Characterisation of enzyme activities in liver parencyhma of diabetic ChREBP-KO mice

Question:

Intraportal pancreatic islet transplantation (IPIT) results in local hyperinsulinism and hyperglycemia in streptozotocin-induced diabetic mice. Metabolic changes emerge in hepatocytes downstream of transplanted islets including an activation of glycolysis and lipogenesis corresponding to glycogen storing clear-cell foci of altered hepatocytes (CCFs). Furthermore, murine CCFs are preneoplastic lesions and progress to hepatocellular adenomas and hepatocellular carcinomas (HCC). To analyse carbohydrate responsive element binding proteins (ChREBP) role in regulating glycolysis, lipogenesis and gluconeogenesis regarding the activities of main metabolic enzymes such as glucose-6-phosphatase (G-6-Pase; gluconeogenesis), glucose-6-phosphate dehydrogenase (G-6-PDH; pentose phosphate pathway) and glucokinase (GK; glycolysis) the transcription factor was knocked out in mice and combined with the IPIT model for hormonally induced hepatocarcinogenesis.

Methods:

Frozen liver tissue specimen of wild type (WT) and ChREBP-KO (KO) mice containing CCFs, HCCs and unaltered liver tissue was cut as serial sections of 10 and 14 µm thickness, mounted on a slide, stained (H&E, PAS, enzyme histochemistry: lead method (G-6-Pase), improved PVA and tetrazolium salt NBT method (G-6-PDH and GK)) and then semiquantitatively compared (18 groups consisting of WT/ChREBP-KO; streptozotocine-induced diabetic/non-diabetic; with/without IPIT; 1, 4, 24 and 48 weeks).

Results:

In non-diabetic WT mice, we observed a regular storage of glycogen from 1 until 48 weeks. Activities of G-6-Pase and G-6-PDH were strongest in acinar zone 1 and faded to zone 3. In contrast, the activity of GK was pronounced in acinar zone 3. Diabetic WT mice revealed a slightly decreased glycogen storage, a strongly increased G-6-Pase and both G-6-PDH and GK were decreased. These patterns are in line with previous observations in rat liver tissue specimen.

In both non-diabetic and diabetic KO mice glycogen storage was higher, activity of G-6-Pase strongly whereas activities of G-6-PDH and GK were only slightly decreased. Moreover, preneoplastic CCFs and HCCs revealed a strong upregulation of GK in WT and KO mice.

Conclusion:

Murine CCFs resemble preneoplastic liver lesions in the rat in terms of carbohydrate related enzyme activity patterns. The altered activities of G-6-Pase, G-6-PDH and GK support the idea of ChREBP as a part in the switch from a glycogenotic to a lipogenic phenotype in preneoplastic lesions.