Liver DNA Research Articles

Preclinical efficacy and safety of adeno-associated virus 5 alpha-galactosidase A gene therapy for Fabry disease

We developed a novel adeno-associated virus 5 gene therapy (AAV5-GLA) expressing human alpha-galactosidase A (GLA) under the control of a novel, small and strong, liver-restricted promoter. We assessed the preclinical potential of AAV5-GLA for treating Fabry disease, an X-linked hereditary metabolic disorder resulting from mutations in the gene encoding alpha-galactosidase A that lead to accumulation of the substrates globotriaosylceramide and globotriaosylsphingosine, causing heart, kidney, and central nervous system dysfunction. Effects of intravenously administered AAV5-GLA were evaluated in 1) alpha-galactosidase A-knockout mice aged 7–8 weeks (early in disease) and 20 weeks (nociception phenotype manifestation) and 2) cynomolgus macaques during an 8-week period. In both species, AAV5-GLA was observed as safe, generated detectable vector DNA and mRNA levels in liver, and produced stable enzyme activity in liver and plasma. In mice, dose-dependent transgene enzyme activity, cross-correction (substrate reduction) in kidney and heart, and improved nociception lasted over 6 months. Moreover, after delayed administration when animals displayed the nociception phenotype, target organ enzyme activity was present, and accumulated substrates were reduced. Given the strong, durable expression of active GLA with this promoter and favorable profile of adeno-associated virus-based gene therapy in humans, AAV5-GLA warrants further investigation in clinical trials for Fabry disease.

Histochemical Analysis of DNA and RNA Identified by Methyl Green - Pyronin ‘Y’ Method and Feulgen’s Reaction Staining in Liver of Channa Punctatus

The light microscopy was to identify cells, storage products, tissue deposits, and microbial pathogens. The Assessment of surrounding tissue with special stains may reveal aspects of interest for the tissue. We illustrate the expected staining characteristics of control Channa punctatus liver tissue with Methyl Green -Pyronin ‘Y’ and Feulgen’s Reaction stains. The aim of this study was to characterize the normal structure of the liver of Channa punctatus, a carnivorous freshwater fish found in Hasanparthy Lake, using gross anatomy, histology and histochemistry. Anatomically, the liver presents C-shaped and only two lobes smaller right and bigger left. The gallbladder is located in right lobe and shows elongated shaped. Histological analysis demonstrated that the hepatic parenchyma is made of two hepatocytes plates surrounded by sinusoids. The fish moreover pancreatic tissue was observed in visceral portion of liver, formed by exocrine pancreatic tissue and islet organ, constituting an extrahepatic pancreas. The nucleic acids content in different tissues in the freshwater fish of Channa punctatus. The fish were scarified and the tissues such as liver and Pancreas removed and processed for determination of nucleic acids. The results obtained in the present study may provide a contribution to the knowledge of the characteristics of nucleic acid as parameters of estimation of DNA and RNA levels in Liver of the fish.

The distinct roles of NEIL1 and XPA in limiting aflatoxin B₁-induced mutagenesis in mice.

Dietary exposure to aflatoxin B₁ (AFB₁) is a risk factor for the development of hepatocellular carcinomas (HCCs). Following metabolic activation, AFB₁ reacts with guanines to form covalent DNA adducts, which induce high-frequency G > T transversions. The molecular signature associated with these mutational events aligns with the single base substitution signature 24 (SBS24) in the Catalog of Somatic Mutations in Cancer (COSMIC) database. Deficiencies in either base excision repair (BER) due to the absence of Nei-like DNA glycosylase 1 (NEIL1) or nucleotide excision repair (NER) due to the absence of xeroderma complementation group A protein (XPA) contribute to HCCs in murine models. In the current study, ultra-low error duplex sequencing was used to characterize mutational profiles in liver DNAs of NEIL1-deficient, XPA-deficient, and DNA repair-proficient mice following neonatal injection of 1 mg/kg AFB₁. Analyses of AFB₁-induced mutations showed high cosine similarity to SBS24, regardless of repair proficiency status. The absence of NEIL1 resulted in an approximately 30% increase in the frequency of mutations, with distribution suggesting preferential NEIL1-dependent repair of AFB₁ lesions in open chromatin regions. A trend of increased mutagenesis was also observed in the absence of XPA. Consistent with the role of XPA in transcription-coupled repair, mutational profiles in XPA-deficient mice showed disruption of the transcriptional bias in mutations associated with SBS24. Implications: Our findings define the roles of DNA repair pathways in AFB₁-induced mutagenesis and carcinogenesis in murine models, with these findings having implications in human health for those with BER and NER deficiencies.

