Hepatic Toxicity
UNC School of Public Health
Friday - Nov. 3 Introduction to the liver
Gary Boorman, NIEHS
Monday - Nov. 6 Molecular aspects of liver injury Dr.
Robert Sills, NIEHS
Wednesday - Nov. 8 Biochemistry of hepatic injury
Dr. LeCluyse, CellzDirect, Inc.
Friday Nov. 10 Acute Responses to Hepatic Injury
Dr. Rich Miller GSK
Monday, Nov 13 Chronic effects of hepatic injury
Dr. Amy Brix, EPL
Why is the Liver
so Important in Toxicology?
Hepatotoxicity is the major reason for
rejecting new drugs in clinical trials
and withdrawal of drugs already in use
Major metabolic organ
Hepatotoxicity is quite common
Cirrhosis - One of top ten causes of death
Model for cancer mechanisms
Liver in Toxicogenomics
The liver is currently the major tissue for
Toxicogenomics
> 6,200 references on gene expression and liver in
last three years
Easily accessible large parenchymal organ
Model for defining toxic responses
Gene changes reflect systemic responses
Lecture Outline
Role of the liver
Anatomy of the liver
Organization of the liver
Cells of the liver
Rodent diseases affecting liver
Liver is Complex Organ
Largest organ in body
1500 g in humans (2% of BW)
15 g in male rat (4% of BW)
1.5 g in male mouse (6% of BW)
25% of cardiac output
2 L/Minute in 70 kg Human
Source of most plasma proteins
Interface between food and energy needs
Largest source of fixed macrophages
Marked circadian rhythm
Role of the Liver
Processes, dietary proteins, carbohydrates, lipids
Stores and releases energy
Exports Glucose from glycogen
Exports Acetoacetate from fatty acids
Detoxification
Endogenous & Exogenous compounds
Oxidation and reduction
Conjugation and hydrolysis
Important role in vitamins
Active synthesis of some forms of B vitamins
Proteins for transport of vitamins
Retinoid storage and metabolism
Role of the Liver
Acute Phase Response
Transient increase or decrease in plasma proteins
Systemic response to local injury
Phagocytosis of particulates
Critical location with blood flow from GI tract
Central role in cholesterol homeostasis
Critical for iron, zinc and copper
metabolism
Maronpot, Pathology of the mouse. 1999
Maronpot, Pathology of the mouse. 1999
Lobe Differences with
Acetaminophen
50, 150, 1500, and 2000 mg/kg
For each rat evaluated both
left (L) and median (M) lobe
Yellow highlights dose
Pink highlights Left Lobe
Acute Phase Proteins show
remarkable Lobe dichotomy
Ttr = Transthyretin
Fgb = Fibrinogen, beta
Ahsg = alpha-2-HS glycoprotein
Tf = transferrin
Significantly DOWN regulated Median (M)
lobe
Significantly UP regulated Left (L) lobe
WHY?
Lobe variability
Gene expression
Copper distribution (R > L)
Cancer (R > L)
Cirrhosis of right lobe
Hypertrophy of left lobe
Cirrhosis of left lobe
No Hypertrophy of right lobe
Why are there lobe
differences?
Umbilical blood flows to left lobe (Fetus)
Right and median lobes receive portal blood
Left better oxygenated during development
Left more directly exposed to maternal toxins
Differential portal flow to lobes (Adult)
Blood from stomach to left lobe
Blood flow from colon to right lobe
Colonic cancer metastatic more to right lobe
Blood flow from spleen to left lobe
Contains hepatic growth factors
May explain cirrhosis and hypertrophy
Organization of Liver
Hepatic plates with bile canicular system
Dual blood supply
75% portal vein (low in oxygen)
25% hepatic artery (high in oxygen)
Blood collects in hepatic vein
Tissues and Organs:a text of scanning electron
microscopy, Kessel, RG and Kardon,RH, 1979
Approximately 1 million classic lobules per liver

