Changes in adipocytes and dendritic cells in lymph node containing adipose depots during and after many weeks of mild inflammation

The time course and cellular basis for inflammation-induced hypertrophy of adipose tissue were investigated over 20 weeks in mature male rats. Mild inflammation was induced by subcutaneous injection of 20 microg lipopolysaccharide into one hind-leg three times/week for 4 or 8 weeks, followed by up to 12 weeks 'rest' without intervention. Mean volume and frequency of apoptosis (TUNEL assay) were measured in adipocytes isolated from sites defined by their anatomical relations to lymph nodes, plus numbers of CCL21-stimulated lymph node-derived and adipose tissue-derived dendritic cells. Experimental inflammation increased dendritic cells and adipocyte apoptosis in the locally stimulated popliteal depot and the lymphoid tissue-associated regions of the contralateral popliteal and mesentery and omentum. Responses declined slowly after inflammation ended, but all measurements from the locally stimulated popliteal depot, and the omentum, were still significantly different from controls after 12 weeks rest. The locally stimulated popliteal adipose tissue enlarged by 5% within 4 weeks and remained larger than the control. We conclude that prolonged inflammation induces permanent enlargement, greater adipocyte turnover and increased dendritic cell surveillance in the adjacent adipose tissue and the omentum. The experiment suggests a mechanism for selective hypertrophy of lymphoid tissue-associated adipose tissue in chronic stress and inflammatory disorders, including impaired lymph drainage, Crohn's disease and HIV-associated lipodystrophy, and a link between evolutionary fitness, sexual selection and aesthetically pleasing body symmetry. It would be useful for further study of molecular mechanisms in inflammation-induced local hypertrophy of adipose tissue and development of specific therapies that avoid interference with whole-body lipid metabolism.     

Vascularisation in adipose depots surrounding immune-stimulated lymph nodes

We report a change in the vascularisation of the adipose depots surrounding the popliteal lymph node that has, and the contralateral node that has not, been exposed to a simulated immune challenge. The percentage of the depot that consists of vessels, as measured by image analysis, decreases over a period of 2 d after immune stimulus, then increases in a biphasic manner over the next 2–3 wk. By 1 mo after the stimulus, the vascularisation has returned to baseline values. The adipose tissue surrounding both the stimulated and the unstimulated lymph nodes shows a similar pattern, but the unstimulated depot lags by 3–6 d in reaching its maximum vascularisation. These data support the hypothesis that perinodal adipose tissue is involved in peripheral immune responses.     

Immunofluorescent localisation of tumour necrosis factor-α receptors on the popliteal lymph node and the surrounding adipose tissue following a simulated immune challenge

We used immunohistochemical techniques to demonstrate the distribution of receptors for the cytokine tumour necrosis factor-α on the popliteal lymph node and the adipose tissue surrounding it for 5 d following a simulated immune challenge to one hind leg in the rat. We found different patterns of expression of receptors on adipocytes surrounding a lymph node to a distance of about 1 mm, and on those more remote from the node. Sites recognised by an antibody to type I tumour necrosis factor receptors appeared on the challenged node and the adipocytes surrounding it within 30 min of an injection of bacterial lipopolysaccharide, but appeared on adipocytes surrounding the unchallenged popliteal node only 24 h later. Adipocytes distant from the node, both within the same depot and in the contralateral depot, showed no response. Sites recognised by an antibody to type II tumour necrosis factor receptors were present at all times on lymph nodes and the adipocytes close to them, but appeared on more distant adipocytes only 24 h after immune challenge, in both challenged and unchallenged legs. These data support the proposal, based on in vitro studies, that the adipose tissue surrounding major lymph nodes is specialised to respond to cytokines derived from lymphoid cells, and participates in the immune responses of the adjacent node.     

