Eva Jennische

Separation of human adipocytes by size: hypertrophic fat cells display distinct gene expression

Enlarged adipocytes are associated with insulin resistance and are an independent predictor of type 2 diabetes. To understand the molecular link between these diseases and adipocyte hypertrophy, we developed a technique to separate human adipocytes from an adipose tissue sample into populations of small cells (mean 57.6±3.54 pm) and large cells (mean 100.1±3.94 pm). Microarray analysis of the cell populations separated from adipose tissue from three subjects identified 14 genes, of which five immune‐related, with more than fourfold higher expression in large cells than small cells. Two of these genes were serum amyloid A (SAA) and transmembrane 4 L six family member 1 (TM4SF1). Real‐time RT‐PCR analysis of SAA and TM4SF1 expression in adipocytes from seven subjects revealed 19‐fold and 22‐fold higher expression in the large cells, respectively, and a correlation between adipocyte size and both SAA and TM4SF1 expression. The results were verified using immunohistochemistry. In comparison with 17 other human tissues and cell types by microarray, large adipocytes displayed by far the highest SAA and TM4SF1 expression. Thus, we have identified genes with markedly higher expression in large, compared with small, human adipocytes. These genes may link hypertrophic obesity to insulin resistance/type 2 diabetes.—Jernås, M., Palming, J., Sjöholm, K., Jennische, E., Svensson, P.‐A., Gabrielsson, B. G., Levin, M., Sjögren, A., Rudemo, M., Lystig, T. C., Carlsson, B., Carlsson, L. M. S., Lönn, M. Separation of human adipocytes by size: hypertrophic fat cells display distinct gene expression. FASEB J. 20, E832–E839 (2006)

Adipose Tissue Has Aberrant Morphology and Function in PCOS: Enlarged Adipocytes and Low Serum Adiponectin, But Not Circulating Sex Steroids, Are Strongly Associated with Insulin Resistance

CONTEXT Comprehensive characterization of the adipose tissue in women with polycystic ovary syndrome (PCOS), over a wide range of body mass indices (BMIs), is lacking. Mechanisms behind insulin resistance in PCOS are unclear. OBJECTIVE To characterize the adipose tissue of women with PCOS and controls matched pair-wise for age and BMI, and to identify factors, among adipose tissue characteristics and serum sex steroids, that are associated with insulin sensitivity in PCOS. DESIGN/OUTCOME MEASURES Seventy-four PCOS women and 31 controls were included. BMI was 18-47 (PCOS) and 19-41 kg/m(2) (controls). Anthropometric variables, volumes of subcutaneous/visceral adipose tissue (magnetic resonance imaging; MRI), and insulin sensitivity (clamp) were investigated. Adipose tissue biopsies were obtained to determine adipocyte size, lipoprotein lipase (LPL) activity, and macrophage density. Circulating testosterone, free testosterone, free 17β-estradiol, SHBG, glycerol, adiponectin, and serum amyloid A were measured/calculated. RESULTS Comparison of 31 pairs revealed lower insulin sensitivity, hyperandrogenemia, and higher free 17β-estradiol in PCOS. Abdominal adipose tissue volumes/distribution did not differ in the groups, but PCOS women had higher waist-to-hip ratio, enlarged adipocytes, reduced adiponectin, and lower LPL activity. In regression analysis, adipocyte size, adiponectin, and waist circumference were the factors most strongly associated with insulin sensitivity in PCOS (R(2)=0.681, P < 0.001). CONCLUSIONS In PCOS, adipose tissue has aberrant morphology/function. Increased waist-to-hip ratio indicates abdominal/visceral fat accumulation, but this is not supported by MRI. Enlarged adipocytes and reduced serum adiponectin, together with a large waistline, rather than androgen excess, may be central factors in the pathogenesis/maintenance of insulin resistance in PCOS.

Regenerating endothelial cells express insulin-like growth factor-I immunoreactivity after arterial injury

SummaryIn the present study the expression of insulin-like growth factor I (IGF-I; somatomedin C) immunoreactivity was examined in endothelial cells during repair after injury to the intima in the femoral artery of adult rats. Two types of injury were examined: (1) endothelial denudation induced by the use of a catheter, and (2) vessel compression by short-term ligation. In untreated rats, arterial endothelial cells showed no or, only infrequently, low IGF-I immunoreactivity in their cytoplasm. Endothelial cells at the border to the denuded area showed increased IGF-I immunoreactivity one day after injury to the intima of the femoral artery. Thrombocytes and fibrin deposits as well as vital endothelial cells, covered by clots, were immunonegative. The maximal intensity of IGF-I immunoreactivity was reached within 3 days after insult. The IGF-I immunoreactivity in the endothelial cells remained elevated for at least 4 weeks, compared to the controls. Intimai thickenings appeared within a week after injury and many cells in these thickenings showed intense IGF-I immunoreactivity as did the covering endothelial cells. Smooth muscle cells in the media were generally immunonegative during control conditions and after endothelial denudation. Spontaneously hypertensive rats (SHR) showed, similarly to their matched controls (WKY), approximately the same patterns of IGF-I immunoreactivity in their endothelial cells both under normal conditions and after injury. It is concluded that IGF-I is likely to be involved in the repair of the intima in injured arteries.

