The authors report the rare association of Prader-Willi syndrome and short-chain acyl-CoA dehydrogenase gene variant. Prader-Willi syndrome, associated with paternal chromosome 15q11-q13 silencing, is characterized by neonatal/infantile hypotonia, growth failure, and neurodevelopmental delays in the first 1 to 2 years of life, typically followed by hyperphagia and obesity. Short-chain acyl-CoA dehydrogenase gene variant, with 625 G-to-A and 511 C-to-T changes, impairs C4-C6 fatty acid metabolism and variably causes neonatal/infantile hypotonia with developmental delays. The authors’ patient continues to exhibit the classic severe growth failure of early infancy Prader-Willi syndrome at 40 months. Extensive laboratory investigations indicate that the short-chain acyl-CoA dehydrogenase gene variant is likely preventing or delaying the normal expression of the Prader-Willi syndrome phenotype.

Prader-Willi syndrome (PWS) is a complex genetic disorder localized to chromosome 15 and is considered the most common genetic cause of the development of life-threatening obesity. Although some morbidities associated with PWS, including respiratory disturbance/hypoventilation, diabetes, and stroke, are commonly seen in obesity, others such as osteoporosis, growth hormone deficiency, and hypogonadism, and also altered pain threshold and inability to vomit, pose unique issues. Various bariatric procedures have been used to cause gastric stasis, decrease gastric volume, and induce malabsorption, with poor results in PWS patients in comparison with normal obese individuals.

An explosion of new research is vastly changing scientists’ understanding of diabetes and giving new clues about how to attack it.

The fifth leading killer of Americans, with 73,000 deaths a year, diabetes is a disease in which the body’s failure to regulate glucose, or blood sugar, can lead to serious and even fatal complications. Until very recently, the regulation of glucose - how much sugar is present in a person’s blood, how much is taken up by cells for fuel, and how much is released from energy stores - was regarded as a conversation between a few key players: the pancreas, the liver, muscle and fat.

Now, however, the party is proving to be much louder and more complex than anyone had shown before.

New research suggests that a hormone from the skeleton, of all places, may influence how the body handles sugar. Mounting evidence also demonstrates that signals from the immune system, the brain and the gut play critical roles in controlling glucose and lipid metabolism. (The findings are mainly relevant to Type 2 diabetes, the more common kind, which comes on in adulthood.)

Focusing on the cross-talk between more different organs, cells and molecules represents a “very important change in our paradigm” for understanding how the body handles glucose, said Dr. C. Ronald Kahn, a diabetes researcher and professor at Harvard Medical School.

The defining feature of diabetes is elevated blood sugar. But the reasons for abnormal sugar seem to “differ tremendously from person to person,” said Dr. Robert A. Rizza, a professor at the Mayo Clinic College of Medicine. Understanding exactly what signals are involved, he said, raises the hope of “providing the right care for each person each day, rather than giving everyone the same drug.”

Last summer, researchers at Columbia University Medical Center published startling results showing that a hormone released from bone may help regulate blood glucose.

When the lead researcher, Dr. Gerard Karsenty, first described the findings at a conference, the assembled scientists “were overwhelmed by the potential implications,” said Dr. Saul Malozowski, senior adviser for endocrine physiology research at the National Institute of Diabetes and Digestive and Kidney Diseases, who was not involved in the research. “It was coming from left field. People thought, ‘Oof, this is really new.’

“For the first time,” he went on, “we see that the skeleton is actually an endocrine organ,” producing hormones that act outside of bone.

In previous work, Dr. Karsenty had shown that leptin, a hormone produced by fat, is an important regulator of bone metabolism. In this work, he tested the idea that the conversation was a two-way street. “We hypothesized that if fat regulates bone, bone in essence must regulate fat,” he said.

Working with mice, he found that a previously known substance called osteocalcin, which is produced by bone, acted by signaling fat cells as well as the pancreas. The net effect is to improve how mice secrete and handle insulin, the hormone that helps the body move glucose from the bloodstream into cells of the muscle and liver, where it can be used for energy or stored for future use. Insulin is also important in regulating lipids.

In Type 2 diabetes, patients’ bodies no longer heed the hormone’s directives. Their cells are insulin-resistant, and blood glucose levels surge. Eventually, production of insulin in the pancreas declines as well.

Dr. Karsenty found that in mice prone to Type 2 diabetes, an increase in osteocalcin addressed the twin problems of insulin resistance and low insulin production. That is, it made the mice more sensitive to insulin and it increased their insulin production, thus bringing their blood sugar down. As a bonus, it also made obese mice less fat.

If osteocalcin works similarly in humans, it could turn out to be a “unique new treatment” for Type 2 diabetes, Dr. Malozowski said. (Most current diabetes drugs either raise insulin production or improve insulin sensitivity, but not both. Drugs that increase production tend to make insulin resistance worse.) A deficiency in osteocalcin could also turn out to be a cause of Type 2 diabetes, Dr. Karsenty said.

