Fat distribution is closely linked to metabolic disease risk. Distribution varies with sex, genetic background, disease state, certain drugs and hormones, development, and aging. Preadipocyte replication and differentiation, developmental gene expression, susceptibility to apoptosis and cellular senescence, vascularity, inflammatory cell infiltration, and adipokine secretion vary among depots, as do fatty-acid handling and mechanisms of enlargement with positive-energy and loss with negative-energy balance. How interdepot differences in these molecular, cellular, and pathophysiological properties are related is incompletely understood. Whether fat redistribution causes metabolic disease or whether it is a marker of underlying processes that are primarily responsible is an open question.

Anatomically separate fat depots differ in size, function, and contribution to pathological states, such as the metabolic syndrome. We isolated preadipocytes from different human fat depots to determine whether the basis for this variation is partly attributable to differences in inherent properties of fat cell progenitors. We found that genome-wide expression profiles of primary preadipocytes cultured in parallel from abdominal subcutaneous, mesenteric, and omental fat depots were distinct. Interestingly, visceral fat was not homogeneous. Preadipocytes from one of the two main visceral depots, mesenteric fat, had an expression profile closer to that of subcutaneous than omental preadipocytes, the other main visceral depot. Expression of genes that regulate early development, including homeotic genes, differed extensively among undifferentiated preadipocytes isolated from different fat depots. These profiles were confirmed by real-time PCR analysis of preadipocytes from additional lean and obese male and female subjects. We made preadipocyte strains from single abdominal subcutaneous and omental preadipocytes by expressing telomerase. Depot-specific developmental gene expression profiles persisted for 40 population doublings in these strains. Thus, human fat cell progenitors from different regions are effectively distinct, consistent with different fat depots being separate mini-organs.

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Fat distribution varies among individuals with similar body fat content. Innate differences in adipose cell characteristics may contribute because lipid accumulation and lipogenic enzyme activities vary among preadipocytes cultured from different fat depots. We determined expression of the adipogenic transcription factors peroxisome proliferator activated receptor-γ (PPAR-γ) and CCAAT/enhancer binding protein-α (C/EBP-α) and their targets in abdominal subcutaneous, mesenteric, and omental preadipocytes cultured in parallel from obese subjects. Subcutaneous preadipocytes, which had the highest lipid accumulation, glycerol-3-phosphate dehydrogenase (G3PD) activity, and adipocyte fatty acid binding protein (aP2) abundance, had highest PPAR-γ and C/EBP-α expression. Levels were intermediate in mesenteric and lowest in omental preadipocytes. Overexpression of C/EBP-α in transfected omental preadipocytes enhanced differentiation. The proportion of differentiated cells in colonies derived from single subcutaneous preadipocytes was higher than in mesenteric or omental clones. Only cells that acquired lipid inclusions exhibited C/EBP-α upregulation, irrespective of depot origin. Thus regional variation in adipogenesis depends on differences at the level of transcription factor expression and is a trait conferred on daughter cells.

. Abundance of two human preadipocyte subtypes with distinct capacities for replication, adipogenesis, and apoptosis varies among fat depots.

Fat mass, adipocyte size and metabolic responsiveness, and preadipocyte differentiation decrease between middle and old age. We show that expression of CCAAT/enhancer binding protein (C/EBP)-alpha, a key regulator of adipogenesis and fat cell function, declined substantially with aging in differentiating preadipocytes cultured under identical conditions from rats of various ages. Overexpression of C/EBP alpha in preadipocytes cultured from old rats restored capacity to differentiate into fat cells, indicating that downstream differentiation-dependent genes maintain responsiveness to regulators of adipogenesis. C/EBP alpha-expression also decreased with age in fat tissue from three different depots and in isolated fat cells. The overall level of C/EBP beta, which modulates C/EBP alpha-expression, did not change with age, but the truncated, dominant-negative C/EBP beta-liver inhibitory protein (LIP) isoform increased in cultured preadipocytes and isolated fat cells. Overexpression of C/EBP beta-LIP in preadipocytes from young rats impaired adipogenesis. C/EBP delta, which acts with full-length C/EBP beta to enhance adipogenesis, decreased with age. Thus processes intrinsic to adipose cells involving changes in C/EBP family members contribute to impaired adipogenesis and altered fat tissue function with aging. These effects are potentially reversible.

Purpose Psychological stress plays a role in the exacerbation of functional lower urinary tract disorders such as painful bladder syndrome and overactive bladder. To better understand the mechanism underlying this relationship, we characterized changes in micturition, anxiety-related behavior, and bladder pathology in rats exposed to repeated water avoidance (WA) stress. Methods Twenty-four Wistar rats were subjected to WA stress or sham. Immediately following acute (day 1) and chronic (day 10) stress or sham, rats were placed in a metabolic cage for a 2-hour voiding behavior assessment. Voiding parameters were compared to baseline values obtained prior to stress. Four animals from each group were sacrificed on day 10 and bladders harvested for histologic and gene expression studies. The remaining 8 animals per group underwent repeated voiding assessment every 3 days for 1 month followed by 10 days of repeat WA stress or sham. Bladder histology and gene expression were studied. Results Rats exposed to WA stress developed a significant increase in micturition frequency and decrease in latency to void, voiding interval and volume of first void compared to sham and baseline. Alterations in micturition persisted for approximately 1 month. Stressed rats showed increased fecal pellet excretion and anxiety-like behavior. Additionally, bladder specimens from stressed animals revealed increased angiogenesis, and increased total and activated mast cells. Conclusions In rats, repeated psychological stress results in lasting alterations in micturition frequency, interval, and volume. This rodent model may represent a valid tool for studying syndromes characterized by increased urinary frequency.

Background and Aims Neurotensin (NT) promotes colon cancer and inflammation via NT receptor-1 (NTR1). MicroRNAs regulate protein synthesis by targeting mRNAs. We determined the microRNA signature of NTR1 stimulation on human colonic (NCM460) epithelial cells. Methods RNA from NT-stimulated NCM460 cells overexpressing NTR1 was used for microarray expression analysis. NF-κB binding sites were identified by sequence homology, ChIP-assay and qPCR. Tumorigenesis was assessed by the soft agar assay and HCT-116 tumor xenografts in SCID mice. Down-stream targets of NT-regulated microRNAs were identified via bioinformatics, real time PCR and Western blot. Results NT stimulated differential expression of 38 microRNAs. We identified NF-κB binding sites on miR-21 and miR-155, previously implicated in tumor growth. NT increased the number of colonies of HCT-116 cells and antisense-microRNAs against miR-21 and/or miR-155 inhibited this response (p<0.001). NT administration (i.p.) increased the rate of tumor growth in xenograft tumors while miR-21 and/or miR-155 antisense attenuated this response. Since potential downstream targets of miR-21 and miR-155 are PTEN and SOCS1, respectively, and both are upstream of Akt, we investigated the effect of NT on Akt activation. NT activated Akt in HCT-116 cells, an effect inhibited by miR-21 and/or miR-155 antisense (p<0.001). We report for the first time PPP2CA phosphatase as a miR-155 target exerting the effects of NT on Akt. Conclusions NT stimulates miR-21 and miR-155 expression in colonocytes via Akt and NF-κB, and both microRNAs mediate colon tumor growth in response to NT. Importantly, this NT-microRNA circuit is found perturbated in human colon cancers and correlates with tumor stage, suggesting its relevance to human colon carcinogenesis.