Kristoffer Ström

Green tea powder and Lactobacillus plantarum affect gut microbiota, lipid metabolism and inflammation in high-fat fed C57BL/6J mice

BackgroundType 2 diabetes is associated with obesity, ectopic lipid accumulation and low-grade inflammation. A dysfunctional gut microbiota has been suggested to participate in the pathogenesis of the disease. Green tea is rich in polyphenols and has previously been shown to exert beneficial metabolic effects. Lactobacillus plantarum has the ability to metabolize phenolic acids. The health promoting effect of whole green tea powder as a prebiotic compound has not been thoroughly investigated previously.MethodsC57BL/6J mice were fed a high-fat diet with or without a supplement of 4% green tea powder (GT), and offered drinking water supplemented with Lactobacillus plantarum DSM 15313 (Lp) or the combination of both (Lp + GT) for 22 weeks. Parameters related to obesity, glucose tolerance, lipid metabolism, hepatic steatosis and inflammation were examined. Small intestinal tissue and caecal content were collected for bacterial analysis.ResultsMice in the Lp + GT group had significantly more Lactobacillus and higher diversity of bacteria in the intestine compared to both mice in the control and the GT group. Green tea strongly reduced the body fat content and hepatic triacylglycerol and cholesterol accumulation. The reduction was negatively correlated to the amount of Akkermansia and/or the total amount of bacteria in the small intestine. Markers of inflammation were reduced in the Lp + GT group compared to control. PLS analysis of correlations between the microbiota and the metabolic variables of the individual mice showed that relatively few components of the microbiota had high impact on the correlation model.ConclusionsGreen tea powder in combination with a single strain of Lactobacillus plantarum was able to promote growth of Lactobacillus in the intestine and to attenuate high fat diet-induced inflammation. In addition, a component of the microbiota, Akkermansia, correlated negatively with several metabolic parameters known to be risk factors for the development of type 2 diabetes.

Hormone-sensitive lipase (HSL) is also a retinyl ester hydrolase: evidence from mice lacking HSL

Here, we investigated the importance of hormone‐sensitive lipase (HSL) as a retinyl ester hydrolase (REH). REH activity was measured in vitro using recombinant HSL and retinyl palmitate. The expression of retinoic acid (RA)‐regulated genes and retinoid metabolites were measured in high‐fat diet fed HSL‐ null mice using real‐time quantitative PCR and triplestage liquid chromatography/tandem mass spectrometry, respectively. Age‐ and gender‐matched wild‐type littermates were used as controls. The REH activity of rat HSL was found to be higher than that against the hitherto best known HSL substrate, i.e., diacylglycerols. REH activity in white adipose tissue (WAT) of HSL‐null mice was completely blunted and accompanied by increased levels of retinyl esters and decreased levels of retinol, retinaldehyde and all‐trans RA. Accordingly, genes known to be positively regulated by RA were down‐regulated in HSL‐null mice, including pRb and RIP140, key factors promoting differentiation into the white over the brown adipocyte lineage. Dietary RA supplementation partly restored WAT mass and the expression of RA‐regulated genes in WAT of HSL‐null mice. These findings demonstrate the importance of HSL as an REH of adipose tissue and suggest that HSL via this action provides RA and other retinoids for signaling events that are crucial for adipocyte differentiation and lineage commitment.—Strom, K., Gundersen, T. E., Hansson, O., Lucas, S., Fernandez, C., Blomhoff, R., Holm, C. Hormone‐sensitive lipase (HSL) is also a retinyl ester hydrolase: evidence from mice lacking HSL. FASEB J. 23, 2307–2316 (2009)

Attainment of Brown Adipocyte Features in White Adipocytes of Hormone-Sensitive Lipase Null Mice

