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Early detection of drug-induced kidney injury is essential for drug development. In this study, multiple low-dose aristolochic acid (AA) and cisplatin (Cis) injections increased renal mRNA levels of inflammation, fibrosis, and renal tubule injury markers. We applied a serum amyloid A3 (Saa3) promoter-driven luciferase reporter (Saa3 promoter-luc mice) to these two tubulointerstitial nephritis models and performed in vivo bioluminescence imaging to monitor early renal pathologies. The bioluminescent signals from renal tissues with AA or CIS injections were stronger than those from normal kidney tissues obtained from normal mice. To verify whether the visualized bioluminescence signal was specifically generated by the injured kidney, we performed in vivo bioluminescence analysis after opening the stomachs of Saa3 promoter-luc mice, and the Saa3-mediated bioluminescent signal was specifically detected in the injured kidney. This study showed that Saa3 promoter activity is a potent non-invasive indicator for the early detection of drug-induced nephrotoxicity.
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Ácidos Aristolóquicos , Luciferasas , Regiones Promotoras Genéticas , Proteína Amiloide A Sérica , Animales , Proteína Amiloide A Sérica/genética , Proteína Amiloide A Sérica/metabolismo , Ratones , Luciferasas/metabolismo , Luciferasas/genética , Ácidos Aristolóquicos/toxicidad , Genes Reporteros , Cisplatino/toxicidad , Cisplatino/efectos adversos , Mediciones Luminiscentes/métodos , Masculino , Enfermedades Renales/inducido químicamente , Enfermedades Renales/genética , Enfermedades Renales/metabolismo , Enfermedades Renales/patología , Riñón/metabolismo , Riñón/efectos de los fármacos , Riñón/patología , Modelos Animales de Enfermedad , Ratones Endogámicos C57BLRESUMEN
The early detection of diabetic nephropathy (DN) in mice is necessary for the development of drugs and functional foods. The purpose of this study was to identify genes that are significantly upregulated in the early stage of DN progression and develop a novel model to non-invasively monitor disease progression within living animals using in vivo imaging technology. Streptozotocin (STZ) treatment has been widely used as a DN model; however, it also exhibits direct cytotoxicity to the kidneys. As it is important to distinguish between DN-related and STZ-induced nephropathy, in this study, we compared renal responses induced by the diabetic milieu with two types of STZ models: multiple low-dose STZ injections with a high-fat diet and two moderate-dose STZ injections to induce DN. We found 221 genes whose expression was significantly altered during DN development in both models and identified serum amyloid A3 (Saa3) as a candidate gene. Next, we applied the Saa3 promoter-driven luciferase reporter (Saa3-promoter luc mice) to these two STZ models and performed in vivo bioluminescent imaging to monitor the progression of renal pathology. In this study, to further exclude the possibility that the in vivo bioluminescence signal is related to renal cytotoxicity by STZ treatment, we injected insulin into Saa3-promoter luc mice and showed that insulin treatment could downregulate renal inflammatory responses with a decreased signal intensity of in vivo bioluminescence imaging. These results strongly suggest that Saa3 promoter activity is a potent non-invasive indicator that can be used to monitor DN progression and explore therapeutic agents and functional foods.
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Nefropatías Diabéticas/metabolismo , Expresión Génica , Genes Reporteros , Luciferasas/genética , Imagen Molecular , Regiones Promotoras Genéticas , Proteína Amiloide A Sérica/genética , Animales , Biomarcadores , Diabetes Mellitus Experimental , Nefropatías Diabéticas/diagnóstico , Nefropatías Diabéticas/etiología , Técnica del Anticuerpo Fluorescente , Regulación de la Expresión Génica , Mediadores de Inflamación/metabolismo , Mediciones Luminiscentes/métodos , Ratones , Imagen Molecular/métodos , TranscriptomaRESUMEN
Cancer stem cells (CSCs) play a vital role in metastasis, recurrence and chemoresistance in breast cancer. ß-catenin, which is a frequently over activated protein in CSCs, binds to T-cell factor/lymphoid enhancer factor (Tcf/Lef) family transcription factors leading to ectopic expression of Wnt pathway responsive genes necessary for the maintenance and action of CSCs. With the aim of identifying a small molecules that can effectively eliminate CSCs, molecular docking studies were performed against the Tcf/Lef binding hotspot on ß-catenin using a library of 100 natural or synthetic small molecules. Small molecule ligands giving docking energy better than - 7 kcal/mol were further investigated by binding interactions analysis and molecular dynamics (MD) simulations. These compounds were then investigated in vitro, for cytotoxicity against CSCs isolated from MDA-MB-231 triple negative breast cancer cells. Alpha-hederin (AH) was identified as the only compound in the selected library that has cytotoxicity against breast CSCs. AH was further investigated for it's ability to regulate Wnt pathway target genes (Cyclin D1 and CD44)and the tumor suppressor p53by real-time quantitative PCR. Absorption, distribution, metabolism, excretion and toxicity properties of the AH was predicted in silico. AH significantly down regulated the transcription of Cyclin D1 and CD44 while up-regulating the transcription of p53. AH was predicted to have acceptable drug likeness. Although AH is currently known to inhibit the growth of various cancer cells in vitro, present study demonstrated for the first time that it is a potent inhibitor of Wnt/ß-catenin signaling pathway and induce apoptosis in breast CSCs.
