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Human adipose cells in vitro are either refractory or responsive to insulin, reflecting host metabolic state.

Lizunov VA, Stenkula KG, Blank PS, Troy A, Lee JP, Skarulis MC, Cushman SW, Zimmerberg J - PLoS ONE (2015)

Bottom Line: Two statistically-defined populations best describe the observed cellular heterogeneity, representing the fractions of refractive and responsive adipose cells.Thus, a two-component model best describes the relationship between cellular refractory fraction and subject SI.Since isolated cells exhibit these different response characteristics in the presence of constant culture conditions and milieu, we suggest that a physiological switching mechanism at the adipose cellular level ultimately drives systemic SI.

View Article: PubMed Central - PubMed

Affiliation: Program in Physical Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States of America.

ABSTRACT
While intercellular communication processes are frequently characterized by switch-like transitions, the endocrine system, including the adipose tissue response to insulin, has been characterized by graded responses. Yet here individual cells from adipose tissue biopsies are best described by a switch-like transition between the basal and insulin-stimulated states for the trafficking of the glucose transporter GLUT4. Two statistically-defined populations best describe the observed cellular heterogeneity, representing the fractions of refractive and responsive adipose cells. Furthermore, subjects exhibiting high systemic insulin sensitivity indices (SI) have high fractions of responsive adipose cells in vitro, while subjects exhibiting decreasing SI have increasing fractions of refractory cells in vitro. Thus, a two-component model best describes the relationship between cellular refractory fraction and subject SI. Since isolated cells exhibit these different response characteristics in the presence of constant culture conditions and milieu, we suggest that a physiological switching mechanism at the adipose cellular level ultimately drives systemic SI.

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Dot plot of basal and insulin-treated cellular activities, GLUT4 storage vesicle (GSV) fusion and mobility rates, in adipose cells isolated from subjects with varying degrees of insulin sensitivity.For each subject (N = 19), eight cells were evaluated in the basal (black) and insulin-stimulated states (red). Subject ID, gender, age, BMI, SI and %Fat is shown below each group of cell measurements. Both GSV mobility and fusion rates were acquired from time-lapse TIRF recordings of isolated adipose cells transfected with GLUT4-mCherry and IRAP-pHlourin The upper panel (a) shows fusion rates measured in individual cells isolated from each subject. GSV fusion events were detected as spikes of IRAP-pHluorin fluorescence representing expansion of the fusion pore and pH equilibration within the lumen of the fusing GSV. Fusion rate for each cell was calculated as the number of IRAP-pHluorin spikes detected per unit area during one minute of recording. The lower panel (b) shows GSV mobility rates measured as the number of mobile GLUT4-mCherry vesicles detected in the TIRF-zone (within ∼200 μm of the cell membrane). GSV mobility rate was measured in isolated basal or insulin-treated cells and quantified as the number of mobile vesicles (with trajectory length >2 μm) detected within a 10x10 μm region of interest during one minute of recording (60 frames). The solid black lines represent average basal rates calculated from the pooled population of basal cells for each subject. The dotted lines represent 95% confidence intervals for pooled basal values. With increasing SI (from left to right), an increasing number of insulin-stimulated cells exhibit GSV mobility and fusion rates outside the 95% confidence interval calculated from the basal values.
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pone.0119291.g001: Dot plot of basal and insulin-treated cellular activities, GLUT4 storage vesicle (GSV) fusion and mobility rates, in adipose cells isolated from subjects with varying degrees of insulin sensitivity.For each subject (N = 19), eight cells were evaluated in the basal (black) and insulin-stimulated states (red). Subject ID, gender, age, BMI, SI and %Fat is shown below each group of cell measurements. Both GSV mobility and fusion rates were acquired from time-lapse TIRF recordings of isolated adipose cells transfected with GLUT4-mCherry and IRAP-pHlourin The upper panel (a) shows fusion rates measured in individual cells isolated from each subject. GSV fusion events were detected as spikes of IRAP-pHluorin fluorescence representing expansion of the fusion pore and pH equilibration within the lumen of the fusing GSV. Fusion rate for each cell was calculated as the number of IRAP-pHluorin spikes detected per unit area during one minute of recording. The lower panel (b) shows GSV mobility rates measured as the number of mobile GLUT4-mCherry vesicles detected in the TIRF-zone (within ∼200 μm of the cell membrane). GSV mobility rate was measured in isolated basal or insulin-treated cells and quantified as the number of mobile vesicles (with trajectory length >2 μm) detected within a 10x10 μm region of interest during one minute of recording (60 frames). The solid black lines represent average basal rates calculated from the pooled population of basal cells for each subject. The dotted lines represent 95% confidence intervals for pooled basal values. With increasing SI (from left to right), an increasing number of insulin-stimulated cells exhibit GSV mobility and fusion rates outside the 95% confidence interval calculated from the basal values.