Background Hormone-sensitive lipase (HSL) is expressed predominantly in adipose tissue, where it plays an important role in catecholamine-stimulated hydrolysis of stored tri- and diglycerides, thus mobilizing fatty acids. HSL exhibits broad substrate specificity and besides acylglycerides it hydrolyzes cholesteryl esters, retinyl esters and lipoidal esters. Despite its role in fatty acid mobilization, HSL null mice have been shown to be resistant to diet-induced obesity. Methodology/Principal Findings Following a high-fat diet (HFD) regimen, energy expenditure, measured using indirect calorimetry, was increased in HSL null mice. White adipose tissue of HSL null mice was characterized by reduced mass and reduced protein expression of PPARγ, a key transcription factor in adipogenesis, and stearoyl-CoA desaturase 1, the expression of which is known to be positively correlated to the differentiation state of the adipocyte. The protein expression of uncoupling protein-1 (UCP-1), the highly specific marker of brown adipocytes, was increased 7-fold in white adipose tissue of HSL null mice compared to wildtype littermates. Transmission electron microscopy revealed an increase in the size of mitochondria of white adipocytes of HSL null mice. The mRNA expression of pRb and RIP140 was decreased in isolated white adipocytes, while the expression of UCP-1 and CPT1 was increased in HSL null mice compared to wildtype littermates. Basal oxygen consumption was increased almost 3-fold in white adipose tissue of HSL null mice and was accompanied by increased uncoupling activity. Conclusions These data suggest that HSL is involved in the determination of white versus brown adipocytes during adipocyte differentiation The exact mechanism(s) underlying this novel role of HSL remains to be elucidated, but it seems clear that HSL is required to sustain normal expression levels of pRb and RIP140, which both promote differentiation into the white, rather than the brown, adipocyte lineage.

Regulation of AMP-activated protein kinase by LKB1 and CaMKK in adipocytes.

AMP‐activated protein kinase (AMPK) is a serine/threonine kinase that regulates cellular and whole body energy homeostasis. In adipose tissue, activation of AMPK has been demonstrated in response to a variety of extracellular stimuli. However, the upstream kinase that activates AMPK in adipocytes remains elusive. Previous studies have identified LKB1 as a major AMPK kinase in muscle, liver, and other tissues. In certain cell types, Ca2+/calmodulin‐dependent protein kinase kinase β (CaMKKβ) has been shown to activate AMPK in response to increases of intracellular Ca2+ levels. Our aim was to investigate if LKB1 and/or CaMKK function as AMPK kinases in adipocytes. We used adipose tissue and isolated adipocytes from mice in which the expression of LKB1 was reduced to 10–20% of that of wild‐type (LKB1 hypomorphic mice). We show that adipocytes from LKB1 hypomorphic mice display a 40% decrease in basal AMPK activity and a decrease of AMPK activity in the presence of the AMPK activator phenformin. We also demonstrate that stimulation of 3T3L1 adipocytes with intracellular [Ca2+]‐raising agents results in an activation of the AMPK pathway. The inhibition of CaMKK isoforms, particularly CaMKKβ, by the inhibitor STO‐609 or by siRNAs, blocked Ca2+‐, but not phenformin‐, AICAR‐, or forskolin‐induced activation of AMPK, indicating that CaMKK activated AMPK in response to Ca2+. Collectively, we show that LKB1 is required to maintain normal AMPK‐signaling in non‐stimulated adipocytes and in the presence of phenformin. In addition, we demonstrate the existence of a Ca2+/CaMKK signaling pathway that can also regulate the activity of AMPK in adipocytes. J. Cell. Biochem. 112: 1364–1375, 2011.

Metabolic effects of whole grain wheat and whole grain rye in the C57BL/6J mouse.

OBJECTIVE A diet rich in whole grain cereals is suggested to protect against type 2 diabetes and facilitate body weight regulation. However, little is known about the impact of different cereals and the underlying mechanisms. The objective of this study was to compare the long-term metabolic effects of diets supplemented with whole grain wheat or whole grain rye in the C57BL/6J mouse. METHODS Mice were fed the whole grain supplements in a low-fat background diet for 22 wk. Oral and intravenous glucose tolerance tests were performed during the study and in vitro insulin secretion assays were performed at the end of the study. Body weight, energy intake, body fat content, and plasma parameters were measured during the study. RESULTS A dietary supplement of whole grain rye suppressed body weight gain and resulted in significantly decreased adiposity, plasma leptin, total plasma cholesterol, and triacylglycerols compared with a supplement of whole grain wheat. Also, a slight improvement in insulin sensitivity was observed in the rye group compared with the wheat group. The decreases in body weight and adiposity were observed in the absence of differences in energy intake. CONCLUSION Long-term administration of whole grain rye evokes a different metabolic profile compared with whole grain wheat in the C57BL/6J mouse, the primary difference being that whole grain rye reduces body weight and adiposity compared with whole grain wheat. In addition, whole grain rye slightly improves insulin sensitivity and lowers total plasma cholesterol.