Rappaport Unit
Zone 1 (central or periportal)
Largest hepatocytes
More mitochondria
More granular ER
Glycogen metabolism
Glucuronidation of xenobiotics
Formation of plasma proteins
Best oxygenated
Highest concentration of bile salts
More Kupffer cells
Portal “triad” (zone 1)
hepatic artery (1-2)
portal vein (1)
bile duct (1-2)
lymphatics
nerves
connective tissue-collagen type I
Pathology of the Liver, MacSween RNM et al, 2002
Glucose-6-phosphatase
Teutsch, et al. 1999. Hepatology 29:494-505
Allyl Alcohol-Induced
Necrosis in Zone 1
PP
THV
THV
PP
Rappaport Unit
Zone 3 (More peripheral - Terminal Hepatic
Venule) (most commonly Centrilobular)
Smaller hepatocytes
Fewer mitochondria
More agranular ER
Fat and pigment storage
Reductase reactions
Enzyme induction may occur
More susceptible to anoxia
Acetaminophen Necrosis
in Zone 3
3
THV
2
1
Sometimes lesions are patchy involving more than a lobule
Rat caudate lobe with resin in the portal vein
demonstrating three dimensional units
Bhunchet and Wake, Hepatology 27:481-487, 1998-487, 1998
Vascular damage following acetaminophen (MRI image)
Malarkey, Ryan, Johnson, Maronpot, 2004
Temporal Aspects are also
important in the liver
Gene Expression for Metallothionein 1a
varies by time during the 24 hour day
CT times; light 0 - 12: Dark 12-24
Phase I enzymes that vary
by age in the rat
Hepatic CYP expression
varies with age of rat
What might this mean for a study where rats are exposed 2 years ?
Hepatic dimensions
 Liver is a heterogeneously complex organ
Space
Variation by lobe
 Zones within lobes
Three-dimensional parenchymal units
Time
Variation by time of day
Variation by age
Heterogeneity of liver
 Epithelial
Cells
Hepatocytes
Cholangiocyes
 Mesenchymal
cells
Kupffer cells
Endothelial cells
Stellate cells
 Other
Hepatic components
Smooth muscle cells (blood vessels)
Mesothelia (capsule)
Nerves (unmyelinated)
Neuroendocrine cells
Hematopoeitic cells
Extracellular matrix
 5-10% of liver is collagen
Heterogeneity of liver
 Hepatocytes
(60%)
 Biliary epithelium (3-5%)
 Endothelia (20%)
 sinusoids
 blood vessels (arteries and veins)
 lymphatics
 Kupffer
cells (15%)
 Hepatic stellate cells (5 -%)
 Lymphocytes (Pit cells)
5 major players
Hepatocytes
80% of mass; 60% of cell numbers
 SER and RER (15% of cell volume)
 Free ribosomes
 Golgi complex
 Lysosomes
(~ 30 per cell)
 Peroxisomes / microbodies (~ 500 per cell)
 Mitochondria (1000 per cell)
 Cytoskeleton (microfilaments, intermediate
filaments, microtubules)
 Glycogen
 Produces bile (~ 15 ml per kg per day)

Lobular Heterogeneity: The Streaming
Liver
Pathology of the Liver, MacSween RNM et al, 2002
Gene expression varies along the hepatic plate
What might happen to Glutamine synthetase (GS)
Transcript levels with zone 3 necrosis?
Endothelial Cells
20% of liver cells, 3.3% of protein content
Discontinuous individual cells/fenestrated
Sieve plates - clustered fenestrate
Direct access of blood to hepatocytes
Gives rise to vascular tumors
Vinyl chloride hemangiosarcomas - human
carcinogen
Sinusoidal
endothelial
cells
Fenestrations
Tissues and
Organs: a text of
scanning electron
microscopy,
Kessel, RG and
Kardon,RH, 1979
Pathology of the Liver, MacSween RNM et al,
2002
Kupffer Cells
15% of liver cell population, 2.5% of liver protein
Precursors arise from circulating monocytes
Major component of fixed macrophage system
Ingest particles
May contribute to liver disease
Mediators of inflammation (TNF-alpha)
Liver showing hepatocytes (H), Kupffer cells (KC),
endothelial cells (EC) and stellate cells (SC)
Stellate Cells
5% - 8% of all parenchymal cells
Vitamin A storage and metabolism
Significant source of collagen, hepatic fibrosis
Major player in hepatic regeneration
Control microvascular tone
Ito cell tumors in mice
Hepatic Stellate Cells (HSC)



Periportal (PP)
small HSCs
 perisinusoidal processes
 small volume of lipid droplets
Midzone
 elongated
 large volume of lipid
 intense desmin
Central vein (CV)
 elongated & IC processes
 vitamin A and desmin reduced
Tissues and
Organs: a text of
scanning electron
microscopy,
Kessel, RG and
Kardon,RH, 1979
Hepatic Stellate cells wrap around Endothelial Cells
Biliary cells
3% to 5% of liver cell population
Form approximately 2 km of tubules
Tight junctions isolate lumen
Modifies bile
Active in secretion and absorption
Effective communicator with other cells
Contains numerous transporters
Prevalence of Rodent
Pathogens that affect Liver
Helicobacter Species
31%
Epidemic Diarrhea of Infant Mice
EDIM Virus
15%
Mouse Hepatitis Virus
Rat Parvo Virus
14%
6%
Dr. Lila Riley (University of Missouri Diagnostic
Laboratory)
Source of Rodent
Pathogens
Transfer of transgenic mice
Sharing of biological specimens
Other animals in colony
Feral animals
Animal care personnel & visitors
Dr. Lila Riley (University of Missouri Diagnostic
Laboratory)
Acute Hepatotoxicity
Dr. Sills (This Monday)
Molecular biology of hepatic injury
Dr. Sills will assume that you know hepatic
lobules before lecture….
Liver structure is also critical for Dr.
Millers lecture the following Friday
Thanks to Dave Malarkey for several new slides