Preparation of rat liver cells

Seglen, Per Ottar

doi:10.1016/0014-4827(73)90414-x

Cited by 317 publications

(51 citation statements)

References 27 publications

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“…Hepatocytes were obtained according to methods previously described. 36,37 Briefly, fed rats were ether anesthetized, and livers were perfused through the portal vein with liberase RH. Dissociated hepatocytes were then collected in William's culture medium supplemented with 5% fetal calf serum (FCS) and insulin (0.1 IU/mL).…”

Section: In Vitro Experimentsmentioning

confidence: 99%

Protein Kinase Activation by Warm And Cold Hypoxia- Reoxygenation in Primary-Cultured Rat Hepatocytes–JNK1/SAPK1 Involvement in Apoptosis

et al. 2000

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Ischemia-reperfusion procedures induced severe hepatic damages owing to different processes related to hypoxia and reoxygenation (H/R) phases, including the consecutive oxygen free radical (OFR) release. Stress-activated protein kinases (SAPKs) could be activated by extracellular stimuli. The aim of this study was to show whether H/R stress conditions could stimulate these kinases, and especially c-jun-N-terminal kinase (JNK 1 /SAPK 1 ), to reveal a potential role of JNK 1 /SAPK 1 in the control of hepatocyte apoptosis. Primary cultured rat hepatocytes, isolated from other liver cells and blood flow, were subjected to warm and cold hypoxiareoxygenation phases mimicking surgical and transplant conditions. The activation status of SAPKs was evaluated by immunoprecipitation or Western-blotting experiments, whereas apoptosis was assessed by measuring caspase activation and internucleosomal DNA fragmentation in vitro and by TUNEL reaction, in vivo. Hypoxia, and especially hypoxia-reoxygenation, significantly increased JNK 1 /SAPK 1 activation in cultured hepatocytes. Either in warm or cold conditions, OFR scavengers (N-Acetylcystein, Di-Phenyleneiodonium, Deferoxamine) decreased this stimulation. Warm ischemia-reperfusion also led to JNK activation. Hypoxia and especially hypoxia-reoxygenation induced programmed cell death in vivo and in vitro. This last phenomenon was inhibited when hepatocytes were treated with SB 202190, which was described as a potent inhibitor of p38 and JNK activities. Altogether, these results confirmed that JNK 1 /SAPK 1 was activated during the hypoxia-reoxygenation process, and that this activity participated in the onset of the apoptosis program. (HEPATOLOGY 2000;32:1029-1036.)

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Section: In Vitro Experimentsmentioning

confidence: 99%

Protein Kinase Activation by Warm And Cold Hypoxia- Reoxygenation in Primary-Cultured Rat Hepatocytes–JNK1/SAPK1 Involvement in Apoptosis

et al. 2000

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“…Rat hepatocytes were isolated the expression and regulation of the p21 gene during hepatousing the two-step collagenase perfusion procedure. 34 Hepatocytes cyte replication. Because of differences in the expression of were plated at a density of 4 1 10 4 cells/cm 2 in 35-mm tissue culture cyclins in the liver after PH in mice and rats, 27,31 it was relewells coated with 8 mg/cm 2 dried type I collagen 35 in Williams E vant to examine both species.…”

Section: Animalmentioning

confidence: 99%

Regulation of cyclin-dependent kinase inhibitor p21WAF1/Cip1/Sdi1 gene expression in hepatic regeneration