Genome-wide DNA methylation sequencing reveals the involvement of ferroptosis in hepatotoxicity induced by dietary exposure to food-grade titanium dioxide

BackgroundFollowing the announcement by the European Food Safety Authority that the food additive titanium dioxide (E 171) is unsafe for human consumption, and the subsequent ban by the European Commission, concerns have intensified over the potential risks E 171 poses to human vital organs. The liver is the main organ for food-grade nanoparticle metabolism. It is increasingly being found that epigenetic changes may play an important role in nanomaterial-induced hepatotoxicity. However, the profound effects of E 171 on the liver, especially at the epigenetic level, remain largely unknown.MethodsMice were exposed orally to human-relevant doses of two types of E 171 mixed in diet for 28 and/or 84 days. Conventional toxicology and global DNA methylation analyses were performed to assess E 171-induced hepatotoxicity and epigenetic changes. Whole genome bisulfite sequencing and further ferroptosis protein detection were used to reveal E 171-induced changes in liver methylation profiles and toxic mechanisms.ResultsExposed to E 171 for 28 and/or 84 days resulted in reduced global DNA methylation and hydroxymethylation in the liver of mice. E 171 exposure for 84 days elicited inflammation and damage in the mouse liver, whereas 28-day exposure did not. Whole-genome DNA methylation sequencing disclosed substantial methylation alterations at the CG and non-CG sites of the liver DNA in mice exposed to E 171 for 84 days. Mechanistic analysis of the DNA methylation alterations indicated that ferroptosis contributed to the liver toxicity induced by E 171. E 171-induced DNA methylation changes triggered NCOA4-mediated ferritinophagy, attenuated the protein levels of GPX4, FTH1, and FTL in the liver, and thereby caused ferroptosis.ConclusionsLong-term oral exposure to E 171 triggers hepatotoxicity and induces methylation changes in both CG and non-CG sites of liver DNA. These epigenetic alterations activate ferroptosis in the liver through NCOA4-mediated ferritinophagy, highlighting the role of DNA methylation and ferroptosis in the potential toxicity caused by E 171 in vivo.

Evaluating the potential impact of sodium-glucose cotransporter-2 inhibitor "canagliflozin" on the hepatic damage triggered by hypertension in rats.

Hypertension (HTN) is a prevalent chronic disease. HTN and liver disease association is extensively noted. Thus, finding a medication that can alleviate HTN and its accompanying liver insult would be promising. This study investigated the potential impacts of canagliflozin "sodium-glucose cotransporter-2 inhibitor" on the liver of the Nω-nitro-L-arginine methyl ester (L-NAME)-induced HTN rat model. Twenty-four adult male rats were divided into four groups; negative control group, canagliflozin group, L-NAME group: 50 mg/kg of L-NAME was injected daily for 5 weeks and L-NAME + canagliflozin group: 1 week after L-NAME injection both L-NAME + canagliflozin (40 mg/kg) were given concomitantly daily for further 4 weeks. Liver functions, serum lipid profile, hepatic oxidative/nitrative stress biomarkers, gene expression of lipogenic enzymes, B-cell lymphoma 2 (Bcl2), and DNA fragmentation, were measured. Besides, hepatic histology and immunohistochemistry of nuclear factor kappa B (NF-κB) and endothelial nitric oxide synthase (eNOS) were assessed. Canagliflozin improved hepatic lipogenesis via the downregulation of fatty acid synthase (FAS) and transcriptional regulatory element binding protein 1c (SREBP1c) genes leading to an improved serum lipid profile. Further, canagliflozin modified the eNOS/inducible nitric oxide synthase (iNOS) pathway and decreased the NF-κB immunoreactivity besides restoring the oxidants-antioxidants balance; increased reduced glutathione concomitant with declined malondialdehyde. This improvement of the liver was mirrored by the significant restoration of liver architecture and confirmed by the preserved liver DNA content and upregulation of the antiapoptotic Bcl2 mRNA level and attenuation of the alanine transaminase, aspartate aminotransferase. In conclusion, canagliflozin is a promising anti-hypertensive and hepatic-supportive medication. RESEARCH HIGHLIGHTS: Canagliflozin's antioxidant, anti-inflammatory, anti-lipogenic, and antiapoptotic characteristics mitigate remote liver compromise caused by hypertension. Canagliflozin can be exploited as a hepatoprotective and antihypertensive medication.

