, Macrophages, Dendritic cells, Mast cells, and other immune cells. Some drugs based on TME and immune infiltration have been approved for the treatment of tumors. Therefore, according to the immune infiltration of different tumors, the corresponding immunotherapy can be developed, and anti-tumor therapy is very important.The cellular components, immune cells, and stromal cells of TME can affect the growth and differentiation of tumor cells . Numerout cancer , gastrict cancer , high mat cancer . CD4 TreRMI2 was related to TME, immune infiltration, and ICPs of pan-cancer, but there are no in vivo or in vitro experiments directly proved that RMI2 affects the survival of tumor patients through these immunological mechanisms. Secondly, we have carried out the integration and evaluation analysis of several databases to obtain the integrity of the results, but the gene chip and sequencing data of different databases may be different, and the way the data are collected may be biased. So it may cause some bias of the research results to some extent. Finally, the exact mechanism by which RMI2 occurs to tumor-associated immunity is still unclear. In the future, study the tumor immune mechanism for specific tumor types in order to explore the relevant mechanism is necessary, and it is helpful to the research and development of anti-tumor drugs. Generally speaking, our study on the relationship between RMI2 and pan-cancer prognosis and immunology provides a new idea for the immune-related treatment of cancer and may be able to do our part to alleviate the physical and mental pain of tumor patients and the national public health financial burden. In the future, more attention may be paid to the study of the immunological mechanism of RMI2 in cancer, and the research on tumor immunity may benefit from this.However, some limitations still exist in this study. This study found that the expression of RMI2 may predict the prognosis and treatment of cancer as a potential biomarker. The high expression lever of RMI2 is related to many tumor types, and the poor prognosis and disease progression of tumors are related to its expression, especially in LIHC, PAAD. The abnormal expression of RMI2 is negatively correlated with immune cells and stromal cells in most types of cancer, and it is closely related to B cells, CD4 T cells, CD8 T cells, T cells follicular helper and other TILs and a variety of immune-related genes . And the enrichment of multiple pathways is also related to it.Returning to the previous question posed in the Introduction, it is now possible to state that http://www.oncomine.org/resource/login.html), which integrates RNA and DNA-seq data profiles from sources TCGA, GEO, and other data that has been made public. To analyze the differential expression of the RMI2 gene in pan-cancer, fold change ≥1.5 and P-value ≤0.05 were set in the Oncomine database. RNA transcript data, mutation data, and clinical data for 33 tumors in the TCGA database were obtained from the University of California Santa Cruz (UCSC) Xena browser (https://xena.ucsc.edu/). The R software package (R version: 4.1.1) was used for data screening and processing. And the data onto the analysis about RMI2 in the TCGA database for different patient samples were listed in http://timer.comp-genomics.org/) mainly provides a presentation of the results of TCGA data analysis, and it can be used to validate the results of differential expression analysis. For cancer types that failed to be matched to normal samples of the TCGA database, we again used the GEPIA (http://gepia.cancer-pku.cn/) database Match TCGA tumor and GTEx normal to facilitate differential expression analysis.Oncomine is a database for oncology (RMI2 (p<0.05). Cox regression was performed to compare RMI2 gene expression with OS, DSS, DFI, and PFI. These items include Hazard ratio (HR), P-values from the log-rank, and 95% confidence intervals set via the maximum selected log-rank statistic, which is based on the grouping of RMI2 gene expression levels. The “forestplot” and “survival” R packages were adopted for Cox analysis and plotting.KM Survival analysis was utilized in evaluating the survival rate of the two groups, which has divided the patients into the high-risk group and low-risk group according to the median expression value of We performed the processing of somatic mutation data from the TCGA by Perl (Perl version: 5.32.1) to count the TMB scores of 33 tumor types and each patient sample.RMI2 expression and TMB and MSI. The “fmsb” R package was used to produce a visual analysis of the radar plot.Spearman rank-sum test was performed to evaluate the relevance between RMI2. The same data processing scheme was used to analyze whether there was a specific association between RMI2 and these two items (DNA methyltransferase as well as m6A-related gene expression levels) in the TCGA. The Spearman correlation method was chosen to