Triple-negative breast cancers (TNBCs) are very heterogenous, molecularly diverse, and are characterized by a high propensity to relapse or metastasize. Clinically, TNBC remains a diagnosis of exclusion by the lack of hormone receptors (Estrogen Receptor (ER) and Progesterone Receptor (PR)) as well as the absence of overexpression and/or amplification of HER2. DNA methylation plays an important role in breast cancer carcinogenesis and TNBCs have a distinct DNA methylation profile characterized by marked hypomethylation and lower gains of methylations compared to all other subtypes. DNA methylation is regulated by the balance of DNA methylases (DNMTs) and DNA demethylases (TETs). Here, we review the roles of TETs as context-dependent tumor-suppressor genes and/or oncogenes in solid tumors, and we discuss the current understandings of the oncogenic role of TET1 and its therapeutic implications in TNBCs. Triple-negative breast cancer (TNBC) is a heterogenous disease that is genetically complex and is defined by lack of estrogen (ER)- and progesterone (PR)-receptor expression and the absence of overexpression and/or amplification of human epidermal growth factor receptor 2 (HER2). TNBC constitutes 15–20% of all breast cancer subtypes. It is more common in African American women, premenopausal women, and in BRCA1/2 mutation carriers. TNBC is also a very aggressive subtype with high tumor grade and high mutation rate and proliferation index compared to other breast cancer types. TNBC has the worst outcome with a relative 5-year survival rate of 91% in localized disease, 65% in locally advanced disease, and 11% in metastatic disease. Patients with TNBC who do not achieve complete pathologic response with chemotherapy and will typically have tumor recurrence or succumb to metastatic disease in less than 5 years of initial diagnosis. Poor outcome in this subtype is also due to lack of available targeted therapies which are commonly used in ER+, HER2+ subtypes .Twenty years ago, using the microarray technique, four intrinsic breast cancer subtypes were identified . This was one of the first reports on basal-like breast tumors . This suAlthough the four TNBC subclassifications are not yet routinely used in the clinical setting, these advances are important steppingstones to making this heterogenous group of cancer more amenable to new therapeutic insights and potentially to personalized medicine. These discoveries also help shift the paradigm from what TNBC is, rather than to what it is not, and possibly to further sub-stratification. In this review we will present the functional role that epigenetic modifiers such as DNA demethylases play in driving this disease and how we envision their therapeutic potential.Epigenetics refers to heritable molecular determinants of gene expression in the absence of changes in DNA sequences. The molecular mechanisms contributing to epigenetic changes in gene expression are DNA methylation, histone modifications, and non-coding RNA regulation. These mechanisms are critically important during normal development and their aberrations have been associated with diseases such as cancer. Histone modifications are post-translational modifications  of the histone tail residues carried by different enzymes such as histone methyltransferases (HMTs), demethylases (HDMs), acetyltransferases (HATs), and deacetylases (HDACs). These different modifications and the expression levels of the modifying enzymes can influence the open and closed states of the chromatin that lead to changes in gene expression. Many aberrations in these modifications, including both increases or depletions in these marks and their respective modifying enzymes, have been found in breast cancer and are associated with breast cancer initiation, progression, and prognosis ,18,19,203) group to the 5-position of cytosine when followed by guanosine (CpG) in DNA. Methylation is initiated by de novo methyltransferases DNMT3A and DNMT3B. These enzymes are highly expressed during embryogenesis and are crucial for establishing methylation patterns. The methyl mark is then maintained by DNMT1 which is the most abundant DNMT in adult mammalian cells and functions as a maintenance methyltransferase that faithfully copies and propagates the methyl mark during replication [DNA methyltransferases (DNMTs) catalyze the transfer of a methyl /alpha-ketoglutarate-dependent oxygenase family enzymes are involved in the oxidation of the thymine base in DNA to 5-hydroxymethyluracil [Compared to gain of methylation, loss of methylation was only thought of as a passive process due to lack of maintenance during replication. However, in early the 2000s, the discovery of tet-eleven translocation (TET) proteins—DNA demethylases (erasers)—revealed the presence of an active demethylation process in early embryogenesis. TET enzymes were discovered by Rao and colleagues when looking for paralogues of the yluracil . The TETyluracil . These myluracil ,48,49.2 and succinate. Accumulation of succinate and fumarate as well as D-2HG which is generated by mutant isocitrate dehydrogenase (IDH) enzymes, can inhibit the TET enzyme demethylases while vitamin C serves as an essential co-factor. At the amino-terminal region, both TET1 and TET3 contain a CXXC domain that targets the protein to unmethylated CpG islands. TET2 does not contain a CXXC domain but pairs up with IDAX protein, an independent CXXC containing protein [All three TET enzymes harbor a common core catalytic domain comprised of a conserved cysteine-rich domain and a double-stranded beta-helix (DSBH) domain that contains key residues that interact with cofactors and substrates. For the oxidation reaction, TET enzymes require 2OG, Fe (II), and molecular oxygen. The oxidation reaction generates CO protein ,51,52,53 protein ,55,56.Besides differences in genomic distribution of TET enzymes, they could also interact with different partners and could demethylate independently of their catalytic activity. For example, catalytic activity of TET1 is important in regulating the expression of PGC7 which displaces UHRF1 and impedes DNMT1 recruitment, leading to DNA demethylation . In in vDNA methylation array studies revealed that epigenetic alterations in breast cancer methylome are not only common but can also be used to stratify different breast cancer subtypes, correlate with cancer stage, and predict clinical outcomes ,59,60. TThere is confounding evidence about the functional roles of TET enzymes particularly in solid tumors . In numeDLit2, ZNF382, HOXA9, and DKK1) and significantly suppressed clonogenicity of cancer cells compared to the catalytically dead mutant [In multiple cancer cell lines and primary tumors, whole genome CpG methylation analysis showed that TET1, but not TET2 or TET3, was downregulated by promoter methylation compared to the normal controls. It is commonly known that DNA methylation alterations, such as global hypomethylation and localized hypermethylation at promoters of tumor-suppressor genes, is a hallmark of cancer . Therefod mutant ,73. Thesd mutant .TP53 and TIMP, which in turn would lead to suppression of tumor development, invasion, and therefore better overall survival.In a meta-analysis of 3100 patients, the association between TET1 expression and prognosis of patients with breast, colorectal, cholangiocarcinoma, endometrial, lung, ovarian, gastric, renal, and liver cancer was studied. In the pooled analysis, higher TET1 expression in cancer was associated with better overall survival (OS) and in subgroup analysis higher TET1 expression was associated with better OS in respiratory tumors as well as in breast cancer from Asian patients. In this analysis, there were four breast datasets, three of which were Asian breast datasets. However, in the pooled subgroup analysis, the non-Asian breast dataset, which showed the opposite association between TET1 and OS, was excluded and therefore the prognostic role of TET1 was only established in the Asian dataset . TherefoAnother study that supports the tumor-suppressive role of TET was shown through the inhibition of alpha ketoglutarate dehydrogenase (KGDH). Inhibition of this enzyme increases the alpha ketoglutarate levels which is a cofactor of TET enzyme in the oxidation reaction. In this study, alpha ketoglutarate increased TET activity in in vivo model of highly aggressive metastatic breast cancer (4T1) and in cell lines. The increase in TET levels was associated with increase in downstream anti-metastatic micro-RNA (mir-200) family expression. The increase in mir-200 consequently downregulated the epithelial mesenchymal transition and lung metastasis .In stark contrast to its tumor suppressing functions, TETs could also activate multiple downstream oncogenic signaling pathways by demethylating the epigenome. TET1 expression was demonstrated to promote cell metastasis in colorectal cancer and activation of PI3K oncogenic signaling in TNBC. TET1 expression was shown to correlate with cell migration, cancer stemness tumorigenicity, and poor survivals in epithelial ovarian cancer and in TNBCs ,78,79.FLI1 and recruits TET1 to demethylate the promoter and increase its expression. Increased FECR1 enhanced invasiveness of the TNBC cell line and it was present in advanced and metastatic breast cancers. Interestingly, FECR1 downregulated DNMT1 which may cause activation of many pro-tumorigenic factors [The oncogenic functional role of TET1 was also described through different interacting partners such as the circular RNA-FECR. FECR1 binds to the promoter of its gene  factors .+) cell proliferation but did not affect the proliferation of the ER-negative cell line [TET2 gene only in ER-positive cells. Moreover, the MLL3 and TET2 correlation was found to be greater and more significant in breast cancer patient samples with ER-positive than in ER-negative tumors [In another study, knock-out of TET2 decreased MCF7  experiments in 2 TNBC cell lines (MDA-MB-231 and Hs578T) using CRISPR/cas9 approach, support the oncogenic potential of this enzyme [TET1 KO cell lines were studied and loss of TET1 resulted in reduced cell migration and proliferation. Transcriptomic analysis of these KO cell lines, revealed that TET1-mediated hypomethylation of oncogenic pathways such as the PI3K pathway were downregulated. These data support TET1/PI3K/mTOR as an important therapeutic target in a subset of TNBC patients with high TET1 levels. Besides, by targeting the oncogenic signaling pathway downstream of TET1, there are potentially additional therapeutic interventions. This subpopulation of TNBCs also had suppressed immune system pathway genes and upon loss of TET1 by CRISPR/cas9, these genes were upregulated. It remains to be determined whether those TNBC patients with high TET1 levels, activated PI3K signaling pathway, and low immune response pathway genes could benefit from treatments targeting TET1 and PI3K immune activators alone or in combination. In the clinic, it is well-known that the basal-like immune-suppressed (BLIS) subgroup of TNBC patients have the worst treatment response and prognosis [Furthermore, s enzyme . Single rognosis . Whetherrognosis . Though In another study that supports the oncogenic role of TET1, it was shown that TET1-mediated hypomethylation upregulated gene expressions that drove self-renewal and expansion of cancer stem cells in TNBC. This TET1-mediated oncogenic function is an alternate mechanism that was shown to be due to the downregulation of the catalase enzyme which increases the hydrogen peroxide levels, or be due to exogenous routes such as systemic inflammation and oxidative stress within the context of obesity in TNBC .ALT) that lacks the CXXC domain but retains its catalytic activity. In fact, the canonical TET1 protein (TET1FL) is the only isoform expressed in embryonic stem cells, while the TET1ALT isoform is predominantly expressed in adult cells and is overexpressed in cancers such as breast, uterine, ovarian, AML and glioblastoma. Furthermore, it was also shown that full-length TET1 (TET1e) and the short isoform (TET1s) were differentially expressed in mouse embryonic stem cells and somatic cells, respectively [TET1 contains a CXXC domain that targets it to unmethylated CGIs. Previously, it has been shown that overexpression of TET1 in HEK293T cells does not induce demethylation in most CpG sites surveyed instead it was suggested to function as a guardian protecting the unmethylated CpG islands from methylation ,84. Howeectively .FL is a large protein (2136 amino acids) that is transcribed from a typical CGI-containing promoter [ALT) that lacks the CXXC domain (exon 2). Indeed, when this alternate region was cloned into the luciferase vector, promoter activity was detected. Furthermore, analysis of the raw RNAseq data for breast cancer from the TCGA database revealed expression from this alternate promoter in many TNBC cases suggesting that TET1 overexpression in TNBC cases is partly driven by activation of TET1ALT promoter. TET1ALT protein was also as efficient as the TET1FL protein in generating 5hmC product in overexpression studies [TET1promoter . On the  studies .FL and TET1ALT in HEK293T cells resulted in demethylation