Ciprofloxacin Affect Sperm Quality and Induced Testicular and Hepatic Injury in Male Mice

Ciprofloxacin (CPFx) is one of a second-generation fluoroquinolones antibiotic and is prescribed a lot for curing bacterial infection. The aim of this study was to investigate of impact CPFx on sperm characteristics, liver DNA and histological structure of liver and testes. Twenty-four male mice aged 7-10 weeks were divided into three groups of eight. The first group (control) was given distilled water. The second and third groups received CPFx with doses 500 and 750 mg/kg respectively. All treatments were administrated orally for 14 days. Three untreated females were put with one treated male for mating. At the end of dosing period, mice were killed, testes and liver were removed and weighed. Sperm parameters, testosterone level and liver DNA were measured. Histopathology of liver and testes was carried out. The results showed that CPFx significantly affected liver DNA, sperm motility and morphology as well as it caused histopathological alterations of hepatic and testicular tissues. Moreover, it led to a significant elevation in the percentage of dead fetuses and also deformity fetuses. The results indicate that CPFX has detrimental effects on sperm quality, liver DNA and histological structure of testes and liver; therefore, it should be used with great caution

Influence of Cu L-Histidinate Schiff Base Derivatives on Structural Features of Irradiated Rat's DNA.

A study of rats liver DNA damages under the influence of X-ray radiation at a dose of 6.5 Gy(LD60) was carried out. The radioprotective properties of newly synthesized Cu(II) L-Schiff Histidinate complexes were also studied. The survival of rats was determined over a 30-day period after exposure to X-rays without pretreatment and also after preadministration of Cu(II) L-Histidinate-Schiff base complexes. The structural defects of rat's liver DNA were detected at 3, 7, 14, and 30 days post-irradiation extracted. The results obtained revealed that irradiation with a 6.5Gy dose in the control group degraded the characteristics of rat liver DNA in comparison to healthy DNA. On all investigated experimental days, a decrease in the melting temperature (Tm), a widening of the melting interval (ΔT), and a decrease in hypochromicity (Δh) were observed in the DNA samples of irradiated animals compared to the norm. The rat's pretreatment by Cu(II) L-Histidinate complexes 1 or 24 hours prior to irradiation improved DNA characteristics. Electrophoretic studies of DNA were in good agreement with the melting data. Based on the study results, it can be concluded that Cu(II) L-Histidinate complexes exhibit radioprotective properties under the studied conditions and can protect DNA from damage.

Choline-An Underappreciated Component of a Mother-to-Be's Diet.

The nutritional status of the mother-to-be has a key impact on the proper development of the fetus. Although all nutrients are important for the developing baby, recent research indicates the importance of adequate choline intake during the periconceptional period, pregnancy, and lactation. Choline plays a key role in the biosynthesis of cell membranes, supporting liver function, neurotransmission, brain development, and DNA and histone methylation. Choline participates in the formation of a child's nervous system, supports its cognitive development, and reduces the risk of neural tube defects. The human body is incapable of producing sufficient choline to meet its needs; therefore, it must be obtained from the diet. Current data indicate that most women in their reproductive years do not achieve the recommended daily intake of choline. The presented narrative review indicates the importance of educating mothers-to-be and thereby increasing their awareness of the effects of choline on maternal and child health, which can lead to a more aware and healthy pregnancy and proper child development.