assess the relationship among RMI2 and these three indicators . The “ggplot2”, “ggpubr”, and “ggExtra” R packages were used for statistical analysis and to graph heat maps.The expression levels of DNMT1, DNMT3A, DNMT3B, and DNMT3 genes related to methylation have been obtained into the TCGA database. Moreover, previous studies have shown that mismatch repairs(MMRs) have a tight relationship with cancer development. The “limma” R package was used to evaluate the link between the expression levels of five MMRs genes  and RMI2 gene. At the same time, its relationship with immune checkpoint-related genes was investigated. The “estimate” R package was chosen to determine the StromalScore and ImmuneScore for each of the 33 types of cancer in the TCGA. The “ e1071” R package was also used to analyze the relationship between various immune-related cell infiltrations in 33 cancers. Spearman’s correlation method was chosen to assess the correlation analysis of RMI2 gene expression with two immune microenvironmental indicators . The method was also chosen to identify the correlation between various types of TILs and checkpoint genes.In recent years, an expanding series of studies have found that tumor immunity is deeply involved in the occurrence and advancement of tumors. We analyzed the two tumor immunity projects, Immune Microenvironment and Immune Cell Infiltration, which are related to the The three R packages “enrichplot”, “org.Hs.eg.db”, and “clusterProfiler” were used for GO and KEGG enrichment analysis and visualization.RMI2 and sample data in pan-cancer. R software (4.1.1) is used for statistical analysis and graphing. Kaplan-Meier method and Cox Regression method were utilized to analyze the relationship between the expression of RMI2 and patient survival and risk factors. Spearman rank-sum test was utilized to assess the correlation between RMI2 expression and TMB and MSI. Spearman correlation analysis was utilized to assess the relationship between RMI2 expression and Methyltransferase, MMR, and immune checkpoint-related genes. We accept such result that R>0.20 is regarded as a positive correlation and P<0.05 is regarded as statistically significant.The Perl (5.32.1) script was used to screen the expression of Our data is collected from public databases and from the following websites:http://genome-asia.ucsc.edu/index.html; http://www.oncomine.org/resource/login.html; http://timer.cistrome.org/; http://gepia2.cancer-pku.cn/#index.Ethics approval and written consent were not needed for this study exploring the public data. The ethics committee of Fuyang Hospital affiliated to Anhui Medical University agreed to the ethical waiver.Supplementary FiguresSupplementary Table 1Supplementary Tables 2-5Supplementary Table 6Supplementary Table 7Supplementary Table 8Supplementary Table 9Supplementary Table 10Supplementary Table 11Supplementary Table 12


Background: Various studies support the inverse correlation between solar exposure and Coronavirus SARS-CoV-2 infection. In Spain, from the Canary Islands to the northern part of the country, the global incidence of COVID-19 is different depending on latitude, which could be related to different meteorological conditions such as temperature, humidity, and ultraviolet index (UVI). The objective of the present work was to analyze the association between UVI, other relevant environmental factors such as temperature and humidity, and the incidence, severity, and mortality of COVID-19 at different latitudes in Spain. Methods: An observational prospective study was conducted, recording the numbers of new cases, hospitalizations, patients in critical units, mortality rates, and annual variations related to UVI, temperature, and humidity in five different provinces of Spain from January 2020 to February 2021. Results: Statistically significant inverse correlations (Spearman coefficients) were observed between UVI, temperature, annual changes, and the incidence of COVID-19 cases at almost all latitudes. Conclusion: Higher ultraviolet radiation levels and mean temperatures could contribute to reducing COVID-19 incidence, hospitalizations, and mortality. The SARS-CoV-2 coronavirus pandemic is believed to have begun in December 2019 in Wuhan, China .The virus is transmitted mainly by exposure to respiratory fluids from an infected person. The three main methods of exposure are: (1) inhalation of fine droplets and aerosol particles, (2) respiratory droplets and particles making contact with exposed mucous membranes, including by direct splashes or sprays, and (3) mucous membranes coming in contact with hands previously soiled by exhaled respiratory fluids containing the virus, or from touching inanimate surfaces contaminated with the virus ,5. Gener4 times higher than UVC, although data confirm that summer UV exposure could provide sufficient UVB and UVA doses for virus inactivation in less than 2 min outdoors.Environmental