of distinct CpG sites. Furthermore, RNAseq analysis of the cells overexpressing either the full length or the alternate isoform revealed that TET1FL and TET1ALT cluster separately and there was moderate overlap between the gene expression targets [Interestingly, overexpression of the TET1 targets . These dWhile research studies have focused on understanding the roles of aberrant DNA methylation events in driving breast carcinogenesis, as prognostic tools and as tools to distinguish breast cancer subtypes, both levels, and alternate isoforms of TET1 have come to shed light on the mechanism of hypomethylation in one of the most hypomethylated tumor types. Moreover, it was recently shown that TET1 has a prognostic potential because TNBCs with high levels of TET1 (the most hypomethylated tumors) had worse overall survival compared to TNBC with low levels of TET1. In fact, high levels of TET1 were associated with worse overall survival in uterine and ovarian cancers as well .In a previous study DNA methylation, particularly hypermethylated differentially methylated regions, were used to stratify TNBCs into survival groups. In contrast to the TET1-associated survival outcome, in this study , the TNBHowever, based on the current literature, the clinical translation of the roles of TETs, even of TET1, particularly in TNBCs is premature. Indeed, current clinical trials are, so far, designed from the mindset that TET enzymes are tumor suppressors and need to be reactivated or their levels should be elevated. These clinical trials  in myelodysplastic syndrome, acute myeloid leukemia, and others have been focused on investigating the efficacy and safety of oral vitamin C, an activator of TET enzymes, in combination with DNMT inhibitors to potentiate the hypomethylating effect in patients with TET2 mutant cancers. However, in the future and with more thorough research on a larger population, it could be important to investigate whether TNBC patients with high TET1 should avoid vitamin C or not. On the other hand, two groups report on discovery of inhibitors of TET enzymes and therefore reflect the need to inhibit the oncogenic roles of TETs. In one of these studies , the cytFurthermore, although currently there is no evidence that TET1 levels in TNBC could be used as a biomarker to predict response to immunotherapy, there is emerging evidence that TET1 mutant samples in several cancers were more immunogenic and showed better responses to immune checkpoint inhibitors . SimilarIn this review, we highlight the role of DNA demethylases in TNBCs. We presented evidence based on the literature that TET enzymes can act as oncogene and tumor suppressors depending on cellular context in breast cancer. The opposing roles of TET enzymes add a layer of complexity in breast cancer, making them more interesting and the target for either inhibition or activation, and the subject of future studies to parse out their roles. Based on the literature review presented in this manuscript, one plausible explanation for the opposing roles of TET is the demethylation and direct or indirect activation of downstream tumor-suppressor genes or oncogenes. This in turn highlights the reliance of TET enzymes, which lack sequence specificity, on interacting partners to be recruited to targeted sites. Therefore, depending on the availability of these various interacting partners, TETs can be targeted to different regions and play different roles. More importantly, the controversial roles of TET1 may be isoform-specific; whereby full-length TET suppresses tumors by protecting CpG islands from gaining methylation while the alternate isoform of TET functions as an oncogene by promoting demethylation of targeted sites based on tissue-specific interacting partners. In fact, the contradictory roles of the TET enzyme as a tumor suppressor and as an oncogene have also been observed in other epigenetic regulators such as DNMT3A and EZH2 ,89,90. I


Heat stress (HS) of dairy cows affects milk protein synthesis partially due to a decline in appetite and dry matter intake (DMI). Several published studies indicate that HS causes oxidative stress (OS) with high levels of free radicals in tissues. Furthermore, the involvement of reactive oxygen species (ROS) in inducing apoptosis, in reducing mammary epithelial cell numbers, and in endocrine disruptions are proposed as the main mechanisms by which HS-induced OS modifies milk protein synthesis. However, challenges remain in determining the levels of apoptosis in vivo, as well as the tracking of the sources of ROS formation. Therefore, further investigations are required.Heat stress (HS) is one of the most important factors posing harm to the economic wellbeing of dairy industries, as it reduces milk yield as well as milk protein content. Recent studies suggest that HS participates in the induction of tissue oxidative stress (OS), as elevated levels of reactive oxygen species (ROS) and mitochondrial dysfunction were observed in dairy cows exposed to hot conditions. The OS induced by HS likely contributes to the reduction in milk protein content, since insulin resistance and apoptosis are promoted by OS and are negatively associated with the synthesis of milk proteins. The apoptosis in the mammary gland directly decreases the amount of mammary epithelial cells, while the insulin resistance affects the regulation of insulin on mTOR pathways. To alleviate OS damages, strategies including antioxidants supplementation have been adopted, but caution needs to be applied as an inappropriate supplement with antioxidants can be harmful. Furthermore, the complete mechanisms by which HS induces OS and OS influences milk protein synthesis are still unclear and further investigation is needed. Exposure of farm animals to high summer environmental temperatures is globally accepted to negatively affect animal husbandry, and no livestock is so more affected than dairy cows. Generally, dairy cows suffer from heat stress (HS) once the heat accumulated surpasses the animals’ capacity to dissipate this heat . HS can 2−), hydrogen peroxide (H2O2), free hydroxyl radicals (OH) [2O2, can act as second messengers for intercellular information exchange through redox signals [Reactive oxygen species (ROS) include superoxide anions , and othals (OH) . These mals (OH) , and som signals . On the  signals ,16. To p signals . When RO signals ,18.The involvement of HS as an inducer of OS in dairy cows has been acknowledged ,20,21. TROS are metabolites produced in the process of mitochondrial electron transport chain reaction , and theFenton reaction:AA = ascorbic acidGSH = reduced glutathioneGSSG = Oxidized glutathioneHaber-Weiss reactionObtaining direct measurements of ROS in vivo are difficult since the ROS has a very short half-life . A few tIndirect methods are widely selected for these tasks because the measuring effects caused by ROS instead of the total amount of generated ROS are easier to use. The techniques including fingerprinting methods and the measurements of antioxidant defense systems have been developed to detect oxidative status . Fingerp+. The ROS produced in mitochondria are mainly superoxide anions, which can go on to generate hydrogen peroxides under the catalysis of superoxide dismutase (SOD) in the inner membrane of the mitochondria [Oxidative stress occurs when the balance between pro-oxidants and antioxidants is disturbed and leads to an elevation in concentrations of free radicals and ROS ,41. ROS chondria .+/NADPH, and it has been shown that heat treatment can enhance this ratio without any changes to NADPH subunits or the NADPH oxidase 1 (NOX1) mRNA expression [Several studies have demonstrated that HS promotes OS and ROS production ,45, whilpression . Furtherpression .2O2, and hydroxyl radicals via the Fenton reaction, as mentioned above [HS leads to an overproduction of transition metal ions (TMI) by increasing the rate of iron release from ferritin . TMI caned above .Hydrogen peroxides are able to diffuse across cell membranes because of their small size and relatively benign reactivity, and they can go on to modify different proteins including phosphatases, transcription factors, ion channels, etc. . While hPrevious studies have shown that Holstein cows under heat exposure have a higher level of thiobarbituric acid-reactive species and malondialdehyde than cows not exposed to heat stress ,58. SincThe heat stress response of cells is a highly conserved cascade of protein activation and gene expression changes which are regulated by heat shock transcription factors and that participate in the defense of stress damage . Heat sh+ gradient through the inner mitochondrial membrane [+ gradient dissipates through the adenosine triphosphate (ATP) synthase complex producing ATP [2− [As summarized above, mitochondria are thermosensitive  and as smembrane . Then thcing ATP . The amocing ATP , it has cing ATP . Consequ ATP [2− , and allAnimals suffer an HS decrease in metabolic heat production to maintain homeothermy, often causing decreased productivity. For dairy cows, HS decreases milk yield as well as milk protein, and this is partly due to the shortage of synthesis precursors in mammary glands ,24. AminFurthermore, signaling pathways dominated by Janus kinase 2/Signal transducers and activators of transcription (JAK2/STAT5) as well as mechanistic target of rapamycin (mTOR) are involved in the regulation of milk protein synthesis . RelatedApoptosis is a process of energy dependent programmed cells death characterized by the specific degradation of cellular DNA. This process is regulated by specific genes and is morphologically manifested as nuclear pyknosis, cell membrane foaming, and apoptotic body formation and ends up with cells break down without any obvious lysis phenomenon . Apoptos2+ seems to assist the release of apoptotic cytochrome c, as was reported in some previous studies [Heat Stress can directly activate the mitochondrial pathway to mediate apoptosis . Hyperth studies ,97.Since cellular stress, including OS, induces mitochondria to generate ROS , this haThe reduction in the amount of lactating mammary epithelial cells can affect the synthesis of milk protein. Results of studies in vitro have shown that hyperthermia as well as the elevated OS contribute to the apoptosis of lactating mammary epithelial cells ,105. HowS binds to eIF4E is also the binding site of eIF4E and eIF4G, which affects the formation of translation initiation complexes [The synthesis of milk protein is regulated by hormones including prolactin (PRL), glucocorticoids, thyroxine, and insulin , while tomplexes . mTORc1 omplexes . S6K1,a omplexes . eEF2 isomplexes . eEF2 kiomplexes .Insulin participates in the regulation of the mTOR signal pathway, as it can activate upstream protein kinase B (PKB) and further act on mTOR ,116. Ins2O2 caused an increase in serine phosphorylation of IRS-1 as well as -2 but also decreased IRS-1 in 3T3-L1 adipocytes [While OS has negative effects on milk production as described above, OS has also been shown to induce the insulin resistance (IR), impaired insulin secretion, and diabetes . IR is dipocytes . The motipocytes . Insulinipocytes ,130. TheTo alleviate the OS induced by HS, strategies to reduce OS by neutralizing the overproduction of ROS or inhibiting its formation should be considered first ,132. ButApart from alleviating heat stress of dairy cows using direct cooling methods (fans and sprays), antioxidants supplementation has been proven useful in HS situations to reduce the occurrence of udder infection as well as improve milk protein content . NumerouThe preceding paragraph detailed the inclusion of the non-enzymatic antioxidants in diets, but the supplementation of plant natural extracts including polyphenols, flavonoids, tannins, and gallic acid has been attracting more attention in recent years due to their high efficiency as well as low residuals ,155,156.The inappropriate use of antioxidant supplements is possible and may lead to the antioxidative imbalance and result in carcinogenesis . As mentHeat stress causes an overproduction of ROS and mitochondrial dysfunction, leading to oxidative stress in dairy cows. Current published studies strongly suggest that oxidative stress as a result of heat stress contributes to a reduction in milk protein. This is caused by an increase in apoptosis in mammary gland tissues that directly reduces the number of mammary epithelial cells, while elevated levels of free radicals also damage milk protein synthesis by regulating signaling pathways. The dietary supplementation of antioxidants has been adopted to alleviate OS including that induced by HS, and this is a strategy that has been proven to be useful in certain circumstances. However, current technology has not been able to establish the real extent of apoptosis in mammary epithelial cells in vivo nor the accurate source of free radical formation in all situations . Further advances are needed to fully understand the effects of HS-induced OS on milk protein reduction, which have the potential to facilitate new methods to reduce the effects of high environmental temperatures on poor milk production in dairy cows.