Evans Blue Permeation of Intestinal Mucosa in the Rat

The azo dye Evans blue (EB; molecular weight, 960.83) is widely used as an indicator of increased capillary permeability. In the present study, however, rat gut absorption of EB was investigated after dye instillation in either the small or large intestine. During a brief period of ether anaesthesia, EB was injected either into jejunal loops with a challenge period of 30 or 60 min or into a proximal and a distal colon loop with a challenge period of 30, 60, or 120 min. After the rats had been killed the intestinal specimens were washed with 6 mM acetylcysteine dissolved in phosphate-buffered saline, which efficiently cleared the tissues of mucus, and thus of EB trapped in mucus. Only EB absorbed by the gut wall remained to be estimated, and this absorption was found to be both dose- and time-dependent in the jejunum and the colon. After instillation in the colon, but not in jejunum, EB could be detected in the blood. EB absorption from the jejunum remained unaffected by the addition of either ouabain (1 mM) or lidocaine (0.38 mM). Either of these compounds inhibited EB uptake in the proximal part of the colon, while enhancing it in the distal part. Fluorescence microscopy showed penetration into the intestinal wall to be a prerequisite for EB to become fluorescent, and EB fluorescence increased with time. It is proposed that EB is transported over the mucosa by the paracellular route and that the amount of absorbed EB reflects epithelial permeability differently in different parts of the gastrointestinal tract.(ABSTRACT TRUNCATED AT 250 WORDS)

Maternal Endotoxemia Results in Obesity and Insulin Resistance in Adult Male Offspring1

Events in utero appear to be important factors contributing to the development of somatic disorders at adult age. The aim of this study was to examine whether maternal immune challenge would be followed at adult age by metabolic and endocrine abnormalities in the offspring. Pregnant rats were given injections of either endotoxin (Escherichia coli lipopolysaccharide; 0.79 mg/kg, ip) or vehicle on days 8, 10, and 12 of gestation. Adult male offspring to lipopolysaccharide-exposed dams were heavier than controls (P < 0.05) and showed increased adipose tissue weights (P < 0.05), elevated food intake (P < 0.05), and increased circulating leptin (P < 0.01). The effect of insulin on glucose uptake was reduced, as measured by an euglycemic hyperinsulinemic clamp technique (P < 0.05). Serum levels of 17β-estradiol and progesterone were elevated (P< 0.01 and P < 0.05, respectively). Baseline levels of corticosterone were normal, but the corticosterone response to stress was attenuated (P < 0.05), and hippocampal gl...

Food induced stimulation of the antisecretory factor can improve symptoms in human inflammatory bowel disease: a study of a concept

BACKGROUND Antisecretory factor (AF), a 41 kDa cloned and sequenced protein, suppresses intestinal inflammation and hypersecretion in animals. Endogenous AF production can be induced by dietary modifications in several animal species, and this feed has been shown to reduce the incidence of diarrhoeal disease in weaning piglets. The role of AF in intestinal disease in humans is not known. AIMS To study the effects of hydrothermally processed cereals, optimised for AF induction in animals, added to the diet of patients with longstanding symptoms of inflammatory bowel disease (IBD). PATIENTS Fifty three patients with IBD (ulcerative colitis and Crohns disease) were entered into the study, and 50 completed follow up. The experimental group consisted of 16 females (mean age 50 (SEM 5) years) and 10 males (41 (4) years) and the placebo group of 12 women (41 (4) years old) and 12 men (51 (5) years). METHODS Patients were randomised to receive either hydrothermally processed cereals (active treatment) or the same amount of ordinary cereals (placebo treatment) for four weeks in a double blind study design. Baseline diet and medications remained unchanged. Bowel symptoms, plasma levels of AF, and colonic biopsies were evaluated before and after treatment. RESULTS The active treatment significantly improved subjective ratings of clinical symptoms and increased plasma AF levels compared with placebo. Plasma lipid levels were unaffected. CONCLUSION Hydrothermally processed cereals can induce AF production in human IBD. This increase in endogenous AF activity is associated with clinical improvement. Further studies are warranted to clarify the exact role of AF in human intestinal disease.