factors, especially solar ultraviolet (UV) radiation, have been reported to play a determinant role in outdoor infection patterns. Around 99% of solar radiation reaching earth’s surface ranges between 0.2 and 3.0 µm, from UV to infrared radiation. UV radiation varies throughout the year, showing greater levels during spring and summer, and lower in autumn and winter . UV radiThe ultraviolet index (UVI) measures UV radiation that potentially causes skin erythema, such as is found during the daytime and at the earth’s surface . BecauseTemperature seems also to be important for SARS-CoV-2, which has been reported to be highly stable at 4 °C but sensitive to heat . SeveralTaking all of this into consideration, our objective was to analyze the associations between UVI, other relevant environmental factors such as temperature and humidity, and the incidence, severity, and mortality of COVID-19 at different latitudes in Spain.https://cnecovid.isciii.es/ (accessed on 14 March 2021), and no ethical approval was necessary to use the data for this work. Meteorological data in terms of daily maximal UVI, mean humidity, and temperature for the five locations were collected from the National Meteorology Agency in Spain (AEMET). Daily data were collected from CNE and AEMET, and are represented in figures by mean data per week, taking information for the first, second, and third weeks from days 1–7, 8–14, and 15–21, respectively. Data for the fourth week were the mean values from day 22 to the end of each month. For comparison between the different latitudes, mean data per season are also represented, as well as the mean of the total study period , Zaragoza (N 41.6563°), Madrid (N 40.4167°), Málaga (N 36.72016°), and Tenerife (N 28.46824°). In order to compare the different cities, we obtained the incidence rates of the four variables per 100,000 inhabitants, according to the number of cases and the total population of each province: Madrid 6,779,888 inhabitants, Málaga 1,685,929, Tenerife 1,044,887, Zaragoza 972,528, and Guipuzcoa 727,212. The data were collected from the National Epidemiology Centre (CNE) at the Official Carlos III Institute of Health. The Institute provides daily detailed information on the demographic, epidemiological, and clinical characteristics of COVID-19 patients. The data are obtained from individualized information from different Spanish Health Centers and are available to the general public. Data are anonymously sourced and can be used for statistical analysis without further requirements. Data were obtained from y period .As the variables did not follow normality, Spearman correlation testing was applied to analyze the correlations between daily meteorologic variables and incidental cases of the different variables of COVID-19. The interpretation level of the correlation coefficient for the different variables analyzed was divided into 0.1–0.39 as weak correlation, 0.4–0.69 as moderate, 0.7–0.89 as strong, and 0.9–1 as very strong correlation .COVID-19 infection is sometimes diagnosed several days after the patient becomes infected, with a mean incubation period of five days . With thMeteorological data  and weekly changes in the incidence of COVID 19  are represented in p < 0.001).The highest mean value for the incidence of COVID-19 cases per 100,000 inhabitants per day over the whole study period was observed in Madrid , followed by Zaragoza , Guipuzcoa . The rate of incidence in Málaga was almost the half that of Madrid  and the lowest rate was in Tenerife . Differences between Tenerife and the other provinces were all statistically significant (p < 0.05) (p = 0.001), Málaga (p = 0.002), and Tenerife (p = 0.006) were statistically significant. Higher mean values for deaths were seen in Zaragoza  and Madrid , followed by Guipuzcoa , Málaga , and Tenerife . Differences in the mean values for deaths between Málaga and Tenerife on one hand and the northern provinces on the other were statistically significant (p < 0.05).The mean number of daily hospitalizations per 100,000 inhabitants was also lower in Tenerife  compared with the rest of the studied cities, with statistically significant differences when compared with all other provinces ( < 0.05) . The meaA statistically significant inverse correlation was observed between UVI and the numbers of daily incidental cases per 100,000 inhabitants, in all the studied provinces . The greThe association between COVID-19 and mean temperature was considered separately, revealing weaker and more heterogenous correlations than were found for UVI . TemperaCorrelations between humidity and COVID-19 variables were very heterogeneous and most were statistically non-significant . InverseThe present study shows that the incidence of COVID-19 was inversely correlated with seasonal variations of UVI in the five provinces studied. Hospitalization and mortality also were inversely correlated with