Another recent suspect in glucose regulation is the immune system. In 2003, researchers from two laboratories found that fat tissue from obese mice contained an abnormally large number of macrophages, immune cells that contribute to inflammation. The finding piqued the curiosity of researchers. “I remember reading the paper and thinking: ‘Wow, look at all those macrophages. What are they doing?’” said Dr. Jerrold M. Olefsky of the University of California, San Diego, School of Medicine.

Scientists have long suspected that inflammation was somehow related to insulin resistance, which precedes nearly all cases of Type 2 diabetes. In the early 1900s, diabetics were sometimes given high doses of aspirin, which is an anti-inflammatory, Dr. Olefsky said.

Only in the past few years has research into the relationship of obesity, inflammation and insulin resistance become “really hot,” said Dr. Alan R. Saltiel, director of the Life Sciences Institute at the University of Michigan.

Many researchers agree that obesity is accompanied by a state of chronic, low-grade inflammation in which some immune cells are activated, and that that may be a primary cause of insulin resistance. They also agree that the main type of cell responsible for the inflammation is the macrophage, Dr. Saltiel said.

But major questions remain, he said: “Why are these macrophages attracted to fat, liver and muscle in the first place? What are they doing? What are they secreting? What other immune cells are in there?”

New research also suggests that “not all macrophages are created equal,” added Dr. Saltiel. There appear to be “good ones and bad ones” competing in fat tissue, with potentially large consequences for inflammation and diabetes.

Meanwhile, the promise of anti-inflammatory compounds as treatment continues to attract attention. “Certain cellular anti-inflammatory proteins may now be important new targets for drug discovery for diabetes treatment,” Dr. Olefsky said. But damping down the immune system is also potentially risky, he noted, adding: “If you’re inhibiting the macrophage inflammatory pathway, that’s good for insulin resistance and diabetes. But it might not be so good for your susceptibility to infections.”

A major goal is to develop a drug that quashes only the specific component of macrophage inflammation that leads to insulin resistance, without causing other side effects. One class of current medications, called thiazolidinediones, may work in part by reducing inflammation, which may in turn improve insulin sensitivity. But an example from this class, the drug Avandia, was also found to increase the risk of heart attacks.

Another participant in the glucose conversation is the brain. Its role has long been suspected. More than a century ago, the French physiologist Claude Bernard suggested that the brain was important in blood sugar regulation. He punctured the brains of experimental animals in specific areas and managed to derange their blood sugar metabolism, making them diabetic. But for years, virtually no one followed up on this finding, said Dr. Kahn, of Harvard. People thought about glucose as a critical fuel for the brain, Dr. Kahn said, but did not explore the brain’s role in glucose regulation.

Only recently, with more advanced laboratory techniques, has this role been definitively established and expanded upon. Today’s genetic techniques, said Dr. Rizza, at the Mayo Clinic, are what have “really driven the process.”

For instance, once scientists developed the ability to manipulate mice so that they lacked particular receptors in specific tissues, they could show that mice without insulin receptors in the brain could not regulate glucose properly and went on to develop diabetes, said Dr. Kahn, whose laboratory published this groundbreaking work in 2000.

Other researchers have shown that free fatty acids, as well as the hormone leptin, produced by fat tissue, signal directly to a part of the brain called the hypothalamus, which also regulates appetite, temperature and sex drive. And several recent papers suggest that direct signaling by glucose itself to neurons in the hypothalamus is also crucial to normal blood sugar regulation in mice.

“If the brain is getting the message that you have adequate amounts of these hormones and nutrients, it will constrain glucose production by the liver and keep blood glucose relatively low,” said Dr. Michael W. Schwartz, a professor at the University of Washington. But if the brain senses inadequate amounts, he continued, it will “activate responses that cause the liver to make more glucose, and new evidence suggests that this contributes to diabetes and impaired glucose metabolism.”

The brain, therefore, appears to be listening to - and weighing and making sense of - a chorus of signals from insulin, leptin, free fatty acids and glucose itself. In response, it appears to send signals to liver and muscle cells by way of several nerves, though additional mechanisms are probably involved. The gut also seems to chime in, said Dr. Rizza, adding that for him, this aspect of sugar regulation came as “the biggest gee whiz of all.”

“Food comes in through the gut, so of course you should look there” for molecules involved in glucose regulation, he said. “But few people realized this until very recently.”

Hormones from the small intestine called incretins turn out to talk directly with the brain and pancreas in ways that help reduce blood sugar and cause animals and people to eat less and lose weight, Dr. Rizza said.

Numerous molecules that mimic incretins or prevent them from being degraded are in clinical trials. Two such drugs have been approved by the Food and Drug Administration: Byetta, an incretin mimic, from Amylin Pharmaceuticals and Eli Lilly; and Januvia, from Merck, which inhibits the destruction of the incretin GLP1. (Dr. Rizza is an adviser to Merck but says all consulting fees go to the Mayo Clinic for education and research.)

Still, it can be hard to predict how different drugs will interact in the body. And many promising candidates will turn out to have side effects - chattering helpfully with one organ, but problematically with another.

“The picture is becoming more and more complicated,” Dr. Saltiel said. “And let’s face it, it was pretty complicated before.”

