Double staining of β-galactosidase with fibrosis and cancer markers reveals the chronological appearance of senescence in liver carcinogenesis induced by diethylnitrosamine

Pacheco-Rivera, R.; Fattel-Fazenda, Samia; Arellanes‐Robledo, Jaime; Silva-Olivares, Angélica; Alemán-Lazarini, Leticia; Rodríguez‐Segura, M. A.; Pérez‐Carreón, Julio Isael; Villa‐Treviño, Saúl; Shibayama, Mineko; Serrano-Luna, Jesús

doi:10.1016/j.toxlet.2015.11.011

Cited by 22 publications

(17 citation statements)

References 46 publications

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“…Experimental parameters including freezing and fixation methods, as well as the thickness of the slides and the pH of the solution were not consistently described (Supplementary Table 1). 5,7–19 Cryopreservation techniques were not detailed, and the section thickness could vary between 4 and 7 µm when mentioned. Fixation methods varied widely, with 0.2% to 0.5% glutaraldehyde and/or 3% to 4% (para)formaldehyde for 1 min to 4 hr, before or after freezing, and at room temperature (RT) or 4C.…”

Section: Introductionmentioning

confidence: 99%

An Optimized Protocol for Histochemical Detection of Senescence-associated Beta-galactosidase Activity in Cryopreserved Liver Tissue

Jannone

Rozzi

Najimi

et al. 2020

J Histochem Cytochem.

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Senescence-associated beta-galactosidase (SA-β-gal) activity assay is commonly used to evaluate the increased beta-galactosidase (β-gal) activity in senescent cells related to enhanced lysosomal activity. Although the optimal pH for β-gal is 4.0, this enzymatic activity has been most commonly investigated at a suboptimal pH by using histochemical reaction on fresh tissue material. In the current study, we optimized a SA-β-gal activity histochemistry protocol that can also be applied on cryopreserved hepatic tissue. This protocol was developed on livers obtained from control rats and after bile duct resection (BDR). A significant increase in β-gal liver activity was observed in BDR rats vs controls after 2 hr of staining at physiological pH 4.0 (6.98 ± 1.19% of stained/total area vs 0.38 ± 0.22; p<0.01) and after overnight staining at pH 5.8 (24.09 ± 6.88 vs 0.12 ± 0.08; p<0.01). Although we noticed that β-gal activity staining decreased with cryopreservation time (from 4 to 12 months of storage at −80C; p<0.05), the enhanced staining observed in BDR compared with controls remained detectable up to 12 months after cryopreservation ( p<0.01). In conclusion, we provide an optimized protocol for SA-β-gal activity histochemical detection at physiological pH 4.0 on long-term cryopreserved liver tissue:

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Section: Introductionmentioning

confidence: 99%

An Optimized Protocol for Histochemical Detection of Senescence-associated Beta-galactosidase Activity in Cryopreserved Liver Tissue

Jannone

Rozzi

Najimi

et al. 2020

J Histochem Cytochem.

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“…And DEN is frequently used for making a series of cancer models in laboratory (Y.-J. Chen et al 2016 ; Jayaprakash et al 2015 ; Pacheco-Rivera et al 2016 ). Hence, DEN was conducted in this study as a representative of NAs.…”

Section: Introductionmentioning

confidence: 99%

Developmental Toxicity of Diethylnitrosamine in Zebrafish Embryos/Juveniles Related to Excessive Oxidative Stress

Huang

et al. 2018

Water Air Soil Pollut

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Diethylnitrosamine (DEN) is present in food, water, and daily supplies and is regarded as a toxicant of carcinogenicity. The developmental toxicity of DEN has been rarely reported as yet. In this study, zebrafish were exposed to different concentrations of DEN at 6 h post-fertilization (hpf) to access embryonic toxicity of the compound. The results show that DEN resulted in negative effects of hatching rate, heartbeat, body length, and spontaneous movement. Deformities, including notochord malformation, pericardium edema, embryonic membrane turbidity, tail hypoplasia, yolk sac deformity, and growth retardation, happened during exposure period. Moreover, production of reactive oxygen species (ROS) significantly increased after DEN treatment. Then, alterations of the expression level of oxidative stress-related genes were observed in our results. To our knowledge, this is the first study concerning the effect of DEN on zebrafish. And from the information of our research, we speculated that development toxicity of DEN should be related to the excessive oxidative stress.

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“…This proposal is supported by the presence of replicative and premature senescence in both parenchymal and nonparenchymal liver cells [58,66,67]. Using a liver injury model to mimic sequential HCC progression (from fibrosis to cirrhosis and HCC) [68], we recently confirmed that CS appearance is intrinsic to the initiation of liver injury, as a chronological increase of CS alongside HCC progression was observed [69]. We did not determine ROS levels in this investigation; however, HCC was induced with continuous diethylnitrosamine administration, a hepatotoxic and potent ROS producer [70], suggesting that the chronological appearance of CS was accompanied by persistent ROS induction.…”

Section: Reactive Oxygen Species and Hepatic Diseasesmentioning

confidence: 70%

“…This hypothetical proposal also raises the question of whether the appearance of senescence/ROS is a transitory, sporadic, or specific event of a determined HCC development stage. We have recently shown that, concomitant to the HCC development induced by diethylnitrosamine, there is also a chronological appearance of CS [69]; however, its association with ROS generation and effect on the outcome of multifocal HCC type has not been investigated.…”

Section: Reactive Oxygen Species and Hepatic Diseasesmentioning

confidence: 99%

Is Senescence Important in Hepatic Diseases?

Rivera¹,

Robledo²,

Serrano-Luna³

2017

Senescence - Physiology or Pathology

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Aging is a condition in which a person gradually loses the ability to maintain homeostasis, due to structural alteration or dysfunction. Aging changes biological processes in many organs and tissues. The loss of regenerative capacity is the most dramatic age-associated alteration in the liver. Cellular damage, if not repaired, leads to apoptosis or senescence. The presence of permanent cell cycle arrest, the acquisition of major morphological change, and expression of senescence-associated β-galactosidase (SA-β-gal) are the characteristics of cellular senescence (CS). Interestingly, CS plays a crucial role in aging of both individual organs and the entire organism; consequently, senescent cells accumulate in organs and decline in organ function. Senescent cells have considerable influence on their microenvironment and exert both beneficial and detrimental effects through secretory associated senescent phenotype (SASP) factors. CS has attracted considerable recent interest with recognition of pathways linking aging, malignancy, and insulin resistance and the current focus on therapeutic interventions to extend healthspan. There are major implications for hepatology in the field of fibrosis and cancer, where cellular senescence of hepatocytes, cholangiocytes, stellate cells, and immune cells has been implicated in chronic liver disease progression.

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Double staining of β-galactosidase with fibrosis and cancer markers reveals the chronological appearance of senescence in liver carcinogenesis induced by diethylnitrosamine

Cited by 22 publications

References 46 publications

An Optimized Protocol for Histochemical Detection of Senescence-associated Beta-galactosidase Activity in Cryopreserved Liver Tissue

An Optimized Protocol for Histochemical Detection of Senescence-associated Beta-galactosidase Activity in Cryopreserved Liver Tissue

Developmental Toxicity of Diethylnitrosamine in Zebrafish Embryos/Juveniles Related to Excessive Oxidative Stress

Is Senescence Important in Hepatic Diseases?

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