Drug-induced liver injury (DILI) is a frequent and dangerous adverse effect faced during preclinical and clinical drug therapy. DILI is a leading cause of candidate drug attrition, withdrawal and in clinic, is the primary cause of acute liver failure. Traditional diagnostic markers for DILI include alanine aminotransferase (ALT), aspartate aminotransferase (AST) and alkaline phosphatase (ALP). Yet, these routinely used diagnostic markers have several noteworthy limitations, restricting their sensitivity, specificity and accuracy in diagnosing DILI. Consequently, new biomarkers for DILI need to be identified.A potential biomarker for DILI is cytokeratin-18 (CK18), an intermediate filament protein highly abundant in hepatocytes and cholangiocytes. Extensively researched in a variety of clinical settings, both full length and cleaved forms of CK18 can diagnose early-stage DILI and provide insight into the mechanism of hepatocellular injury compared to traditionally used diagnostic markers. However, relatively little research has been conducted on CK18 in preclinical models of DILI. In particular, CK18 and its relationship with DILI is yet to be characterised in an in vivo rat model. Such characterization of CK18 and ccCK18 responses may enable their use as translational biomarkers for hepatotoxicity and facilitate management of clinical DILI risk in drug development. The aim of this review is to discuss the application of CK18 as a biomarker for DILI. Specifically, this review will highlight the properties of CK18, summarise clinical research that utilised CK18 to diagnose DILI and examine the current challenges preventing the characterisation of CK18 in an in vivo rat model of DILI. Drug-induced liver injury (DILI) was first described in the 1960’s and was once considered a ‘penalty for progress’  , aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bilirubin (TBIL) still routinely utilised to diagnose DILI  demonstrated ALT > 3 ULN in 1.9% of patients with no reports of acute liver failure or severe hepatocellular injury throughout clinical trials , ebrotidine (H2-receptor antagonist) and troglitazone (PPAR activator), from market due to severe patient morbidity and mortality as a direct result of DILI  to prevent serious or any further progression of hepatocellular injury  is one of a handful of potential biomarkers for DILITherefore, this review will highlight the properties of CK18 that may help to fill current gaps in knowledge left by traditional biomarkers for DILI, provide a brief overview of recent clinical research and discuss the current challenges and limitations surrounding the characterisation of CK18 in an in vivo rat model of hepatotoxicity.The cellular location of CK18, also referred to as KRT18, and the cleavage patterns of the protein make it a potential biomarker for DILI. CK18 is a type-I intermediate filament protein highly concentrated in hepatocytes and cholangiocytes , comprising 5% of total liver protein  fragments in serum or plasma reflects the degree of necrotic hepatocellular injury and/or apoptosis  in both clinical and in vivo mouse models of DILI   and interlukin-1 alpha (IL-1α) (Takeda and Akira 5) Table . With th5) Table .Table 1ACK18 and ccCK18 are not the only novel biomarkers to diagnose DILI, with glutamate dehydrogenase (GLDH), microRNA-122 (miRNA-122), macrophage colony stimulating factor receptor (MCSFR) and osteopontin (OPN) also current candidates . In a handful of these studies, comparisons between the serum level of full-length CK18, ccCK18 fragments and traditional diagnostic markers ALT and AST were made (Table P < 0.05) (Table P < 0.01)  Table . However1) Table . These rP < 0.05)   = 0.83 and 0.78, respectively) were more sensitive and specific in predicting the prognosis of death and liver transplantation compared to serum levels of AST, ALT and ALP    may also be useful to quantitate the levels of full length CK18 and ccCK18 fragments. However, these forms of analyses will only be beneficial once rat CK18 protein has been characterised and added to the respective protein libraries. Once full length CK18 and ccCK18 fragments are able to be quantified, the next step is selecting the appropriate in vivo rat model of DILI. However, this in itself has its own challenges.4) nevertheless, both models have specific limitations in rats, the species commonly used in preclinical drug safety testing (McGill and Jaeschke The two most common model hepatotoxins used in in vivo research are APAP and carbon tetrachloride (CClP > 0.05)   . MTX is an antimetabolite drug used for the treatment and maintenance of inflammatory diseases  and forms part of many chemotherapy regimens  (Sotoudehmanesh et al. In addition to selecting the appropriate in vivo rat model of DILI, one must also understand how liver regeneration in the selected model may affect the release of full-length CK18 and ccCK18 fragments into circulation.4-induced hepatocellular injury (Burr et al. 4 is administered that inhibits liver regeneration and leads to the rapid progression of hepatocellular injury (Mehendale Liver regeneration in the chosen in vivo rat model of DILI can also be considered. The liver has the unusual capacity to repair and regenerate following hepatocellular injury and partial hepatectomy (Clemens et al. CK18 is highly concentrated in hepatocytes and cholangiocytes however, as the protein plays a vital role in maintaining the cytoskeleton of epithelial cells, it can also be found in epithelial cells lining other organs (Church et al. CK18 is also expressed by a variety of adenocarcinomas such as those of the lung, pancreas, prostate, colon and rectum (Kramer et al. As such, it is important for both preclinical and clinical models to determine the level of CK18 present in organs such as the kidney and small intestine and if full-length CK18 and/or ccCK18 fragments are released during drug-induced injury from these organs. This can be addressed in preclinical models of drug-induced kidney, pancreatic and gastrointestinal injury. It may be necessary to define a predetermined ratio or percentage of full-length CK18 and ccCK18 fragments that can be attributed to hepatocellular injury (Church and Watkins DILI is a frequent and serious adverse reaction that can occur during preclinical and clinical drug therapy and for which new biomarkers are required. CK18 is a potential biomarker for DILI and has some desirable properties that may help in detection of early-stage DILI and in the identification of the mechanism of hepatocellular injury. CK18 has been applied as a biomarker for DILI in clinical research, demonstrating both full-length and cleaved versions of the protein are accurate and sensitive in diagnosing DILI. However, for CK18 to be applied as a biomarker for DILI preclinically, it needs to be characterised in an in vivo rat model of DILI, due to being the routine species used in preclinical drug safety assessment. The challenges of characterising CK18 in an in vivo rat model of DILI are well-documented, but, once the appropriate in vivo model of DILI has been identified, it will be possible to undertake validation work on recently released rat-specific M65 ELISAs, and determine the specificity of CK18 to hepatocellular injury. This review has provided evidence to support the characterisation of CK18 in an in vivo rat model of DILI, and to investigate the potential translation of CK18 as a routinely used biomarker for DILI in clinical settings.