Purpose: To study the effects of 8 wk creatine monohydrate (CrM) supplementation on blood and urinary clinical health markers in soccer players. METHODS: 14 soccer players were randomly assigned in a double-blinded fashion to Cre (N = 7) or Pla (N = 7) group. Cre group ingested 15 g/d of CrM during 7 d, and 3 g/d for the remaining 49 d, whereas the Pla group ingested maltodextrin following the same protocol. Soccer-specific training was performed during the study. Total body mass was determined and blood and urine samples were analized for metabolic, hepatic, renal and muscular function markers, before and after supplementation. RESULTS: A gain of total body mass was observed after CrM intake, not with placebo. Blood and urinary markers remained within normal reference values. There were no significant changes in renal and hepatic markers after CrM intake. However, total CK activity significantly increased, and uric acid level tended to decrease after CrM use. Likewise, serum glucose decreased in the Cre group following supplementation. No significant differences in urine parameters were found in either group after supplementation. CONCLUSIONS: 8 wk of CrM supplementation had no negative effect on blood and urinary clinical health markers in soccer players. Properties of CrM may, however, be associated with an increase in CK activity, improving the efficiency for ATP resynthesis, phenomenon indirectly confirmed by the decreasing tendency in uric acid concentration. Furthermore, CrM seems to slightly influence glucoregulation in trained subjects.

Mitochondrial disorders share common cellular consequences: (1) decreased ATP production; (2) increased reliance on alternative anaerobic energy sources; and (3) increased production of reactive oxygen species. The purpose of the present study was to determine the effect of a combination therapy (creatine monohydrate, coenzyme Q(10), and lipoic acid to target the above-mentioned cellular consequences) on several outcome variables using a randomized, double-blind, placebo-controlled, crossover study design in patients with mitochondrial cytopathies. Three patients had mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS), four had mitochondrial DNA deletions (three patients with chronic progressive external ophthalmoplegia and one with Kearns-Sayre syndrome), and nine had a variety of other mitochondrial diseases not falling into the two former groups. The combination therapy resulted in lower resting plasma lactate and urinary 8-isoprostanes, as well as attenuation of the decline in peak ankle dorsiflexion strength in all patient groups, whereas higher fat-free mass was observed only in the MELAS group. Together, these results suggest that combination therapies targeting multiple final common pathways of mitochondrial dysfunction favorably influence surrogate markers of cellular energy dysfunction. Future studies with larger sample sizes in relatively homogeneous groups will be required to determine whether such combination therapies influence function and quality of life.

The developmental regulation of peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) gene expression was studied in mice and compared with that of marker genes of liver energy metabolism. The PGC-1alpha gene was highly expressed in fetal liver compared with that in adults and remained high in neonatal liver. The regulation of PGC-1alpha gene expression during the fetal and early neonatal periods was dissociated from that of gluconeogenic genes, i.e. the phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) genes. Only under the effects of starvation was PGC-1alpha gene expression induced in parallel to phosphoenolpyruvate carboxykinase and G6Pase mRNAs during the perinatal period. Furthermore, the PGC-1alpha gene was not regulated as part of the developmental program of gene expression associated with the maturation of hepatic gluconeogenesis, as revealed by the impaired PEPCK and G6Pase gene expression but unaltered PGC-1alpha mRNA levels in CCAAT/enhancer-binding protein-alpha-null fetus and neonates. Regulation of the PGC-1alpha gene and that of mitochondrial 3-hydroxy-3-methyl-glutaryl-coenzyme A synthase, acyl-coenzyme A oxidase, and long-chain acyl-coenzyme dehydrogenase, marker genes of lipid catabolism, were dissociated in fetuses and neonates. The expression of lipid catabolism genes was down-regulated in fasted neonates, whereas PGC-1alpha was oppositely regulated. The independent regulation of PGC-1alpha and lipid catabolism genes was also found in peroxisome proliferator-activated receptor-alpha-null neonates, in which PGC-1alpha mRNA levels were unaffected whereas gene expression for 3-hydroxy-3-methyl-glutaryl-coenzyme A synthase and acyl-coenzyme A oxidase was impaired. Thus, regulation of the PGC-1alpha gene is partially dissociated from the patterns of regulation of hepatic genes encoding enzymes involved in gluconeogenesis and lipid catabolism during fetal ontogeny and in response to the initiation of lactation.

