Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 1 Lethality in TNF-a/GalN or Fas agonist treated mice. (a) Kaplan– Meier survival curves in TNF-a/GalN treated B6, NOD, and ICR mice. Mice were challenged intravenously with GalN (0.38 mg/g BW) and rmTNF-a (0.024 mg/g BW). The mice were monitored for up to 12 h, and the percent survival was determined. The overall difference in the survival among ICR, NOD, and B6 mice were highly significant (Po0.0001 by the log-rank test). (b) Kaplan–Meier survival curves in Fas agonist (Jo-2) treated B6 and ICR mice. Mice were challenged with Jo-2 (0.3 mg/g BW). The mice were monitored for up to 4 h, and percent survival was determined. The difference in the survival between ICR and B6 mice were highly significant (P ¼ 0.0003 by the log-rank test).

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques:

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 2 Liver injury in TNF-a/ GalN or Fas agonist treated mice. (a) Three mice per group were challenged intravenously with GalN (0.38 mg/g BW) and rmTNF-a (0.024 mg/g BW). The mean sALT activity measured at the time of autopsy is indicated for each group and is expressed as international units per liter (mean7s.e.). (b) Three mice per group were injected intravenously with Jo-2 (0.3 mg/g BW). The mean sALT activity measured at the time of autopsy is indicated for each group and expressed as international units per liter (mean7s.e.). *Po0.05, **Po0.01 in comparison to the value at the same indicated time point of ICR mice.

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Activity Assay, Injection, Comparison

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 3 Induction of apoptosis in the liver of TNF-a/GalN treated mice. B6 mice (a and c) and ICR mice (b and d) were challenged intravenously with GalN (0.38 mg/g BW) and rmTNF-a (0.024 mg/g BW) and killed 2 h (a and b) and 4 h (c and d) after injection. Liver specimen was subjected to an in situ TUNEL assay to detect apoptotic cells (original magnification, 100).

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Injection, In Situ, TUNEL Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 4 Activation of caspases and Bid after TNF-a/GalN injection. (a) Total liver proteins were isolated from the livers of TNF-a/GalN treated mice at the indicated time points. Proteins (50 mg) were subjected to SDS-PAGE, and were probed with specific antibodies against procaspase-8, -3, Bid, and actin. The results shown are representative of at least three independent experiments. The histogram shows quantitative representations of the levels of pro-caspase-8, Bid, and pro-caspase-3 obtained from laser densitometric analysis. Data are the mean7s.e. from at least three independent experiments. *Po0.05 in comparison to the value at 0 h of B6 mice. (b) The caspase activities were monitored by the cleavage of peptide substrates. For the measurement of caspase-3 (like) and -8 (like) protease activities, the substrates Ac-DEVD-pNA and Ac-IETD-pNA were used, respectively. The mean caspase activities measured at the time of autopsy are indicated for each group and expressed by fold increase to those of control (0 h). The data are the mean7s.e. from at least three independent experiments. *Po0.05 in comparison to the value at the same indicated time point of B6 mice.

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Activation Assay, Injection, Isolation, SDS Page, Comparison, Control

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 6 mRNA and protein expressions of DISC forming molecules, and mRNA expressions of pro- and anti-apoptotic genes. (a) Total hepatic RNA (20 mg) was isolated from livers of TNF-a/GalN treated mice at indicated time points and analyzed for FADD, TRADD, and TNFR1 mRNA expressions by RPA. (b) Total proteins were isolated from livers of TNF-a/GalN treated mice at indicated time points. Proteins (50 mg) were subjected to SDS-PAGE, and were probed with specific antibodies against FADD and actin. The results shown are representative of at least three independent experiments. (c) Total hepatic RNA (20 mg) was isolated from the livers at the indicated time points and then was analyzed for Bcl family mRNA expression by RPA.

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Isolation, SDS Page, Expressing

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 7 NF-kB DNA-binding activity in the liver after TNF-a/GalN treatment in B6 and ICR mice. Nuclear extract was isolated from the livers at the indicated time points and 20 mg of nuclear protein was analyzed by electrophoretic mobility shift assay. The results shown are representative of at least three independent experiments. NS; non-specific band.

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Binding Assay, Activity Assay, Isolation, Electrophoretic Mobility Shift Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Intrinsic resistance to TNF-alpha-induced hepatocyte apoptosis in ICR mice correlates with expression of a short form of c-FLIP.

doi: 10.1038/labinvest.3700544

Figure Lengend Snippet: Figure 8 Differential expression of c-FLIPs. (a) Time course of the c-FLIP expression after TNF-a/GalN injection in B6, NOD, and ICR mice. Total proteins were isolated from the livers of the TNF-a/GalN treated mice at the indicated time points. Proteins (50 mg) were subjected to SDS-PAGE, and were probed with specific antibodies against c-FLIPL/S and actin. The results shown are representative of at least three independent experiments. (b) The relationship between the lethality of mice to TNF-a/GalN-induced liver failure and the expression levels of c-FLIPs among different strains of mice. Mice were challenged intravenously with GalN (0.38 mg/g BW) and rmTNF-a (0.024 mg/g BW; upper). The mice were monitored for up to 12 h, and the percent survival was determined. *Po0.05 and **Po0.0001 in comparison to B6 mice by the w2-test (middle). Total proteins were isolated from the livers of TNF-a/GalN untreated mice from B6, ICR, C3H, NOD, and BALB/c strain. Proteins (50 mg) were subjected to SDS-PAGE, and were probed with specific antibodies against c-FLIPL/S and actin. The results shown are representative of at least three independent experiments (lower). The histogram shows quantitative representations of the levels of c-FLIPS obtained from laser densitometric analysis. Data are the mean7s.e. from at least three independent experiments. *Po0.05 in comparison to the value of B6 mice.

Article Snippet: GalN hydrochloride was purchased from Nacalai Tesque (Tokyo, Japan).

Techniques: Quantitative Proteomics, Expressing, Injection, Isolation, SDS Page, Comparison