Liver epigenomic signature associated with chronic oxidative stress in a mouse model of glutathione deficiency

Oxidative stress is intimately involved in the pathogenesis of fatty liver disease (FLD). A major factor contributing to oxidative stress is the depletion of the ubiquitous antioxidant glutathione (GSH). Unexpectedly, chronic GSH deficiency renders glutamate-cysteine ligase modifier subunit (Gclm)-null mice protected from fatty liver injuries. Epigenetic regulation serves as an important cellular mechanism in modulating gene expression and disease outcome in FLD, although it is not well understood how systemic redox imbalance modifies the liver epigenome. In the current study, utilizing the Gclm-null mouse model, we aimed to elucidate redox-associated epigenomic changes and their implications in liver stress response. We performed high-throughput array-based DNA methylation profiling (MeDIP array) in 22,327 gene promoter regions (from −1300 bp to +500 bp of the Transcription Start Sites) in the liver and peripheral blood cells. Results from the MeDIP array demonstrate that, although global methylation enrichment in gene promoters did not change, low GSH resulted in prevalent demethylation at the individual promoter level. Such an effect likely attributed to a declined availability of the methyl donor S-adenosyl methionine (SAM) in Gclm-null liver. Functional enrichment analysis of liver target genes is suggestive of a potential role of epigenetic mechanisms in promoting cellular survival and lipid homeostasis in Gclm-null liver. In comparison with the liver tissue, MeDIP array in peripheral blood cells revealed a panel of 19 gene promoters that are candidate circulating biomarkers for hepatic epigenomic changes associated with chronic GSH deficiency. Collectively, our results provided new insights into the in vivo interplay between liver redox state and DNA methylation status. The current study laid the groundwork for future epigenetic/epigenomic investigations in experimental settings or human populations under conditions of liver oxidative stress induced by environmental or dietary challenges.

Early life exposure to vitamin D deficiency impairs molecular mechanisms that regulate liver cholesterol biosynthesis, energy metabolism, inflammation, and detoxification.

Emerging data suggests liver disease may be initiated during development when there is high genome plasticity and the molecular pathways supporting liver function are being developed. Here, we leveraged our Collaborative Cross mouse model of developmental vitamin D deficiency (DVD) to investigate the role of DVD in dysregulating the molecular mechanisms underlying liver disease. We defined the effects on the adult liver transcriptome and metabolome and examined the role of epigenetic dysregulation. Given that the parental origin of the genome (POG) influences response to DVD, we used our established POG model [POG1-(CC011xCC001)F1 and POG2-(CC001xCC011)F1] to identify interindividual differences. We found that DVD altered the adult liver transcriptome, primarily downregulating genes controlling liver development, response to injury/infection (detoxification & inflammation), cholesterol biosynthesis, and energy production. In concordance with these transcriptional changes, we found that DVD decreased liver cell membrane-associated lipids (including cholesterol) and pentose phosphate pathway metabolites. Each POG also exhibited distinct responses. POG1 exhibited almost 2X more differentially expressed genes (DEGs) with effects indicative of increased energy utilization. This included upregulation of lipid and amino acid metabolism genes and increased intermediate lipid and amino acid metabolites, increased energy cofactors, and decreased energy substrates. POG2 exhibited broader downregulation of cholesterol biosynthesis genes with a metabolomics profile indicative of decreased energy utilization. Although DVD primarily caused loss of liver DNA methylation for both POGs, only one epimutation was shared, and POG2 had 6.5X more differentially methylated genes. Differential methylation was detected at DEGs regulating developmental processes such as amino acid transport (POG1) and cell growth & differentiation (e.g., Wnt & cadherin signaling, POG2). These findings implicate a novel role for maternal vitamin D in programming essential offspring liver functions that are dysregulated in liver disease. Importantly, impairment of these processes was not rescued by vitamin D treatment at weaning, suggesting these effects require preventative measures. Substantial differences in POG response to DVD demonstrate that the parental genomic context of exposure determines offspring susceptibility.

Frequencies and spectra of aflatoxin B1-induced mutations in liver genomes of NEIL1-deficient mice as revealed by duplex sequencing.

Increased risk for the development of hepatocellular carcinoma (HCC) is driven by a number of etiological factors including hepatitis viral infection and dietary exposures to foods contaminated with aflatoxin-producing molds. Intracellular metabolic activation of aflatoxin B1 (AFB1) to a reactive epoxide generates highly mutagenic AFB1-Fapy-dG adducts. Previously, we demonstrated that repair of AFB1-Fapy-dG adducts can be initiated by the DNA glycosylase NEIL1 and that male Neil1-/- mice were significantly more susceptible to AFB1-induced HCC relative to wild-type mice. To investigate the mechanisms underlying this enhanced carcinogenesis, WT and Neil1-/- mice were challenged with a single, 4 mg/kg dose of AFB1 and frequencies and spectra of mutations were analyzed in liver DNAs 2.5 months post-injection using duplex sequencing. The analyses of DNAs from AFB1-challenged mice revealed highly elevated mutation frequencies in the nuclear genomes of both males and females, but not the mitochondrial genomes. In both WT and Neil1-/- mice, mutation spectra were highly similar to the AFB1-specific COSMIC signature SBS24. Relative to wild-type, the NEIL1 deficiency increased AFB1-induced mutagenesis with concomitant elevated HCCs in male Neil1-/- mice. Our data establish a critical role of NEIL1 in limiting AFB1-induced mutagenesis and ultimately carcinogenesis.

Genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc vaccine against SARS-CoV-2 in Sprague-Dawley rats and Beagle dogs.

The emergence of coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to a pandemic, prompting rapid vaccine development. Although vaccines are effective, the occurrence of rare adverse events following vaccination highlights the necessity of determining whether the benefits outweigh the risks posed by the infection itself. The recombinant Vesicular Stomatitis Virus (rVSV) platform is a promising vector for vaccines against emerging viruses. However, limited studies have evaluated the genotoxicity and safety pharmacology of this viral vector vaccine, which is crucial to ensure the safety of vaccines developed using this platform. Hence, the present study aimed to assess the genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc COVID-19 vaccine using micronucleus and comet assays, as well as neurobehavioral, body temperature, respiratory, and cardiovascular assessments in Sprague-Dawley rats and beagle dogs. The intramuscular administration of rVSVInd(GML)-mspSGtc at doses up to 1.5 × 109 PFU/animal did not increase the number of bone marrow micronucleated polychromatic erythrocytes or cause liver DNA damage. Additionally, it had no significant impact on neurobehavioral functions in rats and showed marginal temporary changes in body temperature, respiratory rate, heart rate, and electrocardiogram parameters in rats and dogs, all of which resolved within 24h. Overall, following genotoxicity and pharmacological safety assessments, rVSVInd(GML)-mspSGtc displayed no notable systemic adverse effects in rats and dogs, suggesting its potential as a vaccine candidate for human clinical trials.

The ameliorating effect of Rutin on hepatotoxicity and inflammation induced by the daily administration of vortioxetine in rats

BackgroundVortioxetine (VORTX) is a potent and selective type of selective serotonin reuptake inhibitor (SSRI) that is mainly prescribed for treating major depression along with mood disorders as the first drug of choice. Limited previous findings have indicated evidence of liver injury and hepatotoxicity associated with daily VORTX treatment. Rutin (RUT), which is known for its antioxidant properties, has demonstrated several beneficial health actions, including hepatoprotection. Therefore the current study aimed to evaluate and assess the ameliorative effect of RUT against the hepatotoxic actions of daily low and high-dose VORTX administration.MethodsThe experimental design included six groups of rats, each divided equally. Control, rats exposed to RUT (25 mg/kg), rats exposed to VORTX (28 mg/kg), rats exposed to VORTX (28 mg/kg) + RUT (25 mg/kg), rats exposed to VORTX (80 mg/kg), and rats exposed to VORTX (80 mg/kg) + RUT (25 mg/kg). After 30 days from the daily exposure period, assessments were conducted for serum liver enzyme activities, hepatotoxicity biomarkers, liver antioxidant endogenous enzymes, DNA fragmentation, and histopathological studies of liver tissue.ResultsInterestingly, the risk of liver damage and hepatotoxicity related to VORTX was attenuated by the daily co-administration of RUT. Significant improvements were observed among all detected liver functions, oxidative stress, and inflammatory biomarkers including aspartate aminotransferase (AST), alanine transaminase (ALT), lactate dehydrogenase (LDH), albumin, malondialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), glutathione S-transferase (GST), total protein, acid phosphatase, N-Acetyl-/β-glucosaminidase (β-NAG), β-Galactosidase (β-Gal), alpha-fetoprotein (AFP), caspase 3, and cytochrom-C along with histopathological studies, compared to the control and sole RUT group.ConclusionThus, RUT can be considered a potential and effective complementary therapy in preventing hepatotoxicity and liver injury induced by the daily or prolonged administration of VORTX.