From the full text article:

Introduction 
 
Peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) is a transcriptional coactivator involved in the control of biological responses linked to energy homeostasis in several tissues. It participates in determination of the divergence between white adipocytes (energy storage function) and brown adipocytes (energy expenditure), regulates mitochondrial biogenesis in skeletal muscle, and has recently been reported to play a major role in the control of fuel homeostasis in the liver (1). The action of PGC-1alpha as a master regulator on liver energy metabolism was revealed by its role in the metabolic response to fasting, where it is induced and leads to a coordinate induction of genes involved in hepatic gluconeogenesis. PGC-1alpha coactivates transcription factors and nuclear hormone receptors that control the transcription of genes encoding enzymes involved in this metabolic pathway. PGC-1alpha activates some gluconeogenic genes via its capacity to coactivate hepatic nuclear factor-4 (HNF4) and the glucocorticoid receptor (2, 3). Moreover, PGC-1alpha requires the forkhead transcription factor FOXO1 to activate the gluconeogenic genes in a pathway that is also critical for the inhibitory regulation of gluconeogenesis by insulin (4). It was proposed that the role of PGC-1alpha in the metabolic adaptation of the liver to fasting extends to lipid oxidation pathways, because PGC-1alpha can induce several key genes encoding enzymes of fatty acid oxidation and ketogenesis when overexpressed in hepatic cells (3). The capacity to coactivate peroxisome proliferator-activated receptor-alpha (PPAR-alpha) (5), a master regulator of lipid catabolism, is expected to mediate the effect of PGC-1alpha to induce those genes in the liver in response to fasting, although direct evidence for this is still lacking.

The induction of gene expression by PGC-1alpha in response to fasting is thought to be mediated by a dramatic increase in the amount of this coactivator. This is due to the sensitivity of PGC-1alpha gene expression up-regulation through cAMP-dependent pathways, which are activated by glucagon levels, which rise during fasting (6). Glucocorticoids, which are up-regulated by starvation, also induce PGC-1alpha gene expression (2). Although less studied than fasting, other models of enhanced hepatic gluconeogenesis, such as streptozotocin-induced diabetes and genetic obesity, have confirmed several aspects of the involvement of PGC-1alpha in the control of gluconeogenesis and lipid catabolic pathways in the liver (2).

During rodent development, the activation of gluconeogenic and lipid catabolic pathways, including ketogenesis, takes place at birth. Fuel consumption by fetuses is mainly based on glucose, but birth leads to a sudden decrease in glucose availability associated with the massive appearance of lipids in blood, coming from milk. Thus, the metabolic adaptation to birth requires the induction of gluconeogenesis to maintain glycemia and the activation of lipid oxidation to use this fuel for energy metabolism (7). Accordingly, genes involved in these metabolic pathways are poorly expressed in fetal liver, but are dramatically induced at birth (8, 9). This metabolic and gene expression response, which resembles that elicited by fasting in adults, is induced in neonates by the fed state, because it results from the sudden imbalance between glucose and lipid availability in the transition from the fetal to the neonatal period. Moreover, the induction of metabolic genes in the liver requires the appropriate development of the prenatal program of liver differentiation. Thus, the whole basal transcriptional machinery that allows gene expression to respond to postnatal induction should develop in the liver during the fetal period by an ontogenically programmed mechanism and in the absence of the hormonal stimuli associated with birth. For instance, mice with targeted disruption of CCAAT/enhancer-binding protein-alpha (C/EBPalpha), a master transcription factor for the differentiation of hepatic cells, show impaired expression of gluconeogenic-associated genes, such as phosphoenolpyruvate carboxykinase (PEPCK), in such a way that this gene cannot be induced after birth, and gluconeogenesis cannot be switched on (10).

In the present study the developmental regulation of PGC-1alpha gene expression in the liver was determined in relation to regulation of the expression of genes involved in liver metabolism and was studied in murine models of targeted disruption of master transcription factors controlling gluconeogenic (C/EBPalpha) or lipid oxidative pathways (PPARalpha). We found partial dissociation between gluconeogenic and lipid oxidation pathways and PGC-1alpha gene expression during the perinatal period.

…

Results 
 
Tissue-specific developmental regulation of PGC-1alpha gene expression

PGC-1alpha is highly expressed in the fetal liver. The expression of PGC-1alpha mRNA was determined during the fetal development of mice in the tissues known to express this coactivator in adults. The pattern of expression in fetuses compared with adults was highly specific for every tissue, as shown in Fig. 1. In skeletal muscle, the expression of PGC-1alpha  mRNA was hardly detectable at various stages of the fetal period, whereas it was expressed in adults. In contrast, late fetal brown adipose tissue already expressed relatively high levels of PGC-1alpha mRNA, close to the levels present in adults. Surprisingly, fetal liver expressed much higher levels of PGC-1alpha mRNA than adult liver, in which PGC-1alpha was hardly detectable. This high expression of PGC-1alpha mRNA resulted in the expression of PGC-1alpha protein, as shown by Western blot analysis of nuclear extracts from fetal and neonatal liver (Fig. 1C), whereas PGC-1alpha protein was practically undetectable in adult liver.
 
Hepatic PGC-1alpha mRNA expression is dissociated from the regulation of metabolic genes in the immediate postnatal period

The expression levels of PGC-1alpha mRNA were determined in the transition from the fetal to the neonatal period and compared with the regulation of expression of key genes associated with gluconeogenesis (PEPCK and G6Pase), genes associated with lipid catabolism (mitochondrial HMG-CoA synthase, which is responsible for ketogenesis; ACO, which is involved in peroxisomal fatty acid oxidation; and LCAD, which is involved in mitochondrial fatty acid ß-oxidation) and a marker of mitochondrial biogenesis (the mitochondrial genome-encoded COII). PGC-1alpha mRNA expression hardly varied during the first hours after birth, and PGC-1alpha mRNA levels remained as high as in fetuses (Fig. 2). In contrast, the expression of mRNAs encoding PEPCK and G6Pase as well as that of the three gene markers of lipid catabolism, HMG-CoA synthase, ACO, and LCAD, were high during the immediate postnatal period compared with those in fetal life. COII mRNA expression showed a slight increase during fetal life and remained almost constant during the perinatal period. When birth was blocked by progesterone treatment of pregnant mice, and postmature fetuses were analyzed, the induction of gene markers for gluconeogenesis and lipid catabolism pathways was partially blunted, whereas PGC-1alpha mRNA and COII mRNA expression did not vary (see Table 1).

