18 research outputs found
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Fuelling an immune response: an ultrastructural study of immune-stimulated lymph nodes
A growing body of evidence suggests that adipocytes, and the triacylglycerol fatty acids within them, play an important role in supporting the inflammatory immune response. Inflammatory cytokines produced by an activated lymph node cause the release of fatty acids from neighbouring adipocytes, and the fatty acids are taken up into lymph node lymphoid cells, where they can be used as precursors for eicosanoid and plasma membrane synthesis, or as a fuel source. The mechanism of transport of fatty acids from adipocytes across the lymph node capsule and into lymphoid cells remains to be elucidated. Here we present evidence from light and electron microscopy that small fat-filled cells are found within the lymph node, and that 24 hours after an immune challenge they are associated with dendritic cells. The small adipocytes express S100 protein and insulin receptor, characteristics of mature adipocytes. Our observations suggest a mechanism for delivering triacylglycerol fatty acids directly to their site of use in supplying components for an immune response
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Three-dimensional cell culture
The present invention relates to a three-dimensional cell culture system for culturing adipocytes. The invention also relates to methods for culturing adipocytes and for obtaining adipocytes from preadipocytes in cell culture. Aspects of the invention further relate to methods for evaluating interactions between adipocytes and other cell types, and for evaluating responses of adipocytes to environmental stimuli. This enables a greater understanding of the mechanisms involved in cell and tissue interactions and has implications in the fields of obesity, diabetes, inflammation and related diseases
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Development of a three-dimensional co-culture system to study adipocyte and lymph node cell interactions
White adipose tissue is involved in localised paracrine interactions with lymph node cells. In this adipocyte-lymph node relationship, localised immune challenges stimulate the release of Tumour Necrosis Factor-α, and Interleukins-4,6,8 and10. These cytokines direct neighbouring adipocytes to respond by releasing fatty acids from their triacylglycerol stores. This mechanism is believed to be of great importance as it provides energy and metabolic precursors to lymphoid cells during the immune response. The aim of the study is to investigate the activity exhibited by lymph node cells and adipocytes in a long term in vitro 3-dimensional co-culture system. A number of experimental approaches are being applied to successfully deliver lymph node cells and to increase their viability within the system
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Three-dimensional adipocyte cultures and co-cultures to study tissue interactions
Adipocytes play many important roles within the body, though until recently their importance has frequently gone unrecognised. They are well known for their activities in fat metabolism and storage, but they are also significant secretory cells, and play roles in modulating immune function and provisioning stem cell development. Different adipose depots are specialised to interact in different ways with neighbouring cells, tissues and organs, and there is now an emerging body of evidence demonstrating their roles in areas as diverse as gut function, immune function and cardiac function.
Adipocytes are difficult to study in conventional culture: their buoyant density is such that they float, so are easily lost during medium changes, and their large size makes them too fragile to withstand many routine manipulations. When they lyse they release quantities of triacylglycerols which interfere with many biochemical analyses. Thus much of the work on adipocytes has had to be carried out in vivo.
We have developed a 3D culture system in which preadipocytes are differentiated to functional, mature adipocytes which retain many of the properties of ex vivo adipocytes. The adipocytes remain in place during medium changes, and their secretory activities can be monitored by assaying culture supernatants. Moreover, the culture conditions are mild, and flexible enough to accommodate co-cultures with cells requiring different culture conditions. It is therefore an ideal system for studying the interactions between adipocytes and other cell types that underpin tissue and organ function
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Adipocytes are present inside immune-stimulated lymph nodes
White adipose tissue is involved in localised paracrine interactions with lymph node lymphoid cells. In this adipocyte-lymph node relationship, localised immune challenges stimulate the release of inflammatory cytokines (1,2). These cytokines direct neighbouring adipocytes to respond by releasing fatty acids from their triacylglycerol stores (2,3). This mechanism is believed to be of great importance as it provides precursors for eicosanoid and plasma membrane synthesis, as well as a fuel source to lymph node lymphoid cells (4).
The mechanism of transport of fatty acids from adipocytes across the lymph node capsule and into lymphoid cells remains to be elucidated. Here we present evidence that a population of small adipocytes is found deep within the lymph node. We suggest that these small adipocytes are loaded with triacylglycerol fatty acids which are released during an immune response
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Culture and differentiation of preadipocytes in two-dimensional and three-dimensional in vitro systems
Adipogenesis is a complex process that involves
the differentiation of preadipocytes into mature
adipocytes. We have developed two-dimensional (2D)
and three-dimensional (3D) cell culture systems for
the purpose of culturing and differentiating primary
preadipocytes in vitro. Differentiating preadipocytes
show multiple lipid droplet accumulation and comparable
protein expression patterns to mature adipocytes in
vivo. We report that in both in vitro systems terminally
differentiated adipocytes show characteristics similar to
those of mature adipocytes in vivo, assessed by the expression
of the S100a/b protein, insulin receptor and
caveolin-1, and receptors for inflammatory mediators,
namely tumor necrosis factor-a receptors I and II
(TNFRI and TNFRII) and chemokine receptor 5
(CCR5). Our results demonstrate that the S100 protein,
caveolin-1, and insulin receptor are expressed and upregulated
in differentiating and terminally differentiated
cells. In addition, the receptors for TNFa are not present
in preadipocytes but are expressed in differentiating
preadipocytes and in differentiated adipocytes. Similarly,
CCR5 was exclusively expressed in differentiating
preadipocytes and terminally differentiated adipocytes,
but not in preadipocytes. Both 2D and 3D culture models
are highly robust and reproducible and offer the potential
to study adipogenesis and cellular interactions
closely resembling and comparable to those in vivo. Our
3D collagen system offers a distinct advantage over the
2D system in that the adipocytes remain confined within
the matrix and remain intact during biochemical analysis.
