Investigation of the effects of PKC deletion in mice has yielded several insights into the specific contributions of individual PKC isoforms in the synthesis, esterification or oxidation of fatty acids by different tissues in vivo, as well as their roles in the response to dietary lipid over-supply. In many cases, these studies were in fact designed to investigate PKC-dependent modulation of glucose homeostasis, but, by comparing and contrasting the effects of kinase deletion on pathophysiological outcomes such as obesity, hepatic steatosis and lipidaemia, it becomes evident that the metabolic effects of PKCs extend beyond the induction of insulin resistance upon chronic activation.
Relatively little work has been performed on the involvement of two of the conventional PKC isoforms in lipid metabolism. PKCγ is expressed mainly in the brain and nervous system, and PKCγ-deficient mice have been generated  and extensively studied in terms of behavioural and other neurological effects. However, despite the importance of the brain in regulating many aspects of metabolism , no metabolic phenotypes of PKCγ in metabolism have been reported. In the case of mice lacking PKCα, insulin-stimulated glucose uptake by adipocytes as well as skeletal muscle was enhanced, together with upstream insulin signalling, but lipid metabolism itself was not investigated and the mice were not challenged with a high-fat diet .
In contrast, a major role for PKCβ in energy homeostasis and lipid accumulation has emerged. Initial work was limited to mice fed normal chow, and again demonstrated an enhanced uptake of glucose by muscle and fat cells, although this did not translate into improved glucose tolerance in vivo . Subsequently, an investigation of the effects of PKCβ deletion on broader aspects of metabolism revealed a reduction in adipose tissue depots and in the TAG content of liver and skeletal muscle in chow-fed mice . These changes were accompanied by elevated energy consumption and increased fatty acid oxidation in adipose tissue. Several changes in gene expression may have contributed to this phenotype, including the up-regulation of uncoupling protein-2 and peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC1α), which was associated with mitochondrial biogenesis . These findings were extended by examining the effects of a high-fat diet on the mice . In this case, there was protection against obesity, hyperlipidaemia and hepatic steatosis, and a preservation of insulin sensitivity and hence glucose tolerance. In high fat-fed wild-type mice and in ob/ob mice, PKCβ mRNA and protein expression was enhanced, further supporting a causative role for this isoform in promoting obesity, and it was suggested that this occurred secondary to hyperinsulinaemia, contributing to a vicious circle between insulin resistance and increased tissue TAG accumulation . Conversely, deletion of PKCβ in ob/ob mice resulted in protection against fat accumulation and insulin resistance .
While deletion of the PKCβ gene does not allow selective changes in PKCβI or PKCβII expression, specific over-expression of a constitutively active PKCβII mutant in mouse skeletal muscle demonstrated that this splice variant of PKCβ not only induces insulin resistance, but also affects the levels of several genes involved in lipid metabolism . Thus impairment in the expression of peroxisome proliferator-activated receptor gamma, PGC1α, acyl CoA oxidase and hormone-sensitive lipase, but enhanced expression of the lipogenic transcription factor sterol regulatory element-binding protein 1c (SREBP1c) in skeletal muscle, were associated with decreased lipid oxidation and increased intra-myocellular lipid deposition. In addition to these direct effects in muscle, transgenic over-expression of PKCβII led to indirect defects in insulin action in liver and brain, as well as hepatic lipid accumulation similar to that seen in fat-fed animals. This study is therefore in agreement with those utilizing PKCβ-deficient mice, further supporting a role for this isoform in the regulation of lipogenesis. Evidence that this isoform acts at least in part through the induction of SREBP1c is discussed with other mechanistic investigations further below.
In contrast to the conventional isoforms, several investigators have examined the roles of distinct novel PKCs in the regulation of energy homeostasis and metabolism. The motivation for doing so was the strong association of insulin resistance in skeletal muscle and liver with the translocation/activation of these kinases . However, detailed analysis has frequently revealed that, while the kinases do play causative roles in the generation of insulin resistance, their ablation also leads to unexpected effects on hepatic and intramyocellular fat accumulation.
