A. Nature of Liver Regeneration. I. Phenomenology. 1. Key Questions. This chapter focuses on three central questions: 1) What is the evidence that glucagon ...
CHAPTER 21
Glucagon and Liver Regeneration H. L. LEFFERT, K. S. KOCH, P.J. LAD, B. DE HEMPTINNE, and H. SKELLY
A. Nature of Liver Regeneration I. Phenomenology 1. Key Questions This chapter focuses on three central questions: 1) What is the evidence that glucagon controls liver regeneration? 2) What growth control parameters does glucagon affect? 3) How does glucagon work? Because glucagon acts together with other endogenous hepatoproliferogens, and because the proliferative response is complex, a brief review is given to put these questions into their proper perspective. 2. Background Liver regeneration is a compensatory response to liver ablation or injury (SLATER 1978). It consists of an explosive but limited proliferation ofhepatocytes and nonparenchymal cells. For example, using adult rats 1 as experimental models (BUCHER and MALT 1971), the near-maximal stimulus of70% hepatectomy converts the Go remnant, wherein ~ 5/10,000 and 5/100,000 hepatocytes synthesize DNA semiconservatively (S-phase) and divide (M-phase), respectively, - into a tissue whose labeled nuclear and labeled mitotic indices (L.I. and L.M.I.) are 600-fold higher within 24 h (Table 1 defines these and other "growth control" terms). In other words, regeneration is so fast that a normal intact liver would take almost 600 days to produce the numbers of new hepatocytes formed by regenerating liver within 24-48 h. Liver cells near portal tracts proliferate sooner than midzonal cells. The latter, in turn, proliferate before centrilobular hepatocytes (GRISHAM 1962). These observations, together with the results of cross-circulation (SAKAI 1970), heterotopic autograft (LEONG et al. 1964), and portal blood flow reversal studies (SIGEL et al. 1968) were for many years the basis of the controversial notion that blood-borne factors, especially those of portal origin, control liver regeneration (FISHER et al. 1971). During the last decade, unequivocal evidence has accumulated to support this concept (for review, see LEFFERT et al. 1979). One of the controlling factors is glucagon (molecular weight 3,600 daltons), which originates from and is secreted by pancreatic islet A-cells. Glucagon's known and postulated role as a hepatoproliferogen is the subject of this chapter. 1 Unless noted otherwise, all experimental work discussed in this chapter refers to studies with rats and rat hepatocytes P. J. Lefèbvre (ed.), Glucagon I © Springer-Verlag Berlin Heidelberg 1983
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H. L. LEFFERT et at.
Table 1. Growth control terminology Term
Definition a
G
A temporal gap in the "cell cycle"
Go
Quiescent time interval, when cellular progression toward S is at a low or minimal value
Radioactive dT labeling c; kinetic studies C
Time interval between M (or Go) and S; its duration varies, depending on cell type and environmental conditions
Radioactive dT labeling; kinetic studies; DNA dye binding
Phase of semiconservative nuclear DNA replication, when genetic material is duplicated, that follows G1 and precedes M
Radioactive dT labeling; kinetic studies; DNA dye binding
Time interval between Sand M
Radioactive dT labeling; kinetic studies; DNA dye binding
M
Phase when chromosomes form and segregate into daughter cells (mitosis); it follows G2 and precedes Go or G1
Cytochemical staining
L.I.
Nuclear labeling index: the fraction of (mononucleated) cells whose nuclei contain radioactive thymidine ([3HJdT) because they are in S
Radioactive dT pulse labeling; radioautography
M.l.
Mitotic index: the fraction of cells in M
Cytochemical staining
L.M.l.
Labeled mitotic index: the fraction of cells in M that previously passed through S
Radioactive dT pulse labeling; radioautography; cytochemical staining
Duration oftime between the application of a known proliferogenic stimulus and a detectable L.I. change d
Radioactive dT pulse labeling; radioautography; kinetic studies
Fraction of cells entering S-phase per unit timed
Radioactive dT pulse labeling; radioautography; kinetic studies
Duration of time between the application of a known proliferogenic stimulus and a detectable L.M.I. changed
Radioactive dT pulse labeling; radioautography; cytochemical staining; kinetic studies
Fraction of cells entering M-phase per unit timed
Radioactive dT pulse labeling; radioautography; cytochemical staining; kinetic studies
S
Method of measurement b
See LEFFERT et al. (1982) See KOCH and LEFFERT (1979, 1980) C Both methods are indirect (no specific Go "markers" are known currently) d See Fig. 1 and KOCH and LEFFERT (1980) for more details a
b
3. Kinetic Aspects
To understand the relation between glucagon and hepatic proliferation, some fundamental kinetic properties of regenerative responses following conventional stimuli like 70% hepatectomy will be outlined. The interested reader should consult the excellent reviews of this subject by BUCHER and MALT (1971), and BECKER (1973).
Glucagon and Liver Regeneration
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Hepatocytes
50
::JL.O f-
D
l.
30
eo Old medium .. " Fresh medium _0 Fresh medium insulin. glucagon. EGF
"Nonhepatocytes"
50 +
40
30
C1J
>
:g
20
20
(5 10
10
::J
E
o
12
12
24
36
48
Time (h) Growth transition constants Hepatocytes
"Nonhepatocytes"
..... N.D.
........... ......... ___ ........... ......... ___ ...........
21 11 0.2 1.0 1.0 N.D. 26
......... 22
___ N.D. ...-.. 0.25 ......... 0.7
o--
