Activin A, p15INK4b signaling, and cell competition promote stem/progenitor cell repopulation of livers in aging rats.

BACKGROUND & AIMS: Highly proliferative fetal liver stem/progenitor cells (FLSPCs) repopulate livers of normal recipients by cell competition. We investigated the mechanisms by which FLSPCs repopulate livers of older compared with younger rats. METHODS: Fetal liver cells were transplanted from DPPIV(+) F344 rats into DPPIV(-) rats of different ages (2, 6, 14, or 18 months); liver tissues were analyzed 6 months later. Cultured cells and liver tissues were analyzed by reverse transcription polymerase chain reaction, immunoblot, histochemistry, laser-capture microscopy, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling analyses. RESULTS: We observed 4- to 5-fold increases in liver repopulation when FLSPCs were transplanted into older compared with younger rats. Messenger RNA levels of cyclin-dependent kinase inhibitors increased progressively in livers of older rats; hepatocytes from 20-month-old rats had 6.1-fold higher expression of p15INK4b and were less proliferative in vitro than hepatocytes from 2-month-old rats. Expression of p15INK4b in cultured hepatocytes was up-regulated by activin A, which increased in liver during aging. Activin A inhibited proliferation of adult hepatocytes, whereas FLSPCs were unresponsive because they had reduced expression of activin receptors (eg, ALK-4). In vivo, expanding cell clusters derived from transplanted FLSPCs had lower levels of ALK-4 and p15INK4b and increased levels of Ki-67 compared with the host parenchyma. Liver tissue of older rats had 3-fold more apoptotic cells than that of younger rats. CONCLUSIONS: FLSPCs, resistant to activin A signaling, repopulate livers of older rats; hepatocytes in older rats have less proliferation because of increased activin A and p15INK4b levels and increased apoptosis than younger rats. These factors and cell types might be manipulated to improve liver cell transplantation strategies in patients with liver diseases in which activin A levels are increased.

Glycemic control in critically ill patients before and after institution of an intensive insulin infusion protocol: circadian rhythm and the quality duration calculator.

INTRODUCTION: A circadian rhythm of blood glucose values has been recently reported in critically ill patients, but there are no reports of how this rhythm is altered by a continuous intensive insulin infusion therapy protocol (IIT). We wished to examine the effect of IIT on this rhythm as well as to describe the use of the quality duration calculator (QDC) for the evaluation of glycemic control before and after IIT. METHODS: This was a retrospective multihospital observational study that took place in the medical and surgical intensive care units (ICUs) of 2 tertiary care hospitals. Cohorts of consecutively admitted critically ill patients from 2-year periods before and after institution of an IIT protocol were examined. Laboratory, demographic, and outcome data were extracted from hospital databases. RESULTS: We studied 167,645 blood glucose measurements from 8,327 patients. We observed a circadian rhythm of blood glucose control in the pre-IIT cohort that was greatly attenuated in the post-IIT cohort. The difference between the morning and the average daily blood glucose in the pre-IIT cohort was 3.53 mg/dL (P < .001), and the difference between these values in the post-IIT cohort was 1.10 mg/dL (P = .031). In addition, the circadian nature of hyperglycemia incidence observed in the pre-IIT cohort was not seen in the post-IIT cohort. The amount of time spent in goal glycemic range increased from 23.69% (95% CI 23.01-24.38) in the pre-IIT cohort to 29.67% (95% CI 29.04-30.31) in the post-IIT cohort as estimated by the QDC. The amount of time spent in the hyperglycemic decreased from 20.17% (95% CI 19.33-20.99) in the pre-IIT cohort to 14.80% (95% CI 14.15-15.39) in the post-IIT cohort. CONCLUSIONS: The circadian rhythm of blood glucose control confirmed in our pre-IIT cohort was lost after institution of IIT. The morning blood glucose value appears to be a reasonable surrogate of overall glycemic control in a critically ill population on IIT, although this may vary based on the degree of control achieved. The QDC method is useful for analyzing glycemic control in patients on IIT.

Repopulation of rat liver by fetal hepatoblasts and adult hepatocytes transduced ex vivo with lentiviral vectors.

Recent studies have shown that nondividing primary cells, such as hepatocytes, can be efficiently transduced in vitro by human immunodeficiency virus-based lentivirus vectors. Other studies have reported that, under certain conditions, the liver can be repopulated with transplanted hepatocytes. In the present study, we combined these procedures to develop a model system for ex vivo gene therapy by repopulating rat livers with hepatocytes and hepatoblasts transduced with a lentivirus vector expressing a reporter gene, green fluorescent protein (GFP). Long-term GFP expression in vivo (up to 4 months) was achieved when the transgene was driven by the liver-specific albumin enhancer/promoter but was silenced when the cytomegalovirus (CMV) enhancer/promoter was used. Transplanted cells were massively amplified ( approximately 10 cell doublings) under the influence of retrorsine/partial hepatectomy, and both repopulation and continued transgene expression in individual cells were documented by dual expression of a cell transplantation marker, dipeptidyl peptidase IV (DPPIV), and GFP. In this system, maintenance or expansion of the transplanted cells did not depend on expression of the transgene, establishing that positive selection is not required to maintain transgene expression following multiple divisions of transplanted, lentivirus-transduced hepatic cells. In conclusion, fetal hepatoblasts (liver stem/progenitor cells) can serve as efficient vehicles for ex vivo gene therapy and suggest that liver-based genetic disorders that do not shorten hepatocyte longevity or cause liver damage, such as phenylketonuria, hyperbilirubinemias, familial hypercholesterolemia, primary oxalosis, and factor IX deficiency, among others, might be amenable to treatment by this approach.