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OBJECTIVES: Muscle wasting is a common complication in patients with nonalcoholic fatty liver disease (NAFLD). In this study, we investigated the effect of NAFLD on satellite cell (SC) content and skeletal muscle repair. METHODS: Male CD-1 mice fed a choline-deficient diet for 4 wk were used as an NAFLD model. We performed histologic and mRNA expression analyses, immunochemical staining with single muscle fibers to assess the effect of NAFLD on muscle Pax7+ SCs, and muscle regeneration by intramuscular injection of cardiotoxin. RESULTS: We found that the total number of Pax7+ SCs in the extensor digitorum longus and tibialis anterior muscles of mice with NAFLD was significantly decreased when compared with that in the control group, in which the depletion of the SC pool possibly impaired muscle regeneration, as evidenced by the smaller size of the regenerating myofibers. Importantly, we found that NAFLD significantly impaired the differentiation ability of SCs, as shown by a decreased number of SCs expressing a myogenic marker, MyoD. Finally, this study indicated that molecular mechanisms underlying a decline in SC numbers may be attributed to the upregulation of proinflammatory cytokines (tumor necrosis factor α [TNFα]) and an oxidative stress marker (NADPH oxidase-2 [NOX2\) in mice with NAFLD. CONCLUSIONS: The findings demonstrate that a decrease in SC content in the skeletal muscle is an important factor that contributes to muscle wasting in NAFLD. Thus, preservation of the muscle SC pool is a potential therapeutic strategy to reduce NAFLD-associated muscle wasting.
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Enfermedad del Hígado Graso no Alcohólico , Células Satélite del Músculo Esquelético , Animales , Diferenciación Celular , Modelos Animales de Enfermedad , Humanos , Masculino , Ratones , Desarrollo de Músculos , Músculo Esquelético/metabolismo , Atrofia Muscular/patología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Células Satélite del Músculo Esquelético/metabolismo , Células Satélite del Músculo Esquelético/patologíaRESUMEN
AIMS: Streptozotocin (STZ)-induced diabetic animal models have been widely used to study diabetic myopathy; however, non-specific cytotoxic effects of high-dose STZ have been discussed. The purpose of this study was to compare diabetic myopathy in a high-STZ model with another well-established STZ model with reduced cytotoxicity (high-fat diet (HFD) and low-dose STZ) and to identify mechanistic insights underlying diabetic myopathy in STZ models that can mimic perturbations observed in human patients with diabetic myopathy. MAIN METHODS: Male C57BL6 mice were injected with a single high dose of STZ (180 mg/kg, High-STZ) or were given HFD plus low-dose STZ injection (STZ, 55 mg/kg/day, five consecutive days, HFD/STZ). We characterized diabetic myopathy by histological and immunochemical analyses and conducted gene expression analysis. KEY FINDINGS: The high-STZ model showed a significant reduction in tibialis anterior myofiber size along with decreased satellite cell content and downregulation of inflammation response and collagen gene expression. Interestingly, blood corticosteroid levels were significantly increased in the high-STZ model, which was possibly related to lowered inflammation response-related gene expression. Further analyses using the HFD/STZ model showed downregulation of gene expression related to mitochondrial functions accompanied by a significant decrease in ATP levels in the muscles. SIGNIFICANCE: The high-STZ model is suitable for studies regarding not only severe diabetic myopathy with excessive blood glucose but also negative impact of glucocorticoids on skeletal muscles. In contrast, the HFD/STZ model is characterized by higher immune responses and lower ATP production, which also reflects the pathologies observed in human diabetic patients.