Moreover, the collagen matrix allows adipocytes to
closely simulate morphological characteristics and behavior
as in vivo whilst permitting manipulation of the
microenvironment in vitro to study adipogenesis
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Adipocytes: a role in immunological memory?
Many reports in the literature suggest the involvement of perinodal adipocytes in inflammatory immune responses taking place in immune-challenged lymph nodes. Triacylglycerols (TAG) from the diet accumulate in adipocytes, and pass to lymph node lymphoid cells where they are used to provision immune responses. These adipocytes are specialised: TAG in perinodal adipose tissue are relatively enriched in polyunsaturated fatty acids compared with adipocytes that are not in proximity to lymphoid tissue. Polyunsaturated fatty acids can be used as precursors of eicosanoids and other regulatory molecules, as well as for membrane biosynthesis and energy. Thus perinodal adipocytes are specialized to provision immune cell metabolism.
Dendritic cells are well known as antigen presenting cells. They are migratory and are found in lymph nodes, and in significant numbers among perinodal adipocytes. Their numbers are modulated by dietary fatty acids. Dendritic cells can induce primary immune responses and allow the establishment of immunological memory. They are known to interact with adipocytes in various situations such as Crohn’s disease, and it has been reported that developing dendritic cells accumulate lipid, which may derive from adipocytes. Thus dendritic cells are candidates for involvement in the transfer of TAG to lymphoid cells mounting an immune response.
Here we show by light and electron microscopy that small adipocyte-like cells are normal residents of the lymph node, and that following an immune challenge they are observed in association with dendritic cells deep within the stimulated node.
We also show in an in vitro co-culture system that both adipocytes and lymphoid cells exhibit altered metabolic activities when they are cultured together compared to when they are cultured alone. This is indirect evidence for a functional molecular interaction in vivo.
We propose that this co-localisation and putative interaction is the basis for provisioning a rapid immune response by dendritic cells. This phenomenon may therefore represent part of the mechanism for the development and/or deployment of immunological memory
Prodromal symptoms and temperamental characteristics in first episode psychotic mania: re-looking the cynosure
Background: Prodrome has recently been proposed as target for early intervention but their phenomenology remains obscure and often certain temperamental characteristics are confused with prodromal symptoms leading to the possibility of over-reporting. Methods: Fifty-one consecutively admitted patients, between the age group of 18-60 years, fulfilling the ICD-10 criteria for mania with psychotic symptoms were included. They were rated on Young Mania Rating Scale (YMRS) and were started on appropriate medications. After they attained remission (YMRS<12), they were interviewed on the Bipolar Prodrome Symptom Scale- Retrospective version (BPSS-R) and the General Behaviour Inventory (GBI). Twenty-five normal controls were assessed using the same tools. Results: Prodromal symptoms were reported by 58.8% of the patients, as compared to eight per cent of controls. Irritability (28.33%) was the most commonly reported prodromal symptom followed by anxiety (13.33%) and sleep disturbance (ten per cent). All the three frequently reported prodromal symptoms were of moderate severity and recurrent in frequency in three-fourth of the patients. The mean number of prodromal symptoms among the patients was 1.14 (SD 1.44), which was significantly higher than the normal controls (p<0.001). Hypomanic/ depressive temperaments were significantly higher in the patients with prodrome as compared to normal controls (p<0.01) as well as patients without prodrome (p<0.01). Discussion: Significantly higher number of prodromal symptoms was present in the patients and these symptoms were mostly moderate in severity and recurrent in nature. Also, the number of patients who reported prodromal symptoms was much higher than the number of controls. However, when a comparison was made between the reported prodromal symptoms in the patient and control group, a significant difference was noted only in ‘irritability’. Further, the proportion of prodromal symptoms reported in our study was lower when compared to previous studies. This could be because most of these studies had looked for these symptoms over the lifetime leading to likelihood of over-reporting. Individually, these symptoms may not have predictive significance but the occurrence of these clinical characteristics in patients who have hypomanic or depressive temperament could be a sign of impending first episode mania
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Deleterious effects of a cafeteria diet on the livers of non-obese rats
Rats were fed from weaning on three diets. Those fed a cafeteria diet had livers that were enlarged and abnormal by visual inspection. The rats themselves appeared healthy, had a normal growth rate and were not significantly different in weight from control animals. Histological examination revealed the livers of these rats to be rich in lipids and glycogen. Liver function tests showed a depressed level of alanine transaminase and an abnormal HDL: LDL. Dietary lipids generate free radicals which can interact with, and damage, DNA. However, when DNA was extracted from the livers and examined for the presence of the adduct M1-dG, there were no significant differences in adduct levels in livers from animals fed any of the diets. We conclude that the cafeteria diet can have long term adverse effects on liver function even though overt measures of health may be unimpaired, body mass maintained within normal limits, and liver DNA not adversely affected