Klotho rs577912, rs564481, rs9536314; FGF23 rs7955866; IGF1 rs35767; TNFA rs1800629; IL6 rs1800795; MIF rs755622, rs1007888) chosen in relation to their possible involvement with renal disease and concomitant pathologies. Analysis of the raw data did indicate that IL6 rs180795 and MIF rs755622 SNPs might be markers of genetic susceptibility to ESRD. In particular, the C positive genotypes of MIF rs755622, (dominant model) seem to be an independent risk factor for ESDR patients . Stratifying results according to age MIF rs755622 C positive genotype frequencies are increased in both the two age classes considered (<59 and ≥59-year-old subjects). Analyses of data according to gender allowed us to observe that ESRD women shoved a significantly reduced frequency of genotypes bearing IL6 rs180795 C allele. In addition, MIF rs755622 might interact with diabetes or hypercholesterolemia in increasing susceptibility to ESRD. In conclusion, our data indicate that some polymorphisms involved in the regulation of both renal function and inflammatory response can influence the evolution of chronic kidney disease and suggest that the modulation of the activities of these and other genes should also be considered as therapeutic targets on to intervene with innovative therapies.Chronic kidney disease (CKD) is characterized by an increased risk of kidney failure and end-stage renal disease (ESRD). Aging and comorbidities as cardiovascular diseases, metabolic disorders, infectious diseases, or tumors, might increase the risk of dialysis. In addition, genetic susceptibility factors might modulate kidney damage evolution. We have analyzed, in a group of ESRD patients and matched controls, a set of SNPs of genes ( The NatIn recent years, many studies have been carried out aimed at identifying the molecules to be used as the most sensitive biomarkers of kidney function, many of which are proteins produced by different segments of the nephrons, cytokines, receptors that might be implied in the kidney acute or chronic damage ,4. In adRecently, the scenario of possible biomarkers in chronic renal failure has been enriched by new players such as Fibroblast Growth Factor-23 (FGF-23) and Klotho. The progressive deterioration of renal function is characterized by a progressive reduction of Klotho protein and an increase of the FGF-23 levels ,7,8.Klotho gene related to premature aging have also been identified [Klotho has anti-inflammatory and anti-fibrotic activity, preventing vascular calcifications. In humans, the circulating levels of Klotho decrease with age in serum, and polymorphisms of the entified ,10.Klotho expression in the kidney has a direct correlation with the reduction of GFR-23 in CKD subjects [Circulating Klotho can be considered an early diagnostic marker of CKD. Already for estimated glomerular filtration rate (eGFR) values below 90 mL/min, there is a progressive reduction. Clinical and experimental studies have shown that a significant decrease in subjects . High lesubjects  in CKD prs577912 polymorphism of the Klotho gene sequence might affect the expression of the protein and therefore its serum levels. The CC genotype is associated with a 16–21% lower Klotho expression compared to the AA/AC genotype with an association with an increased risk of mortality at 1 year from the start of dialysis [rs7955866 in exon 3 of FGF23 has been related to renal damage [FGF23-239M plasmid release higher concentrations of FGF-23 [Klotho polymorphisms involved in metabolic disorders and cardiovascular pathologies might play a role in favoring progression to ESRD. In particular, rs564481 and rs9536314 might play a role, being implied in metabolic diseases and potentially influencing the prognosis of CKD [rs56481 might influence hypertension susceptibility and creatinine serum levels in different populations [rs9536314 might influence the kidney failure susceptibility impinging on the kidney function decline due to aging. Actually, a pathophysiological modification associated with chronic kidney disease mimics that of elderly individuals, which underlies shared clinical characteristics such as an increased risk of infection and atherosclerosis [rs9536314 was the polymorphism most frequently involved in association studies on cardiovascular and non-cardiovascular mortality. This may be particularly relevant in patients with chronic kidney disease who have decreased Klotho levels in tissues and blood [In particular, dialysis . In addil damage . The minf FGF-23 . On the s of CKD ,16,17,18ulations ,16,17 whclerosis . Moreovend blood .IGF1 gene [IGF1 rs35767 increases the risk of renal disease.More recently, the interactions between the Klotho/FGF-23 system and the growth hormone (GH)/insulin-like growth factor-1 (IGF-1) axis have been analyzed. The GH/IGF-1 system appears relevant for glomerular and tubular function, as well as the synthesis of Klotho . On the GF1 gene . MoreoveGF1 gene , IGF1 rsActually, the chronic inflammation state that characterizes CKD and ESRD induces an up-regulation of proinflammatory cytokines and the associated biomarkers (interleukin (IL)-1β, IL-1 receptor antagonist, IL-6, tumor necrosis factor (TNF)-α, C-reactive protein, and fibrinogen) ,27.Proinflammatory markers are shown to be important in the development of atherosclerotic complications and inflammation. In addition, it has been associated with hypertension in both experimental animal models and human studies ,30,31,32TNF SNPs [IL6 gene SNPs [MIF gene variants [rs1800629 and rs1800795 polymorphisms of TNFA and IL6 genes can selectively affect spontaneous production of inflammatory cytokines influencing kidney function decrease and consequent development and progression of chronic kidney disease.On the other hand, an aging kidney can be considered a paradigmatic model of immunosenescence characterized by defective immune responses and increased systemic inflammation. In the kidney, resident macrophages and fibroblasts are continuously exposed to environmental stimuli, and the effects of cellular reprogramming induced by local immune responses, which accumulate with age, might have a role in the increased susceptibility to kidney disease among elderly individuals ,33 and pTNF SNPs ,39, IL-6TNF SNPs ,41, and ene SNPs ,43,44, avariants ,46,47,48Herein, data on some genetic variants of key cytokine, mediator, and receptors involved in the complex scenario of kidney chronic failure have been reported and their relevance as risk profile markers useful in the prediction of kidney disease development and progression have been evaluated.In collaboration with the Department of Nephrology and Dialysis with C.R.R. for arterial hypertension, “Paolo Giaccone” University Hospital, a population of 93 patients with end-stage renal chronic failure (ESRD) who were routinely undergoing dialysis therapy was recruited. Two control populations, for a total of 213 individuals, were recruited, one of healthy young people and one of the people over 59 years old, at the Department of Medical Biotechnology of the University of Palermo and in collaboration with the U.O.C of Transfusion Medicine of the Paolo Giaccone” University Hospital. The cut-off for age between the two control population was chosen on the basis of the median age of ESRD patients. The ≥59-year-old control population included both healthy subjects and people with hypercholesterolemia, type 2 diabetes, or hypertension without renal impairment . Persons with autoimmune or tumor pathologies or with GFR < 90 were excluded from the control group. The clinical and demographic characteristics of the populations studied are shown in Our study was performed in accordance with ethical standards of the Helsinki Declaration of the World Medical Association and Italian legislation, and was approved by the local institutional review board . All participants gave their informed consent and the data has been encoded to ensure the protection of patient privacy and controls. The EDTA peripheral blood samples used for molecular typing respectively were collected and stored at 70 °C until their use.https://www.ncbi.nlm.nih.gov/snp/, accessed on 18 January 2022).DNA were extracted using a salting-out protocol and DNA quality was verified by 260/280 nm absorbance ratio and gel electrophoresis as previously reported . SNP typhttps://sample-size.net/, accessed on 17 February 2022). Significant differences in genotype distributions between groups were calculated using chi-square or Fisher’s exact test applied to the appropriated 2 × 2 or 3 × 2 contingency tables. Multiple logistic regression models were applied using overdominant , codominant , dominant , and recessive  of heredity and adjusting the results for age, sex, and concomitance of existing pathologies. The choice of the most suitable model was based on the Bayesian Information Criterion (BIC) and the Akaike Information Verification Test (AIC). Odds ratio (OR) values and 95% confidence intervals (95% CI) were calculated and a p-value < 0.05 was considered statistically significant. Statistical analyzes were conducted using version 3.06 of GraphPadInStat software  and online statistical analysis tools  devoted to the evaluation of the association of SNPs with diseases in the presence of other biological, genetic, or clinical variances.Allele and genotype frequencies were evaluated by gene count. Pearson’s test was applied to test the Hardy–Weinberg equilibrium. Power of calculation has been evaluated for all comparisons made using an online tool  of inheritance and adjusting the results for biological variables  and concomitant comorbidities .MIF rs755622, (dominant model) is an independent risk factor for ESDR patients . In addition, a rough significant influence of both TNFA rs1800629 homozygous genotypes have been detected  and clinical variables , genotype frequencies of ESRD patients and controls were stratified and compared according to the different variables. Complete results of these analyses are reported in MIF rs755622 C positive genotype frequencies are increased both in <59 and ≥59-year-old patients than in controls  is the result of the progressive reduction of renal function and the onset of a progressively more severe chronic renal failure whose evolution is strongly influenced by the underlying disease, age, sex, and genetic background of the patients. If we consider the progressive aging trend of the population and the increase in the prevalence of the main diseases predisposing to chronic kidney failure (CKF) , identifying predicting genetic markers that allow early identification of patients with an increased risk to develop severe CKF and reach the ESRD appears to be mandatory ,53.Klotho, in spite of the above-mentioned data on the role of rs577912 polymorphism as risk factors of mortality in dialyzed patients [rs9536314 in the so-called longevity trait [FGF23 rs7955866 and IGF1 rs35767 were not found to be associated with ESRD. On the other hand, more recently, other FGF23 SNPs have been associated with CKD [rs35767 is known to contribute to the development of diabetes in various populations [Analyses of alleles and genotypes crude frequencies of nine SNPs of genes codifying for key cytokine and receptors involved in kidney damage pathogenesis do not allow to identify associations with the patients  and the ty trait . In a stty trait  but we wwith CKD . So furtulations .IL6 rs1800795 are lower in patients than in controls allowing us to hypothesize that the presence of the C allele might be protective against progression to ESRD. This was not confirmed by adjusting the odds ratio for biological variances and clinical comorbidities (Interleukin-6 (IL-6), another important proinflammatory cytokine has been shown to play a critical role in the development of atherosclerosis and atherosclerotic disease including kidney diseases ,56, and bidities , whereasrs1800795G/C variant located in the gene promoter (−174G/C) is associated with changes in transcriptional expression [It has long been known that the pression .rs1800795 and gender are not conclusive. Studies