Mentions: To analyze individual adipose cell responses to insulin, we have utilized statistical methods that avoid data averaging and allow us to identify underlying distributions of cellular responses per subject and among the pooled data from a group of 19 subjects with SI ranging from 0.16 to 11. The data presented in Fig. 1 show GSV mobility and fusion rates measured in individual adipose cells in the basal state (black) and in response to 0.1 IU/ml insulin (maximal stimulation, red), plotted for each subject, with SI increasing from left to right. In order to show all the individual cell responses without masking or overlapping of the individual data points, we also plotted these parameters for each subject in a separate graph (Figs. 2 and 3). Interestingly, the major difference between cells from insulin-resistant subjects (low SI<2) and insulin-sensitive subjects (SI>4) is not the individual cell response amplitude, but rather the number of cells that exhibit a 3–4 fold response. Simultaneously, in almost every subject, we observed cells that do not exhibit any insulin response that could be statistically distinguished from the typical basal range of values for mobility and fusion rates (Fig. 1, symbols between the solid black lines representing the average basal rate and the dotted lines representing the 95% confidence intervals). This observed heterogeneity in the insulin response of individual adipose cells strongly indicates that the underlying distribution is far from normal and thus that simple averaging of the cellular data is not appropriate.


Human adipose cells in vitro are either refractory or responsive to insulin, reflecting host metabolic state.

Lizunov VA, Stenkula KG, Blank PS, Troy A, Lee JP, Skarulis MC, Cushman SW, Zimmerberg J - PLoS ONE (2015)

Dot plot of basal and insulin-treated cellular activities, GLUT4 storage vesicle (GSV) fusion and mobility rates, in adipose cells isolated from subjects with varying degrees of insulin sensitivity.For each subject (N = 19), eight cells were evaluated in the basal (black) and insulin-stimulated states (red). Subject ID, gender, age, BMI, SI and %Fat is shown below each group of cell measurements. Both GSV mobility and fusion rates were acquired from time-lapse TIRF recordings of isolated adipose cells transfected with GLUT4-mCherry and IRAP-pHlourin The upper panel (a) shows fusion rates measured in individual cells isolated from each subject. GSV fusion events were detected as spikes of IRAP-pHluorin fluorescence representing expansion of the fusion pore and pH equilibration within the lumen of the fusing GSV. Fusion rate for each cell was calculated as the number of IRAP-pHluorin spikes detected per unit area during one minute of recording. The lower panel (b) shows GSV mobility rates measured as the number of mobile GLUT4-mCherry vesicles detected in the TIRF-zone (within ∼200 μm of the cell membrane). GSV mobility rate was measured in isolated basal or insulin-treated cells and quantified as the number of mobile vesicles (with trajectory length >2 μm) detected within a 10x10 μm region of interest during one minute of recording (60 frames). The solid black lines represent average basal rates calculated from the pooled population of basal cells for each subject. The dotted lines represent 95% confidence intervals for pooled basal values. With increasing SI (from left to right), an increasing number of insulin-stimulated cells exhibit GSV mobility and fusion rates outside the 95% confidence interval calculated from the basal values.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4359092&req=5

