Insufficient autophagy in prolonged critical illness may cause inadequate removal of damaged proteins and mitochondria. Such incomplete clearance of cellular damage, inflicted by illness and aggravated by hyperglycemia, could explain lack of recovery from organ failure in prolonged critically ill patients. These data open perspectives for therapies that activate autophagy during critical illness.

Context: Responses to critical illness, such as excessive inflammation and hyperglycemia, may trigger detrimental chain reactions that damage cellular proteins and organelles. Such responses to illness contribute to the risk of (nonresolving) multiple organ dysfunction and adverse outcome. Objective: We studied autophagy as a bulk degradation pathway able to remove toxic protein aggregates and damaged organelles and how these are affected by preventing hyperglycemia with insulin during critical illness. Design and Setting: Patients participated in a randomized study, conducted at a university hospital surgical/medical intensive care unit. Patients: We studied adult prolonged critically ill patients vs. controls. Interventions: Tolerating excessive hyperglycemia was compared with intensive insulin therapy targeting normoglycemia. Main Outcome Measures: We quantified (ultra)structural abnormalities and hepatic and skeletal muscle protein levels of key players in autophagy. Results: Morphologically, both liver and muscle revealed an autophagy-deficiency phenotype. Proteins involved in initiation and elongation steps of autophagy were induced 1.3- to 6.5-fold by critical illness (P < 0.01), but mature autophagic vacuole formation was 62% impaired (P = 0.05) and proteins normally degraded by autophagy accumulated up to 97-fold (P < 0.03). Mitophagy markers were unaltered or down-regulated (P = 0.05). Although insulin preserved hepatocytic mitochondrial integrity (P = 0.05), it further reduced the number of autophagic vacuoles by 80% (P = 0.05). Conclusions: Insufficient autophagy in prolonged critical illness may cause inadequate removal of damaged proteins and mitochondria. Such incomplete clearance of cellular damage, inflicted by illness and aggravated by hyperglycemia, could explain lack of recovery from organ failure in prolonged critically ill patients. These data open perspectives for therapies that activate autophagy during critical illness.

Acute kidney injury frequently complicates critical illness and increases mortality; maintaining normoglycemia with insulin has been shown to reduce the incidence of intensive care unit (ICU)-acquired kidney injury. Here we tested the mechanisms by which this intervention might achieve its goal, using a rabbit model of burn-induced prolonged critical illness in which blood glucose and insulin were independently regulated at normal or elevated levels. Hyperglycemia caused elevated plasma creatinine and severe morphological kidney damage that correlated with elevated cortical glucose levels. Renal cortical perfusion and oxygen delivery were lower in hyperglycemic/hyperinsulinemic rabbits, compared to other groups, but this did not explain the elevated creatinine. Mitochondrial respiratory chain activities were severely reduced in the hyperglycemic groups (30-40% residual activity), and were inversely correlated with plasma creatinine and cortical glucose. These activities were much less affected by normoglycemia, and hyperinsulinemia was not directly protective. Mitochondrial damage, evident at day 3, preceded the structural injury evident at 7 days. Our study found that hyperglycemia evoked cellular glucose overload in the kidneys of critically ill rabbits, and this was associated with mitochondrial dysfunction and renal injury. Normoglycemia, independent of insulinemia, protected against this damage.

In a rabbit model of critical illness, HG evokes cellular glucose overload in liver and myocardium inducing mitochondrial dysfunction, which explained the HG-induced organ damage. Maintenance of normoglycemia, but not HI, protects against such mitochondrial and organ damage.

Our observations suggest tissue-dependent attempts of compensatory activation of mitochondrial repair mechanisms during critical illness. Considering the previously observed persistent mitochondrial damage, this activation may be insufficient and contribute to mitochondrial dysfunction. Suppressed activation of fusion/fission when excessive hyperglycemia is prevented with insulin may reflect reduced need for diluting (less) damage during normoglycemia or, alternatively, a suppressive effect of insulin on repair.

IIT in PICU did not evoke neurological damage detectable by circulating S100B and NSE, despite increased incidence of hypoglycemia. Elevated markers in patients with hypoglycemia were not caused by hypoglycemia itself but rather reflect an increased incidence of hypoglycemia in the most severely ill. This hypoglycemia risk appears difficult to capture by classical illness severity scores.

Promiscuous hormone mRNA expression in the pituitary remains poorly understood. We examined by means of RT-PCR and immunostaining whether glycoprotein hormone alpha-subunit (alphaGSU) could be coexpressed with proopiomelanocortin (POMC) in vivo and under pressure of CRH in vitro. Cells coexpressing alphaGSU and POMC mRNA amounted to 2.6% of the cells in ex vivo rat pituitary at birth [postnatal d 1 (P1)], fell to much lower level at P14, and were undetectable in adulthood. In cultured pituitary aggregates of P14 rats, alphaGSU/POMC cells remained scarce but represented up to 6.6% after chronic treatment with CRH but not leukemia inhibitory factor. CRH was less effective in aggregates from P1 and adult rats. The total alphaGSU population ex vivo at P1 was two times smaller than at P14, but in culture it expanded 2.5 times, concomitantly with a reciprocal change in POMC cell abundance. Tpit transcripts were detected in POMC-only and alphaGSU/POMC cells but not in alphaGSU-only cells. Cells coexpressing alphaGSU and POMC mRNA were relatively abundant in P14 chicken pituitary and aggregate cultures, but occurrence was not affected by CRH. Immunostaining showed alphaGSU and POMC colocalization in sporadic cells in intact rat pituitary and CRH-treated cultures at P1 but not at P14 and adult age. The data demonstrate the occurrence of cells coexpressing alphaGSU and POMC in rat and chicken pituitary. The developmental dynamics of this cell population and its response to CRH in vitro in the rat suggest a relationship of these cells with the embryonic branching of the POMC and alphaGSU cell lineages and their mutually opposite developmental course during early postnatal life.

Although the G-protein coupled receptor GPR10 is highly expressed in the anterior pituitary, the action of its ligand prolactin-releasing peptide-31 (PrRP) in this tissue is controversial. The present study examined the acute effect of this peptide on prolactin secretion in perifused rat pituitary reaggregate cell cultures from adult male rats. PrRP readily and dose-dependently stimulated prolactin release at concentrations of 10 and 100 nM, although with a magnitude several times lower than that of thyrotropin-releasing hormone. Surprisingly, PrRP inhibited prolactin release at 0.1 and 1 nm in a pertussis toxin-sensitive manner. Inhibition was markedly favoured by long-term culture. Stimulation and inhibition were differentially affected by the presence of hormones during culture: dexamethasone favoured the inhibitory effect and decreased the magnitude of the stimulatory effect, while oestradiol and triiodothyronine strongly reduced stimulation, as well as inhibition. PrRP, even at 1 nm, counteracted the inhibition of prolactin release by dopamine. There was no effect of PrRP on growth hormone release in aggregates cultured either in the absence or presence of hormones. The present results confirm the prolactin-releasing capacity of PrRP at nanomolar doses and reveal a hitherto unrecognized inhibitory activity of this peptide. Furthermore, dopamine inhibition of prolactin release is antagonized by PrRP, irrespective of the PrRP dose.