Genome-wide analysis of hepatic DNA methylation reveals impact of epigenetic aging on xenobiotic metabolism and transport genes in an aged mouse model

Hepatic xenobiotic metabolism and transport decline with age, while intact xenobiotic metabolism is associated with longevity. However, few studies have examined the genome-wide impact of epigenetic aging on these processes. We used reduced representation bisulfite sequencing (RRBS) to map DNA methylation changes in liver DNA from mice ages 4 and 24 months. We identified several thousand age-associated differentially methylated sites (a-DMS), many of which overlapped genes encoding Phase I and Phase II drug metabolizing enzymes, in addition to ABC and SLC classes of transporters. Notable genes harboring a-DMS were Cyp1a2, Cyp2d9, and Abcc2 that encode orthologs of the human drug metabolizing enzymes CYP1A2 and CYP2D6, and the multidrug resistance protein 2 (MRP2) transporter. Cyp2d9 hypermethylation with age was significantly associated with reduced gene expression, while Abcc2 expression was unchanged with age. Cyp1a2 lost methylation with age while, counterintuitively, its expression also reduced with age. We hypothesized that age-related dysregulation of the hepatic transcriptional machinery caused down-regulation of genes despite age-related hypomethylation. Bioinformatic analysis of hypomethylated a-DMS in our sample found them to be highly enriched for hepatic nuclear factor 4 alpha (HNF4α) binding sites. HNF4α promotes Cyp1a2 expression and is downregulated with age, which could explain the reduction in Cyp1a2 expression. Overall, our study supports the broad impact of epigenetic aging on xenobiotic metabolism and transport. Future work should evaluate the interplay between hepatic nuclear receptor function and epigenetic aging. These results may have implications for studies of longevity and healthy aging.

Gene targeting in adult organs using in vivo cleavable donor plasmids for CRISPR-Cas9 and CRISPR-Cas12a

The CRISPR-Cas system for in vivo genome editing is a powerful tool for gene therapy against several diseases. We have previously developed the pCriMGET_9-12a system, an in vivo cleavable donor plasmid for precise targeted knock-in of exogenous DNA by both Cas9 and Cas12a. Here, we show that the pCriMGET_9-12a system can be applied for in vivo in-frame knock-in of exogenous DNA in adult mouse liver by hydrodynamic delivery of the targeting plasmids. The in vivo cleavable pCriMGET_9-12a donor plasmids significantly increased the knock-in efficiency of both CRISPR-Cas9 and CRISPR-Cas12a in the adult mouse liver compared to uncleavable donor plasmids. This strategy also achieved in-frame reporter gene knock-in without indel mutations. Therefore, in vivo gene targeting using the pCriMGET_9-12a system may contribute to the establishment of safer, more precise, versatile and efficient gene therapy methods in adult organs.

Dietary zinc deficiency increases damage rate and copy number of mitochondrial DNA in the mouse liver

Zinc is an essential trace element involved in many physiological processes, ranging from cellular growth and immune functions to enzymatic activity and gene expression. Inadequate dietary zinc levels can disrupt the functioning of numerous organs, including the liver. Given the pivotal role played by the liver, in-depth understanding of the mechanisms driving disorders in this organ caused by zinc deficiency is of great importance. In the present study, the effects of a four-week low-zinc diet on the integrity and copy number of mitochondrial DNA (mtDNA) in the liver was assessed using a mouse model. The research revealed a significantly elevated damage rate and an increased copy number of mtDNA in the livers of mice subjected to a low-zinc diet when compared to control animals. These findings indicate that a zinc deficiency, by promoting DNA damage in mitochondrial genomes, increases a potential risk of harmful mutations that could compromise ATP production in the liver. The rise in the mtDNA copy number suggests an initial compensatory response to the detrimental effects of the zinc deficiency, which is likely to diminish with a chronic insufficiency of this element. The study confirmed the significant role of mitochondria in the processes leading to liver dysfunction induced by a zinc deficiency. It showed additionally that mtDNA is a very sensitive indicator of the liver's condition that is responsive to environmental changes such as a micronutrient deficiency in the diet.