Broad Up-Regulation of Innate Defense Factors during Acute Cholera

ABSTRACT We used a whole-genome microarray screening system (Affymetrix human GeneChips covering 47,000 different transcripts) to examine the gene expression in duodenal mucosa during acute cholera. Biopsies were taken from the duodenal mucosa of seven cholera patients 2 and 30 days after the onset of diarrhea, and the gene expression patterns in the acute- and convalescent-phase samples were compared pairwise. Of about 21,000 transcripts expressed in the intestinal epithelium, 29 were defined as transcripts that were up-regulated and 33 were defined as transcripts that were down-regulated during acute cholera. The majority of the up-regulated genes characterized were found to have an established or possible role in the innate defense against infections; these genes included the LPLUNC1, LF, VCC1, TCN1, CD55, SERPINA3, MMP1, MMP3, IL1B, LCN2, SOCS3, GDF15, SLPI, CXCL13, and MUC1 genes. The results of confirmative PCR correlated well with the microarray data. An immunohistochemical analysis revealed increased expression of lactoferrin in lamina propria cells and increased expression of CD55 in epithelial cells, whereas increased expression of the SERPINA3 protein (α1-antichymotrypsin) was detected in both lamina propria and epithelial cells during acute cholera. The expression pattern of CD55 and SERPINA3 in cholera toxin (CT)-stimulated Caco-2 cells was the same as the pattern found in the intestinal mucosa during acute cholera, indicating that the activation of the CD55 and SERPINA3 genes in intestinal epithelium was induced by CT. In conclusion, during acute cholera infection, innate defense mechanisms are switched on to an extent not described previously. Both direct effects of CT on the epithelial cells and changes in the lamina propria cells contribute to this up-regulation.

Dynamic changes in insulin-like growth factor. I: Immunoreactivity correlate to repair events in rat ear after freeze-thaw injury

The distribution of the trophic peptide insulin-like growth factor I (IGF-I) in the rat ear was mapped after freeze-thaw injury. Immunocytochemical methods and antisera specific to human IGF-I were used. In the ear of normal adult rats scattered basal epidermal cells and a few cells in the underlying connective tissue and elastic cartilage expressed IGF-I immunoreactivity. Within 1 day after injury and reaching maximum in 3 days, all epidermal cells became stained as did invading macrophages and some of the other inflammatory cells. Concomitantly, there were hypertrophic changes. The staining leveled off after 1-2 weeks. Perichondrial cells became IGF-I immunoreactive in increasing frequency during the first week, reached maximal intensity and frequency in 2 weeks, and remained stained for at least 4 weeks. New cartilage was formed concomitantly on both sides of the old one. It is proposed that IGF-I is a substance of general importance for cell maintenance and tissue repair.

Influence of Topical Rectal Application of Drugs on Dextran Sulfate-Induced Colitis in Rats

A rat model of colitis [dextran sulfate (DSS)]was used to study the permeation of Evans blue (EB) fromthe lumen into the wall of proximal and distal colonicloops after exposure to the dye for 2 hr. Topical application of drugs used in human ulcerativecolitis (lidocaine, mesalazine, prednisolone, orsucralfate) was given daily during induction of colitisto protect the mucosa. The mucosal changes wereevaluated with special regard to peptidergic innervation[substance P (SP) and neuropeptide Y (NPY)], invasion ofantigen-presenting polydendritic cells, andmucin-containing goblet cells. DSS-treatment caused a significantly increased permeation of EB. Inthe proximal loops a significant inhibition was obtainedafter treatment with lidocaine, prednisolone, orsucralfate. In the distal loops only treatment with lidocaine had a preventive effect.Immunocytochemically there was a clear hyperplasia ofboth mucosal SP- and NPY-immunoreactive nerve fibers inregions with crypt abnormalities. In these regions alsomost of the goblet cells were devoid of mucus. Likethe changes in permeation, these morphological changeswere most prominent in the distal loops. With inductionof colitis, the mucosa and lamina propria were invaded by polydendritic cells; the visualscore was markedly decreased in the proximal loopstreated with lidocaine, prednisolone, or sucralfate. Inthe distal loops similar effects were obtained after treatment with lidocaine or prednisolone.Prevention of the influx of antigens in both loops afterlidocaine treatment with reduced recruitment ofpolydendritic cells into the lamina propria issuggested. The nerve hyperplasia may thus be secondary toluminal challenge with antigens during induction ofcolitis. The discrepancy between increased permeationand absence of polydendritic cell response in the distal loops after prednisolone may reflectseparate actions of steroids on the intestinalepithelium and the immune cells.

Antisecretory factor suppresses intestinal inflammation and hypersecretion

Background—Antisecretory factor (AF) is a recently identified regulatory protein which inhibits the intestinal fluid secretion induced by cholera toxin. Aims—To test the effect of AF on: (a) inflammation and hypersecretion induced by toxin A fromClostridium difficile; and (b) morphological changes and hypersecretion induced by okadaic acid (the blue mussel toxin) in rat intestinal mucosa. Methods—Morphological changes and fluid accumulation were observed in intestinal loops challenged with 1 μg of toxin A or 3  μg of okadaic acid administered before or after injection of 0.1  μg of recombinant AF (rAF). Results—The cytotoxic and inflammatory reaction caused by toxin A was abolished after treatment with rAF given either intraveneously or intraluminally prior to the toxin or one hour after the toxin. The intestinal fluid response induced by toxin A and okadaic acid was reduced 55–80% by rAF. However, the characteristic increase in goblet cells at the tips of villi in the okadaic acid treated mucosa was not inhibited by rAF. Conclusion—Results suggest that AF might be involved in protection against inflammation and in counteracting dehydration caused by enterotoxins. Both effects are probably mediated via the enteric nervous system.