UVI, but not in all the studied provinces. Correlations between temperature and incidental cases were weaker and less consistent, however, a clear negative correlation between temperature and hospitalization was observed in the warmer provinces (Málaga and Tenerife). Deaths were correlated with mean temperatures in Guipuzcoa, Zaragoza, and Málaga. The role played by humidity remains uncertain, because the correlations found with the different COVID-19 variables were inconsistent.The greatest mean value of daily incidence of COVID-19 cases per 100,000 inhabitants across the study period was found in Madrid , followed by Zaragoza , Guipuzcoa , Málaga , and Tenerife . Differences were statistically significant between Tenerife and the rest of the provinces. This is in agreement with studies that confirm a lineal correlation between latitude and COVID-19 incidence, without a clear association with temperature or humidity . The higThis study’s most relevant and consistent finding is the negative correlation between incidence of COVID-19 cases and the UVI at every studied latitude. These results agree with those published by Páez et al. , which wTang et al. also found a correlation between the average percentage of positive cases of five human coronaviruses  and sunlight UV radiation, but only in the eastern United States and not in the western side . Accordip < 0.01).In a study performed in Spain during the early months of the pandemic, temperature was described as the main meteorological factor involved in COVID-19 transmission, with Spanish regions that had the highest temperatures showing lower incidences of infection . AnotherAccording to our results, it can be stated that the COVID-19 outbreaks that occurred in the spring and at the end of summer support a relationship between UVI, temperature, and incidence of COVID-19. In this context, the end of the spring outbreak was related to UVI over 4 and temperatures over 15 °C, while the summer–autumn outbreak coincided with UVI under 5–6 for all latitudes and temperatures that were again under 20 °C.Relative humidity may potentially affect COVID-19 transmission. Our results show that in northern provinces, relative humidity correlated negatively with COVID-19 incidence five days later; meanwhile, relative humidity showed a positive correlation in the southern provinces of Málaga and Tenerife. In a recent global analysis of different meteorological factors affecting SARS-CoV-2 transmission, humidity did not show a statistically significant association . HoweverAdmission to ICU was apparently not related to any of the studied meteorological factors, in agreement with several studies, the severity of the disease being mainly conditioned by individual patient comorbidities . Fang etAccording to our results, higher mean temperatures and higher levels of ultraviolet radiation correlated with deaths caused by COVID-19. In a retrospective study carried out in Wuhan, temperature was also inversely associated with COVID-19 deaths, whereas diurnal temperature range was positively associated .Environmental data from our work and from other related studies could indicate two distinct effects of seasonal variations on the different COVID-19 outbreaks in 2020 and at the beginning of 2021. First, high UVI and temperature make the air and the earth’s surface comparatively free of the virus on days with UVI over 4–5 and temperatures over 20 °C. Second, spring and summer in Spain, due to their mild temperatures, invite to population to spend more time outside and thus social distancing is increased. According to other studies, the immunomodulatory role of vitamin D, which is clearly influenced by solar UV seasonality, is an evident indirect factor affecting COVID-19 infection rates and severity. From mid-spring to late summer, mean maximal UVI ratings over 8 are to be found in our study locations in Spain, and it is well established that spring and summertime are related to increases in Vitamin D serum levels in humans. Accordingly, autumn and winter, when higher incidences and severity of COVID-19 were manifest, are related to lower outdoor UV levels and lower levels of vitamin D ,8. RecenTaking all these limitations into consideration, meteorological factors, especially UV radiation and less probably temperature, seem to have played an important role in incidence levels of COVID-19 in Spain, similar to observations in other countries around the world.UV radiation seems to be the most relevant meteorological factor affecting the incidence and hospitalization rates of COVID-19 in Spain throughout the pandemic. Higher UV levels can contribute to lower rates of COVID-19 incidence and hospitalizations. Temperature seems to have a lesser effect that differs in warmer places (inverse correlation) and colder places (direct correlation), while the effect of humidity remains unclear. ICU admissions caused by COVID-19 infection seem not to be influenced by meteorological factors. Higher mean temperatures and higher ultraviolet radiation levels could contribute to reducing numbers of deaths caused by COVID-19.