Endogenous lipid antigens for invariant natural killer T cells hold the reins in adipose tissue homeostasis

The global obesity epidemic and its associated co‐morbidities, including type 2 diabetes, cardiovascular disease and certain types of cancers, have drawn attention to the pivotal role of adipocytes in health and disease. Besides their ‘classical’ function in energy storage and release, adipocytes interact with adipose‐tissue‐resident immune cells, among which are lipid‐responsive invariant natural killer T (iNKT) cells. The iNKT cells are activated by lipid antigens presented by antigen‐presenting cells as CD1d/lipid complexes. Upon activation, iNKT cells can rapidly secrete soluble mediators that either promote or oppose inflammation. In lean adipose tissue, iNKT cells elicit a predominantly anti‐inflammatory immune response, whereas obesity is associated with declining iNKT cell numbers. Recent work showed that adipocytes act as non‐professional antigen‐presenting cells for lipid antigens. Here, we discuss endogenous lipid antigen processing and presentation by adipocytes, and speculate on how these lipid antigens, together with ‘environmental factors’ such as tissue/organ environment and co‐stimulatory signals, are able to influence the fate of adipose‐tissue‐resident iNKT cells, and thereby the role of these cells in obesity and its associated pathologies.     

Chemical composition of the infrapatellar fat pad of swine

The porcine infrapatellar fat pad is a structure composed of adipocytes and adipose connective tissues. Limited information is available concerning its chemical composition. Samples of the fat pad collected from young hogs were dissected into two portions: a relatively hard core of the pad with cushioning properties (inner tissue), and a soft adipose tissue surrounding the core (outer tissue). The inner tissue contained less moisture and nitrogen than did the outer tissue. The yield of dry-delipidated tissue was also lower in the inner tissue, indicating a higher content of lipid in this tissue. Fatty acid analysis showed that the proportions of C18: 1, C16: 1 and C18: 2n-6 are higher, and the proportion of C16: 0 is lower in the inner than in the outer tissue. Collagen is the major protein, with relatively small amounts of glycosaminoglycans in both tissues. The content of hyaluronic acid relative to sulphated galactosaminoglycan was lower in the inner than in the outer tissue. The electrophoresis pattern of sulphated galactosaminoglycan was also different between the two tissues. These results suggest that chemical composition varies between adipose tissues with different biomechanical function.     

Oxygen consumption in undifferentiated versus differentiated adipogenic mesenchymal precursor cells

To date, no adequate implant material for the correction of soft tissue defects such as after extensive deep burns, tumor resections or in congenital defects is available. A biohybrid composed of viable adipose precursor cells and an optimised matrix could help towards a solution. Morphologically, preadipocytes resemble fibroblasts and have not yet built a large cytoplasmic lipid droplet as found in differentiated adipocytes. Additionally, preadipocytes are smaller than mature adipocytes allowing a quicker revascularization after transplantation. Furthermore, transplanted preadipocytes can form adipose tissue in vivo whereas the transplantation of mature adipocytes often gives poor results, i.e. oil cysts or shrinkage of the transplant. Since these observations point to differences in metabolic activity between preadipocytes and adipocytes, we investigated the oxygen consumption of preadipocytes stimulated to undergo differentiation, and fibroblasts, by measuring the respiration with a Clark-type oxygen electrode. Preadipocytes had a significantly lower oxygen consumption than mature adipocytes. This advantage in respiration and the better revascularization of undifferentiated adipose tissue cells allow the development of innovative transplants and point to preadipocytes as promising tool to improve transplantations in adipose tissue reconstruction.     

Alpha-syntrophin deficient mice are protected from adipocyte hypertrophy and ectopic triglyceride deposition in obesity