Hepatic PGC-1alpha mRNA expression is up-regulated during fasting in the perinatal period

To ascertain whether PGC-1alpha mRNA expression in the perinatal period is sensitive to the known physiological stimuli acting in adults, fetuses and neonates were studied under the stimulus of starvation. Hepatic PGC-1alpha mRNA was dramatically up-regulated after 24-h starvation in adult mice (Fig. 3), in agreement with a previous report (2). When fetuses at term or even immature fetuses were born by cesarean sections and kept for several hours without milk intake, the expression of PGC-1alpha mRNA was up-regulated despite the already high basal levels of expression. We obtained the same results in neonates born spontaneously and not allowed to initiate suckling as well as in 2-d-old neonates that had been suckling and were suddenly withdrawn from their mothers (Fig. 3). This response was similar to that of gluconeogenic genes, such as PEPCK and G6Pase, but was the opposite of the behavior of gene markers for lipid catabolism. Whereas the mRNA levels of HMG-CoA synthase, ACO, and LCAD were induced by starvation in adults, the expression of these three mRNAs in the liver was down-regulated in response to fasting during the perinatal period (Fig. 3). COII mRNA levels were not significantly modulated by starvation in fetuses, neonates, or adults.

We also determined to what extent the changes in gene expression in the liver of neonates due to starvation are specifically blunted by carbohydrates or lipids (Table 2). Pups at birth were withdrawn from their mothers before initiating suckling and were intragastrically infused with equivalent caloric amounts of a glucose solution or a lipid emulsion. PGC-1alpha mRNA induction due to starvation was partially reduced by both glucose and lipid administration. The metabolic genes responded differentially. PEPCK mRNA and G6Pase mRNA induction by fasting was reduced by both treatments, whereas the down-regulation of HMG-CoA synthase mRNA and ACO mRNA and, to a lesser extent, LCAD mRNA, was specifically attenuated by lipid intake. The model employed for food intake depletion in cesarean-derived fetal and newborn mice also involved fluid restriction, and this may add stress to the strict nutrition depletion, which can contribute to changes in gene expression. However, the differential effects of nutrients on gene expression suggest that fluid restriction, even if potentially involved, is not the main element determining changes in gene expression in this model.
 
C/EBP-null mice show impaired expression of hepatic gluconeogenic genes, but unaltered PGC-1alpha gene expression during the perinatal period

Because C/EBP is a master regulator of liver differentiation and, in particular, of gluconeogenic gene expression in the liver, we determined the effects of C/EBPalpha disruption in mice on PGC-1alpha gene expression (Fig. 4). Given the postnatal mortality of C/EBPalpha-null mice (10), only fetuses at several stages of fetal development and recently born neonates were studied. The data indicated that PEPCK mRNA expression, which is only detectable by Northern blotting in the late fetal and neonatal periods of wild-type mice, was blunted in C/EBPalpha-null mice, and the same was observed for G6Pase mRNA, which was dramatically reduced in fetal and neonatal C/EBPalpha-null mice. These observations were in agreement with the already established impairment of gluconeogenic gene expression in these mice (10). PGC-1alpha mRNA expression was unaffected by C/EBPalpha-targeted disruption. Neither HMG-CoA synthase mRNA levels nor LCAD mRNA levels were significantly altered in C/EBPalpha-null mice. However, the expression of ACO mRNA was significantly reduced in C/EBPalpha-null fetuses and neonates, suggesting that C/EBPalpha may be involved in developmental regulation of gene expression for peroxisomal enzymes. COII mRNA expression was also unchanged in liver from C/EBPalpha-null mice.

PPAR disruption impairs hepatic expression of genes of lipid catabolism during the perinatal period without altering PGC-1alpha mRNA expression

To further explore the link between PGC-1alpha mRNA expression and lipid catabolism in perinatal liver, neonatal mice with targeted disruption of PPARalpha were studied (Fig. 5). The expression of PPARalpha mRNA in control mice was high at birth, as previously reported (8), and was not essentially affected in the first hours after birth in either the 16-h fed (135 ± 14% vs. 0-h-old pups) or 16-h fasted (98 ± 8% vs. 0-h-old pups) condition. Disruption of PPARalpha caused a marked down-regulation of mitochondrial HMG-CoA synthase mRNA and ACO mRNA expression in pups at birth, after 16 h of suckling, and after 16 h of postnatal starvation. LCAD mRNA was not significant reduced, in agreement with previous findings on the expression of this gene in adult PPARalpha-null mice (17). This confirmed that PPARalpha acts a master regulator of several genes of mitochondrial and peroxisomal lipid catabolism after birth and can contribute to the switch-on of lipid oxidation pathways during development. PEPCK mRNA, G6Pase mRNA, and COII mRNA were essentially unaffected, as was PGC-1alpha mRNA, pointing to a dissociation between PGC-1alpha gene expression and the PPARalpha-regulated pathways of gene expression in the liver during this period.