Background. Periodontitis is an inflammatory disease of the tooth-supporting structures. Current data suggest that periodontal disease may be a risk factor for asthma. The present study aimed to assess the prevalence of periodontitis and its relationship with the severity of asthma in asthmatic patients. Methods. This study was conducted on 70 newly diagnosed asthmatic patients as the case group and 70 healthy subjects as the control group, aged 20‒50. The asthma was diagnosed by a pulmonologist according to Global Initiative for Asthma (GINA) guideline. All the participants underwent periodontal examinations, which included measuring the pocket depth (PD), attachment loss (AL), gingival index (GI), and plaque index (PI) in one tooth from each sextant, including the incisor/canine and left and right premolar/molar regions for both the maxillary and mandibular dental arches.Results. Periodontal disease was significantly more prevalent in newly diagnosed asthma patients. Patients with asthma had significantly higher PI, GI, PD, and AL scores (P<0.001). Furthermore, dry mouth in asthmatic patients with cough and mucosal changes in asthmatic patients with wheeze were significantly more common than in non-asthmatic patients (P<0.05). The median AL in wheezing patients and the median AL and PD in participants who had asthma attacks within the previous month were significantly higher than in other patients. Furthermore, there was a significant negative correlation between AL with Forced expiratory volume in one second (FEV1) and forced vital capacity (FVC) and PD with FEV1 and FVC. Conclusion. Our results showed that periodontal diseases were more prevalent in newly diagnosed asthmatic patients, and asthma was more severe in periodontitis patients. Bronchial asthma is a chronic inflammatory disease of the airways that affects people of all ages around the world.8 For example, previous studies reveal that periodontitis as a disease is associated with a higher risk of atherosclerosis, myocardial infarction, respiratory disease, diabetes, and preeclampsia.9 Periodontitis is a chronic inflammation of the periodontium (tooth-supporting tissues) caused by a bacterial infection.10 Bacterial biofilm on the teeth, genetics, environmental factors, and some inappropriate behavioral habits are involved in the development and progression of this disease.11 In addition, bronchodilators and inhaled medicines used by asthmatic patients are other risk factors for oral hygiene.12 Current data suggest that periodontal disease may be a risk factor for asthma, emphasizing the correlation between oral hygiene and general systemic health.13found a possible association between asthma and periodontal disease in adults after comparing periodontal disease indicators such as plaque index (PI), gingival index (GI), calculus index (CAI), pocket depth (PD), attachment loss (AL), and bleeding on probing (BOP) between asthmatics and healthy individuals. However, they suggested further research to determine the interrelationship of these disorders. Although the exact association between asthma and periodontal disease is unclear, it may involve either pathological activation of the immune and inflammatory responses or side effects of anti-asthma medications, which need to be further considered.13 On the other hand, some results are contradictory due to differences in sample size, sociodemographic factors, and diagnostic criteria used to diagnose periodontal disease. In this regard, some studies have found higher microbial plaque deposition, gingivitis, periodontitis, and reduced salivary flow in asthmatic patients compared to healthy individuals. For example, Kelly et al Therefore, this study aimed to investigate periodontal indices in Iranian asthmatic patients and determine the relationship between these indices and disease severity. The novelty of the research is periodontitis in newly asthmatic patients without inhalation of bronchodilator spray, such as inhaled corticosteroids that affect the orodental system. This experimental study was conducted under the Helsinki Declaration to treat human subjects. The study protocol was also approved by the Ethics Committee of Mashhad University of Medical Sciences, Mashhad, Iran, under the code IR.MUMS.DENTISTRY.REC.1399.113. The participants were informed about the research objectives and written informed consent was obtained. Furthermore, participants were guaranteed that their information would be kept confidential, the research protocol would not interfere with their asthma care, and they would not be charged.14 The inclusion criteria were: newly diagnosed with bronchial asthma, an age range of 20‒50, having natural teeth, and the ability to give informed