on aging frailty or longevity trait have demonstrated the possibility that genetically determined high production of IL-6 and women gender are risk factors for age-related disease [rs1800795 C allele might be protective against ESRD for CKD-affected women. On the other hand, the relatively small size of the patient sample does not allow to consider this observation conclusive.Even if IL-6 tracks with disability and age-related diseases, published data on the interaction among  disease ,59. In tIn diabetes, the C allele is associated with a low risk of nephropathy ,61. On tThese apparently conflicting results are not easy to interpret. On the other hand, genetically determined low production of IL-6 might be a vantage in the contrasting progression of systemic diseases characterized by a chronic low rate of the inflammatory response , as CKD,TNFA rs1800629 genotypes has been detected. Cytokines of the TNF family and their receptor has been recently proposed as biomarkers of kidney disease. Actually, TNF signaling pathways play important roles in the progression of atherosclerotic and kidney disease [As reported in  disease ,32,33 an disease ,67.TNFA SNPs have suggested that rs1800629, associated with an increased TNFA gene transcription, might be a risk factor for multifactorial diseases [rs1800629 might be a risk factor for CKD in particular in diabetic patients but data have been not conclusive [Studies on diseases , in partnclusive ,39. Genenclusive .Arterial hypertension is one of the main causes of kidney damage leading to end-stage renal disease (ESRD) ,71, and In spite of this, our data do not allow us to identify the association of hypertension and SNP genotype studied as risk or protective factors in our group of ESRD patients.MIF rs1007888, which was found associated with an increased risk of diabetes [rs755622, located at −173bp before the transcription initiation site of MIF gene and associated with a higher production of the cytokine [MIF rs755622 seem to be associated with an increased risk to develop an ESRD both in patients ≥59 and <59 years old.Typing of diabetes , does no2), the risk was relevant only when eGFR had fallen below 45 mL/min. In this view, the possibility to refine the analyses combining age and significant SNP markers could be useful for the best management of CKD prevention and therapy. However, our data indicate that the MIF rs755622 C allele is an age and gender independent risk factor for ESRD progressionAging is per se a risk factor for kidney diseases and in particular for CKD. However, it is well known that glomerular filtration rate (eGFR) declines with aging and this might impinge on the risk of CKD evolution to terminal stage and mortality. As recently reported by Delanaye et al. , for peoOur results are in good agreement with data of other groups indicating that high production of MIF is correlated with an increased susceptibility for a large number of inflammatory and metabolic diseases including chronic kidney failure ,77,78,79rs755622 on CKD predisposition and ESRD progression might be influenced by individual biological factors such as aging [On the other hand, in chronic kidney diseases (CKD), MIF limits the pro-fibrotic and proinflammatory activation of tubular cells by counteracting the arrest of the cell cycle and promoting its regeneration; In addition, in acute kidney damage (AKI), MIF also counteracts the apoptosis of tubular cells with a protective effect ,82,83. Sas aging  or the pas aging .As it is well known, diabetes is one of the major comorbidities associated with CKD. CKD can be a consequence of the alteration of the glucose status or we can have a kidney disease on which type II diabetes is superimposed. In diabetic nephropathy, the endothelial involvement and local induction of cytokines and chemokines induce immune cell recruitment accompanying and promoting tissue remodeling and nephron atrophy . MoreoveKlotho rs564481 [MIF rs755622 genetic polymorphism might play a role in ESRD progression in subjects affected by these pathologies. However, our data were obtained analyzing a relatively low number of subjects bearing diabetes or hypercholesterolemia, so a definitive conclusion cannot be drawn, even if some literature reports seem to support this view [The analysis of the association between the polymorphisms we typed and the risk of dialysis in subjects suffering from hypercholesterolemia or diabetes do not allow us to identify associations with polymorphisms known to be associated with changes in lipid and carbohydrate metabolism as rs564481 ,16, wherhis view ,89,90.IL6rs1800795 and MIF rs755622, involved in the regulation of both renal function and inflammatory response can influence the evolution of chronic kidney disease. In particular, IL6rs1800795G genotype and MIF rs755622C alleles associated with increased production of the respective cytokines inducing increased inflammatory damages of kidney parenchyma might be useful as a marker for evaluation of ESRD evolution risk.In conclusion, CKD might be considered a paradigm of the multifactorial diseases in which preexisting pathological conditions and genetic susceptibility factors might interplay with each other, resulting in different trajectories of damage evolution to the final stage as ESRD. In this view, we have analyzed the role of some genetic variants of key cytokines, mediators, and receptors involved in the complex scenario of kidney chronic failure and their interaction with comorbidities. Our data indicate that some polymorphisms, MIF rs755622, in the progression of end-stage renal disease.In addition, it is hypothesized that the modulation of the activities of these and other genes can also be considered therapeutic targets for innovative therapies. In this light, further studies will certainly be needed to investigate the role and interaction of these SNPs, in particular


COVID-19 during pregnancy is associated with an increased risk of maternal death, intensive care unit admission, and preterm birth; however, many people who are pregnant refuse to receive COVID-19 vaccination because of a lack of safety data.The objective of this preliminary study was to assess whether Twitter data could be used to identify a cohort for epidemiologic studies of COVID-19 vaccination in pregnancy. Specifically, we examined whether it is possible to identify users who have reported (1) that they received COVID-19 vaccination during pregnancy or the periconception period, and (2) their pregnancy outcomes.We developed regular expressions to search for reports of COVID-19 vaccination in a large collection of tweets posted through the beginning of July 2021 by users who have announced their pregnancy on Twitter. To help determine if users were vaccinated during pregnancy, we drew upon a natural language processing (NLP) tool that estimates the timeframe of the prenatal period. For users who posted tweets with a timestamp indicating they were vaccinated during pregnancy, we drew upon additional NLP tools to help identify tweets that reported their pregnancy outcomes.We manually verified the content of tweets detected automatically, identifying 150 users who reported on Twitter that they received at least one dose of COVID-19 vaccination during pregnancy or the periconception period. We manually verified at least one reported outcome for 45 of the 60 (75%) completed pregnancies.Given the limited availability of data on COVID-19 vaccine safety in pregnancy, Twitter can be a complementary resource for potentially increasing the acceptance of COVID-19 vaccination in pregnant populations. The results of this preliminary study justify the development of scalable methods to identify a larger cohort for epidemiologic studies. COVID-19 during pregnancy is associated with an increased risk of maternal death, intensive care unit admission, and preterm birth . SurveysIn the United States, 42% of people aged 18-29 years and 27% of people aged 30-49 years use Twitter . Our priThe Institutional Review Board of the University of Pennsylvania reviewed this study and deemed it exempt from human subjects research under Category 4 of Paragraph b of the US Code of Federal Regulations Title 45 Section 46.101 for publicly available data sources (45 CFR §46.101(b)(4)).To facilitate a preliminary assessment of self-reports of COVID-19 vaccination on Twitter, we developed 6 handwritten, high-precision regular expressions designed to match tweets mentioning that the user received at least one dose of COVID-19 vaccination . In prioFor users who posted tweets with a timestamp indicating they were vaccinated during pregnancy or the periconception period, we drew upon additional automated NLP tools, developed in our prior work -11, thatWe manually verified the content of tweets detected automatically, identifying 150 users who reported on Twitter that they received at least one dose of COVID-19 vaccination during pregnancy or the periconception period. Our study demonstrates that there are users who report on Twitter that they were vaccinated during pregnancy, including in early pregnancy and the periconception period, and that many of them report their pregnancy outcomes. Therefore, the results of this study justify the development of scalable methods to identify a larger cohort on Twitter for epidemiologic studies of COVID-19 vaccination in pregnancy. The 150 users in this study were identified based on tweets posted through the beginning of July 2021. Since identifying these users, we have redeployed the 6 regular expressions for detecting tweets that self-report COVID-19 vaccination and our NLP tool that estimates the timeframe of the prenatal period  on usersOur preliminary results suggest that reports of preterm birth and miscarriage are largely unaffected by a potential reporting bias, given that we detected a gestational age of at least 37 weeks for 39 of the 40 (98%) completed pregnancies for which we did not identify a preterm birth or miscarriage. However, reports of low birth weight may be affected by a potential reporting bias, given that we detected a birth weight of at least 5 pounds and 8 ounces for only 7 of the 44 (16%) completed pregnancies for which we did not identify a low birth weight. Given our initial small sample of Twitter users, it is not surprising that we did not detect any reports of birth defects or stillbirth, which have an incidence in the United States of 3%  and lessGiven the limited availability of data on COVID-19 vaccine safety in pregnancy, Twitter can be a complementary resource for continued monitoring and potentially increasing the acceptance of COVID-19 vaccination in pregnant populations. Directions for future work include developing methods to detect a larger cohort, and performing an epidemiologic study comparing their pregnancy outcomes to those of users who have announced their pregnancy on Twitter  but gave