Alpha-syntrophin (SNTA) is a molecular adapter protein which is expressed in adipocytes. Knock-down of SNTA in 3T3-L1 preadipocytes increases cell proliferation, and differentiated adipocytes display small lipid droplets. These effects are both characteristics of healthy adipose tissue growth which is associated with metabolic improvements in obesity. To evaluate a role of SNTA in adipose tissue morphology and obesity associated metabolic dysfunction, SNTA deficient mice were fed a standard chow or a high fat diet. Mice deficient of SNTA had less fat mass and smaller adipocytes in obesity when compared to control animals. Accordingly, these animals did not develop liver steatosis and did not store excess triglycerides in skeletal muscle upon high fat diet feeding. SNTA?/? animals were protected from hyperinsulinemia and hepatic insulin resistance. Of note, body-weight, food uptake, and serum lipids were normal in the SNTA null mice. SNTA was induced in adipose tissues but not in the liver of diet induced obese and ob/ob mice. In human subcutaneous and visceral fat of seven patients SNTA was similarly expressed and was not associated with body mass index. Current data demonstrate beneficial effects of SNTA deficiency in obesity which is partly attributed to smaller adipocytes and reduced white adipose tissue mass. Higher SNTA protein in fat depots of obese mice may contribute to adipose tissue hypertrophy and ectopic lipid deposition which has to be confirmed in humans.     

Angiomyomatous hamartoma of a popliteal lymph node: an unusual cause of posterior knee pain

Angiomyomatous hamartoma is a primary vascular tumor primarily found in the inguinal and femoral lymph nodes characterized by the replacement of nodal tissue by smooth muscle cells and fibrous tissue in sclerotic lymphatic stroma. There has been 1 report of an angiomyomatous hamartoma of a cervical lymph node, and this is the first reported case occurring in an extremity. We present a case of angiomyomatous hamartoma occurring in a single popliteal lymph node.     

Effects of different fatty acids and dietary lipids on adiponectin gene expression in 3T3-L1 cells and C57BL/6J mice adipose tissue

Obesity is positively correlated to dietary lipid intake, and the type of lipid may play a causal role in the development of obesity-related pathologies. A major protein secreted by adipose tissue is adiponectin, which has antiatherogenic and antidiabetic properties. The aim of this study was to evaluate the effects of four different high-fat diets (enriched with soybean oil, fish oil, coconut oil, or lard) on adiponectin gene expression and secretion by the white adipose tissue (WAT) of mice fed on a selected diet for either 2 (acute treatment) or 60 days (chronic treatment). Additionally, 3T3-L1 adipocytes were treated for 48 h with six different fatty acids: palmitic, linoleic, eicosapentaenoic (EPA), docosahexaenoic (DHA), lauric, or oleic acid. Serum adiponectin concentration was reduced in the soybean-, coconut-, and lard-enriched diets in both groups. Adiponectin gene expression was lower in retroperitoneal WAT after acute treatment with all diets. The same reduction in levels of adiponectin gene expression was observed in epididymal adipose tissue of animals chronically fed soybean and coconut diets and in 3T3-L1 cells treated with palmitic, linoleic, EPA, and DHA acids. These results indicate that the intake of certain fatty acids may affect serum adiponectin levels in mice and adiponectin gene expression in mouse WAT and 3T3-L1 adipocytes. The effects appear to be time dependent and depot specific. It is postulated that the downregulation of adiponectin expression by dietary enrichment with soybean oil or coconut oil may contribute to the development of insulin resistance and atherosclerosis.     

Hibernoma: a possible model of brown fat histogenesis

Hibernomas are composed of cells remarkably similar to those observed in brown adipose tissue. A surgically resected hibernoma was observed to contain cells at all stages of maturation from immature, relatively lipid free cells to multiloculated and finally uniloculated adipocytes. The ultrastructural features of adipocytes contained in the hibernoma were compared to previously reported features of differentiating preadipocytes and lipid depleted mature adipocytes derived from human white adipose tissue. This comparison confirmed the existence of distinct mitochondrial and cytoplasmic membrane component differences at all developmental stages and suggests that brown and white adipose tissue are two unique histologic types.     

Microcirculation of the lymph node with metastases.