consent. The exclusion criteria for patients were: wearing partial or complete dentures, nonsteroidal anti-inflammatory drugs (NSAIDs) use in the past three months, smoking and opium addiction, history of diabetes mellitus, collagen vascular diseases, rheumatic diseases, epilepsy and seizures, cardiovascular diseases, hypertension, thyroid disease, hyperlipidemia, chronic renal failure, liver cirrhosis, hematologic disorders, malignancy, and endocrine diseases. The control group included healthy volunteers with no history of bronchial asthma, natural teeth, and an age range of 20‒50. At the beginning of the test, the participants’ demographic data were recorded in the sociodemographic characteristic form. This control‒case study was conducted on 70 patients with bronchial asthma, who were admitted to the Lung Diseases Research Center, Mashhad University of Medical Sciences, Mashhad, Iran, between 2020 and 2021. The participants were enrolled by convenience sampling. The sample size was calculated using Epi-info software, based on a 40% prevalence of the periodontal disease among asthmatic patients as indicated by Khassawneh et al.15 The participants were subjected to a spirometry test using a Micro Medical Spirometer  to determine their forced expiratory volume in one second (FEV1). The FEV1 values >80%, 80‒60%, and <60% were considered mild, moderate, and severe, respectively. In addition, forced vital capacity (FVC) and FEV1/FVC were calculated for patients. Asthma was diagnosed by a pulmonologist based on spirometry results according to the Global Infinitive for Asthma (GINA) guidelines. The ACT is a 5-item, self-administered questionnaire used to assess asthma control, independent of pulmonary function measurements. For asthma control, the ACT questionnaire items (14) are rated on a five-point Likert scale from 1  to 5 (never), as well as 1  to 5 (completely controlled). The ACT scores vary from 5 (poorly managed asthma) to 25 (well-controlled asthma), with a score of 19 or above suggesting well-controlled asthma. Furthermore, the incidence of asthma symptoms, including dyspnea, coughing, wheezing, chest tightness, pain, the number of attacks per month, and the amount of medication used, were recorded. Spirometry and ACT questionnaires were performed by one experienced pulmonologist (FR).16 All measurements were made in four areas for each tooth  using a Williams probe  according to the standard method. The probe was placed in the periodontal pocket parallel to the longitudinal axis of the tooth; therefore, it did not pass through the junctional epithelium. Plaque index and gingival index (Silness and Loe)17 were measured, and the means of the recorded values were considered as the plaque index and gingival index and registered for each subject. Furthermore, dry mouth and decreased saliva, changes in the oral mucosa, the presence of oral candidiasis, gingivitis, oral hygiene status, dental caries clinically , and the total number of teeth were examined. Periodontal examinations were carried out by a skilled dentist (AA). They were qualified, calibrated, and approved by a reliable senior professor of periodontology (AM). Patients were subjected to a clinical periodontal examination, including measuring the community periodontal index for treatment needs (CPITN), PD, AL, and GI. A dentist examined one tooth (CPITN index teeth) from each sextant, including the incisor/canine and left and right premolar/molar regions for both the maxillary and mandibular dental arches. CPITN was measured using a standardized protocol to investigate gingival bleeding, calculus, and periodontal pockets. The following ten teeth: upper right: 17 and 16; upper anterior: 11; upper left: 26 and 27; lower right: 47 and 46; lower anterior: 31; and lower left: 36 and 37, were examined, and each tooth was given its highest CPITN score ranging from 0 to 4. Statistical analyses were performed by SPSS 21.0 . Following the Shapiro–Wilk test to determine the normality of the data, intergroup comparisons were performed using the Student’s t-test for parametric data and the Mann–Whitney U test for nonparametric data. The chi-squared test was also used to compare qualitative variables. The quantitative data were expressed as median (interquartile range) and qualitative data as a percentage. A P-value of <0.05 was considered statistically significant. The periodontal indices of 70 asthmatic patients were evaluated in this study to determine the relationship between periodontitis and asthma. The mean ± SD ages of asthmatic patients and healthy controls were 37.67±9 and 38.37±9.7 years, respectively. Furthermore, the asthmatic group consisted of 32 (45.7%) males and 38 (54.3%) females, while the control group included 34 (48.6%) males and 36 (51.4%) females. However, no statistically significant difference in gender distribution was found between asthmatic patients and healthy controls