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Quantitative analysis of endoplasmic reticulum and cytochrome P-450 in hepatocytes from rats injected with
T Tanaka1, J Watanabe, Y Asaka
1Department of Anatomy, Kansai Medical University, Osaka, Japan.
Abstract:
To examine whether the smooth endoplasmic reticulum (SER) proliferates in hepatocytes from animals treated with methylcholanthrene (MC) frequently used as an inducer for the enzymes of the microsomal mono-oxygenase system, we estimated the area of (smooth and rough) ER per unit cytoplasmic volume by morphometry in periportal, midzonal and perivenular hepatocytes from rats injected with 25 mg/kg MC once a day for 3 days. In addition, immunostaining intensity of major MC-inducible cytochrome P-450 (P-450) forms (1A1/1A2) and total P-450 content in the cytoplasm of hepatocytes in the three zones were measured by microphotometry to ascertain whether P-450 is sufficiently induced in each sublobular zone by the administration. In spite of significant increase in the staining intensity of P-450 1A1/1A2 and amount of total P-450, the proliferation of SER (and RER) did not occur in the three-zone hepatocytes from rats injected with MC. In perivenular hepatocytes, constitutive forms of P-450 other than 1A1/1A2 decreased (to 10%) instead of marked increase in P-450 1A1/1A2 (about 20 times), while the constitutive forms decreased to 50% in midzonal hepatocytes and remained unchanged in periportal hepatocytes after MC administration. In addition, the present results show divergence between biochemical and immumohistochemical results previously reported on MC-inducible P-450 after MC administration to be due primarily to a curvilinear relationship between content and intensity.
Insights
Methylcholanthrene (MC) treatment in rats did not induce smooth endoplasmic reticulum (SER) proliferation in hepatocytes, despite increasing cytochrome P-450 enzyme levels. This suggests SER proliferation is not a necessary response to MC-induced P-450 enzyme activity.
Area of Science:
- Hepatocyte biology
- Drug metabolism
- Endoplasmic reticulum function
Background:
- Methylcholanthrene (MC) is a known inducer of microsomal mono-oxygenase system enzymes.
- The smooth endoplasmic reticulum (SER) is involved in various metabolic processes, including detoxification.
- Hepatocytes exhibit zonal heterogeneity in enzyme expression and function.
Purpose of the Study:
- To investigate whether methylcholanthrene (MC) treatment leads to smooth endoplasmic reticulum (SER) proliferation in rat hepatocytes.
- To assess the zonal distribution of SER and rough endoplasmic reticulum (RER) in hepatocytes after MC administration.
- To correlate the induction of cytochrome P-450 (P-450) enzymes with changes in ER volume.
Main Methods:
- Morphometric analysis of SER and RER volume per cytoplasmic volume in periportal, midzonal, and perivenular hepatocytes.
- Administration of methylcholanthrene (MC) at 25 mg/kg/day for 3 days to male rats.
- Immunohistochemical staining and microphotometry to quantify P-450 1A1/1A2 and total P-450 content.
Main Results:
- Despite significant increases in P-450 1A1/1A2 and total P-450 content, SER (and RER) proliferation was not observed in any hepatocyte zone.
- Constitutive P-450 forms decreased in perivenular and midzonal hepatocytes, while remaining unchanged in periportal zones after MC treatment.
- A divergence was noted between biochemical and immunohistochemical results for MC-inducible P-450, possibly due to a curvilinear relationship between content and intensity.
Conclusions:
- Hepatocyte SER proliferation is not a requisite response to methylcholanthrene (MC)-induced cytochrome P-450 enzyme induction.
- MC administration alters the balance of constitutive and inducible P-450 forms in a zone-dependent manner.
- The relationship between P-450 enzyme levels and immunohistochemical intensity may not be linear.