Discussion 
 
Our results indicate that PGC-1alpha gene expression is under powerful tissue-specific developmental regulation. The high levels of PGC-1alpha mRNA expression in fetal brown adipose tissue are consistent with the postulated role for this coactivator in brown fat differentiation and the fact that brown adipocyte differentiation in vivo takes place during the late fetal period in rodents (11, 18). Moreover, the progressive induction of PGC-1alpha mRNA in skeletal muscle, with very low levels during the fetal period that become much higher in adults, is also in agreement with a major role for PGC-1alpha in this tissue, i.e. the control of mitochondrial biogenesis (19), which develops postnatally in mice (20). In contrast, the high expression of PGC-1alpha in fetal liver was a surprising finding. The up-regulation of PGC-1alpha in liver has been attributed to cAMP-dependent activation due to the action of glucagon or the effects of glucocorticoids (2, 6). These mechanisms are responsible for the high levels of expression of PGC-1alpha in the liver of fasted adults, but they can hardly be involved in the high expression in the fetal liver, when stress signaling elicited by birth and mediated by glucagon or glucocorticoids is still not active. However, the PGC-1alpha gene in late fetal development appears to be already sensitive to those pathways when they are suddenly activated, as revealed by the up-regulation elicited by delivery plus starvation even in immature fetuses. This is in agreement with the reported capacity of fetal liver to activate gluconeogenesis in response to glucocorticoid infusion (21).

A second aspect to be considered is how the high PGC-1alpha expression in the fetus is compatible with the low expression of gluconeogenic genes. Two facts should be taken into account. First, fetuses are hyperinsulinemic (7), and insulin is known to behave as an inhibitor of the gluconeogenic gene expression program mediated by PGC-1alpha (1). Second, it was recently reported that HNF4 is a key transcription factor required for the coactivation of gluconeogenic genes by PGC-1alpha (3). HNF41, the major isoform of HNF4, is hardly expressed in fetal liver, and it is first expressed around birth (22). Moreover, when HNF4 expression is reduced by targeted gene disruption, PGC-1alpha expression is dramatically up-regulated, suggesting that HNF4 is not only a target of this coactivator, but is also a negative regulator of PGC-1alpha gene expression (3). The low hepatic expression of HNF41 in fetuses may mediate the high PGC-1alpha gene expression and the dissociation between high PGC-1alpha levels and low gluconeogenic gene expression during the fetal period. Moreover, several reports have recently identified negative regulators of PGC-1alpha action, such as sterol regulatory element-binding protein-1, which interacts with HNF4 and interferes with PGC-1alpha recruitment to suppress hepatic gluconeogenic genes (23), or p160 Myb-binding protein, which inhibits the mitochondriogenic action of PGC-1alpha (24). Additional studies are required to test the involvement of these transcription factors and coregulators in the dissociation between the expression of PGC-1alpha and that of target genes in the late fetal development.

In contrast, the potential of PGC-1alpha as a mitochondriogenic coactivator has been stressed in muscle cells and brown adipocytes, but a recent report also indicates that overexpression of PGC-1alpha in hepatic cells up-regulates mRNA expression for mitochondrial components (25). High PGC-1alpha gene expression may have been involved in the high COII mRNA expression in the fetal liver. However, there were no signs of marked regulation of COII mRNA levels even in situations such as perinatal starvation, in which PGC-1alpha expression is induced.

In addition to the fetal period, the early neonatal period revealed a dissociation between the regulation of PGC-1alpha gene expression and the activation of gluconeogenic and lipid oxidation pathways. After birth and initiation of suckling, glucagon- and glucocorticoid-mediated pathways of regulation are rapidly activated, and insulinemia declines suddenly; thus, metabolic target genes involved in gluconeogenesis and fat oxidation are dramatically induced in the first hours after birth (7). This is associated with the sudden appearance of fatty acids in milk as the major source of metabolic energy and the requirement of lipid oxidation and glucose synthesis to maintain glycemia. However, again in this situation, PGC-1alpha expression is high, but not higher than in fetuses despite the induction of gluconeogenesis. Only if neonates are not allowed to suckle, starvation leads to a parallel overinduction of PGC-1alpha, PEPCK, and G6Pase expression. This indicates that the regulation of PGC-1alpha expression is not associated with the naturally occurring induction of gluconeogenesis after delivery, although it can be involved in the overinduction of this pathway in response to perinatal starvation, as in adults. Apart from starvation, the dissociation between PGC-1alpha gene expression and the modulation of gluconeogenesis during the perinatal period is also evidenced in postmature fetuses, in which the up-regulation of PEPCK and G6Pase is blunted, but PGC-1alpha remains unaltered. Although progesterone may affect the expression of metabolic genes in fetuses, the absence of the stress and nutrient shift associated with delivery was probably the cause of the impaired PEPCK and G6Pase induction in postmature fetuses. Additional evidence of the dissociation between PGC-1alpha gene expression and the gluconeogenic program during the perinatal period comes from C/EBPalpha-null mice, in which blockage of the whole program of hepatic gluconeogenic gene expression in the late fetal and early neonatal periods does not modify PGC-1alpha gene expression. All of these findings are consistent with a recent report indicating that PGC-1alpha acts a transcription amplifier, but is not essential for basal and hormone-induced PEPCK gene expression (26), and support the idea that in a particular physiological situation, such as the perinatal period, the extent of activation of target metabolic pathways by PGC-1alpha does not necessarily require a parallel regulation of PGC-1alpha gene expression.