Interestingly, authors highlighted important challenges and provided a sharp focus on technical aspects of current research. Those works, which include multiple plant systems , focused on many and different developmental processes, such as flower morphogenesis, gametogenesis, plastid differentiation, root development, and seed germination, which overall could orient the direction of future research and efforts in this exciting field.This special issue includes different research papers and reviews that studied the role of signaling cascades controlling both plant developmental processes and plant response mechanisms to biotic and abiotic stresses. It is not possible to include all relevant areas in a single issue, but this special issue includes excellent research and thought-provoking review articles covering aspects of signal transduction impacting development: transcriptional regulation, hormonal control, cell wall homeostasis, ROS/CaTETRATRICOPEPTIDE THIOREDOXIN-LIKE 1 (TTL1). The new study reports novel insights into the function of TTL1. Based on mutant analyses, TTL1 is involved in determining the stiffness of the cell wall of the cells from the root elongation zone. The authors concluded that TTL1 is involved in anisotropic root growth during osmotic stress adaptation [An emerging new field in plant development concerns the study of signaling by cell wall (CW) modifications, which are responsible for many aspects of plant life. Plant development is extensively studied at many levels. At the cellular level, an important aspect of plant development is the CW. Developmental processes can influence CWs, but also CWs can affect plant development. In this issue, two reviews ,2 and a aptation .Arabidopsis thaliana as a model system [VviZFP family). They identified 98 potential VviZFP genes, and tissue-specific expression pattern analysis allowed them to identify a group of interesting ZFP candidates linked to pollen development. The authors performed elegant expression analysis and described the spatio-temporal expression pattern of two ZFPs during anther and mature pollen grain maturation. These results constitute an important basis for further agronomic studies that can exploit the role of VviZFPs in grapevine reproductive development [Transcription factors (TFs) are crucial regulators of plant development. This issue includes several articles that discuss the role of TFs in development in distinct plant systems such as Arabidopsis and grapevine ,5,6. TFsl system  is an exelopment .gun1-102 ftsh5-3 double mutant, Jeran et al., 2021 [PUB4-7 inactivation, in the gun1-102 ftsh5-3 background, is able to restore functional chloroplasts, although with an altered phenotype, thus suggesting that the variegated phenotype of gun1-102 ftsh5-3 is largely dependent on PUB4 activity [This issue also includes an interesting review on reproductive development . The autl., 2021 , identifactivity .Ceratopteris richardii (Ceratopteris), a subtropical fern which represents a plant model system for developmental biology studies. The results of this work support the hypothesis of a high mitotic rate of the root apical cell, while suggesting the lack of a quiescent center in the stem cell niche of Ceratopteris roots [Interesting views are also provided in this issue by including developmental studies using other model systems, such as ferns . A cruciis roots .2+ signaling pathways also determine gametophyte functioning, sexual reproduction, and embryo formation in plants and animals. In this issue, Lodde et al. [2+ in both plant and animal developmental biology studies to further elucidate these crucial signaling pathways. The field is well explored in animals, and, in recent years, multiple advances in plant science have been made concerning signal transduction via ROS and Ca2+ signaling into developmental processes and in response to biotic and abiotic stresses. The emphasis on the reproductive system provided in the work is interesting, since this had not previously been reviewed. The review described the basis for ROS production, metabolism, and detoxification systems used by plants and animals to control ROS homeostasis below toxicity levels. Interestingly, the authors reviewed recent developments in the use of genetically engineered sensors to monitor concentration fluxes and localization of ROS/Ca2+ in vivo [2+ signaling pathways involved in animal embryo and seed development and discussed “omics” data, providing a list of potential targets affecting ROS in reproductive processes in plant development [Reactive oxygen species (ROS) control multiple developmental processes, and in this issue two complementary contributions reviewed these relationships ,11. DiffWe would like to express our great appreciation for the efforts of the 64 authors from 20 institutions (in nine different countries) that have participated in this special issue, even more so considering the difficult circumstances that have accompanied the pandemic  and the limitations researchers have suffered to keep science active in the laboratories. We also thank enormously the work of the reviewers involved in the revision of the 11 articles that constitute this issue.In summary, the current Special Issue “New Insights into Plant Development and Signal Transduction” provides a state-of-the-art perspective of experts in the field and also provides suggestions for potential industrial applications and challenges for plant biologists.


Available information on the radiological findings of the 2019 novel coronavirus disease (COVID-19) is constantly updated. Ground glass opacities (GGOs) and consolidation with bilateral and peripheral distribution have been reported as the most common CT findings, but less typical features can also be identified. According to the reported studies, SARS-CoV-2 infection is not limited to the respiratory system, and it can also affect other organs. Renal dysfunction, gastrointestinal complications, liver dysfunction, cardiac manifestations, and neurological abnormalities are among the reported extrapulmonary features. This review aims to provide updated information for radiologists and all clinicians to better understand the radiological manifestations of COVID-19.Radiological findings observed in SARS-CoV-2 virus infections were explored in detail in PubMed and Google Scholar databases.The typical pulmonary manifestations of COVID-19 pneumonia were determined as GGOs and accompanying consolidations that primarily involve the periphery of the bilateral lower lobes. The most common extrapulmonary findings were increased resistance to flow in the kidneys, thickening of vascular walls, fatty liver, pancreas, and heart inflammation findings. However, these findings were not specific and significantly overlapped those caused by other viral diseases, and therefore alternative diagnoses should be considered in patients with negative diagnostic tests.  Radiological imaging plays a supportive role in the care of patients with COVID-19. Both clinicians and radiologists need to know associated pulmonary and extrapulmonary findings and imaging features to help diagnose and manage the possible complications of the disease at an early stage. They should also be familiar with CT findings in patients with COVID-19 since the disease can be incidentally detected during imaging performed with other indications. Coronavirus disease (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). After COVID-19 was first reported in Wuhan, China, in December 2019, it spread rapidly worldwide and was declared a pandemic by the World Health Organization (WHO) on March 12, 2020 [1]. The SARS-CoV-2 virus has had devastating effects on human life, health systems, and economies. SARS-CoV-2 is a beta-coronavirus that infected more than 167 million people as of June 2021, resulting in more than 3 million deaths worldwide. A significant proportion of those with COVID-19 infection remain asymptomatic (40%–50%) or show relatively mild symptoms (40%) [2]. COVID-19 disease caused by SARS-CoV-2 typically presents with weakness, fever, cough, and sore throat but can progress to pneumonia, bronchitis, and acute respiratory distress syndrome (ARDS). However, since none of these symptoms are specific to COVID-19, and the disease can rapidly progress to severe pneumonia, multiple organ involvement including renal failure, vascular thrombosis, ischemia, respiratory distress, and gastrointestinal symptoms, there is an urgent need for rapid diagnostic tests [3]. Although the reverse transcriptase-polymerase chain reaction (RT-PCR) test for viral nucleic acids is the gold standard in the diagnosis of COVID-19, radiological imaging, especially computed tomography (CT) has become increasingly important in diagnosis due to the false-negative results of the RT-PCR test [4]. Studies have shown that the sensitivity of CT is between 60% and 98% . This range is relatively higher compared to the RT-PCR test (60%–70%), but its specificity (25%–53%) is low. The positive and negative predictive values of chest CT for COVID-19 are estimated at 92% and 42%, respectively. The relatively low negative predictive value reduces the value of CT as a screening test for COVID-19 in the early stages of the disease [7]. In addition, considering that it contains ionizing radiation, CT should be used as a problem-solving method in patients with a negative RT-PCR test rather than as a screening method [8]. Imaging plays an essential role in the management of COVID-19. In this study, we aimed to review the areas of use and imaging findings of various imaging methods, such as chest radiographs, CT, and ultrasonography in the diagnosis and treatment of COVID-19. Chest radiography is often the first imaging modality of choice for patients with known or suspected COVID-19 pneumonia. In the literature, the diagnostic value of chest X-ray in COVID-19 pneumonia is reported to vary between 30% and 60%, which is relatively low. Although it is possible to detect some abnormalities in the presence of viral pneumonia on chest X-ray, normal findings obtained from this modality do not exclude the disease. Moreover, if the disease progresses, the possibility of an abnormal chest radiograph increases .Chest radiography often does not show any abnormality in the early stage of COVID-19 infection. Therefore, radiography has not been recommended as a first-line method for the investigation of chest abnormalities in patients with suspected COVID-19 [11]. However, in mild COVID-19 conditions, chest radiography may demonstrate local irregular radiopacities in the outer parts of the lungs and the subpleural region [12]. COVID-19 lung involvement occurs in bilateral involvement of the peripheral and lower lobe basal segments [13]. In severe cases, there is a widespread, sometimes patchy, multiple consolidation with a small amount of pleural effusion. In critically ill patients, areas of consolidation covering the entire lung are referred to as white lung [14]. In conclusion, the diagnostic contribution of chest radiography is lower than that of chest CT, and noncontrast chest CT should be considered for the early diagnosis of viral disease in suspected patients with normal chest radiography findings [10].USG is one of the most commonly used imaging methods in clinical practice and allows for the systematic and rapid examination of multiple organs at the bedside without using radiation. The general opinion is that USG cannot detect lung lesions; however, many lung pathologies can be imaged with this modality. The main signs of lung ultrasound in COVID-19 can be defined as A-lines, B-lines, subpleural consolidation, and lung deviation signs [15]. In COVID-19-infected cases, the occurrence of pulmonary lesions predominantly in the peripheral regions of the lung and the subpleural areas of both lungs facilitates sonographic imaging [16]. However, deeply located lung lesions cannot be detected on USG, and it is difficult to use this imaging method in the differential diagnosis of pneumonia. USG is not recommended in the diagnosis of mild cases but it can be considered as an additional pulmonary and cardiac evaluation method, especially in critically ill patients who should not be moved and for whom a CT examination is not possible [17].USG is particularly important in examining critically ill patients and determining treatments, particularly hemodynamic therapy. It is helpful in functional imaging rather than anatomical imaging, and its applications can be summarized as follows : First, USG is an imaging method that offers the rapid evaluation of the disease. An increase in the number of B-lines and the area of consolidation may indicate lung disease progression [20]. Second, respiratory support is required in severe and critical cases, but the time point for the transition from noninvasive ventilation to invasive mechanical ventilation is essential [21]. Lung USG showing progressive diffuse B-lines and significantly increased diaphragm activity may indicate severe lung damage. When mechanical ventilation cannot improve oxygenation, extracorporeal membrane oxygenation (ECMO) may be a treatment option. USG provides visual management in the ECMO process [22]. Third, echocardiography (cardiac ultrasound) can be