by chi-squared analysis. Furthermore, none of the participants smoked or were drug addicts . The ACT questionnaire was employed to assess asthma severity. According to the results, asthma was well controlled in 2 (2.9%) patients, partially in 28 (40%) patients, and uncontrolled in 40 (57.1%) patients. Three (4.3%) of the patients had also experienced an emergency in the last month. Furthermore, coughing, wheezing, dyspnea, and chest tightness were also reported in 50 (71.4%), 35 (50%), 64 (91%), and 30 (42.9%) patients, respectively. Twenty-eight (40%) patients had a history of nocturnal asthma attacks. In addition, 55 patients had no asthma attacks in the previous month; five had one, one had two, one had three, and eight had four attacks. On the other hand, spirometry values revealed that the mean ± SD of FEV1, FVC, and FEV1/FVC ratio were 2.21±1.30 L/s, 2.9±1.1 L/s 70%±14.88%, respectively. According to the results, FEV1/FVC was abnormal in 48 (86.6%) patients. In contrast, the percentage of people in the asthmatic patients’ group with CPITN scores of one, two, and four was significantly higher (P<0.001). Furthermore, most participants had gingivitis, and the groups did not differ significantly in this regard. In addition, candidiasis was also not observed in any of the participants. It should be noted that asthmatic patients had a significantly higher incidence of dry mouth than healthy subjects (P=0.014), and their oral hygiene was considerably worse . Accordi Furthermore, we investigated the periodontal indices and oral hygiene in asthmatic patients based on gender. The results revealed that males had significantly weaker oral hygiene status than females, with 62.5% having poor oral hygiene compared to 34.2% (P<0.05). Moreover, 21.9% of males had moderate oral hygiene, and 15.6% had good or excellent oral health. These values were 52.6% and 13.2% in female patients, respectively, which was significantly higher than in males (P<0.05). However, there was no statistically significant difference between male and female patients regarding other parameters. Based on asthmatic variables, we evaluated the periodontal indices and oral hygiene in asthmatic patients. The results of a chi-squared analysis showed that dry mouth was significantly more common in asthmatic patients with cough (46%) than in those without cough (15%) (P<0.05). Furthermore, the total number of teeth in asthmatic patients who did not have wheezing was 28, which was significantly higher than the 24 teeth in asthmatic patients with wheezing (P=0.008). In addition, the AL and mucosal changes were significantly higher in asthmatic patients with wheeze (P<0.05). Moreover, in patients who had not experienced an asthma attack in the previous month, AL and PD parameters increased from 2.5 and 1.7 to 2.9 and 1.8 in patients who had an asthma attack in the previous month, which was significant . On the other hand, dyspnea, chest tightness, nocturnal asthma, and asthma control had no significant effect on periodontal parameters. In addition, correlation analysis of respiratory variables and periodontal indices revealed a significant negative correlation between AL and FEV1 and FVC . There w The present study aimed to assess the incidence of periodontitis in asthmatic patients. This research investigated periodontal indicators in asthmatic patients and their relationship with asthma severity. The importance of inflammation and infection as risk factors for bronchial asthma has been considered. Asthma appears to be associated with immune cells and inflammatory mediators like cytokines, leukotrienes, and immunoglobulins. In this regard, periodontal diseases are inflammatory conditions of the oral cavity that cause inflammation of the tissues surrounding the teeth and chronic immune system disturbance. Periodontal diseases aggravate inflammatory and immunological responses by transferring bacteria into the bloodstream, which affects the progression of the disease in patients with bronchial asthma by altering the airway epithelium. The present study found a significant increase in periodontal disease in newly diagnosed asthma patients. Despite having a higher number of decayed teeth and higher CPITN scores in healthy controls (P<0.001), asthmatic patients had significantly higher PI, GI, PD, and AL scores (P<0.001). In addition, dry mouth in asthmatic patients with cough and mucosal changes in asthmatic patients with wheeze was significantly more common than in other patients who did not have these symptoms (P<0.05). The median AL in patients with wheeze, and the median AL and PD in participants who had asthma attacks in the previous month, were significantly higher than in other patients.8 Harshita et al18 reported that periodontal diseases are more common in asthmatic patients compared with healthy people. Furthermore, the oral hygiene index in healthy individuals was better than in