Chronic activation of PKCδ has been observed in skeletal muscle  and liver  upon fatty acid over-supply, and increased hepatic expression of the kinase has also been linked to obesity in animals and humans . As may be expected from these findings, ablation of PKCδ has beneficial effects on insulin sensitivity and glucose homeostasis in fat-fed mice. However, these studies also indicate that the kinase exerts major effects on the expression of genes involved in hepatic lipid metabolism [19, 20]. PKCδ-deficient mice exhibit down-regulation of genes involved in lipogenesis, associated with reduced liver steatosis, plasma triglyceride levels, muscle TAG accumulation and adipose tissue mass [19, 20]. Interestingly, acute deletion of PKCδ in liver, through adenovirally mediated Cre recombinase expression in PKCδ-floxed mice, reversed insulin resistance in fat-fed animals without improving lipid abnormalities, indicating the independence and also the complexity of these effects of PKCδ .
In the case of PKCθ, manipulation of kinase expression or activity by independent investigators, using different models to assess its role in metabolism, has yielded disparate results, which may be reconciled by the conclusion that the phenotypes observed are dependent on the duration of exposure to lipid over-supply in the absence of the enzyme. On the one hand, a short-term elevation in plasma fatty acids induced by lipid infusion in mice showed that PKCθ-deficient mice are protected against the resulting acute whole-body insulin resistance. This was chiefly explained by the preservation of insulin signalling and glucose uptake in skeletal muscle, whereas hepatic glucose output was unaffected . This is in good agreement with the relatively high expression of PKCθ in muscle and its frequently reported translocation in this tissue upon lipid excess , and with the inhibition of insulin signalling at the level of IRS-1 mediated by this kinase . Interestingly, glucose uptake into brown but not white adipose tissue was also conserved , although the role of PKCθ in this tissue has not been explored.
On the other hand, longer-term studies have revealed additional roles for PKCθ in metabolic control that are essential for normal glucose and lipid homeostasis. Transgenic over-expression of a dominant-negative PKCθ mutant in skeletal muscle resulted in obesity in older mice, associated with impaired glucose tolerance and diminished insulin sensitivity . It was speculated that PKCθ may participate in cross-talk between muscle and fat, resulting in the observed accumulation of visceral fat. Similarly, the same line of PKCθ-deficient mice that exhibited protection of glucose homeostasis when subjected to acute lipid infusion  was found to become obese with increasing age, associated with reduced energy expenditure, defective fat oxidation and muscle insulin resistance . These phenotypes were exacerbated by high-fat feeding, leading to greater insulin resistance in muscle, fat and liver, although glucose intolerance was averted by compensatory hyperinsulinaemia . Additional effects of PKCθ on adipose tissue function were suggested by adverse changes in adiponectin expression and plasma fatty acid levels in fat-fed mice. Overall, it is clear that, while acute activation of PKCθ by fatty acid excess may inhibit insulin signalling and subsequent glucose disposal, especially in skeletal muscle, the normal function of the enzyme is important in the maintenance of body composition and energy homeostasis in the longer term.
PKCε is another novel PKC isoform that has been extensively linked to lipid-induced insulin resistance, both in skeletal muscle [17, 26-29] and liver [30, 31]. Its activation in models of hepatic insulin resistance is now frequently used as a indicator of the underlying mechanism causing defective insulin action [32-37]. A number of studies have examined the effect of its deletion or ablation not only directly on insulin signalling, where it plays an inhibitory role as expected, but also on its broader metabolic role in several tissues and under various nutritional conditions. While these have indicated more complex interactions, a unifying theme is the promotion of lipid partitioning away from β-oxidation and towards TAG storage.