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Adenosina Trifosfato/metabolismo , Complicaciones de la Diabetes/patología , Diabetes Mellitus Experimental/complicaciones , Dieta Alta en Grasa , Enfermedades Musculares/patología , Animales , Glucemia/metabolismo , Complicaciones de la Diabetes/etiología , Complicaciones de la Diabetes/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Enfermedades Musculares/etiología , Enfermedades Musculares/metabolismoRESUMEN
BACKGROUND: Since the index case was reported in China, COVID-19 has led to the death of at least 4 million people globally. Although there are some vaccine cocktails in circulation, the emergence of more virulent variants of SARS-CoV-2 may make the eradication of COVID-19 more difficult. Nsp16 is an S-adenosyl-L-Methionine-dependent methyltransferase that plays an important role in SARS-CoV-2 viral RNA cap formation-a crucial process that confers viral stability and prevents virus detection by cell innate immunity mechanisms. This unique property makes nsp16 a promising molecular target for COVID-19 drug design. Thus, this study aimed to identify potent phytocompounds that can effectively inhibit SARS-CoV-2 nsp16. We performed in silico pharmacokinetic screening and molecular docking studies using 100 phytocompounds-isolated from fourteen Nigerian plants-as ligands and nsp16 (PDB: 6YZ1) as the target. RESULTS: We found that only 59 phytocompounds passed the drug-likeness analysis test. However, after the docking analysis, only six phytocompounds (oxopowelline, andrographolide, deacetylbowdensine, 11, 12-dimethyl sageone, sageone, and quercetin) isolated from four Nigerian plants (Crinum jagus, Andrographis paniculata, Sage plants (Salvia officinalis L.), and Anacardium occidentale) showed good binding affinity with nsp16 at its active site with docking score ranging from - 7.9 to - 8.4 kcal/mol. CONCLUSIONS: Our findings suggest that the six phytocompounds could serve as therapeutic agents to prevent viral survival and replication in cells. However, further studies on the in vitro and in vivo inhibitory activities of these 6 hit phytocompounds against SARS-CoV-2 nsp16 are needed to confirm their efficacy and dose.
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ETHNOPHARMACOLOGICAL RELEVANCE: Disturbed mitochondrial function and energy crisis serve as key mechanisms for the development of liver injury. Hence, targeting cellular mitochondria in liver diseases might serve as a therapeutic option. Tapinanthus globifer (A.Rich.) Tiegh. has been used in traditional medicine in the management of liver disease. However, there is no scientific evidence supporting such use. AIM OF THE STUDY: The current investigation was designed to evaluate the protective role of Tapinanthus globifer treatment on the liver mitochondrial function after the induction of hepatotoxicity by the hepatotoxic agent Fe2+in vitro. MATERIALS AND METHODS: In this study, isolated mitochondria from rats' liver was incubated with Fe2+ (10⯵M) for 1â¯h in the absence or presence of T. globifer (50, 100 and 200⯵g/mL) metanolic extract (MVA). Mitochondrial viability, mitochondrial membrane potential (ΔΨm), mitochondrial swelling (MPTP)., total thiol content, lipid peroxidation (TBARS) and reactive oxygen species (ROS) production were measured. HPLC-DAD was used to identify potential phytochemicals in MVA. RESULTS: (MVA) was able to improve mitochondrial dysfunction induced by Fe2+, by attenuating MTT reduction, increased ΔΨm and mitochondrial swelling. Reduced total thiol and non-protein thiol contents which were associated with increased lipid peroxidation and ROS generation in Fe2+-treated mitochondria were significantly improved by MVA co-treatment. HPLC-DAD analysis revealed the presence of gallic acid, catechin, epigallocatechin, caffeic acid, rutin, glycoside flavonoid and quercetin in MVA that can be responsible for its beneficial effect. CONCLUSION: MVA phyto-compounds enhance mitochondrial redox signaling and possess mitochondrial function improving potential, thereby, providing scientific basis for its use in traditional medicine.