Abstract 759: Soy protein diet changes metabolic profile in liver of 7,12-dimethylbenz(α)anthracene (DMBA)-induced mammary tumors in obese zucker rats

Abstract Obesity is associated with the risk of certain types of cancers development including breast cancer. Previously, we reported that obesity increased mammary tumor development using obese Zucker rat model. Several studies have shown the health benefit of soy protein consumption. However, the effects of soy protein diet on liver of mammary tumor for oxidative stress and DNA reaction is less known. We used DMBA induced-mammary tumor model to investigate the effects of soy protein diet on pro-oxidative environment and oxidative DNA modification in liver of obese DMBA mammary tumors rats. Female obese zucker rats (n=45) were either fed casein as control (OC; n=20) or soy protein isolate (OS; n=25), received DMBA (65mg/kg BW) at the age 50 days and sacrificed 20 weeks later. We used HPLC-ECD, HPLC-UV and LC-MS techniques for liver metabolic analysis. Comparing liver of OS vs. OC, soy protein 1) decreased the level of Methionine (P<0.02), increased 2) free reduced Homocysteine (P<0.02), 3) S-Adenosylmethionine (SAM, P<0.03), 4) S-Adenosyhomocysteine (SAH, P<0.001) and 5) 5methyl-Cytosine (5mC, P<0.03). Also, soy protein decreased (P<0.003) level of free oxidized Glutathione (GSSG), increased (P<0.02) GSH/GSSG oxidative stress ratio, without any changes a level of free reduced Glutathione (GSH), followed decrease (P<0.04) in liver DNA level of 8-OH-Guanosine (8-OH-G) and level (P<0.02) of 5hmethyl-Cytosine (5hmC). Our results shows soy protein diet capable change liver metabolism: a. Modify a methylation status by increasing levels of SAM (universal methyl group donor) following global liver DNA hypermethylation (5mC), and SAH (methylases inhibitor); b. Alleviate level of oxidative stress in liver by decreasing level of GSSG (not active glutathione), increasing GSH/GSSG ratio (antioxidative capacities), following lower ROS liver DNA damages by decrease levels of 8-OH-G and 5hmC. In conclusion, using obese DMBA-induced mammary tumor Zucker rat model, soy diet making a complex and significantly modification of metabolic conditions in liver affecting both methionine cycle and transsulfuration metabolic pathways. Citation Format: Reza Hakkak, Stepan Melnyk. Soy protein diet changes metabolic profile in liver of 7,12-dimethylbenz(α)anthracene (DMBA)-induced mammary tumors in obese zucker rats [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 759.

Methylation changes of liver DNA during the formation of gallstones.

Aim: To explore the overall methylation changes in liver tissues during the formation of gallstones, as well as the key pathways and genes involved in the process. Methods: Reduced-representation bisulfite sequencing and RNA sequencing were conducted on the liver tissues of mice with gallstones and control normal mice. Results: A total of 8705 differentially methylated regions in CpG and 1410 differentially expressed genes were identified. The joint analysis indicated that aberrant DNA methylation may be associated with dysregulated gene expression in key pathways such as cholesterol metabolism and bile secretion. Conclusion: We propose for the first time that methylation changes in some key pathway genes in liver tissue may be involved in the formation of gallstones.

Fatty liver disease and DNA replication stress

Fatty liver disease and DNA replication stress

In Vivo Anti-Hepatocellular Carcinoma Effects of the Chloroform Root Extract of Clausena excavata Burm.

Liver cancer is the most common cancer among males in Africa. The disease has a poor prognosis and its treatment is associated with toxicity and resistance. For this reason, numerous herbal combinations are being subjected to anticancer screening to circumvent the shortcomings of the conventional anticancer drugs. In the current study, the in vivo anti-cancer effects of the chloroform root extract of the herb, Clausena excavata Burm were investigated. Liver cancer was induced in mice by a single intraperitoneal injection of diethylnitrosamine (DEN) followed by oral administration of the promoter of carcinogenesis, 2-aminoacetyl fluorine that was mixed with the mice feed. The cytotoxicity of the root extract of C. excavata on liver cancer cells was investigated using liver enzyme, histology, DNA fragmentation and caspases assays. Real time qPCR was conducted to evaluate the effect of the extract on apoptotic genes. The findings revealed that the extract of C. excavata significantly decreased the progression of hepatocarcinogenesis and the toxicity-induced production of the liver enzymes, alanine and aspartate aminotransferases. The histological analyses of the liver tissues revealed evidence of apoptotic cell death. The extract also provoked significant (p < .05) expressions of caspase 9 protein and gene as well as other apoptotic genes (P53, P27, Apaf-1, cytochrome C, bax and bid). Therefore, we postulate that the chloroform root extract of C. excavata induces apoptosis of liver cancer in mice.