pone.0119291.g001: Dot plot of basal and insulin-treated cellular activities, GLUT4 storage vesicle (GSV) fusion and mobility rates, in adipose cells isolated from subjects with varying degrees of insulin sensitivity.For each subject (N = 19), eight cells were evaluated in the basal (black) and insulin-stimulated states (red). Subject ID, gender, age, BMI, SI and %Fat is shown below each group of cell measurements. Both GSV mobility and fusion rates were acquired from time-lapse TIRF recordings of isolated adipose cells transfected with GLUT4-mCherry and IRAP-pHlourin The upper panel (a) shows fusion rates measured in individual cells isolated from each subject. GSV fusion events were detected as spikes of IRAP-pHluorin fluorescence representing expansion of the fusion pore and pH equilibration within the lumen of the fusing GSV. Fusion rate for each cell was calculated as the number of IRAP-pHluorin spikes detected per unit area during one minute of recording. The lower panel (b) shows GSV mobility rates measured as the number of mobile GLUT4-mCherry vesicles detected in the TIRF-zone (within ∼200 μm of the cell membrane). GSV mobility rate was measured in isolated basal or insulin-treated cells and quantified as the number of mobile vesicles (with trajectory length >2 μm) detected within a 10x10 μm region of interest during one minute of recording (60 frames). The solid black lines represent average basal rates calculated from the pooled population of basal cells for each subject. The dotted lines represent 95% confidence intervals for pooled basal values. With increasing SI (from left to right), an increasing number of insulin-stimulated cells exhibit GSV mobility and fusion rates outside the 95% confidence interval calculated from the basal values.
Mentions: To analyze individual adipose cell responses to insulin, we have utilized statistical methods that avoid data averaging and allow us to identify underlying distributions of cellular responses per subject and among the pooled data from a group of 19 subjects with SI ranging from 0.16 to 11. The data presented in Fig. 1 show GSV mobility and fusion rates measured in individual adipose cells in the basal state (black) and in response to 0.1 IU/ml insulin (maximal stimulation, red), plotted for each subject, with SI increasing from left to right. In order to show all the individual cell responses without masking or overlapping of the individual data points, we also plotted these parameters for each subject in a separate graph (Figs. 2 and 3). Interestingly, the major difference between cells from insulin-resistant subjects (low SI<2) and insulin-sensitive subjects (SI>4) is not the individual cell response amplitude, but rather the number of cells that exhibit a 3–4 fold response. Simultaneously, in almost every subject, we observed cells that do not exhibit any insulin response that could be statistically distinguished from the typical basal range of values for mobility and fusion rates (Fig. 1, symbols between the solid black lines representing the average basal rate and the dotted lines representing the 95% confidence intervals). This observed heterogeneity in the insulin response of individual adipose cells strongly indicates that the underlying distribution is far from normal and thus that simple averaging of the cellular data is not appropriate.

Bottom Line: Two statistically-defined populations best describe the observed cellular heterogeneity, representing the fractions of refractive and responsive adipose cells.Thus, a two-component model best describes the relationship between cellular refractory fraction and subject SI.Since isolated cells exhibit these different response characteristics in the presence of constant culture conditions and milieu, we suggest that a physiological switching mechanism at the adipose cellular level ultimately drives systemic SI.

View Article: PubMed Central - PubMed

Affiliation: Program in Physical Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States of America.

ABSTRACT
While intercellular communication processes are frequently characterized by switch-like transitions, the endocrine system, including the adipose tissue response to insulin, has been characterized by graded responses. Yet here individual cells from adipose tissue biopsies are best described by a switch-like transition between the basal and insulin-stimulated states for the trafficking of the glucose transporter GLUT4. Two statistically-defined populations best describe the observed cellular heterogeneity, representing the fractions of refractive and responsive adipose cells. Furthermore, subjects exhibiting high systemic insulin sensitivity indices (SI) have high fractions of responsive adipose cells in vitro, while subjects exhibiting decreasing SI have increasing fractions of refractory cells in vitro. Thus, a two-component model best describes the relationship between cellular refractory fraction and subject SI. Since isolated cells exhibit these different response characteristics in the presence of constant culture conditions and milieu, we suggest that a physiological switching mechanism at the adipose cellular level ultimately drives systemic SI.

Show MeSH
Related in: MedlinePlus