1997

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during proliferation by phosphorylating numerous protein WAF1/Cip1/Sdi1 (p21) is the prototype of a family of targets. 1 The activity of the cyclin/CDK kinase complexes is proteins that inhibit cyclin-dependent kinases and regutightly controlled by several mechanisms, including synthesis late cell cycle progression in eukaryotic cells. In addiof the cyclin and CDK proteins, assembly of the subunits, tion to normal cell cycle progression, p21 is involved in phosphorylation of the CDK, and binding of CDK-inhibitory growth suppression mediated by p53 and transforming proteins (CKIs). 2 Properly coordinated activation of the growth factor b (TGFb), differentiation, and apoptosis.cyclin/CDK complexes leads to orderly progression through To gain insight into the possible involvement of p21 in the normal cell cycle, while aberrant regulation of these comliver cell growth, the expression and regulation of the plexes is associated with malignant transformation. 3 For exp21 gene was evaluated in rodent models of liver regenample, overexpression or amplification of cyclin D1, cyclin E, eration and specimens of human liver diseases. Little p21 mRNA was detected in normal liver tissue. After and CDK4 occurs in numerous human cancers, while deletion growth stimulation in vivo by 70% partial hepatectomy or mutation of the CKI genes p15 INK4B and p16 INK4A also ap-(PH), the p21 transcript was upregulated in a biphasic pears to contribute to carcinogenesis. 4,5 manner, with enhanced expression during G1 phase andThe CKIs play an important role in regulating CDK activfollowing S phase. The induction of p21 after PH was ity and cell cycle progression in response to a wide variety regulated primarily at the post-transcriptional level and of stimuli, including DNA damage, irradiation, nutritional was due to enhanced mRNA stability. Inhibition of pro-depletion, and antimitogenic factors such as TGFb. 6 At least tein synthesis with cycloheximide rapidly induced p21 seven mammalian CKIs have been identified. These include expression, primarily by post-transcriptional stabiliza-the related p15 INK4B , p16 INK4A , p18, and p19 proteins, which tion of the transcript. Hepatic p21 mRNA was also in-appear to specifically inhibit CDK4 and CDK6. 2 The p21, duced by dietary protein deprivation in normal mice. p27 KIP1 , and p57 KIP2 proteins comprise a distinct family of Expression of the p21 gene after PH was similar in p53-CKIs that can regulate CDK1, CDK2, CDK4, CDK6, and posdeficient (p53 0/0) and wild-type mice, but was p53-de-sibly other CDKs. 2,7,8 Of the CKIs, p21 has been most extenpendent following protein deprivation. Primary hepato-sively characterized, after its independent discovery by sevcytes in culture demonstrated increased p21 expression eral groups. 9 The p21 promoter contains p53 binding sites, after treatment with hepatocyte growth factor, TGFb, and p21 transcription can be induced by the p53 tumor-supand activin A. p21 mRNA was upregulated in human pressor protein; p21 is thought to be a key downstream mediliver diseases,...

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“…Rainbow trout hepatocytes were isolated by a modification of the two-step perfusion method (Seglen, 1972). Fish were killed by a blow on the head and a ventral incision was made from the pelvic fins to the gills.…”

Section: Animals Rainbow Trout (Oncorhynchus Mykiss)mentioning

confidence: 99%

DNA adduct formation and 7-ethoxyresorufin O-deethylase induction in primary culture of rainbow trout hepatocytes exposed to benzo[a]pyrene

Masfaraud¹,

Devaux²,

Pfohl‐Leszkowicz³

et al. 1992

Toxicology in Vitro

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Abstract--The formation of DNA adducts, using the 32p-postlabeiling assay, and induction of 7-ethoxyresorufin O-deethylase (EROD) were investigated in a primary culture of rainbow trout hepatocytes exposed to benzo[a]pyrene (B[a]P). Concentrations of 0.1 and 1/zM-B[a]P were shown to induce EROD whereas 10 #M was an inhibitory concentration. DNA adducts were detected for 12 hr to 72 hr after exposure to 1 #M-B[a]P whereas EROD activity was increased 36 hr after treatment. The pattern of adducts was shown to be identical to that obtained after B[a]P treatment of rainbow trout in vivo, as demonstrated by co-chromatography of the adducts. Pre-exposure of hepatocytes for 48 hr to fl-naphthoflavone (~NF) and subsequent 24-hr exposure to 1 ~uM-B[a]P did not lead to increased DNA adduct formation although flNF treatment led to a 3.4-fold induction of EROD activity at the time of B[a]P addition. This study suggests that primary culture of rainbow trout hepatocytes provides a suitable method for studying DNA adduct formation and its modulating factors/n vitro.

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Preparation of rat liver cells

Cited by 317 publications

References 27 publications

Protein Kinase Activation by Warm And Cold Hypoxia- Reoxygenation in Primary-Cultured Rat Hepatocytes–JNK1/SAPK1 Involvement in Apoptosis

Protein Kinase Activation by Warm And Cold Hypoxia- Reoxygenation in Primary-Cultured Rat Hepatocytes–JNK1/SAPK1 Involvement in Apoptosis

Regulation of cyclin-dependent kinase inhibitor p21WAF1/Cip1/Sdi1 gene expression in hepatic regeneration

DNA adduct formation and 7-ethoxyresorufin O-deethylase induction in primary culture of rainbow trout hepatocytes exposed to benzo[a]pyrene

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