Some of the present findings are not in total agreement with a recent report by Lin et al. (25). These researchers also observed a higher expression of PGC-1alpha mRNA in late fetuses compared with adults, although the differences they found were fewer. They did not observe such an increase in younger fetuses, and they also report a spontaneous induction in neonatal mice after birth. Those studies were performed using mice from the C57BL/6 strain, which is the background strain we used in our experiments with mice bearing targeted disruption of PPAR. We did not observe postnatal induction in wild-type, fed pups from this strain; therefore, additional research is required to establish the reasons for this discrepancy.

The dissociation between PGC-1alpha regulation and lipid catabolism in the neonate was even more marked than that for gluconeogenesis. Genes for lipid oxidation and ketogenesis are induced by overexpressing PGC-1alpha in hepatic cells (3, 25). Because the expression of PGC-1alpha and lipid oxidation genes is up-regulated in adults in response to starvation, a major role for PGC-1alpha has been proposed in the control of lipid catabolism, similarly to gluconeogenesis. During the perinatal period, lipid oxidation genes, either peroxisomal or mitochondrial, as well as genes involved in ketogenesis, are poorly expressed in fetal liver and are induced around birth to cope with the massive influx of fat from milk (Refs.8 and 9 and the present results). This pattern is not associated with parallel changes in PGC-1alpha expression. In contrast to adult starvation, perinatal starvation down-regulates lipid oxidation genes. This differential response may be due to the fact that the fasting of neonatal rodents results in the cessation of fatty acid supplies, which can come only from milk, not from white adipose tissue stores that have not yet developed. In contrast, fasting in adults is linked to the increased availability of free fatty acids from white fat lipolysis. The up-regulation of PGC-1alpha expression in fasted neonates is the opposite of the down-regulation of fat catabolism genes, which rules out a role for PGC-1alpha in the control of this pathway.

PPARalpha acts as a master transcriptional regulator of genes encoding enzymes involved in fatty acid catabolism (27) and has been proposed as the major mediator of coactivation by PGC-1alpha to induce the genes involved in lipid catabolism (5). Here we show that the targeted disruption of PPARalpha impairs the expression of HMG-CoA synthase and ACO, marker genes for fatty acid catabolism, during the perinatal period. However, this does not affect PGC-1alpha gene expression in the fed or fasted state. The depletion of fatty acid-derived metabolites may explain why high PGC-1alpha levels in association with the high PPARalpha in neonates (8) did not elicit a high expression of lipid oxidation genes in the liver of fasted neonates. Fatty acids provide not only the fuel supply for energy metabolism, but also the ligand molecules required for the PPARalpha-dependent activation of target genes (28). In the presence of high PPARalpha and PGC-1alpha expression, low availability of lipid ligands in fasted neonates can result in low gene expression of lipid oxidation genes. The preferential reactivation of expression of lipid catabolism genes when fasted pups were treated with a single dose of a lipid emulsion compared with equivalent amounts of glucose supports this possibility.

In summary, PGC-1alpha gene expression is partially dissociated from the activation of gluconeogenic genes during the perinatal period depending on developmental stage and nutritional status. It appears that developmental regulation of transcription factors such as HNF4, FOXO1, or other targets of PGC-1alpha coactivation can be essential regulatory elements to build up the appropriate regulatory machinery for the gluconeogenic program in which PGC-1alpha is involved as a coactivator. The regulation of PGC-1alpha gene expression is completely dissociated from the extent of activation of genes of lipid catabolism during the perinatal period, which is highly dependent on PPARalpha and nutritional status. Other coactivators, such as PGC-1ß, a recently identified member of the PGC-1 family (25) that shows preferential coactivation activity for PPARalpha (29), deserve additional research as more preferential candidates to regulate lipid oxidation pathways through transcriptional coactivation. The growing identification of novel coregulators, such as p160 Myb-binding protein (24), or transcription factors and nuclear receptors, such as sterol regulatory element-binding protein-1 (22), estrogen-related receptor-alpha (30, 31), or farnesoid X receptor (32), which act as repressive or activating modulators of the metabolic effects of PGC-1alpha, indicates that the action of PGC-1alpha on target metabolic genes results from cross-talk with all of these factors. Developmental regulation of the expression of these factors together with PGC-1alpha gene expression would be essential to determine the activity of PGC-1alpha at a given stage of development.