Microangiographic and histologic examination of the popliteal lymph node was performed on 49 rabbits 3 to 55 days after V2 tumor implantation into the hind paw. Control animals received subcutaneous tissue extract from normal rabbit donors. During the first 10 days after the tumor implant from an allogeneic animal donor, the draining lymph node exhibited a hypervascular response which after 2 weeks gradually subsided. Subsequently, during the early stages of lymph node metastasis there was still hypervascularity adjacent to the metastatic deposit. In about 4 weeks the metastases became more established and surrounded by layers of plasma cells. The hypervascular changes of the surrounding lymph node subsided by this time. In lymph node metastases the microvasculature could be an indicator of the immunologic activity of the host.     

Adipocytes and macrophages secretomes coregulate catecholamine-synthesizing enzymes

Obesity associates with macrophage accumulation in adipose tissue where these infiltrating cells interact with adipocytes and contribute to the systemic chronic metabolic inflammation present in immunometabolic diseases. Tyrosine hydroxylase (TH) and phenylethanolamine N-methyltransferase (PNMT) are two of the main enzymes of catecholamines (CA) synthesis. Adipocytes and macrophages produce, secrete and respond to CA, but the regulation of their synthesis in the interplay between immune and metabolic systems remains unknown. A model of indirect cell coculture with conditioned medium (CM) from RAW 264.7 macrophages with or without LPS-activation and 3T3-L1 adipocytes and preadipocytes was established to study the effect of cellular secretomes on the expression of the above enzymes. During the adipocyte differentiation process, we found a decrease of TH and PNMT expression. The secretome from LPS-activated macrophages downregulated TH and PNMT expression in preadipocytes, but not in mature adipocytes. Mature adipocytes CM induced a decrease of PNMT levels in RAW 264.7 macrophages. Pre and mature adipocytes showed a similar pattern of TH, PNMT and peroxisome proliferator-activated receptor gamma expression after exposure to pro and anti-inflammatory cytokines. We evidenced macrophages and adipocytes coregulate the expression of CA synthesis enzymes through secretome, with non-inflammatory signaling networks possibly being involved. Mediators released by macrophages seem to equally affect CA production by adipocytes, while adipocytes secretome preferentially affect AD production by macrophages. CA synthesis seems to be more determinant in early stages of adipogenic differentiation. Our results suggest that CA are key signaling molecules in the regulation of immune-metabolic crosstalk within the adipose tissue.     

Efficient proliferation and adipose differentiation of human adipose tissue-derived vascular stromal cells transfected with basic fibroblast growth factor gene

Human vascular stromal (VS) cells obtained from mature adipose tissue were transfected with an adenovirus vector carrying the basic fibroblast growth factor (bFGF) gene. bFGF protein was observed in VS cell nuclei 24 h after transfection and in the cytoplasm and extracellular space 72 h after transfection. Naive VS cells were almost static in vitro and proliferated in a dose-dependent manner on stimulation with recombinant bFGF (rbFGF). However, bFGF-transfected VS cells proliferated spontaneously to the same extent as naive VS cells when stimulated with rbFGF at 100 ng/ml. The former cells started to proliferate on day 3 after transfection and the proliferation pattern was similar to that of the latter cells, although only a slight amount of bFGF protein was detected in the culture medium when the bFGF-transfected cells started to proliferate. The proliferation of bFGF-transfected VS cells was completely inhibited by bFGF neutralizing antibody, which also completely inhibited the proliferation of naive VS cells stimulated with rbFGF. Under conditions favoring differentiation to adipocytes, bFGF-transfected VS cells stopped proliferating and started to accumulate lipid in the cytoplasm. bFGF-transfected VS cells, which spontaneously and efficiently proliferate while preserving their ability to differentiate into adipocytes, may be an adequate cell source for human adipose tissue regeneration.     