used to evaluate circulatory functions. The most frequently reported complication in critically ill patients with COVID-19 is ARDS, followed by acute heart damage and secondary infection [23]. Echocardiography can also quickly assess hemodynamics and determine whether there are segmental wall motion abnormalities, stress-induced cardiomyopathy, or acute right heart dysfunction that may guide treatment . Finally, coagulation abnormalities have been reported in approximately 20% of critically ill COVID-19 cases . Sudden worsening of the condition often suggests a state of hypercoagulation with a dramatic increase in the D-dimer level [27]. Asymmetrical pain or swelling may indicate the risk of lower extremity deep vein thrombosis (DVT) for patients staying in bed for more than three days. For patients with central venous catheters, local swelling of catheterized limbs may indicate the risk of upper limb DVT. In all these cases, vascular structures can be evaluated with color Doppler USG [28]. In this way, effective and timely interventions can be provided for patients at high risk of embolism [12].The main limitation of USG is that in general it has poor specificity, and its findings overlap with those of other pneumonia etiologies or incidental chronic findings [29].CT is sensitive the early signs of COVID-19 and changes that may not be seen on chest radiography [30]. It is essential to know that in 50% of patients with COVID-19 symptoms, an abnormal chest CT finding may be expected in the first days after symptoms appear [13]. On the contrary, lung changes due to COVID-19 can be monitored on CT in patients with no symptoms and are usually seen incidentally.Chest CT can have characteristic features of COVID-19 during active infection, and therefore it can help monitor the course of the disease and provide timely treatment. The hallmark of COVID-19 is the bilateral presence of patchy GGOs that can merge into dense, consolidative lesions with a predominantly peripheral distribution in the subpleural areas, as well as bronchovascular bundles . As the disease progresses, the number of lesions can rapidly increase and extend into central regions [33]. During the recovery of the disease, lesions gradually regress over two weeks, during which time fibrotic changes may occur [34]. In addition to GGOs and consolidations, many other CT findings, such as interstitial thickening, halo sign, inverted halo sign, and airway-vascular changes can be seen in COVID-19 pneumonia. These findings and distribution patterns may help distinguish COVID-19 pneumonia from other types of pneumonia [35].GGOs are nonspecific findings defined as hazy lung opacities that do not cover the underlying vascular or bronchial boundaries and are assumed to be related to partial airspace filling or interstitial thickening  [36]. The predominant CT pattern in COVID-19 is bilateral GGOs associated with consolidations, but findings may differ from patient to patient or depending on the stage of the disease . It has been reported that GGOs are seen at a rate of 50%–95%, and they are isolated in 50% of cases and accompanied by consolidations in 30%–40% [37–39].Consolidations result from the complete displacement of alveolar air spaces by pathological fluids or cells, leading to an increase in parenchymal density that obscures the underlying vessels and bronchial walls [36]. Consolidations have been reported in 20%–63% of cases and may be multifocal, irregular, or segmental with subpleural or peribronchovascular distribution . Consolidations occur with the progression of the disease within two weeks after the onset of the disease and often accompany GGOs [42].The reticular pattern consists of a complex network of linear opacities associated with interlobular and intralobular septal thickening and is caused by lymphocyte infiltration  . With an incidence of 27%, it is the third most common finding in COVID-19 after GGOs and consolidations , and its incidence increases over the course of the disease [42].This pattern is defined as the presence of GGOs with overlapping interlobular septal thickening, giving the appearance of irregular cobblestones and may be associated with alveolar edema combined with the inflammation of interstitial structures [36]. In COVID-19, this finding is less frequent than GGOs and consolidations alone [34]. However, this sign has been reported in 5%–36% of patients and may be considered an indication of progression or the highest stage of the disease when accompanied by GGOs and consolidations [41].Air bronchograms, in which air-filled bronchi can be seen in a highly attenuated parenchymal background, have been recorded in series of patients affected by COVID-19  . However, some autopsy studies indicate that the bronchi are actually filled with gelatinous mucus plugs; therefore, the term bronchiolectasis may be more appropriate than air bronchogram because the bronchi are not filled with air and are associated with mild bronchial dilatation [46]. Similar studies have also emphasized that the high viscosity of mucus can cause bronchiolar damage, and thus lead to bronchiolectasis and characteristic dry cough in patients with COVID-19 [35].Airway changes reported in association with COVID-19 occur as a result of bronchiectasis in some patients and bronchial wall thickening in 10%–20% of cases . The pathological mechanism is bronchial obstruction and inflammatory damage of the bronchial wall, both of which lead to the destruction of the bronchial wall structure and development of fibrosis, and consequently bronchiectasis [36]. In addition, the percentage of bronchial wall thickening has been determined to be significantly higher in patients with critical COVID-19 [41].The most common pleural change in patients with COVID-19 is not pleural effusion (5%) but pleural thickening (32%) [42]. The presence of pleural thickening was confirmed by autopsy findings [46]. The presence of pleural effusion has been suggested to be a poor prognostic factor .Subpleural lines refer to thin curvilinear opacities of approximately 1–3 mm in thickness, located in the subpleural region and distributed parallel to the pleural surface . They are a nonspecific sign seen in 33% of COVID-19 cases, but they may also result from atelectasis, pulmonary edema, and fibrosis [47]. In addition, subpleural lines may be associated with the predominantly peripheral location of parenchymal changes in COVID-19 [48].Fibrotic lesions may occur in patients with the proliferative disease and those recovering from chronic pulmonary inflammation due to the replacement of cellular components by scar tissue [49]. Fibrosis is usually seen at two to three weeks after the onset of COVID-19 when interstitial changes begin to occur [50]. Some studies have suggested that the presence of fibrosis indicates advanced COVID-19, which can lead to bronchial deformation. However, the general belief is that fibrosis is a part of the recovery process .A pulmonary nodule is defined as a round or irregular parenchymal opacity of less than 3 cm in diameter and often associated with the presence of viral pneumonia . Multiple solid nodules with irregular contours have been found in 3%–13% of patients with COVID-19, sometimes accompanied by a halo sign .This is described as a nodule or mass surrounded by GGOs [36]. The pathological mechanism underlying this sign remains unclear. However, apart from perilesional hemorrhage being a possible sign of viral infections and organized pneumonia, the halo sign has also been associated with angioinvasive fungal infections or hypervascular metastases .The presence of a ring-like consolidation zone with an overlapping circular GGO is defined as an inverted halo sign  [36], which has been reported to be associated with COVID-19. It is considered as a healing lesion with a low-density core or a new lesion that develops around a preexisting GGO . In addition, it has been previously described in association with cryptogenic organizing pneumonia and other pulmonary infections .The dilation of the pulmonary vessels can be caused by proinflammatory factors resulting in capillary wall damage and edema. The onset of the disease can occur in as early as one to seven days [51].Cavitation is extremely rare in patients with COVID-19 [60]. The air bubble sign, defined as the small air-containing areas associated with bronchiolectasis, is also uncommon but can be seen in COVID-19 cases  [61].When the short-axis diameter of the mediastinal lymph node is more than 1 cm, it is defined as lymphadenopathy [36]. This finding has been reported in 4%–8% of patients with COVID-19 and considered to be an essential risk factor for severe pneumonia [40–42]. However, bacterial superinfection should be suspected when lymphadenopathies are accompanied by pleural effusion and small lung nodules .Pericardial effusion has been reported in approximately 5% of patients with COVID-19 and assumed to be associated with the presence of severe inflammation .CT showing high sensitivity (97%) but poor specificity (25%) for the diagnosis of COVID-19 has been primarily attributed to the COVID-19 pneumonia findings overlapping those of other viral types of pneumonia, especially influenza viruses, parainfluenza virus, adenovirus, and respiratory syncytial virus [64–66]. Similar to COVID-19, most viral types of pneumonia have a predominant distribution in the posterior and peripheral parts of the lungs, affecting more than one lung lobe. However, some findings may be useful in the differential diagnosis. For example, the main feature of pneumonia caused by the respiratory syncytial virus is small centrilobular nodules and consolidation areas, usually distributed asymmetrically in the lungs. Adenovirus pneumonia on chest CT shows bilateral multifocal GGOs with irregular consolidations and may have a lobar or segmental distribution. In parainfluenza virus pneumonia, the presence of centrilobular nodules with bronchial wall thickening may help distinguish it from other viral types of pneumonia [33]. Coronaviridae, and they have similar underlying pathological features. However, unlike the reported SARS cases, COVID-19 pneumonia shows a multifocal tendency of distribution and a peripheral distribution of GGOs in the upper lobes and a basilar or subpleural preference in the lower lobes.The radiological appearance of COVID-19 is not much different from pneumonia associated with the other two coronaviruses, SARS and MERS, possibly because they belong to the same family of The consolidation frequency and severity score of COVID-19 pneumonia are also much lower than SARS, which may explain the lower mortality rates of the former. The more peripheral distribution of COVID-19 pneumonia in the upper lobes can be used to distinguish it from MERS [67]. In contrast to COVID-19 pneumonia, SARS and MERS pneumonia usually have a single focus, and no reference to halo or reverse halo signs has been made in the literature [68]. Furthermore, unlike COVID-19, neither SARS nor MERS has been associated with the findings of lymphadenopathy, pleural effusion, or nodules [67].The CT features of COVID-19 also show some similarities with other pulmonary pathologies, such as pulmonary edema, pulmonary hemorrhage, bronchiolitis obliterans, chronic obstructive pulmonary disease, and drug-induced lung disease [66]. In pulmonary edema, chest CT displays GGOs with central distribution, usually associated with smooth interlobular septal thickening, pleural effusion, and cardiomegaly, indicating congestive heart failure. In diffuse pulmonary hemorrhage, CT usually reveals patchy or diffuse GGOs associated with consolidations or poorly defined centrilobular opacities [69]. Air trapping is the main CT feature in bronchiolitis, while bronchial wall thickening, bronchiectasis, and centrilobular opacities can also be seen. In chronic obstructive pulmonary disease, bronchial wall thickening can be seen in addition to pulmonary emphysema. Drug-induced lung diseases are very diverse and can show various lung presentations ranging from ARDS to pulmonary