Perilipin is present in islets of Langerhans and protects against lipotoxicity when overexpressed in the beta-cell line INS-1.

Lipids have been shown to play a dual role in pancreatic beta-cells: a lipid-derived signal appears to be necessary for glucose-stimulated insulin secretion, whereas lipid accumulation causes impaired insulin secretion and apoptosis. The ability of the protein perilipin to regulate lipolysis prompted an investigation of the presence of perilipin in the islets of Langerhans. In this study evidence is presented for perilipin expression in rat, mouse, and human islets of Langerhans as well as the rat clonal beta-cell line INS-1. In rat and mouse islets, perilipin was verified to be present in beta-cells. To examine whether the development of lipotoxicity could be prevented by manipulating the conditions for lipid storage in the beta-cell, INS-1 cells with adenoviral-mediated overexpression of perilipin were exposed to lipotoxic conditions for 72 h. In cells exposed to palmitate, perilipin overexpression caused increased accumulation of triacylglycerols and decreased lipolysis compared with control cells. Whereas glucose-stimulated insulin secretion was retained after palmitate exposure in cells overexpressing perilipin, it was completely abolished in control beta-cells. Thus, overexpression of perilipin appears to confer protection against the development of beta-cell dysfunction after prolonged exposure to palmitate by promoting lipid storage and limiting lipolysis.

Expression of Phosphofructokinase in Skeletal Muscle Is Influenced by Genetic Variation and Associated With Insulin Sensitivity

Using an integrative approach in which genetic variation, gene expression, and clinical phenotypes are assessed in relevant tissues may help functionally characterize the contribution of genetics to disease susceptibility. We sought to identify genetic variation influencing skeletal muscle gene expression (expression quantitative trait loci [eQTLs]) as well as expression associated with measures of insulin sensitivity. We investigated associations of 3,799,401 genetic variants in expression of >7,000 genes from three cohorts (n = 104). We identified 287 genes with cis-acting eQTLs (false discovery rate [FDR] <5%; P < 1.96 × 10−5) and 49 expression–insulin sensitivity phenotype associations (i.e., fasting insulin, homeostasis model assessment–insulin resistance, and BMI) (FDR <5%; P = 1.34 × 10−4). One of these associations, fasting insulin/phosphofructokinase (PFKM), overlaps with an eQTL. Furthermore, the expression of PFKM, a rate-limiting enzyme in glycolysis, was nominally associated with glucose uptake in skeletal muscle (P = 0.026; n = 42) and overexpressed (Bonferroni-corrected P = 0.03) in skeletal muscle of patients with T2D (n = 102) compared with normoglycemic controls (n = 87). The PFKM eQTL (rs4547172; P = 7.69 × 10−6) was nominally associated with glucose uptake, glucose oxidation rate, intramuscular triglyceride content, and metabolic flexibility (P = 0.016–0.048; n = 178). We explored eQTL results using published data from genome-wide association studies (DIAGRAM and MAGIC), and a proxy for the PFKM eQTL (rs11168327; r2 = 0.75) was nominally associated with T2D (DIAGRAM P = 2.7 × 10−3). Taken together, our analysis highlights PFKM as a potential regulator of skeletal muscle insulin sensitivity.