OBJECTIVE: Basal cortisol and cortisol stress responsivity are valuable biological characteristics of children with disruptive behavior disorder (DBD). In this study, the predictive value of cortisol to outcome of intervention was investigated. METHOD: Basal cortisol levels and cortisol levels under stress were studied in 22 children with DBD before the start of a psychotherapeutic treatment. The disruptive behavior of the child was assessed before treatment and after cessation (9 months later). RESULTS: Children with DBD with relatively high and low basal cortisol levels differed in the severity of problem behavior at pretreatment, with the low basal cortisol group having more severe problems. During stress, children with DBD showed either increasing or decreasing cortisol values. Although these cortisol responsivity groups were similar in the severity of behavioral problems at pretreatment, the behavioral problems of the group with high cortisol stress responsivity were significantly lower after the intervention than the behavioral problems of the group with low cortisol stress responsivity. CONCLUSIONS: In children with DBD, the basal cortisol level was related to the severity of behavioral problems at pretreatment but not to the severity of behavioral problems after treatment. The cortisol response pattern during stress was related to treatment outcome.

Dehydroepiandrosterone (DHEA) and its sulfate derivative DHEA-S are neurosteroids, produced in the brain, and neuroactive steroids, produced in the adrenals and affecting the brain. We compared the ratios of serum cortisol/DHEA or DHEA-S in schizophrenia patients with normal subjects, and determined the correlation of these ratios with psychopathology and distress. Early morning plasma concentrations of DHEA, DHEA-S, and cortisol were determined by radioimmunassay in 40 medicated schizophrenia inpatients, and 15 healthy subjects with similar age and sex distribution. Subjects were assessed for psychopathology using the Positive and Negative Syndrome Scale (PANSS) and the Montgomery and Asberg Depression Rating Scale (MADRS), anxiety, anger, emotional and somatic distress levels. Schizophrenia inpatients demonstrated significantly higher levels of state and trait anxiety, anger expression index, emotional and somatic self-reported distress scores. Cortisol/DHEA and cortisol/DHEA-S ratios were significantly higher in schizophrenia patients than in healthy comparison subjects. Both ratios correlated positively with age and duration of illness; cortisol/DHEA-S ratio also showed positive association with age of illness onset. When age, illness duration and age of onset were controlled, cortisol/DHEA-S ratio significantly correlated with severity of depression (MADRS, r=0.33, p=0.048), state and trait anxiety (r=0.43, p=0.008 and r=0.40, p=0.014, respectively), trait anger (r=0.41, p=0.012), angry temperament (r=0.46, p=0.004), anger expression index (r=0.36, p=0.033), and hostility (r=0.42, p=0.010). No significant association was found between these ratios and severity of psychopathology, and type or dosage of antipsychotic agents. Thus, elevated cortisol/DHEA and/or cortisol/DHEA-S ratios in schizophrenia patients are positively associated with higher scores for anxiety and anger, depression and hostility, age and age of onset/duration of illness, but are independent of severity of psychopathology (PANSS) and antipsychotic treatment.

The normal low-density lipoprotein (LDL) cholesterol range is 50 to 70 mg/dl for native hunter-gatherers, healthy human neonates, free-living primates, and other wild mammals (all of whom do not develop atherosclerosis). Randomized trial data suggest atherosclerosis progression and coronary heart disease events are minimized when LDL is lowered to <70 mg/dl. No major safety concerns have surfaced in studies that lowered LDL to this range of 50 to 70 mg/dl. The current guidelines setting the target LDL at 100 to 115 mg/dl may lead to substantial undertreatment in high-risk individuals.

Patterns of lower autonomic nervous system (ANS) and hypothalamic-pituitary-adrenal (HPA) axis activity have been found in children with oppositional defiant disorder (ODD). The aim of the present study was to investigate whether children with attention-deficit/hyperactivity disorder (ADHD) differ from ODD children with (OD/AD) or without comorbid ADHD in ANS and HPA axis activity under baseline and stressful conditions. The effects of stress on cortisol, heart rate (HR), and skin conductance level (SCL) were studied in 95 children (26 normal control [NC] children and 69 child psychiatric patients referred for externalizing behavior problems [15 ODD, 31 OD/AD, and 23 ADHD]). No baseline differences were found in cortisol between the four groups. However, the ODD and OD/AD groups showed a significantly weaker cortisol response to stress compared to the ADHD and NC groups; the ADHD group had a similar cortisol response as the NC group. Within the ODD group this pattern of low cortisol responsivity was most clearly present in the more severely affected inpatients. With respect to HR, the ODD group had a significantly lower HR during baseline and stressful conditions. The higher HR levels in the OD/AD and ADHD groups were likely to be caused by methylphenidate. The externalizing groups had significantly lower SCL levels, and no differences were found between these groups. It was concluded that differences in cortisol responsivity during stress exposure are important in distinguishing within a group of children with externalizing behavior between those with ODD and ADHD.