fibrosis. For example, the most common chest CT features in methotrexate-induced lung disease are diffuse parenchymal opacification, reticular opacities, and centrilobular nodules with a pattern of nonspecific interstitial pneumonia [66].Since COVID-19 can show similar clinical and radiological features with many lung pathologies, it is essential to be aware of the characteristic findings of the disease in both clinical and imaging to determine its severity, predict its prognosis, and perform a differential diagnosis.Although the respiratory spread of COVID-19 is well documented in the literature, it has also been shown that the virus is not limited to the lungs [70]. There are many studies on the extrapulmonary involvement of COVID-19, and it is accepted that some of these involvements can seriously affect the disease prognosis [71]. It is considered that SARS-CoV-2 uses angiotensin-converting enzyme 2 (ACE2) as the cell receptor for cellular access in humans [72]. ACE2 receptor is highly expressed in the lungs, kidneys, testicles, gastrointestinal system (GIS), liver, vascular endothelial cells, and arterial smooth muscle cells [73]. Therefore, all these organs and systems with a high expression of ACE2 receptors can be considered to be the targets of SARS-CoV-2 infection [74]. Kidneys are one of the most frequently affected extrapulmonary organs in patients infected with SARS-CoV-2, especially in severe cases [75–77]. The etiology of kidney dysfunction associated with COVID-19 disease is probably multifactorial [78]. The SARS-CoV-2 virus may have a direct cytopathic effect on kidney tissue or cause damage through systemic inflammatory response syndrome, cytokine release, and sepsis, as observed in other viral diseases. Renal cell damage can cause tubular atrophy, renal interstitial fibrosis, and acute kidney injury (AKI), which is strongly associated with increased mortality and morbidity [79]. Studies conducted with patients affected by COVID-19 have shown that AKI is mainly characterized by tubular injury, increased serum creatinine and urea-nitrogen concentrations . The severity of tubular damage is also higher in patients with severe COVID-19 than in those less affected by the disease. CT imaging findings in patients with AKI include inflammation and edema of the renal parenchyma characterized by lower CT attenuation values than normal kidneys . In addition, multiple wedge-shaped parenchymal defects compatible with infarction can be seen . Increased parenchymal echogenicity, loss of cortex-medulla separation, and thickening of the collecting system wall can also be seen in renal USG. A significant increase in the resistive index value (above 0.9) on the color Doppler USG examination indicates tubular damage. However, similar changes due to rejection can be seen in kidney transplant patients. In these cases, if clinical findings do not allow for a differential diagnosis, a renal biopsy is required [84–86].Since ACE2 is produced in testicular cells, it plays a critical role in fertility. Therefore, due to potential testicular damage caused by the virus, infertility can be seen as one of the late complications of the disease. Additionally, viral orchitis may occur [87].Cardiac involvement in COVID-19 disease may be due to the direct invasion of the myocardium. However, it may also be secondary to a hyperinflammatory condition, which results in atherosclerotic plaques becoming prone to rupture. As a result, endothelial dysfunction and increased procoagulant activity increase the risk of thromboembolism. In addition, 40% of patients requiring hospitalization due to COVID-19 have cardiovascular disease, which may worsen due to COVID-19 involvement .Myocarditis is among the major complications of SARS-CoV-2 infection, accounting for 7% of deaths associated with COVID-19 [90]. In the echocardiography of patients infected with COVID-19, half of the patients show ventricular abnormalities, mainly ventricular dysfunction patterns, myocardial infarction, and signs of myocarditis [91]. Right ventricular abnormalities are common in those with the more severe disease [90]. Other findings include global or regional myocardial systolic dysfunction with or without pericardial effusion . Right ventricular abnormalities have been reported in these patients, also reflecting the presence of severe respiratory illness, including subclinical or clinical thromboembolism [91].CT coronary angiography can help rule out coronary artery disease in all these cases. It is also used to differentiate the pathologies of nonobstructive coronary artery myocardial infarction, cardiomyopathy, and acute myocarditis [91]. Cardiac magnetic resonance imaging (MRI) is an ideal imaging method in the diagnosis of acute myocarditis. Cardiac involvement has been detected in 78% of those who have had COVID-19 and undergone cardiac MRI, while ongoing myocardial inflammation is reported to be as high as 60% [92]. Imaging findings in these patients are abnormal T1 and T2 signals and late contrast enhancement indicating regional damage due to myocardial inflammation. The majority of high T2 signals are associated with myocardial edema in the interventricular septum and the anterior and anterolateral walls [93].Vascular involvement in COVID-19 infection may not be limited to the cardiovascular system. This disease can cause a transient state of hypercoagulation, resulting in the development of vascular thromboembolic events, such as deep vein thrombosis and pulmonary embolism if proper medication is not administered [94–96]. Although the exact pathogenesis of COVID-19-associated hypercoagulopathy remains unclear, high D-dimer levels and coagulation abnormalities have been reported in patients with the disease. These findings have been associated with a poor clinical course and higher mortality rate [97–100]. GIS symptoms are common in COVID-19 and may be present in up to 26% of patients. GIS symptoms usually occur one to two days after pulmonary symptoms and include anorexia (26.8%), nausea and vomiting (10.2%), diarrhea (12.5%), and abdominal pain (9.2%) . GIS symptoms can sometimes be seen at an early stage of the disease, preceding the other typical symptoms of COVID-19 [103]. The spectrum of cross-sectional imaging findings of intestinal pathologies in patients with COVID-19 ranges from inflammation to ischemia and necrosis. Imaging findings include intestinal wall thickening, contrast enhancement changes in the intestinal wall, ileus, pneumatosis, and gas in the portal venous system .COVID-19 involvement of the liver, gall bladder, pancreas, and spleen has also been described. In liver injury associated with SARS-CoV-2, increased parenchymal echogenicity and decreased parenchymal density can be seen on the CT examination due to hepatic steatosis [104]. Infarcts may occur in the spleen, which can be visualized as wedge-shaped, low-density areas on CT [107]. Examples of pathologies in the gall bladder and tracts of COVID-19 cases are distension of the gallbladder, thickening of the bladder wall, and sludge formation without intraluminal stones [108]. Concerning the pancreas, an increase in the pancreas volume can be seen, indicating pancreatitis without stones in the biliary tract  [109]. USG or CT can easily detect the presence of these findings.Approximately one-quarter of patients with COVID-19 have some form of neurological pathology [110]. In hospitalized patients, the rate of neurological complications varies between 6 and 36%, with acute cerebrovascular diseases being the most common and serious [111]. The most common symptoms are altered mental states, focal deficits, and seizures due to ischemic stroke [112]. Other nonspecific clinical symptoms include headache, confusion, and dizziness. Loss of smell and taste in the early stages of COVID-19 disease occurs in approximately 15% of patients [113]. Most patients with neurological involvement have at least one preexisting comorbidity, such as cardiovascular disease and diabetes [112].Nervous system imaging findings associated with COVID-19 disease can be categorized as follows: cerebrovascular disorder, encephalitis, other encephalopathies, and peripheral nervous system abnormalities. Cerebrovascular disease may refer to the presence of large or small vessel ischemic infarction or hemorrhagic infarction due to venous sinus thrombosis, as well as microhemorrhages involving the juxtacortical and callosal white matter or gross hemorrhage [114]. Encephalitis is diagnosed based on cortical FLAIR signal abnormality regardless of diffusion imaging feature. Although some publications indicate that medial temporal lobe involvement is more common, there is no consensus on this issue [115]. In addition, there may be an abnormal leptomeningeal enhancement in encephalitis, which is sometimes only seen in the postcontrast FLAIR sequence [116]. Depending on encephalitis, differences in signal and enhancement and diffusion abnormalities can also be seen in the basal ganglia, especially the substantia nigra, globus pallidus, and striatonigral pathways [113].Another form of neurological involvement seen in COVID-19 is diffuse leukoencephalopathy characterized by symmetrical and multifocal hyperintense lesions in the supratentorial white matter and diffusion restriction in which the juxtacortical white matter is partially preserved. Sometimes the lesions can be hemorrhagic . These findings, possibly due to hypoxia, are seen in critically ill patients receiving long-term ventilator support and associated with increased mortality [117]. Acute necrotizing encephalopathy, which is included in some case reports and considered to be caused by the cytokine storm, is among the neurological complications. This type of involvement is characterized by rim-shaped symmetrical hemorrhagic lesions in the thalamus, medial temporal lobe, and subinsular region [118]. Widespread brainstem involvement may also be seen. Similar findings have been reported in patients with COVID-19 presenting with hypoxic-ischemic encephalopathy or posterior reversible encephalopathy syndrome [112].In patients with anosmia, abnormal hyperintensity and enhancement may be seen in the olfactory bulbus and gyrus rectus, despite imaging findings being completely normal in the early stage of the disease [119]. Lastly, in Guillain–Barré syndrome associated with SARS-CoV-2, enhancement can be seen in the caudal nerve roots [120].Radiological imaging plays a supportive role in the care of patients with COVID-19. Imaging findings are very diverse, and depending on the stage of the disease, their frequency and distribution vary. Comprehensive studies on the imaging of COVID-19 pneumonia emphasize the importance of CT, although other imaging methods can also be used to diagnose and treat COVID-19 infection. Pulmonary magnetic resonance imaging is not typically considered among the first-line modalities in investigating suspected lower respiratory tract infections, but it may be a viable alternative for patient groups where excessive or repeated exposure to ionizing radiation should be avoided.COVID-19 pneumonia progresses with bilateral GGOs, primarily distributed in the peripheral parts of the lower lung lobes and often accompanying consolidations on CT scans. However, these findings are not specific and significantly overlap those related to the other causes of viral pneumonia, and alternative diagnoses should be considered in patients with negative diagnostic tests. The respiratory tract is the primary system affected by COVID-19, but the cardiovascular system, GIS, hepatobiliary system, GUS, and the nervous system can also show signs and symptoms associated with COVID-19. Therefore, both clinicians and radiologists need to know the extrapulmonary findings and imaging features of COVID-19 to help diagnose and manage the possible complications of the disease in an early stage. They should also be familiar with CT findings that can be seen in patients with COVID-19 since the disease can be incidentally detected during imaging performed with other indications.