asthmatic patients, consistent with the current study. They found that the type of asthma medication used by asthmatics affected the extent of periodontitis rather than the severity of asthma. Their findings also showed that, despite the lack of differences in GI between asthmatics and healthy individuals, the mean PD and AL were significantly higher in asthma patients, which was consistent with our findings.18 It should be emphasized that measures for AL and PD in the current study were taken between 6 teeth in the CPITN index rather than all teeth. Another study by Shen et al19showed that the incidence of periodontal disease was significantly higher in patients with newly diagnosed asthma than in healthy subjects. They suggested that asthmatic patients are more likely to develop periodontal disease.19 Furthermore, Yaghoobi et al20 reported that periodontal parameters in asthmatic patients were weaker than in healthy subjects, providing additional support to previous studies that advocated an association between respiratory disease and periodontal health status.20 In a study on a representative sample of Korean adults, Lee et al21 discovered a relationship between asthma and periodontitis in patients with a recent asthma diagnosis. Patients taking anti-asthmatic drugs regularly were also less likely to be diagnosed with periodontitis. According to their findings, poorer oral hygiene was associated with older age, female gender, less physical activity, lower socioeconomic status, smoking, and alcohol consumption.21 In a similar study, Moeintaghavi et al22found a strong negative correlation between FEV1 and FVC with PI and AL levels in patients with COPD. There was also a significant positive association between the COPD severity score with PI and AL. Similarly, FEV1/FVC was also found to have a negative correlation with PD and AL.22 To explain the difference between Moeintaghavi’s work and the present study, it can be concluded that COPD usually has a chronic course and is associated with older age groups who have a history of smoking. Also, evidence indicated that the prevalence of periodontitis in patients with severe asthma was much greater than in the healthy control group. Recent research has suggested that periodontal involvement in asthma may be due to immunological and inflammatory processes, asthma drug adverse effects, or both. The main cause of periodontal disorders is likely a decrease in the protective impact of saliva during the dry mouth. Mouth breathing, changes in salivary composition, or decreased salivary flow can all contribute to this issue. A dry mouth can diminish the concentration of IgA in saliva and disrupt the balance of bacterial and immunological components in the oral cavity. Higher levels of calcium and phosphate in saliva and increased IgE concentrations in gingival tissues can contribute to periodontal diseases.24 Inhaled corticosteroids used to treat asthma, on the other hand, may lower bone mineral density, including the mandible, and increase the risk of pathological fractures, especially in those using moderate to high doses.26 Alveolar mucosal dehydration, altered immune response with increased IgE and IgA concentrations, decreased salivary flow, increased mass deposition due to increased salivary calcium and phosphorus levels, and reduced jaw bone density as a result of steroid use are all possible causes of periodontal disease in asthma patients.27 Generally, the findings of this study are consistent with previous research on the association between asthma and periodontal disease. Patients with newly diagnosed asthma are far more likely than healthy individuals to develop periodontal diseases and poor dental hygiene. This study also revealed that periodontal disease is more common in individuals with severe asthma. These findings emphasize the significance of regular oral health monitoring in asthmatic patients, specifically those with severe asthma. Due to the impossibility of obtaining dental radiographs for all patients, dental caries was evaluated only clinically . Therefore, we may not be able to accurately evaluate the results of caries prevalence . This may be the reason for the discrepancy between the results of caries prevalence and oral health status in the two groups of asthmatic and healthy subjects. This study was financially supported by the Research Vice-Chancellor, Mashhad University of Medical Sciences . The authors declare no conflict of interest related to the study. AM, AA, and FR designed and carried out the research, collected the data, interviewed the patients, coordinated the study, participated in the research and prepared the manuscript. AA performed the statistical analyses and revised the English manuscript. All authors have read and approved the content of the manuscript. There was no funding support for the study. The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request. .