Knockdown of PKCε using specific oligonucleotides reduced expression of the kinase in certain tissues, including liver and adipose tissue but not skeletal muscle . This protected mice against the insulin resistance caused by a 3-day high-fat diet, with a major effect on suppression of glucose output by the liver as assessed by the euglycaemic clamp technique, and this was associated with improved insulin receptor signalling . This indicates that, as for several other PKC isoforms, an important acute action of PKCε is the inhibition of insulin signal transduction upon lipid over-supply. A similar beneficial effect was observed in mice in which the PKCε gene had been deleted, when fed a high-fat diet for up to 3 weeks . In these mice, glucose tolerance was unimpaired by the diet, whereas wild-type mice became glucose-intolerant. Insulin levels were not elevated under these conditions by PKCε deletion, consistent with the maintenance of insulin sensitivity, and pyruvate tolerance tests again demonstrated an effect at the level of liver insulin action .
While these findings support a direct role of PKCε in the inhibition of glucose disposal in response to insulin, additional studies indicated that this enzyme also contributes to the early response of the liver to dietary lipid excess. First, the sustained insulin action observed in the absence of PKCε during the early phase of a high-fat diet was unexpectedly associated with an increase in hepatic TAG content and a defect in lipid oxidation [39, 40]. Thus, although liver lipid content was still much lower than in long term fat-fed mice, it was up to 50% greater in the PKCε-deficient mice compared to wild-type mice, and this was associated with reduced energy consumption despite normal switching to fat as the major oxidative fuel . Second, PKCε-deficient mice fed high-fat diets for over 6 weeks no longer exhibited protection against insulin resistance [39, 41]. These studies suggest that when activated, PKCε functions as a early lipid sensor, promoting β-oxidation in the liver when dietary fat is more prevalent. Although insulin resistance may also result in the shorter term, either through inhibition of insulin signalling  or through mechanisms related to accelerated β-oxidation such as increased production of reactive oxygen species , the overall effect may be beneficial when the nutritional excess is temporary.
Interestingly, the lipid partitioning observed in the liver that results from the absence of PKCε is not confined to this tissue. PKCε-deficient mice fed a high-fat diet for longer periods are still protected against glucose tolerance despite exhibiting insulin resistance, which is explained by an enhancement of insulin release that fully compensates for the impairment in insulin action [39, 41, 42]. This is associated with altered lipid partitioning in pancreatic islets upon chronic lipid exposure , and an enhancement of the production of esterification products that may act as signals to promote the amplification pathway of glucose-stimulated insulin secretion [42, 43]. Importantly, insulin secretion is not increased in an unregulated manner in PKCε-deficient mice, but only upon demand when glucose levels become elevated, avoiding β-cell exhaustion. This phenotype shows a different temporal dependence to the protection against insulin resistance, in that it is only observed under conditions of longer-term dietary lipid excess , suggesting that it depends on accumulation of specific lipids or other factors in the pancreatic islets. Thus islets from mice fed a high-fat diet for only 1 week secrete insulin in a similar manner to islets from wild-type mice, and glucose tolerance is protected only at the level of insulin sensitivity .
As well as modulating lipid partitioning during over-nutrition, PKCε also affects the supply of lipid from adipose tissue to the liver in mice upon chronic fasting. Normally, lipolytic signals such as glucagon and catecholamines promote the release of free fatty acids from adipocytes into the circulation under these conditions, which may be taken up by the liver to generate ketone bodies such as β-hydroxybutyrate, for use as alternative fuels by muscle and the brain when glucose levels are reduced . In PKCε-deficient mice, fat pad mass is not reduced to the same extent as in wild-type mice after a 48 h fast, plasma free fatty acids are not as elevated, and liver lipid stores are not supplemented to the same extent. This reduced supply of ketogenic precursors to the liver results in plasma β-hydroxybutyrate levels that are almost 50% lower than in wild-type mice . This prevention of fatty acid release by adipose tissue under fasting conditions, together with the alterations in lipid flux exhibited by the liver and pancreatic islets in the fed state, contributes to the emerging theme of lipid partitioning towards TAG storage in the absence of PKCε.