Quantitative image analysis in adipose tissue using an automated image analysis system: differential effects of peroxisome proliferator-activated receptor-alpha and -gamma agonist on white and brown adipose tissue morphology in AKR obese and db/db diabetic mice

Morphometric analysis of adipocytes is widely used to demonstrate the effects of antiobesity drugs or anti-diabetic drugs on adipose tissues. However, adipocyte morphometry has been quantitatively performed by manual object extraction using conventional image analysis systems. The authors have developed an automated quantitative image analysis method for adipose tissues using an innovative object-based quantitative image analysis system (eCognition). Using this system, it has been shown quantitatively that morphological features of adipose tissues of mice treated with peroxisome proliferator-activated receptor (PPAR) agonists differ dramatically depending on the type of PPAR agonist. Marked alteration of morphological characteristics of brown adipose tissue (BAT) treated with GI259578A, a PPAR-alpha agonist, was observed in AKR/J (AKR) obese mice. Furthermore, there was a 22.8% decrease in the mean size of adipocytes in white adipose tissue (WAT) compared with vehicle. In diabetic db/db mice, the PPAR-gamma agonist GW347845X decreased the mean size of adipocytes in WAT by 15.4% compared with vehicle. In contrast to changes in WAT, GW347845X increased the mean size of adipocytes in BAT greatly by 96.1% compared with vehicle. These findings suggest that GI259578A may activate fatty acid oxidation in BAT and that GW347845X may cause adipocyte differentiation in WAT and enhancement of lipid storage in BAT.     

Lymphoepithelial cyst of the pancreas in a 65-year-old man.

A case of an extremely rare cystic lesion of the pancreas is presented. The multilocular cyst was found adjacent to the upper border of the pancreatic body, and the cyst contained bean curd lees-like substances. Histologically, the cyst wall consisted of mature keratinizing squamous epithelium and surrounding lymphoid tissue stroma, and the cyst was filled with keratinized materials. A histopathologic diagnosis of typical lymphoepithelial cyst of the pancreas, proposed by Truong et al (Am J Surg Pathol 11:899-903, 1987), was made. Its histogenesis is still unknown; however, we hypothesize that it might arise from a benign epithelial inclusion of a peripancreatic lymph node, followed by squamous metaplasia of the epithelial inclusion. We recently found a retropyloric lymph node with a squamous epithelial inclusion, which might support this hypothesis regarding the histogenesis of the cyst.     

The Effects of Estrogens on Regional Adipose Tissue Cellularity in the Rat

The effects of high and low dose estrogen administration on adipose tissue cellularity and metabolism were investigated and compared with the effect of total starvation in female rats. Total starvation in female rats resulted in local fat cell size decrease which was, in different adipose tissue regions, directly proportional to the primary fat cell size. Thus, the largest fat cell size decrease was observed in the parametrial depot of adipose tissue with primarily the largest adipocytes and the smallest decrease of fat cell size in the subcutaneous adipose tissue, with primarily the smallest adipocytes. High doses of estrogen (Estradurin®—120 μg/km/gonth) resulted in a body and adipose tissue weight decrease after 10 weeks treatment. Contrary to the starvation effect the greatest fat cell size decrease was observed in the subcutaneous depot with primarily the smallest adipocytes. Lower estrogen dose (60 μg/kg/month) resulted, after 4 weeks treatment, in weight increase of adipose tissue. Fat cell size increased most in the parametrial depot—an adipose tissue region with primarily the biggest adipocytes. This is in contrast to the situation after insulin treatment and overfeeding which produce the largest size increase in the region with the smallest fat cells, and can be interpreted as evidence for regional specificity of the estrogen effect on fat cells.     

Mesenteric lymph node stroma cells in the generation of intestinal immune responses

Lymph nodes at different anatomical locations share similar architecture and operate on the basis of identical principles. Still, the quality of immune responses is modified substantially by the local peculiarities at the site of its induction. Here, we discuss how lymph node stroma cells contribute to functional differences between various lymph nodes, thus helping to explain why and how an immune response induced in skin draining peripheral lymph nodes differs from that elicited in the gut draining mesenteric lymph nodes. Stroma cells constitute a major part of the lymph node scaffold and control the flow of immune cells as well as soluble substances within the organ. Moreover, stroma cells express cytokines, chemokines as well as adhesion factors and thereby actively influence immune status. Lymph node transplantations and adoptive transfers of dendritic cells demonstrated that regional lymph node stroma cells differ in their ability to support mucosal tolerance, the induction of tissue tropism, and humoral immunity. This suggests that stroma cells shape tissue-specific immune responses and equip lymph nodes with unique functional properties that might originate during lymph node organogenesis.     

