Hormonal Regulation of Phosphoenolpyruvate Carboxykinase in Cultured Foetal Hepatocytes from the Rat (original) (raw)
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European Journal of Biochemistry
The involvement of glucocorticoids and insulin in the induction of cytosolic Ppyruvate carboxykinase was studied in cultured hepatocytes derived from 19-day foetal rat liver. Dexamethasone did not increase Ppyruvate carboxykinase activity, but had a synergistic effect on the induction of the enzyme by N 6 , 0 2'-dibutyryladenosine 3',5'-monophosphate (BbcAMP). Insulin partially inhibited the induction of Ppyruvate carboxykinase by glucagon, epinephrine or BtZcAMP. The inhibitory effect of insulin was dose-dependent : significant inhibition was obtained with 0.1 nM insulin. However, the effect of insulin was independent of the dose of glucagon or epinephrine added to the culture medium.
In Vitro Cellular & Developmental Biology, 1985
A completely defined medium (EHM-1), which reflects the amino acid composition of fetal rat serum and contains albumin as the sole proteinaceous compound, allows the accumulation of carbamoylphosphate synthetase and phosphoenolpyruvate carboxykinase in the presence of dexamethasone, dibutyryl cyclic AMP, and triiodothyronine to approximately twice the level attained in a standard culture medium (RPMI 1640) supplemented with 10% fetal bovine serum (and hormones). Using the EHM-1 medium we could show that the capacity of hepatocytes to synthesize phosphoenolpyruvate carboxykinase in the presence of hormones is manifest as soon as the cells differentiate from the embryonic foregut {embryonic Day 11). Furthermore we could show that embryonic hepatocytes can become binuclear or polyploid when cultured in the presence of thyroid hormone.
European Journal of Biochemistry, 1984
The primary appearance of phosphoenolpyruvate (P-pyruvate) carboxykinase RNA transcripts in fetal liver was induced by a number of different stimulii. This may occur as rapidly as an hour after injection in utero of N 6 , 0 2 'dibutyryl-adenosine 3',5'-monophosphate (Bt,cAMP) to fetuses, suggesting that all stimulii predominantly affect activation of the P-pyruvate carboxykinase gene. Bt,cAMP treatment induces the appearance of the enzyme RNA transcripts, predominantly of the mature type in the cytoplasm. However, insulin deficiency by streptozotocin treatment causes the appearance of large-size as well as mature mRNA in the nucleus, in addition to the appearance of P-pyruvate carboxykinase mRNA in the cytoplasm. Insulin treatment of such diabetic fetuses, prior to causing the disappearance of P-pyruvate carboxykinase mRNA, reduces nuclear transcripts but increases the abundance of mature cytoplasmic enzyme mRNA. Bt,cAMP treatment of insulin-deficient fetuses causes an additive effect, increasing the abundance of not only the mature but the large P-pyruvate carboxykinase RNA transcripts as well.
Histochemistry, 1990
The postnatal appearance of phosphoenolpyruvate carboxykinase activity (PEPCK) and acinar heterotopy was investigated in newborn rats aged 2 h, 12 h, 24 h and 3 days, as well as in juvenile rats aged 25 days. The livers showed an almost homogeneous distribution of activity along the sinusoidal length at the beginning of extrauterine life where energy needs are greatest. Compared to rats aged 2 h, the PEPCK activity was higher in the livers from rats aged 12 h. The increase in activity was most pronounced in the intermediary zone. After 24 h of extrauterine life the activity decreased again creating a homogeneous acinar activity pattern. By day 3 activity had increased in the periportal zone, while decreasing in the perivenous zone, resulting in a periportal to perivenous gradient. By day 25 total activity had reached highest values both in males and females, due to a relatively high perivenous activity. The more prominent acinar gradient corresponded approximately to the one seen in adult animals.
Biochemistry, 1975
T he activity of hepatic phosphoenolpyruvate carboxyki-nase (GTP) (EC 4.1.1.32)' is regulated by glucagon, acting via CAMP, by insulin, and by glucocorticoids (Shrago et al., 1963; Foster et al., 1966; Reshef et al., 1969; Reshef and Hanson, 1972). Dibutyryl cAMP has been ...
European Journal of …, 1978
The CCAAT/enhancer-binding protein ␣ (C/EBP) is a transcription factor that trans-activates a number of metabolically important genes. Previous work has demonstrated that C/EBP␣ and C/EBP have the potential to mediate the cAMP responsiveness of phosphoenolpyruvate carboxykinase (PEPCK) in liver cells. However, these studies used GAL4 fusion proteins and artificial promoter-reporter gene vectors in transfection experiments; as a result, these studies only indicated that both isoforms had the potential to mediate the hormonal response and not which isoform actually participated in vivo. To address this issue, we produced hepatoma cell lines that stably expressed either a dominant negative inhibitor or antisense RNA for these two main liver C/EBP isoforms. Inhibition of all C/EBP isoforms via expression of the dominant negative protein eliminated cAMP responsiveness, and reduced glucocorticoid responsiveness, of the endogenous PEPCK gene in hepatoma cells. Antisense directed against C/EBP␣ mRNA, which reduced C/EBP␣ protein levels by nearly 80%, also significantly reduced the cAMP responsiveness of the endogenous PEPCK promoter, whereas antisense directed against C/EBP was without effect. There was no major alteration in cAMP signaling in the C/EBP␣ antisense cells, as cAMP induction of the C/EBP gene was similar to that in wild-type H4IIE cells. These data suggest that the ␣-isoform of C/EBP is specifically utilized for mediating the cAMP responsiveness of the PEPCK gene.
Journal of Histochemistry & Cytochemistry, 1990
We used immunohistochemical techniques to analyze the cell distribution of phosphoenolpyruvate carboxykinase (PEPCK) in adult and developing mouse tissues. PEPCK immunoreactivity was detected in many tissues, including some that had not been previously reported to contain PEPCK enzyme activity (bladder, stomach, ovary, vagina, parotid gland, submaxillary gland, and eye). In some multicellular tissues, PEPCK immunoreactivity was observed in multiple cell types. Several tissues (spleen, thyroid, and submaxillary gland) contained no detectable PEPCK immunoreactivity. During development, PEPCK immunoreactivity was associated with the developing nervous system and somites in 15-day embryos. At prenatal day 18, PEPCK immunoreactivity was detected only in the nervous system. At prenatal day 20, PEPCK immunoreactivity was observed in many of the tissues that contain PEPCK in the adult, with the exception of liver, lung, and stomach. PEPCK immunoreactivity was detected in liver at postnatal ...
Pyruvate kinase isoenzyme transitions in cultures of fetal rat hepatocytes
Cell differentiation and development : the official journal of the International Society of Developmental Biologists, 1988
Changes in the expression of two isoenzymic forms of pyruvate kinase in fetal hepatocyte cultures derived from 15- and 19-day gestation rats are studied by immunocytochemical localization of the respective antigens. Initially, in cultures established from 15-day gestation rats only the 'embryonic' form of the enzyme (M2-PK) is detected in all cells. Cells which stain positively for the liver specific form of the enzyme (L-PK) are not observed. After 2 days' culture, a significant number of cells have become positive for L-PK. All the positive cells have a morphology which is typical of liver parenchymal cells. However, the majority of parenchymal cells remain negative for L-PK while retaining M2-PK. In contrast, all cells which display a fibroblastic morphology, as well as clear epithelial cells are M2-PK positive, but L-PK negative. In 5-day-old cultures, all hepatocytes have become L-PK positive. Hepatocytes derived from 19-day gestation rat liver stain positively for ...