Altered Desaturation and Elongation of Fatty Acids in Hormone-Sensitive Lipase Null Mice

Background Hormone-sensitive lipase (HSL) is expressed predominantly in adipose tissue, where it plays an important role in catecholamine-stimulated hydrolysis of stored lipids, thus mobilizing fatty acids. HSL exhibits broad substrate specificity and besides acylglycerides it hydrolyzes cholesteryl esters, retinyl esters and lipoidal esters. Despite its role in fatty acid mobilization, HSL null mice have been shown to be resistant to diet-induced obesity. The aim of this study was to define lipid profiles in plasma, white adipose tissue (WAT) and liver of HSL null mice, in order to better understand the role of this multifunctional enzyme. Methodology/Principal Findings This study used global and targeted lipidomics and expression profiling to reveal changed lipid profiles in WAT, liver and plasma as well as altered expression of desaturases and elongases in WAT and liver of HSL null mice on high fat diet. Decreased mRNA levels of stearoyl-CoA desaturase 1 and 2 in WAT were consistent with a lowered ratio of 16∶1n7/16∶0 and 18∶1n9/18∶0 in WAT and plasma. In WAT, increased ratio of 18∶0/16∶0 could be linked to elevated mRNA levels of the Elovl1 elongase. Conclusions This study illustrates the importance of HSL for normal lipid metabolism in response to a high fat diet. HSL deficiency greatly influences the expression of elongases and desaturases, resulting in altered lipid profiles in WAT, liver and plasma. Finally, altered proportions of palmitoleate, a recently-suggested lipokine, in tissue and plasma of HSL null mice, could be an important factor mediating and contributing to the changed lipid profile, and possibly also to the decreased insulin sensitivity seen in HSL null mice.

Rose hip exerts antidiabetic effects via a mechanism involving downregulation of the hepatic lipogenic program

The aim of this study was to investigate the metabolic effects of a dietary supplement of powdered rose hip to C57BL/6J mice fed a high-fat diet (HFD). Two different study protocols were used; rose hip was fed together with HFD to lean mice for 20 wk (prevention study) and to obese mice for 10 wk (intervention study). Parameters related to obesity and glucose tolerance were monitored, and livers were examined for lipids and expression of genes and proteins related to lipid metabolism and gluconeogenesis. A supplement of rose hip was capable of both preventing and reversing the increase in body weight and body fat mass imposed by a HFD in the C57BL/6J mouse. Oral and intravenous glucose tolerance tests together with lower basal levels of insulin and glucose showed improved glucose tolerance in mice fed a supplement of rose hip compared with control mice. Hepatic lipid accumulation was reduced in mice fed rose hip compared with control, and the expression of lipogenic proteins was downregulated, whereas AMP-activated protein kinase and other proteins involved in fatty acid oxidation were unaltered. Rose hip intake lowered total plasma cholesterol as well as the low-density lipoprotein-to-high-density lipoprotein ratio via a mechanism not involving altered gene expression of sterol regulatory element-binding protein 2 or 3-hydroxymethylglutaryl-CoA reductase. Taken together, these data show that a dietary supplement of rose hip prevents the development of a diabetic state in the C57BL/6J mouse and that downregulation of the hepatic lipogenic program appears to be at least one mechanism underlying the antidiabetic effect of rose hip.

The hormone-sensitive lipase C–60G promoter polymorphism is associated with increased waist circumference in normal-weight subjects

Objective:Hormone-sensitive lipase (HSL) is a key enzyme in the mobilization of fatty acids from triglyceride stores in adipocytes. The aim of the present study was to investigate the role of the HSL gene promoter variant C-60G, a polymorphism which previously has been associated with reduced promoter activity in vitro, in obesity and type 2 diabetes.Design:We genotyped two materials consisting of obese subjects and non-obese controls, one material with offspring-parents trios, where the offspring was abdominally obese and one material with trios, where the offspring had type 2 diabetes or impaired glucose homeostasis. HSL promoter containing the HSL C-60G G-allele was generated and tested against a construct with the C-allele in HeLa cells and primary rat adipocytes. HSL mRNA levels were quantified in subcutaneous and visceral fat from 33 obese subjects.Results:We found that the common C-allele was associated with increased waist circumference and WHR in lean controls, but there was no difference in genotype frequency between obese and non-obese subjects. There was a significant increased transmission of C-alleles to the abdominally obese offspring but no increased transmission of C-alleles was observed to offspring with impaired glucose homeostasis. The G-allele showed reduced transcription in HeLa cells and primary rat adipocytes. HSL mRNA levels were significantly higher in subcutaneous compared to visceral fat from obese subjects.Conclusion:The HSL C-60G polymorphism is associated with increased waist circumference in non-obese subjects.