Resident and migratory adipose immune cells control systemic metabolism and thermogenesis

Glucose is a vital source of energy for all mammals. The balance between glucose uptake, metabolism and storage determines the energy status of an individual, and perturbations in this balance can lead to metabolic diseases. The maintenance of organismal glucose metabolism is a complex process that involves multiple tissues, including adipose tissue, which is an endocrine and energy storage organ that is critical for the regulation of systemic metabolism. Adipose tissue consists of an array of different cell types, including specialized adipocytes and stromal and endothelial cells. In addition, adipose tissue harbors a wide range of immune cells that play vital roles in adipose tissue homeostasis and function. These cells contribute to the regulation of systemic metabolism by modulating the inflammatory tone of adipose tissue, which is directly linked to insulin sensitivity and signaling. Furthermore, these cells affect the control of thermogenesis. While lean adipose tissue is rich in type 2 and anti-inflammatory cytokines such as IL-10, obesity tips the balance in favor of a proinflammatory milieu, leading to the development of insulin resistance and the dysregulation of systemic metabolism. Notably, anti-inflammatory immune cells, including regulatory T cells and innate lymphocytes, protect against insulin resistance and have the characteristics of tissue-resident cells, while proinflammatory immune cells are recruited from the circulation to obese adipose tissue. Here, we review the key findings that have shaped our understanding of how immune cells regulate adipose tissue homeostasis to control organismal metabolism.     

Long-term intermittent compressive stimulation improves the composition and mechanical properties of tissue-engineered cartilage

Tissue engineering of articular cartilage is a promising alternative for cartilage repair. However, it has been difficult to develop tissue in vitro that mimicks native cartilage. Cartilaginous tissue formed in vitro does not accumulate enough extracellular matrix, is deficient in collagen, and possesses only a fraction of the mechanical properties of native cartilage. In this study, we investigated whether long-term intermittent compressive stimulation would improve the quality of the generated tissue. Chondrocyte cultures were established on the surface of porous calcium polyphosphate substrates and allowed to form cartilaginous tissue. In vitro-formed tissues were subjected to different stimulation protocols for 1 week. The optimal mechanical stimulation parameters identified in this short-term study were then applied to the cultures for up to 4 weeks. Mechanical stimulation applied at a 5% compressive amplitude at a frequency of 1 Hz for 400 cycles every second day resulted in the greatest increase in collagen synthesis (37 +/- 9% over control) while not significantly affecting proteoglycan synthesis (2 +/- 8% over control). This condition, applied to the chondrocyte cultures for 4 weeks, resulted in a significant increase in the amount of tissue that formed (stimulated, 2.4 +/- 0.2 mg dry wt; unstimulated, 1.61 +/- 0.08 mg dry wt). Stimulated tissues contained approximately 40% more collagen (stimulated, 590 +/- 58 microg; unstimulated, 420 +/- 42 microg), and 30% more proteoglycans (stimulated, 393 +/- 34 microg; unstimulated, 302 +/- 32 microg) as well as displaying a 2- to 3-fold increase in compressive mechanical properties (maximal equilibrium stress: stimulated, 10 +/- 1 kPa; unstimulated, 5 +/- 1 kPa; maximal equilibrium modulus: stimulated, 80 +/- 23 kPa; unstimulated, 24 +/- 6 kPa). The results of this study demonstrate that intermittent mechanical stimulation can increase collagen synthesis and, when applied over a 4-week period, can accelerate extracellular matrix accumulation as well as improve the material properties of the developed tissue. Interestingly, only short periods of mechanical stimulation (6 min every second day) were needed to affect the quality of cartilaginous tissue formed in vitro.